Leica Application Suite

LAS User Manual

Living up to Life
LAS User Manual
All reasonable steps have been taken to ensure that this publication is correct and complete, but
should any user be in doubt about any detail, clarification may be sought from Leica Microsystems
(Switzerland) Ltd., or their accredited representative. The information in this document is subject to
change without notice and should not be construed as a commitment by Leica Microsystems
(Switzerland) Ltd.. Leica Microsystems (Switzerland) Ltd.accepts no responsibility for any errors
that may appear in this document.

© 2011 Leica Microsystems (Switzerland) Limited

No part of this publication may be reproduced, transmitted, transcribed, stored in any retrieval
system or translated into any human or computer language by any means or in any form, without
the prior written permission of Leica Microsystems (Switzerland) Ltd.

Due to a policy of continuous development, we reserve the right to change specifications without
notice.

Microsoft and MS-DOS are registered trademarks and Windows, the Windows logo, the Windows
Vista logo, the Windows 2000 logo and the Windows XP logo are trademarks of Microsoft
Corporation.

Date: April 2011

Software Version: LAS V3.8

Leica Microsystems (Switzerland) Ltd.

Stereo and Macroscope Systems
CH 9435 Heerbrugg
Switzerland

Telephone: +44 1223 411411

FAX +44 1223 210692
Hotline: +44 1223 401824
[email protected]
www.microscopy-imaging.com
Contents

Leica Application Suite Overview 1

The Core 2
Optional Modules 3

Getting Started 4
Documentation 5
Hardware Setup 8
Starting LAS 18
The User Interface 19
Shortcut Keys: General 22
Dual Monitors 29

The Core 36
Functions Widely Available 37
The Setup Workflow 104
The Acquire Workflow 156
The Browse Workflow 298
The Process Workflow 346
The Analysis Workflow 367

Optional Modules 368

Optional Modules: Demo Licence and Enabling 369
Archives: Basic and Standard 371
Extended Annotation 478
MultiTime 493
MultiFocus 520
Montage Multifocus 540
Leica Map 571
Image Overlay 582
MultiStep 603
Power Mosaic and Power Mosaic Plus 615
Live and Interactive Measurements 709
Image Analysis 766
Macro Runner 912
Phase Expert 913
Grain Expert 945
Reticule 973
Web Sharing 984
Multi-User Package 997
 Leica Application Suite Overview:

The Leica Application Suite (LAS) comprises: When the Leica Application Suite is first installed on the
computer, all of the Optional Modules are also installed but
The Core: The basic part of the Suite which includes not enabled. Selecting Demo mode will allow each one to
the Framework and the Setup, Acquire, Browse and be enabled ready for evaluation.
Process Workflows. These are the essential tools
needed to refine, capture and present images from Some of the modules may not be appropriate to the tasks
the microscope. The Core utilities are always available the microscope will be required to perform, so they need
and do not have to be licensed. not be enabled initially. Instead, they can be turned on at a
later date to start the 60 day evaluation. However, once
Optional Modules: Powerful, specialised programs evaluation is started the 60 days will start to run and
that perform specific functions to enhance, augment cannot be turned off, even if the module itself is disabled.
and extend the Core. Optional Modules are provided
free for an evaluation period of 60 days. After that, LAS installation is described separately in the Installation
they have to be purchased and licensed to continue to Guide.pdf on the LAS DVD.
work.

LAS User Manual 1

 The Core:

The Core provides the basic software for configuration Scale Bar displayed on the live image to indicate
and control of the selected microscope as well as for the image size.
acquisition, analysis and processing of high quality digital
images. The Core components include: Digital Image Acquisition into the familiar Image
Explorer tree and folder structure. There is an
Microscope and Digital Camera configuration and Optional Module to include database Archive.
control - all fully integrated.
Thumbnail Gallery of acquired images for quick and
Auto and Manual Exposure adjustments to allow easy review.
optimised imaging.
Text, Scale Bar and Distance Tools for
Image Calibration based on data read directly from straightforward image annotation.
Leica microscopes and cameras.
Link to the Core: 36

2 © 2011 Leica Microsystems (Switzerland) Limited

 Optional Modules:

The powerful features of the Core can be

enhanced with a range of advanced Optional
Modules, each providing the flexibility to tailor a
system to suit individual needs. The range of
Optional Modules includes:

LAS Archives: Basic and Standard.

Image Analysis
Extended Depth of Focus
Movie Recording
Image Measurements
Autofocus
Image Overlay
Power Mosaic...

..and more.

Link to Optional Modules: 368

Link to installing the 60-day Demo evaluation

licence and enabling the Optional Modules:
369

LAS User Manual 3

Getting Started

The following topics describe how to make effective use The User Interface Go there... 19
of LAS when you first start to use the software after Get familiar with the concept and terminology of the user
installation. The following topics are covered: interface

Documentation Go there... 5 Short-cut keys Go there... 22

Please read the Release Notes for details of new These help to improve your productivity
features and restrictions in use. Check you have
supported hardware and PC specification in the System Dual Monitors Go there... 29
Requirements If you have dual-monitors, this topic shows how to make
best use of them
Configure the hardware Go there... 8
When LAS is first used, you must specify the microscope
and camera hardware being used

Starting LAS Go there... 18

Once your system is configured, the usual starting
procedure follows these notes.

4 © 2011 Leica Microsystems (Switzerland) Limited

 Documentation

Documentation comprises LAS User Help and

Manual and related documents concerning the
installation, operation and restrictions on the
use of Leica Application Suite. Please consult
them before using the software.

Install Guide
A detailed description of the installation
procedure for Leica Application Suite is in the
LAS Install Guide.pdf.

Release Notes
Recent information about the version of LAS is
described in the document LAS Release
Notes.pdf. This describes features of the
software that have recently changed,
operational limitations and other technical
information.

System Requirements
The supported hardware - the microscopes,
macroscopes and cameras that can be used
with LAS - is described in the document
System Requirements.pdf. This document
also describes the appropriate computer
specification. Please ensure that your
computer specification corresponds to the
recommendations made. Other factors that
influence the performance of LAS are also The help documents are found on the LAS DVD or after installation as
noted in the same document. follows:

For Leica DM microscopes, refer to the 1: Click on Windows Start.

operator's manual supplied with the
microscope for detailed guidance on 2: Click on the All Programs arrow...
configuring and operating the microscope.
3: ...and on Leica Application Suite V3.

4: Click on the Release Notes option.

Continued... 6

LAS User Manual 5

 Getting Started: Documentation: Release Notes:

The Release Notes contain information

about changes and revisions that came too
late to be included in this documentation.
The Notes may be found on the original
installation DVD or on the computer hard
drive.

Release Notes on the DVD:

Load the DVD to the drive on the computer.
In most cases it should start up
automatically. If it does not:

1: Click on the My Computer icon on the

computer desktop.

2: From the dialog, right click on the DVD

drive. The disk name will reflect the
LAS Software Version and may not be
the same as the illustration.

3: From the popup menu select AutoPlay

and the DVD should start to run.

4: Click on the Release Notes button.

Continued... 7

6 © 2011 Leica Microsystems (Switzerland) Limited

 Getting Started: Documentation: Release Notes: Continued:

Leica Application Suite is supplied on a

single DVD.

1: Click on the Release Notes button.

2: The Release Notes require that Adobe

Acrobat reader is installed on the
machine. If it is not, the program is
also available on the DVD. Click on the
Install Adobe Acrobat button. After the
program has installed, return to the
DVD and once again click the Release
Notes button to display the notes (3).

4: If the program DVD is not available,

the same Release Notes file may be
accessed from the hard drive.
Launch Microsoft Word and open:

C:\Program Files\
Leica Microsystems\
Leica Application Suite\
Release Notes.pdf

LAS User Manual 7

 Hardware Setup:

The Hardware Setup facility allows

Administrators to create configurations for
either a single microscope and camera
combination or multiple combinations that are
sharing the same personal computer.

Although several microscope/camera

combinations can be plugged into the
computer, only one can be active and
connected to Leica Application Suite. Hardware
Configurations allow a specific combination to
be selected and connected.

As well as the microscope and camera

combination, the Configuration saves the last
used camera settings and the calibration values
so when the Configuration is recalled all of the
settings are automatically applied. An overwrite
is available that will apply the settings
established by the Administrator rather than
recall those last used by the current User.

Only computer Administrators can create

Hardware Setup Configurations, but both
Administrators and Users can select and use a
Configuration.

Continued... 9

8 © 2011 Leica Microsystems (Switzerland) Limited

 Hardware Setup: Reaching the Setup Dialog:

A: Administrators with User Rights turned

OFF:
1: Press and hold down the keyboard Shift
key.

2: Double-left click on the LAS desktop

icon. The Application Suite Framework
loads ready to open the Hardware
Setup dialog: Go there… 10

B: Administrators with User Rights turned

ON:
3: Right-click on the LAS desktop icon.

4: From the drop-down menu left-click on

Run as administrator. The Application
Suite Framework loads ready to open
the Hardware Setup dialog: Go there…
10

Selecting a Configuration to connect to

LAS:
C: Administrators and Users:
5: Press and hold down the Ctrl key.

6: Double-click the LAS desktop icon.

The Application Suite Framework loads
with the Configuration Selection dialog
displayed: Go there… 17

LAS User Manual 9

 Hardware Setup: Administrators: Starting Hardware Selection:

For Administrators:
Computers that have just one microscope
attached must also have the hardware set
up and saved as a configuration - usually, for
convenience the default Standard
Configuration name is used although any
appropriately named configuration can be
saved

1: Click on Options on the main header.

2: From the drop-down menu click to 4: Image Source provides for camera selection.
select Hardware Setup. The Setup
dialog appears. It is divided into three 5: Hardware Configuration provides the facility for storing the
sections: selected microscope and camera combination as a Configuration
file with an appropriate name. This can be retrieved by any
3: Microscope in which a microscope Administrator or User at a later date and the microscope/camera
stand can be selected. Additional combination ‘connected’ to LAS.
controls appear depending upon the
microscope chosen.
Continued… 17

10 © 2011 Leica Microsystems (Switzerland) Limited

 Hardware Setup: Administrators: Microscope Stand and Connection:

To select the microscope stand: Select the Connection type:

1: Click on the small arrows to the right of the 4: For microscopes that have a connection option, click
Microscope header and… on the small arrows to the right of the Connection
header and…
2: …from the drop down list click to select the stand
model. 5: …click to select the Connection type. Several
microscopes have a ‘virtual’ or Demo mode – a
3: If necessary, use the up/down arrows to scroll the microscope stand is not actually present but the
list. Leica Application Suite software can emulate limited
microscope behaviour. Demo modes are ideal for
training and evaluation.

Continued… 12

LAS User Manual 11

 Hardware Setup: Administrators: Stage Controller and Connection Test:

Stage Controller setup: Microscope Connection Test:

1: Click on the small arrows to the right of the Stage 3: To test the connections to the microscope and any
Controller header and… other associated hardware (excluding the Image
Source), click on the Test button.
2: …from the drop down list click to select the installed
controller. If the microscope does not have a 4: Click the OK button if the Connection Test is
motorised stage, click to select the None option. successful, otherwise check all connecting plugs and
power connectors and re-try.

Continued… 13

12 © 2011 Leica Microsystems (Switzerland) Limited

 Hardware Setup: Administrators: Log Communication File:

It is possible to keep a log of all the

communications between the microscope
and Leica Application Suite which could be of
use to a service engineer. Enable
communications traffic by:

1: Click on the small arrow to the right of

the Microscope menu header.

2: On the Microscope Advanced Settings

dialog, click to enable the traffic logging
check box. A small tick mark should
appear.

3: Click on the View Log Files button to

display the log data (usually in Notepad:
4).

5: If required, clear the log data by clicking

the Clear all log files button.

6: Click the Close button.

Continued… 15

LAS User Manual 13

 Hardware Setup: Administrators: Image Source Selection:

Choose a camera by:

1: Click on the small arrows to the right of the Image 3: Click on the Test button (does not apply to Demo
Source drop down header. Camera) to check that the device selected is
communicating with the software.
2: Click to select the image capture device from the
drop down menu. Leica Demo Camera uses an 4: If the device is found it will be confirmed on the Test
image installed with the Application Suite and is ideal Camera Connection dialog. Click OK.
for evaluating some of the optional modules without a
camera or frame grabber attached to the
microscope. Continued… 16

14 © 2011 Leica Microsystems (Switzerland) Limited

 Hardware Setup: Administrators: Create a New Configuration:

If the default Standard Configuration is to be used (usually

for a single microscope/camera attached to the computer),
skip to Saving the Configuration: Go there... 16 3: Click OK.

Creating a New Configuration: 4: The new configuration name appears in the

1: Click on the Create Configuration button. Hardware Configuration window.

2: On the dialog, click inside the text box and type a Continued… 11
new, unique name for the configuration.

LAS User Manual 15

 Hardware Setup: Administrators: Saving the Configuration:

With the microscope and image source selected: The process of creating and saving a configuration does
1: Click on the Apply button. not automatically apply it. Configurations have to be
selected and loaded specifically. See the following page.
2: The Overwrite dialog appears warning that the
current hardware setup will overwrite any existing
values in the Configuration file. This safety warning Continued… 17
appears even if the named Configuration is newly
created and does not yet contain data. Click Yes to
save the Hardware Configuration.

16 © 2011 Leica Microsystems (Switzerland) Limited

 Hardware Setup: Administrators: Select a Hardware Configuration:

A: Administrators already working in

Hardware Setup and the Framework is open:
1: Click on Options on the main header.

2: Click on the Select Hardware

Configuration option.
Go to item (7) below.

B: Administrators and Users from the

Desktop starting with the Shift + Icon double-
click:
3: Click on Options on the main header.

4: The drop-down menu has fewer options

compared to the Administrator only
menu above. Click on the Select
Hardware Configuration option.

5: The Administrators dialog has an

addition button – Hardware Setup (11)
that will launch the Hardware Setup
sequence described on previous pages.

6: The Users dialog.

9: Click OK.
To select a Hardware Configuration:
7: Click on the small arrows to the right of 10: To automatically load the prevailing camera settings – exposure,
the configurations header and… live and capture types etc., - click to enable the Overwrite Current
Settings check box. If left unchecked the camera settings stored
8: …click to select the required with the Configuration will be loaded.
configuration.
12: The selected Hardware Configuration is displayed bottom left of
the LAS interface.

LAS User Manual 17

Getting Started: Starting LAS:

Launching Leica Application Suite:

In most cases the Application Suite can be
launched by double-clicking on the
desktop icon:

To start in Administrator mode, right-click

on the icon, and select 'Run as
Administrator'.

Alternatively:

1: Click the Start button on the Windows

Task Bar (bottom left).

2: From the popup click on All

Programs.

3: Locate and click Leica Application

Suite on the All Programs list and...

4: Click on Leica Application Suite from

the dropdown. LAS should load and
run.

18 © 2011 Leica Microsystems (Switzerland) Limited

 The User Interface:

The illustration is a graphical representation of the LAS display showing the principal features:

1: Main Tool Bar: Come here for File, Options and Help menus.
2: Workflows: Select Setup, Acquire, Browse, Process and Analysis here.
3: Module Launcher: Click to reveal the installed modules and launch them if they are enabled.
4: Control Panels: All of the power tools for the selected Workflow.
5: Tabbed Control Panels: Click tabs to select additional tools for the Workflow and running module.
6: Image Data Form: Displays and edits selected data for the current image.
7: The Image Viewer: Display and working area for the current image: Press keyboard F5 to show full screen.
8: Side Tool Bar: Common working tools control all aspects of the display and tasks.
9: The Grid: Displays the current folder image data in a scrollable grid format. Available only with LAS Archives.
10: The Gallery: Displays a thumbnail of all the images in the selected folder.
11: Fast Search Controls: Create filters and fast search for images. (Archive Option dependant).
12: Gallery Browser: Locate rapidly thumbnails and display in the Viewer.
13: Gallery Thumbnail Scaler: Click and drag to re-size the thumbnails.
14: Acquire: Universal capture button.
15: Status Bar: Displays Hardware Configuration, RGB Intensity, Stage Position and Magnification data.

LAS User Manual 19

The User Interface: Main Areas:

The Leica Application Suite User Interface

is divided into 5 main areas.

1: Module Launcher: Click here to open

the Launcher and click to select the
required module.

2: Menu bar: Select items here to

access administrative options and
Preferences

3: Workflow bar: The Workflow creates

the appropriate panels and controls
for the selected application. Click on
a Workflow to open it.

4: Control Panels: The programme

controls are displayed on tabbed
panels. Click on the tab to display the
panel.

5: Image Viewer and Gallery: The

remaining part of the screen
application is devoted to the Image
Viewer , Grid and Gallery. The
thumbnail images in the Gallery can
be re-sized by clicking on the slider
(6) and dragging it up to increase the
size or down to decrease it.
.

20 © 2011 Leica Microsystems (Switzerland) Limited

 The User Interface: The Side Toolbar:

The Side Tool Bar is situated on the right-

hand edge of the Viewer and provides the
essential working tools for many of the
tasks concerned with managing images -
Export, Delete and Print - and customising
the environment - Hide and Reveal display
features, Fit to Screen - and so on.

If optional modules are installed, where

appropriate the Side Tool Bar changes to
reflect the additional functions that the
modules provide (*). In the illustration the
additional tools are loaded with module
Image Analysis.

LAS User Manual 21

Shortcut Keys: General Shortcuts:

Leica Application Suite has a wide range of Keyboard Keys and Mouse combinations to
simplify actions for the user and so speed throughput and improve productivity. Some
shortcuts are applicable only to particular optional modules and if those modules are not
installed the shortcuts will not function.

Shift + Left click LAS desktop icon: Start LAS framework only so hardware configuration and
firmware download, licensing can be used. Note: Requires administrator privileges.

Control + Left click LAS desktop icon: Show Hardware Configuration dialog.

Show help files for the module currently active.

Acquire single image / current mode image sequence.

Show image full screen size - live and browse image.

Return from image full screen to normal LAS layout.

With Dual Monitor mode enabled - switch to second monitor to show image.

With Dual Monitor mode enabled - revert to single monitor to show image.

Left click and drag on live image: Used to set Region of Interest for spot exposure, zoom
focus, etc.

Show camera refresh rate at top right hand of image.

Show actual image size.

Show Preferences dialog.

Mouse wheel on image: Image zoom in and out.

Slider control is selected: Mouse wheel adjusts slider is single steps - for example in
Exposure.

22 © 2011 Leica Microsystems (Switzerland) Limited

 Shortcut Keys: General Shortcuts: Continued:

If Measurements is licensed: On/off - show a zoom region around the mouse position. For
example, when setting calibration end points or drawing in Live Measurements.

When the Gallery or Grid have focus: Selects all images in Gallery.

When the Gallery or Grid have focus: Copy the selected image and its metadata to the
clipboard

Paste the selected image and its metadata to the folders indicated in the navigator. Note the
folder must be a folder that the user can access and also not a sequence folder.

Control plus left click on an image in the Gallery or Grid entry: Add this image to the
selected images.

Shift plus left click on an image in Gallery or Grid entry: Select all the images between a
previously selected image and the clicked image.

Right click on an image in Gallery or Grid entry: Show pop-up context menu with Open,
Open with, Send to and Export options.

Right click on an image in Browse: Show pop-up context menu with Open, Open with, Send
to, Export, Print, Copy, Zoom and Pan window options.

Select all data in the Record Details table.

Select and individual record in the Record Details table.

Select all the records between a previously selected record in the Record Details table and
the clicked one.

Copy all of the records in the Record Details table to the clipboard. They can then be pasted
into another application, Word for example.

LAS User Manual 23

Shortcut Keys: Live and Interactive Measurements Shortcuts:

Alt Gr held down + draw measurement: Hide the labels so that it is easier to see where the
measurement is drawn. Also Ctrl + Alt for keyboards that do not have the AltGr key.

Shift held down: Show a zoom region around the mouse position.

Space bar held down: Temporarily switch to Edit mode after drawing.

With the Select tool ‘on’, click and drag rectangle around measurements: Selected
measurements in the rectangle can be grouped. Or edited collectively.

Double click on Parameter name in Grid: Label all measurements with this Parameter if
possible. Note: Some measurements don’t have all parameters.

Click on Grid header near small arrow: Re-orders the results in increasing or decreasing
values.

Selects all of the measurements in the Grid.

Selects an individual measurement in the Grid.

Selects all of the measurements between and including the currently selected measurement
in the Grid and this one.

Copies the selected measurements in the Grid so they can be pasted into another
application such as Word or Notepad.

24 © 2011 Leica Microsystems (Switzerland) Limited

 Shortcut Keys: Annotation Shortcuts:

Start a new line in a Text Box or Rectangle.

If Measurements is licensed, toggles On/Off a zoom region around the mouse position.

With an annotation selected, copy the annotation to the clipboard.

Paste the contents of the clipboard into a Word document.

Select all of the annotations in the Grid.

Select an individual annotation in the Grid.

Selects all of the annotations between and including the currently selected annotation in the
Grid and this one.

Copy the selected annotations to the clipboard so that they can be pasted into another
application such as Word or Notepad.

LAS User Manual 25

Shortcut Keys: Image Analysis Shortcuts:

Double click on Parameter name in Grid: Label features with this parameter where possible.

Click on Grid header near small arrow: Re-orders the results in increasing or decreasing
values

Shift held down: Show a zoom region around the mouse pointer whilst in the Binary Edit
step.

Click on a feature in the Measurement step: Select the feature and move it to the top of the
Grid.

Click on a row on the Grid: Selects and highlights the feature on the image.

Whilst in Measurement > Edit, deletes the select feature.

Selects all measurements.

Selects an individual measurement.

Selects a range of measurements between and including the currently selected

measurement and this one.

Copies the selected measurement(s) to the clipboard so that they can be pasted into
another application such as Word or Notepad.

26 © 2011 Leica Microsystems (Switzerland) Limited

 Shortcut Keys: Montage 3D Viewer Shortcuts:

Drag the model to a different viewing angle: Rotate and tilt.

Drag the model to a new location: Pan.

Zoom the model.

Mouse wheel click and drag: Select a region of the model to zoom.

Scale the model height.

LAS User Manual 27

Shortcut Keys: Power Mosaic Shortcuts:

Mouse Interactive (on side tool bar) enabled: Double click Mouse Wheel: Return directly
from a zoom to show mosaic in 'Fit to Window' mode.

Move Pattern (on side tool bar) enabled: Hold down Control: Click and drag Mouse on the
mosaic Pattern to rotate it.

Move Pattern (on side tool bar) enabled: Hold down Shift: Halt mosaic Pattern rotation.

Mouse Interactive (on side tool bar) enabled: Right Mouse click on Map to display Map
Properties dialog.

Power Mosaic Plus Shortcuts:

Mouse Interactive (on side tool bar) enabled: Right click on Workspace to display
Workspace properties.

Move Pattern (on side tool bar) and Pattern Navigator enabled: Click and drag Workspace
or Pattern.

28 © 2011 Leica Microsystems (Switzerland) Limited

 Dual Monitors:
With the necessary Dual Screen Video card and software The Side Tool Bar is automatically displayed on the
installed (*), Leica Application Suite can be configured appropriate monitor and Dual Monitor Mode can be
quickly and easily to show all of the usual controls - enabled or disabled with a single keystroke.
including optional modules - on one monitor (Primary)
whilst using the entire viewable area of the other ( (*) Use of dual monitors requires that the graphics card in
Secondary) for the image - live or captured. the PC supports this option. Some graphics cards lack
performance that slows down the movement of the mouse
The extra-large image area provides greater precision cursor. If this occurs, the drivers of the graphics card may
and ease of working for capture, analysis and need updating or it may not have the necessary
measurement, whilst the greater area given to Gallery performance.
Thumbnails means many more can be displayed at once
or individuals enlarged to examine fine detail.

LAS User Manual 29

Dual Monitors: Swap Monitors:

The conventional layout for dual monitors

is primary monitor (displaying Windows
controls) in front of the user, and the
secondary monitor to the right. This is
initially setup in the video driver software,
but can be changed in Windows so that
the secondary monitor is moved to the left
without affecting the smooth transition of
the cursor from one screen to the other.

LAS users may find swapping the

monitors a convenient way of
reducing the distance the cursor has
to be moved to place it on the image.

1: Right-click on the Desktop.

2: On the menu, for Windows XP users,

click on Properties. For Vista users
click on Personalise.

3: On the dialog, Windows XP users

click on the Settings tab and Vista
users the Display Settings option.

4: The illustration shows the Vista dialog

but the Windows XP version is very
similar.

5: The two monitors are shown as icons

– 1 being the primary display and 2
the secondary.

6: Click and drag the secondary display

to the left of the primary display and
release the mouse button.

7: Click OK.

Continued... 31

30 © 2011 Leica Microsystems (Switzerland) Limited

 Dual Monitors: Enabling and Disabling:
The secondary monitor is turned on or off
either from the Options menu or by pressing
keyboard function key F5.

1: Click on Options on the main tool bar.

2: On the drop down menu, click to

enable the check box to the left of the
Use Second Monitor option. Clicking
the checkbox again will disable the
second monitor and move the Image
Viewer back to the primary monitor –
the usual LAS layout.

3: Function key F5 performs the same

operation with fewer mouse clicks.
Press F5 to enable the second monitor
and press again to disable it.

Continued… 32

LAS User Manual 31

 Dual Monitors: Primary Monitor:
With the Viewer moved to the Secondary Monitor, all of All of the tools (except image zoom and sizing) are
the Primary Viewer area can be occupied by the Gallery available on the Side Bar to show and hide the Record
Thumbnails which means they can be larger and clearer. Form, Record Information, Data Grid and Data Items to
Display.
The smooth Thumbnail Sizing slider has been moved to
the top edge - click and drag to the left for smaller, more- Export, Print, Reporting (with some images) and Delete
on-view Thumbnails, or to the right for larger, fewer are also present. See Side Bar Tools: Go there... 34
images.

32 © 2011 Leica Microsystems (Switzerland) Limited

 Dual Monitors: Secondary Monitor:
The Secondary Monitor is dedicated totally to the Viewer 2: A Zoom Level readout on the Viewer top edge.
so that images can fill the entire screen. Taking
measurements or analysing the image is easy and precise 3: The monitor can also display the Floating Navigator
with such a large working area. which allows users to move between folders without
having to return to the Browse Workflow.
1: The Image Control Tools - detailed on the following
page - are automatically moved to the Side Tool Bar.

LAS User Manual 33

 Dual Monitors: Side Bar Tools:
With the second monitor enabled and displaying the image,
the right-hand Tool Bar is divided appropriately between the
two displays.

On the primary monitor displaying the LAS controls:

1: Export image.

2: Print image.

3: Prepare Report depending upon the image type.

4: Delete image (Trash Can).

5: Show/hide the Record form.

6: Show/Hide the image, disabled in Dual Monitor mode.

7: Show/hide the Grid.

8: Show/hide the Gallery Thumbnails, disabled in Dual

Monitor mode.

9: Show Record Details.

10: Show Select Visible Fields dialog.

On the secondary monitor displaying the Image Viewer:

A: Show the Annotations Options.

B: Open/Close the Floating Navigator (Browse). E: Image to Fill Viewer.

C: Pan the Image. The Pan Window appears on the F: Display Image at Actual Size.
primary monitor.
G: Viewer Options (Dual Viewer etc).
D: Zoom Into/Out of image.
H: Save the Output Image.

Continued… 35

34 © 2011 Leica Microsystems (Switzerland) Limited

 Dual Monitors: Image Options:

A wide range of options is available by

right-clicking the image in the Viewer or its
Thumbnail in the Gallery. The options
vary depending upon which item was
clicked, the operating system and the
software installed on the computer.

1: Right-click the Thumbnail for the

context menu of basic options and...

2: ...click it to select.

3: Some options have additional

possibilities displayed as a sub-menu.

4: Right-clicking the image in the Viewer

displays a different context menu.

5: Additional functions are available

some of which will also have sub-
menus (6).

For detailed help on the available options,

see the Browse Workflow: Go there... 298

LAS User Manual 35

 The Core Features:

This chapter describes the Workflow organization of the Each is selected by clicking on the Workflow bar.
unique Leica Application Suite user interface and its basic
capabilities. Selecting a Workflow displays the appropriate controls
arranged on one or more panels that allow the user to
LAS Workflow describes the order and grouping of tasks perform the selected action.
for image documentation and analysis. While the Workflow
suggests an order for the tasks, versatility is retained so Because the Workflow arrangement is so versatile, in
that the operation of the software is not constrained to contrast to many Windows programs, LAS does not
fixed steps. Grouping tasks into related operations makes employ a menu bar for the main operation of the software.
working with LAS intuitive and easy.
Installed and enabled Optional Modules are listed on a
Click on ( ) to link to a Workflow: menu revealed by clicking the Select Acquisition Mode
icon to the left of the Workflows.
Setup 104 .

Acquire 156

Browse 298

Process 346 and optionally...

Analysis 367

36 © 2011 Leica Microsystems (Switzerland) Limited

 Functions Widely Available:

There are functions within Leica Application Suite that

apply to several Workflows or have a more global
application. These are available from a drop-down menu
that appears when Options on the main menu bar is
clicked.

Clicking the Acquire option captures an image and is the

equivalent to pressing the function key F3 on the
keyboard or clicking on the Acquire button on the Acquire
or Browse Workflows.

Use Second Monitor, or function key F5 displays the

image on the second monitor (if fitted). Click again to
return to single screen. Works on all Workflows. If only a
single monitor is in use, pressing F5 displays the image
full screen with a 'Return to normal LAS layout' button

Additionally the Scale Bar can be accessed from nearly all

steps and the Export Images feature is also widely
available so both are included in this section:

Setting Preferences: 38 .
Scale Bar: 61

Update Calibration: 69

Image Comparison: 79

Export Images: 84

Printing: 91

Gallery Docking: 103

LAS User Manual 37

 Preferences:

The Preferences dialog allows the user to select preferred When LAS is launched by an administrator, the tabs will
options for Leica Application Suite and most of the vary to include options that only administrators are
Optional Modules to reflect the fastest and most permitted to set:
convenient working methods.
1: Administrators launch LAS by right-clicking the
As Optional Modules are added to Leica Application Suite, desktop icon and...
tabs on the Preferences dialog are also added to include
settings that are specific to the modules. 2: ...left-clicking the Run as administrator option on the
context menu.
Preferences may be altered at any time – even while a
module is running. 3: When LAS starts and the Preferences dialog is
opened, the Admin tab is present. This is not
available to other users.

38 © 2011 Leica Microsystems (Switzerland) Limited

 Launching Preferences:

Preferences may be altered at any time – even while a

module is running.

1: On the Main Toolbar, click on the Options label.

2: From the drop down options, click to select

Preferences.

3: The Preferences dialog appears. Click on a tab along

the top of the dialog to reveal the options and settings
required.

4: When changes have been made, click on the OK Quick links to the Preferences tabs:
button to save them or…
Defaults: What happens after image capture.
Admin: Settings that only administrators can change.
Image: Formats, captions and other image settings.
5: …click on the Cancel button to keep existing settings. Warnings: Turns individual warnings on or off.
Movie Settings: Controls the size of movies.
Store & Recall: Save and retrieve image data.
Status Bar: Display helpful information and calibrate
monitors.

Click a tab on the illustration for more information.

LAS User Manual 39

 Preferences: The Defaults Tab:

The Defaults tab has three panels that specify the actions Default Action: What should happen when an image is
that will occur in various situations. The Default actions are captured.
user defined:
Image Table Field for Description: Specify the data
1: Click on the Defaults tab to reveal the panels. field to display as the annotation on the image.

Default Action When Image Rejected: The capture

location is not currently active so specify the options.

Click on a panel on the illustration for more information:

40 © 2011 Leica Microsystems (Switzerland) Limited

 The Defaults Tab: Default Action:

1: Click on the Defaults tab to reveal the

panels.

2: Default Action that determines the action

when the shortcut key F3 is pressed with
an archive selected.

3: Click on the arrows to the right of the

Default Action drop-down header.

4: Click to select the required option that will

automatically occur when key F3 is
pressed.

LAS User Manual 41

 The Defaults Tab: Field for Image Description & Image Rejected:

1: Click on the Defaults tab to reveal the

panels.

2: An archive field from the Image Table can

be used as part of the image description.

3: Click on the small arrows to the right of

the header and...

4: ...from the drop-down menu click to select

the field to use with the image. Use the
scroll slider to reveal the entire list.

Image Rejected:

5: If an image cannot be saved in the current

folder - usually because it has not been
nominated as the fixed capture location -
chose a default action as follows:

6: Clicking to enable the Display dialog

check box (Tick mark visible) displays a
Capture Options dialog to the user. It is
recommended that this check box is
enabled.

7: Choose to either Show or Not Show the

image by clicking the appropriate radio
button.

42 © 2011 Leica Microsystems (Switzerland) Limited

 Preferences: The Admin Tab:

Administrators can restrict access to panels on the 1: Click on the Admin tab to reveal the Setup Tab
Setup Workflow to prevent unauthorised changes Availability and Administrator panels.
being made to essential settings by standard users.
2: When changes have been made click the OK button.
Once a microscope has been calibrated,
administrators can prevent further changes by Click on a panel for more information.
standard users can be prevented by locking the
calibrations.

Users can be allowed to save images to a network

drive as well as the local computer.

LAS User Manual 43

 The Admin Tab: Hide Setup Tab:

Administrators can restrict access to panels

on the Setup Workflow to prevent
unauthorised changes being made to essential
settings by Standard Users.

Each of the available tabs on Setup can be

hidden by checking the check box and so
prevent display and access.

1: Click on the Admin tab on the

Preferences dialog.

2: On the Setup (Workflow) Tab Availability

dialog...

3: ...click the required check box to hide

(checked) or reveal (un-checked) a tab. In
the example only the Archive tab will be
displayed in the Setup Workflow;
Microscope, Reticule and MUP will be
hidden and their panels inaccessible.

44 © 2011 Leica Microsystems (Switzerland) Limited

 The Admin Tab: Lock calibrations& Capture to Network:

1: Click on the Admin tab to reveal the

Administrator panel.

2: Once a microscope has been calibrated,

administrators can prevent further
changes by standard users can be
prevented by checking (tick mark visible)
the Lock calibrations check box.

3: Allow standard users to save images to a

networked drive or folder as well as a
local computer by clicking to enable the
Capture to network drive/folder check
box.

Points to consider before enabling Capture

to network...

Is the network accessible.

Is it fast enough to capture extensive
sequences like time-lapse.
Movies cannot be saved using the
network.
A slow network will slow down LAS
startup.

LAS User Manual 45

 Preferences: The Image Tab:

The controls on the Image tab determine the details of Measurement Displays sets the number of decimal
the image capture and what should happen after capture. places for some measurement labels.

Administrators can also set up the Camera panels both to Camera panel configuration. Determines those controls
avoid modification and avoid clutter. that are displayed or hidden on the Acquire > Camera
panel.
Click on the Image tab (1) to reveal the dialog. The tab
has 5 individual panels: Click on a panel on the illustration for more information.

Save Images which determines the names and Click OK to save the settings and close the Preferences
format of saved images. dialog.

Application to launch After Image Capture.

The application to launch for the Movie File player.

46 © 2011 Leica Microsystems (Switzerland) Limited

 Image Tab: Save Images:

1: Click to check and get a prompt for an

image name before saving a captured
image.

2: Check the Fixed Capture Location

checkbox to have images saved in the
folder selected as ‘fixed’ on the Browse
Workflow.

3: With the Always Create Thumbnail File

checkbox enabled, when the image is
captured a separate thumbnail file with the
extension .thb will also be created. Whilst
this does occupy additional disk space,
thumbnails are loaded to the Gallery far
quicker. Applicable to all folders.

4: The Default Image Name is a prefix for

each image saved. Click and swipe on the
text box and type a new name appropriate
to the work in hand.

5: Leading Zeroes automatically pads the

Image Name sequence with zeroes so that
all image names are the same length. Use
the up/down arrows to the right of the
Leading Zeroes to set the value.

6: To select the Image Format and

compression, click on the arrows to the
right of the In this Format header and...

7: ...from the drop down menu click to select

the format required.

8: Image resolution measured in Dots Per

Inch (DPI), can be set using the up/down
arrows to the right of the DPI box. This
function is particularly useful to ensure that
3rd party applications such as Word or
PowerPoint®, display images correctly.

The DPI setting does not affect the

capture format or the capture resolution
set up in Acquire > Camera.

LAS User Manual 47

 Image Tab: Configure Camera Panels:

The Acquire > Camera panel has a wealth of

tools and controls but for some users who do
not require all of the available facilities, it can
become 'cluttered' especially if Optional
Modules are added. The number of panels on
display can be reduced by setting them as
hidden on the Configure camera Panels
dialog. The controls are not lost only hidden.

Any user can hide or reveal a panel but only

an Administrator or member of the LAS
Administrator Group can lock or unlock it. If a
panel is locked it cannot be hidden or revealed
- its state when the lock was applied remains
in force until the lock is released.

1: Click on the Camera > Configure Panels

button.

2: Control panels on the Camera tab can be

Hidden - the selected panel will not
appear - and...

3: ...also Locked which locks the selected

panel in position on the panel sequence.
Click on a check box to the right of the
selected panel to enable hiding and
locking.

4: Enabling the Lock Settings Window will

prevent the camera setting dialog from
being displayed or altered.

5: Click OK to save the settings and close

the dialog.

48 © 2011 Leica Microsystems (Switzerland) Limited

 Image Tab: After Capture:

After an image is captured there are three

possible automatic options that can aid speed
and efficiency especially if the image required
post-capture work:

1: Do Nothing. The image and data will be

saved and the current Workflow will
remain unaltered. Click on the radio
button if a number of images need to be
captured in quick succession without any
intervening editing process.

2: Open In will automatically divert to the

selected Workflow:

3: Click on the arrows to the right of the

window and from the drop down click to
select the required Workflow.

Continued: 50

LAS User Manual 49

Image Tab: After Capture: Continued:

1: Open Image Using allows a (usually)

third-party application to be launched to
display and/or edit the image. Click on the
radio button and then...

2: ... on the Browse button to reveal…

3: …the Windows Navigator.

4: Navigate to and select the application

required and click Open. The selected
application will appear in the Open Image
Using text box.

50 © 2011 Leica Microsystems (Switzerland) Limited

 Image Tab: Measurement Display & Play Movie Using:

1: Measurement Display sets the number of decimal Play Movie Using...

places that will be displayed on some of the Workflows
and Optional Modules: 3: After a movie has been created, it may be played
using a nominated application.

4: Click on the Browse button to the right of the Play

Movie Files text box and...

5: ... from the Windows Navigator navigate to...

6: .. and select the application required. Click on the

Open button and the application name will appear in
2: Click on the Up/Down arrows to the right of the the Play Movie Files Using text box.
Measurements Display widow to increase/decrease the
number of decimal places.

LAS User Manual 51

 Preferences: The Warnings Tab:

The major confirmation and warning messages can be

turned on or off on this tab. 3: The Set All button will check all of the messages and...

1: Click on the Warnings tab to reveal the options panel. 4: ...the Set None button will clear them all.

2: Click to check (message on) or un-check (message off 5: Click OK.

) the check box to the left of each warning.

52 © 2011 Leica Microsystems (Switzerland) Limited

 Preferences: The Movie Settings Tab:

Because movies can be disk-space 'hungry’, the Movie

Settings tab provides two ways of limiting movie size:

Maximum Movie Size limits file size in terms of free

disk space whereas:

Limit Movie Size prevents files exceeding a physical

size measured in Megabytes (MBytes).

If Limit Movie Size is enabled, both features will run

together to control movie size.

1: Click on the Movie Settings tab to reveal the Disk

Usage panel.

Limit the size of movies as disk space:

Limit movie files to a specific size:
2: Click on the up/down arrows to the right of the
Maximum Movie Size window to increase/decrease the 3: Click on the Limit Movie Size checkbox to enable size
percentage of disk space that can be allocated to limiting. The checkbox will become red with a white
movies. tick.
Note that at least 1 Gigabyte (GByte) of free disk
space is required simply to run the movie application. 4: Click on the up/down arrows to the right of the Limit
Movie Size window to increase/decrease the maximum
file size. Each click is a 1MByte step.

LAS User Manual 53

 Preferences: The Store & Recall Tab:

The Store and Recall feature allows the microscope and Of the five functions available:
camera setting to be stored with an image so that precisely
the same conditions may be repeated at a later date. It can Store Single Images,
also provide consistency across a range of different
Store Sequences,
specimens.
Recall and
Fully automated microscopes can automatically adjust to
Workflow After Recall...
the settings; The settings display can be used to adjust
manual models.
...each have options selected by clicking a button. The
Click on a panel on the illustration for more information. buttons are mutually exclusive - only one may be active
within a function.

Undo Recall....

...is switched on or off by clicking a checkbox. This is a

toggle action.

54 © 2011 Leica Microsystems (Switzerland) Limited

 Store & Recall: Store Single Images & Sequences:

1: To reveal the Store and Recall options, click on the

tab.

2: There is a settings panel for storing Single Images

and...

3: ...another for storing Sequences.

Both panels have the same options:

Always will always store the settings.

Ask will prompt you to store or not when the image is

saved.

Never switches off settings store. The store facility

adds a very short delay to storing images and the
files are longer.

Click a button to select the option.

LAS User Manual 55

Store & Recall: Recall, Image Mode & Undo Recall:

1: Recall determines if stored settings are also recalled

when an image is retrieved from disk.

Always will always recall any settings stored with the

image. Automated microscopes will adopt the
settings.
Ask prompts if the settings should be retrieved or not.

Never switches off settings recall.

2: When a stored image is selected two options are

available on the Workflow After Recall panel:

Acquire will switch to the Acquire Workflow and

display the live image from the microscope, whilst...

Browse will remain in the Browse Workflow and

display the selected image in the Viewer.

3: If Undo Recall > Enable Undo After Recall is

checked, the current microscope and camera
settings are saved before any recalled settings are
applied to the microscope and camera. Reverting to
the current settings is then possible.

56 © 2011 Leica Microsystems (Switzerland) Limited

 Preferences: Status Bar Data Display:

The Status Bar is located along the bottom edge of the The Status Bar panel also allows the user to set up the
Viewer and displays data relating to: resolution both for the PC monitor and a second monitor
allowing the software to calculate the magnification.
The current Hardware Configuration.
To reveal the Status Bar panels:
The intensity of the image RGB (Red, Green, Blue)
components beneath the mouse. 1: Click on the Status Bar tab.
Stage Position is the X/Y co-ordinates when the
2: Save settings by clicking the OK button.
image was captured.
Viewer Magnification. Click on a panel on the illustration for more information.

LAS User Manual 57

Status Bar: Data Display Selection:

1: The Show Status Bar Data panel determines the items

to be displayed along the Status Bar.

2 to 5: Enable a data item by clicking the associated

check box. A tick mark indicates the item display is
enabled. Click again to turn off the display.

2: Hardware Configuration shows the currently selected

and active configuration.

3: RGB Intensity displays the Red(R), Green(G) and Blue

(B) values of the image pixel directly below the mouse.
The mouse position is indicated by the X/Y co-
ordinates. Greyscale images are represented as a
single value - Intensity.

4: Stage Position is the X, Y and Z stage co-ordinates

when the selected image was captured or current 5: Magnification represents the monitor display
values for the live image. The values are stored with magnification - not the microscope magnification. As
the image when it is captured. the user clicks the Zoom buttons on the side tool bar
the magnification value changes. The value will only
display if the monitor has been set up: 59

58 © 2011 Leica Microsystems (Switzerland) Limited

 Status Bar: Monitor Setup:

The microscope magnification can be read

from the microscope or from Leica Application
Suite, but the magnification on the computer
monitor(s) is different because LAS scales the
image to the user’s demands and the monitor’s
capabilities.

LAS can indicate the image magnification by

calculation from the live or acquired image pixel
size and that of the monitor. The image
calibration is established as described in the
section Camera > Calibration.

1: Click on the Status Bar tab.

2: If necessary, change the measurement

type – millimetres, centimetres or inches –
by clicking on the arrows to the right of the
Screen Width header and...

3: ...click to select the measurement type

required

Continued: 60

LAS User Manual 59

Status Bar: Monitor Setup: Measuring the Monitor(s):

1: Measure the monitor(s) horizontally

across the entire viewable area of the
screen (not just the displayed image) with
a ruler using the measurement type
selected.

2: Click in the Monitor 1 text box and type

the monitor width.

3: Repeat the process if a second monitor is

fitted, in the Monitor 2 text box. Click OK.

The magnification factor is shown bottom

right on the screen. It will change as the
microscope objective zoom or the image
The magnification value is not displayed if a monitor size has not
zoom - either on the Side Tool Bar or
been entered or if the Viewer is in Dual View mode.
mouse wheel - is changed. It will also
update if the Viewer available display area
is resized by opening, closing or re-sizing
the Grid or Gallery.

60 © 2011 Leica Microsystems (Switzerland) Limited

 Scale Bar: Introduction:

The Scale Bar is available on Acquire, Browse,

Process and Analysis Workflows.

1: Clicking the Show Annotations button on

the Side Tool Bar...

2: ...displays the Annotations and Scale Bar

Quick Launch menu.

3: Click to select the Scale Bar option.

4: The Scale Bar dialog can be dragged by

the header and ‘parked’ on any part of the
Viewer.

5: Close the dialog and return it to the

Workflow by clicking the ‘X’ on the dialog
caption.

6: If the dialog is obscuring the image or

controls, collapse it by clicking on the small
arrow to the right of the dialog caption.
Expand it by clicking the arrow again.

7: The Scale Bar is revealed or hidden by

clicking the Show check box...

8: ... and can be Merged so that it is a

permanent part of a captured image.

Scale Bar Features: Click the link ( ) for more

information:

Modes Selection: 62 . The Scale Bar dialog will be active only when the Show check
Style Selection: 63 ! box is enabled.

End Bars, Thickness, Digits: 64

Placement and Font Change: 65

Scale Bar and Font Colour: 66

Background Colour: 67

Merging: 68

Continued 62

LAS User Manual 61

 Scale Bar: Mode Selection:

Three Scale Bar Modes are available to

determine how it behaves on the image.

Fixed Pixel Size: Acts as a known distance

‘ruler’. As the image is zoomed the Scale
Bar length remains constant whilst the
distance value changes to reflect the zoom
level.

User Length: In contrast to the Fixed Pixel,

the numeric value of the User Length
option remains unchanged as the zoom is
changed, but the line length on the display
is adjusted. Values typed in to the Size text
box will change to ‘Undersize’ if too small or
revert to the overall image width/height if
too large.

Adaptive: Displays a ‘true’ calibrated Scale

Bar at about 20% of the image width or
height depending upon the bar orientation
and as the zoom is changed, the numeric
distance and the pixel size of the scale bar
both change to maintain a 'reasonable' size
on the image.

1: Click on the small arrow to the right of the

Mode header and…

2: …click to select the required option.

Change the measurement units for Fixed Pixel

Size and User Length modes:

3: On the Camera tab...

4: ... in Calibration Settings.

The Adaptive mode ‘inherits’ the last

measurement units used.

5: To enter a distance in either Fixed Pixel

Size or User Length modes, click in the
Size text box and type a value.

Continued 63

62 © 2011 Leica Microsystems (Switzerland) Limited

 Scale Bar: Style Selection:

The orientation and shape of the Scale Bar is

set on the Style menu.

1: Click on the small arrows to the right of the

Style header and…

2: …click to choose the required style from

the drop down list.

3: Conventional Horizontal style.

4: Vertical style.

5: Combined Horizontal and Vertical styles (

Both). Although only one dimension is
shown both ‘legs’ represent the same
distance.

Continued 64

LAS User Manual 63

 Scale Bar: End Bar, Thickness and Significant Digits:

There are four options for the type of Scale Bar

ends:

1: Click on the small arrows to the right of the

End Bar header and…

2: …from the drop down list click to select the

type required:

3: None,
4: Small,
5: Medium or
6: Large.

Scale Bar Thickness:

7: Click on the Up/Down arrows to the right of

the Thickness text box to increase/
decrease the Bar thickness. The End Bars
are not affected.

Significant Digits:

8: Increase or decrease the number of digits

displayed by clicking the Up/Down arrows
to the right of the Sig Fig text box. The
value determines the total number of digits
displayed not just those after the decimal
point. So, in Figure (6) the Sig Fig value
would be 5. Whole numbers before the
decimal point are not affected.

Continued 65

64 © 2011 Leica Microsystems (Switzerland) Limited

 Scale Bar: Placement and Font Change:

The Scale Bar can be placed quickly and

precisely at the four corners of the image by
using the Placement buttons.

1: The Placement buttons will position the

Scale Bar at top-left, top-right, bottom-left
or bottom-right of the image, live or
captured, accurately and precisely simply
by clicking the appropriate button. The
Scale Bar can still be clicked and dragged if
needed.

Changing the Font:

2: Click on the Font button.

3: On the Font dialog select the Font, Font

Style and Size as required and…

4: …click OK.

Continued 66

LAS User Manual 65

 Scale Bar: Scale Bar and Font Colour:

Change the Scale Bar and Font colour by:

1: Click on the Colour button and…

2: …on the Select Colour dialog choose a

new colour from the swatches or from the
colour wheel. Alternatively, click in the
Current Colours text boxes and type the
Red, Green and Blue values.

3: Adjust the colour intensity by clicking and

dragging the slider on the Intensity bar.

4: Click OK.

5: The new colour is shown on the Colour

button.

Continued 67

66 © 2011 Leica Microsystems (Switzerland) Limited

 Scale Bar: Background Colour and Transparency:

The Scale Bar is displayed against a small

panel the colour and transparency of which may
be changed to suit the image:

1: On the Scale Bar dialog click on the

Background button. The Select Colour
dialog appears with additional controls to
change the transparency of the
background.

2: Choose a new background colour by

clicking a Basic Colour Swatch or…

3: …clicking and dragging the ‘target’ on the

Colour Wheel.

4: Adjust the colour intensity by clicking and

dragging the slider on the Colour Bar or
typing new values for Red, Green and Blue
or Hue, Saturation and Intensity in the
appropriate text boxes.

Background Transparency:

5: The Scale Bar background transparency

can be altered by clicking and dragging the
slider or typing a value in the Transparency
text box. A value of 255 results in a solid
colour (6) and a value of 0 makes the
background panel disappear. The
illustration (7) shows the result of a value of
138.

8: Click OK. The selected colour appears in

the Background check box (9).

Continued 68

LAS User Manual 67

 Scale Bar: Merging:

Merging is the process of combining the Scale Bar and its 3: Two options are available: Replace the captured
caption with the saved image. image including the Scale Bar with it, or …

Once merged, the image will always appear with the Scale 4: …Duplicate which makes a copy of the existing
Bar that cannot be altered. captured image and merges the Scale Bar with the
copy.
1: For live images Merge is a check box – click to enable
merging when the image is captured. Click the required option button.

2: For previously captured images Merge is a

conventional LAS button – click to merge the Scale
Bar with the image. The Merge Annotations dialog
appears.

68 © 2011 Leica Microsystems (Switzerland) Limited

 Update Calibration:

If the system calibration has changed or images

are being used that do not reflect the current
calibration values, Update Calibration provides
a simple and quick way to bring images up-to-
date.

Four options for the calibration source are

available:

Use the current image: 71 71 Uses the

calibration values of the selected image..

Use system calibration: 72 72 Uses the

prevailing calibration settings.

Manual from the measurement line: 73

73 Allows new calibration values to set up
directly from a known distance on the
displayed image.

Automatic from Calibration Slide: 74 74

The calibration is calculated automatically
from an image of a calibration slide
captured at the same time as those to be
updated.

Update Calibration is used on captured images

including those that have a Scale Bar ‘burnt
in’ (merged) but in those cases the merged
displayed value will not change.

1: Click to select the Browse Workflow.

2: Click on Options on the main tool bar and

from the drop-down menu,...

3: ... click to select Update Calibration. The

Update Calibration dialog (4) appears.

LAS User Manual 69

 Update Calibration: Selecting Images:

There are several ways to select images that will updated: 2: Press and hold down the Shift key.

A: Range of images: 3: Click on the image from which the calibration will be
1: Click on the first image to be selected. copied. All of the images between the two will also be
selected.

B: Individual Images: 3: Click individually on all the other images to be included

1: Click on the first image to be selected. in the selection. The last image will be used as the
calibration source.
2: Press and hold down the Ctrl key.

C: All of the images in the Gallery: 3: To select the calibration source, still holding down the
1: Click on the first image. Ctrl key double-click the source image.

2: Press and hold down the Ctrl key and then press and
release the 'A' key (All).

70 © 2011 Leica Microsystems (Switzerland) Limited

 Update Calibration: Use Current Image Calibration Values:

To use an image as the calibration source for all

of the other selected images, choose the
images to be updated making sure that the
calibration source is the last to be selected.
More information 70

1: Click the Current image calibration button.

2: If necessary, change the measurement

units by clicking the small arrows to the
right of the units header and from the drop-
down list, clicking to select the units.

In the Pending Calibration panel the 1

! Pixel = nnn value refers to the calibration
of the selected source image. The
reference to Pixel represents the unit used
to store the image not the Camera or the
Monitor pixels.

3: To update the measurement units for the

selected image but not to change their
calibration values, click to enable (tick mark
visible) the Update units only check box.

4: If the selected images are part of a

sequence the Update all images in
sequence check box becomes available.
Click to enable it and update all of the
sequence images.

5: Click the Apply button.

Click the Close button to exit Update
Calibration.

LAS User Manual 71

 Update Calibration: Using the System Calibration:

Apply the current System Calibration settings to

an image or range of images by selecting the
images to be updated. More information 70

1: Click to select Current system calibration.

2: If necessary, change the measurement

units by clicking the small arrows to the
right of the units header and from the drop-
down list, clicking to select the units.

In the Pending Calibration panel the 1

! Pixel = nnn value refers to the system
calibration. The reference to Pixel
represents the unit used to store the
image not the Camera or the Monitor
pixels.

3: To update the measurement units for the

selected image but not to change their
calibration values, click to enable (tick mark
visible) the Update units only check box.

4: Update the image microscope properties to

the current system settings by clicking to
enable the check box.

5: If the selected images are part of a

sequence the Update all images in
sequence check box becomes available.
Click to enable it and update all of the
sequence images.

6: Click the Apply button.

Click the Close button to exit Update
Calibration

72 © 2011 Leica Microsystems (Switzerland) Limited

 Update Calibration: Using a Measurement Line:

A new calibration value can be set by using a

line extended across a feature with a known
dimension on the image. Best accuracy is
achieved by using a Calibration Slide.

Select the images to be updated. More

information 70

1: Place a specimen with a known and precise

dimension on the stage. A clean Calibration
Slide is the best option. Focus the
specimen or slide.

2: If necessary, change the measurement

units by clicking the small arrows to the
right of the units header and from the drop-
down list, clicking to select the units.

In the Pending Calibration panel the 1

! Pixel = nnn value refers to the system
calibration. The reference to Pixel
represents the unit used to store the
image not the Camera or the Monitor
pixels.

3: Click to select the Manual from

measurement line option.

4: The measurement line appears. Click on

the centre of the line and drag it so that the
left end stroke aligns with the edge of the
specimen or slide.

Hold down the Shift key to display the

magnifier, moving it with the mouse to
assist alignment.

5: Aim to place the end stroke on the outside

of the left edge.
Click and drag the right 'handle' so that the
right stroke aligns with the inside of the right
edge.

6: Click inside the Length text box and type

the known length of the line in the chosen
measurement units.

7: The calibration value is automatically

calculated.

8: Enable the Update units only check box to

apply changes in the measurement units
only.

Click Apply to update the selected images.

LAS User Manual 73

 Update Calibration: Automatic from Calibration Slide:

Automatic calibration offers a fast method of

applying precise calibration to selected images.
It uses an image of a calibration slide that was
captured using the same microscope settings
as the images to be updated.

The software can accurately and automatically

detect the slide image and calculate the
calibration from a known interval between the
divisions, providing the image is sharp and the
division lines clearly defined.

Select the images to be updated making sure

that the calibration slide image is the last
selected and is displayed in the Viewer. More
information 70

1: Click to select Automatic from calibration

slide.

2 & 3: If necessary, change the measurement

units for both the calibration slide and the
calibration value by clicking the arrows to
the right of the headers and selecting from
the drop-down list.

The measurement units do not have to

match.

The calibration slide image does not have

to be perfectly horizontal or vertical, but the
closer it is then the faster the detection.

4: Click inside the Slide Interval text box and

type the interval value of the calibration
slide - that is the distance between two
adjacent divisions in the selected
measurement units.

Continued 75

74 © 2011 Leica Microsystems (Switzerland) Limited

 Update Calibration: Automatic Calibration: Continued:

The software will now try to find a calibration

slide and verify that it is precise and suitable as
a calibration source:

1: Click on the Get Calibration button.

2: If a calibration scale is found and verified,

the colour of the division strokes on the
scale will change, a new calibration value is
automatically calculated and...

3: ...the Apply button becomes active. Click it

to apply the new calibration value to the
selected images.

4: If a scale is not detected or does not

conform to the verification parameters, an
error message appears. The message
changes to reflect the error.

Calibration slide scale verification

parameters can be changed: 76 .

The calibration scale detected colour can

be changed:

Continued 76

LAS User Manual 75

 Update Calibration: Automatic Calibration: Continued:

The verification parameters check that a

calibration slide image is sufficiently accurate to
be used as a calibration source. For example,
random fibres on an calibration slide image
could be interpreted as part of the scale and
have to be 'filtered' out.

Users can change the settings but should be

aware that significant changes can result in
compromised calibration accuracy. If in doubt
revert to the factory optimised defaults.

1: Click on the 'spanner' icon to reveal the

Auto Calibration Settings dialog.

2: The Patch size refers to the spread of

pixels leading to a discernible edge. In the
illustration there are several pixels ranging
from white to dark grey before the black
central 'edge' appears. Increasing the patch
size could 'create' spurious edges. Keep
the patch size as small as possible.

3: Angle tolerance determines the how much

the angle between two adjacent scale
strokes can vary from the parallel. The
software is looking for a series of parallel
strokes at a consistent 'pitch' or interval.

4: The interval of the scale strokes must be

close to the value entered by the user. The
Error threshold determines how much it
can be allowed to vary.

5: Outliers are scratches and debris that may

be present on the image and could be
interpreted as part of the scale. The Outlier
acceptance sets the % level at which the
interval mean (a central 'average' for all of
the detected intervals) can vary. Strokes
falling below the Outlier acceptance are
removed.

6: If changes are made to the settings click

the Apply button to save them.
Continued 77
7: To restore the factory settings click the
Defaults button.

76 © 2011 Leica Microsystems (Switzerland) Limited

 Update Calibration: Automatic Calibration: Continued:

Change the colour of a detected calibration

slide image by:

1: Click on the Colour button on the Slide

Interval panel.

2: From the Windows Colour dialog choose a

colour by clicking a swatch, dragging the
crosshairs on the wheel or typing Red,
Green and Blue (RGB) values.

3: Click OK.

LAS User Manual 77

 Update Calibration: Display Preferences:

The number of decimal places displayed in

Interactive Measurements and Image Analysis,
can be set by:

1: Click on the Display Preferences tab.

2: Using the Up/Down arrows, increase or

decrease the number of digits to be
displayed after the decimal place.

3: Click Apply.

Return to the calibration options by clicking the

Calibration tab.

78 © 2011 Leica Microsystems (Switzerland) Limited

 Image Comparison - Dual Viewer:

The Dual Viewer option is available in the Acquire, 3: In Acquire the image currently being viewed will
Browse and Process Workflows. The Viewer area can be usually appear in the left-hand pane.
split to show two images simultaneously:
4: ..To display a captured image in either pane, click
In Acquire the current live image usually appears in anywhere in the pane and then...
the left pane) with a previously captured image
selected from the Gallery in the right pan). 5: ...click a thumbnail in the Gallery.

In Browse and Process two previously captured

images selected from the Gallery appear left and Dual Viewer features and quick links ( ):
right.
Unlock and Lock views: 80 .
1: On the Side Tool Bar click the Viewer Options button. Pan Window: 81

2: Click to enable (tick mark visible to the left) the Dual Comparing Features: 83

Viewer option. The Viewer will then divide into two

panes.

LAS User Manual 79

 Dual Viewer: Unlock and Lock Views:

Lock View Disabled: Lock View enabled:

With the Lock View option disabled – no tick mark against Follow steps (1) and (2) above but click to enable the Lock
it, both images can be scaled independently: View – tick mark visible.

1: Click on the View Options button on the Side Tool Bar

and…

2: …from the context menu click to disable Lock View –

no tick mark.

Both images automatically scale to the smaller view.No

need to click on an image because both are now
synchronised – all of the Side Tool Bar options affect both
images simultaneously.

Continued 81

3/4: Click to select the image in either pane.

5: Use the Side Tool Bar buttons to Zoom In or Zoom

Out,

6: …Fit to Screen or…

7: …display at Original Size.

80 © 2011 Leica Microsystems (Switzerland) Limited

 Dual Viewer: Pan Window:

The Pan tool allows detailed areas of an image

that exceeds the visible area of the Viewer to be
examined. It will not work if Fit to Viewer is
enabled because all of the image is being
displayed.

Dual Viewer will work in either locked or

unlocked modes when Pan is being used.

Pan with Lock View disabled: Only one of the

images is panned

1: Click the View Options button on the Side

Tool Bar.

2: From the context menu click to disable

Lock View – no tick mark visible. The
images are now independent.

3: Click on the image to pan and, using the

Zoom In button enlarge it.

4: Click on the Pan Window button and the

window appears with...

5: ... a red-outlined Pan Area. Click inside the

Pan Area and drag it to the required
position. The selected image tracks the
movement
.
6: The Pan Window can be moved to any
convenient position in the Viewer by
clicking and dragging its header bar.
Either image can be displayed in the Pan
Window simply by clicking on it.

Continued: 82

LAS User Manual 81

 Dual Viewer: Pan Window Continued:

Pan with Lock View enabled:

1: Follow the sequence on the previous page

but click to select Lock View - tick mark
visible. Both images are automatically
scaled to the same size and both pan in
unison.

Pan with Lock Pan enabled:

2: Follow the steps on the previous page and

click to enable Lock Pan - tick mark visible.
The images can be displayed at different
sizes but will pan in unison as the Pan Area
is dragged.

82 © 2011 Leica Microsystems (Switzerland) Limited

 Dual Viewer: Comparing Images:

To compare both live and captured images:

4: Click on the View Options button again (1) and this
1: Click on the View Options button on the Side Tool Bar time click to disable the Lock View.
and…
5 & 6: Use the Scroll Bars to independently move the
2: …from the menu click to enable the Lock View option images so that the features are close enough to be
– tick mark visible. compared.

3: Enlarge the images (now scaling in unison) to the

desired size. In the illustration the two bolt heads are
being compared and so the images were enlarged to
view them easily.

LAS User Manual 83

 Export Images:

The Export function copies a selected image or

multiple images to a location of the users
choice.

Options allow complete meta data files or

individual fields of data to be attached to, and
exported with the image(s). Meta data contains
the settings for the Scale Bar and annotations
so is useful only for other LAS installations.

Additionally, the image type - jpg, png etc - can

be changed during the export process. Image
names can be changed as well or image
sequences exported with the same name but
with an automatically applied incremental suffix.

1: On the Export Images dialog, there is a...

2: ...Don't ask again checkbox which, when

enabled (a tick mark visible) will skip the
dialog options. This is very useful if a large
number of images are to be exported and
continually completing the dialog becomes
a chore.

Enabling the checkbox clears the Show

Export Options in Preferences, so to
display the Export dialog again the option
has to be re-enabled in Preferences 85 .

84 © 2011 Leica Microsystems (Switzerland) Limited

 Export Images: Enable Show Export Options:

Enabling the 'Don't ask again' checkbox clears the Show 3: On the Preferences panel click the Warnings tab
Export Options in Preferences, so to display the Export and...
dialog again:
4: ...click to enable the Show Export Options check box.
1: Click on Options on the Main Menu.
5: Click OK and the next time Export is invoked the
2: Click to select Preferences. dialog will re-appear.

LAS User Manual 85

 Export Images: Selecting Images:

There are several ways to select images that will exported: 2: Press and hold down the Shift key.

A: Range of images: 3: Click on the image from which the calibration will be
1: Click on the first image to be selected. copied. All of the images between the two will also be
selected.

B: Individual Images: 3: Click individually on all the other images to be included

1: Click on the first image to be selected. in the selection. The last image will be used as the
calibration source.
2: Press and hold down the Ctrl key.

C: All of the images in the Gallery: 3: To select the calibration source, still holding down the
1: Click on the first image. Ctrl key double-click the source image.

2: Press and hold down the Ctrl key and then press and
release the 'A' key (All).

86 © 2011 Leica Microsystems (Switzerland) Limited

 Export Images: Select the Destination Folder:

1: If necessary, click to select the File option

on the Image Destination panel.

2: To change the destination folder, click on

the Browse for Folder button and...

3: ...navigate to the destination folder. Create

a new folder if required (4).

5: Click the OK button.

6: To include all of the Meta Data with the

image, click to enable the Include all meta
files check box.

Continued 88

LAS User Manual 87

 Export Images: Rename File and Add Fields:

The images can be renamed before they are stored in the 4: The new name is displayed in the lower text box and
destination folder: the Image Data panel opens on the right.

1: Click to enable the Rename File check box. 5: To include a field and its data, click to select the Field
Name.
2: Click in the text box and type a new name for the
image(s). 6: The Field Name is added to the image name in the
upper text box contained in tag marks <>, and
3: To include selected Data Fields with the image, click relevant data to the lower text box within brackets [].
on the right-facing arrow and on the Image Data
prompt. Continued 89

88 © 2011 Leica Microsystems (Switzerland) Limited

 Export: Change Image Type:

The image type can be changed before it is

saved:

1: Click to enable the Change image type

check box.

2: Select the new type by clicking on the

small arrows to the right of the Image
Type header and from the drop down
menu...

3: ...click to select the Image Type required.

4: When the jpeg option is selected the

quality menu is enabled. Click on the
small arrows to the right of the header
and...

5: ...click to select the required quality or

compression.

6: Click the OK button.

Continued 90

LAS User Manual 89

 Export Image: Options Dialog:

If an image with the same name already exists

in the destination folder, a warning will appear.
There a three options:

1: Overwrite the existing with the new

export. The existing image is lost.

2: Abort the export - Don't copy.

3: The Copy but keep both files option

exports the image but adds a numeric
suffix to the new copy to distinguish it.
This is the option to use if multiple images
are being exported and a common name
has been chosen for them.

4: Click to enable the Do this... check box to

automatically employ the chosen option in
the future.

90 © 2011 Leica Microsystems (Switzerland) Limited

 Printing:

The Printing facility allows the user to print individual Include multiple Header and Footer comments: 96 .
images quickly and simply. The built-in formatting provides
flexible styling resulting in professional-looking documents. Print landscape for maximum image size or portrait for
extra comment space: 95
Printing's most powerful feature is direct access to the
image data, including all of the microscope settings, that Automatically insert image information - no need to
can be included with just a mouse click to display type it in: 98
alongside the users own text.
Choose the Font, Style and Size to personalise the
Printing features and quick links ( ): report: 96

Print LAS images in high resolution colour.. Comments can be re-used with other images - no
need to re-type.
Include Annotations and Scale Bar on the image:
101 See a full-screen preview before printing: 101

Include Measurement Drawings on the image.

LAS User Manual 91

 Printing: Select the Printer:

To print a single image together with a wide range of data: 3: Click on the arrow to the right of the Name text box
and from the drop down list click to select and
1: Click on the Print icon. The Print Properties dialog alternative printer.
appears.
4: Change the print properties by clicking on the
2: The currently selected printer (if there is more than one Properties button.
connected to the machine or network) is displayed in
the Printer window. To change the printer, click on the 5: Click OK to apply the new printer and its properties.
Select button and the Windows Printer dialog appears.

92 © 2011 Leica Microsystems (Switzerland) Limited

 Printing: Page Margins:

The print page is structured to provide the user with a 1, 2, 3 & 4: Top, bottom, left and right margins can all be
quick and simple way of adding text and data to an image. set independently using the Page Setup dialog (5).
The layout has been designed to provide maximum
flexibility and a professional presentation. The dialog also allows the page Size and Orientation
to be set.
There are 4 margins that can be setup by the user to
accommodate different printers and bindings:

LAS User Manual 93

 Printing: Text & Image Areas:

The printable area inside the margins is divided into 3 The columns are further divided:
columns each of which can contain justified text:
A: Text appearing above the image called Header Text,
1: The left-hand column contains left-aligned text. and...

2: The centre column displays centred text, and... B: ...text below the image referred to as Footer Text.

3: ...the right-hand column right-aligned text. The fonts are selected and the text entered using two
dialogs - Header Text and Footer Text setups (4).

94 © 2011 Leica Microsystems (Switzerland) Limited

 Printing: Page Setup:

To change the page size, margins and the orientation: 4: Click to select the page Orientation - portrait or
landscape.
1: From the Print Properties dialog click on the Page
Setup button. 5: Set the margins by clicking inside the appropriate text
box and typing a value.
2: On the Page Setup dialog, if required change the
page Size by clicking the arrow to the right of the Size 6: Click OK.
header and selecting from the drop-down menu.

3: If the printer software allows, the printer Paper Source

can be set by clicking the arrow to the right of the
header and selecting from the menu.

LAS User Manual 95

 Printing: Fonts & Sizes:

Different Fonts, Style and Type Size can be used for 3: On the Header or Footer Text dialog, click the Select
Header and Footer text but cannot be mixed across the button to the right of the Font text box.
columns.
4: From the Windows Font dialog, select the Font, Style
To select a Font, Style and Size: and Size from the lists.

1 & 2: On the Print Properties dialog click the 5: Click OK.

appropriate Edit button - either Header or Footer text.

96 © 2011 Leica Microsystems (Switzerland) Limited

 Printing: Centred Text:

In this example the document title, the originator and the 4: The document title is typed...
department are to be displayed in the centre column above
the image. It will look like this:

5: ...followed by the key combination Ctrl + Enter: Press

and hold down the Ctrl key and then press the Enter
key. This will insert a new line break.

1: On the Print Properties dialog click the Page Header

check box to display a tick mark. This will ensure the
header text prints.

2: Click the Page Header Edit button.

3: On the Header Text Setup dialog click inside the 6: The originator text is added, again followed by the new
Centre Justified Text box. line break combination Ctrl + Enter.

7: Finally, the department text is added in the same way.

LAS User Manual 97

 Printing: Left-Aligned Text with Data:

LAS stores a wealth of data about an image. Individual

data items can be selected from a list and added
automatically to the page.

The date of the image capture and the image name are to
be added to the left and right Header Text respectively (1)
. The captions will be typed into the 'Justified' text boxes
but the actual values are imported from the image data.

To add the image capture date:

2: On the Header Text Setup dialog click inside the Left

Justified Text box. In this example the text 'Image
Date: ' is typed. Do not add a new line break.

3: The image Capture Date is going to be added to this

text. Click on the arrow to the right of the Left
Justified Text box.

4: Click to select Print from the data menu, and...

The Image Name is displayed in the same way but using
5: ...from the drop-down list, click to select the data item the Right Justified Text box instead and selecting File
required - in this case Capture Date. Data items are Name from the list. It will appear as:
represented in the text box enclosed in tags:
Image Name: <File Name>
Capture Date: <Capture Date>

98 © 2011 Leica Microsystems (Switzerland) Limited

 Printing: Left-Aligned Text: Continued:

The illustration shows the Header Text Setup dialog as it When the page is printed the data inside the tags (<>) is
would appear from the example on the previous page. retrieved and formatted. When this image was captured it
was given the name CircuitBoard.jpg.

LAS User Manual 99

 Printing: Footers & Image Data:

Footers - text appearing below the image - is entered in 3: To add an Image Data item, click on the arrow to the
the same way as described for the Headers with left, right of the text box and...
centre and right text boxes.
4: ...select Image Data from the options.
Image data - information about the hardware, exposure
and illumination - is also included using the same method 5: Click to select the required item from the drop-down
as for print data. list. In this example the text would appear as:

In the example the final piece of information is the Microscope: <Image Data.Microscope Stand Name>
microscope name which will be centred below the image.
On the Print Properties dialog click the Footer Edit button Any number of data items can be added to a printout
and... providing space allows.

1: ...the Footer Text Setup dialog appears.

2: For centred text click inside the Centre Justified Text

box and type - in this example the word 'Microscope: '.

100 © 2011 Leica Microsystems (Switzerland) Limited

 Printing: Print Preview and Annotations:

1: To preview how the page will look when printed, on 4: Close the Preview by clicking the Close button.
the Print Properties dialog click the Preview button.
5: If a Scale Bar and Annotations have been added to
2: The Print Preview window appears with a scaled down the image but not merged, they can be included on
representation of the page with the Header, Footer the printout by clicking to enable the Include Scale
and image in relative positions. Bar... check box.

3: Use the Preview Zoom to enlarge or reduce the 6: Click the Print button on the Print Properties dialog to
preview or display it full-screen. print the page(s).

LAS User Manual 101

 Printing: The Result:

This is the printed result of the example described in the

text.

102 © 2011 Leica Microsystems (Switzerland) Limited

 Gallery Docking:

Available in Acquire, Browse and the Process Workflows, 2: ...from the drop-down menu, left-click to select the
the thumbnail Gallery can be 'docked' either horizontally - Toggle Gallery Position option.
along the bottom edge of the Viewer - or vertically - along
the left-hand edge of the Viewer - to suit the user. The action toggles between horizontal and vertical
docking.
1: Right-click on a thumbnail or on the spaces around
the thumbnails and... Scroll bars, if required, are placed automatically.

LAS User Manual 103

The Setup Workflow:

The Setup Workflow provides the method

of specifying the Leica microscope
components connected to Leica
Application Suite and, when the optional
LAS Archive modules are used, creating
archives to capture images and data.

Components such as objective types and

filter descriptions can be readily selected,
saved and subsequently recalled.

In addition, ‘fine tuning’ of some of the

microscope features can be performed
such as parfocality, setting the focus step
size for each lens.

When leaving the Workflow, the new

items/settings are permanently stored.

The major types of Leica microscope that

are used with LAS are described in
separate sections.

104 © 2011 Leica Microsystems (Switzerland) Limited

 Setup: Image Explorer and LAS Archive

If you are an LAS Core user with no

optional LAS Archive modules installed,
please ignore this topic.

If LAS optional module Archive – either

Basic or Standard – is installed, the
Archive tab appears on the Setup
Workflow.

1: If necessary click on the tab to reveal

the control panels.

2: Because two image storage methods

are available – Image Explorer and
LAS Archive – the Select Storage
Method facility allows either to be
selected. The buttons are mutually
exclusive so it is not possible to use
both methods at the same time. Click
LAS Image Explorer to use tree-and-
folders navigation or…

3: …click Folder Archive to use

archives.

4: The Archive Toolbar and…

5: …the Archive List window are active

only when Archive is selected.

Using Image Explorer: Go there... 298

Using LAS Archives: Go there... 385

LAS User Manual 105

 Stereo Microscope Setup

Stereo Microscope setup and

configuration is a fast and simple exercise
due to the intuitive interface and concise
option lists.

The setup program is part of Leica

Application Suite that is able to 'read' data
from the microscope and sort and display
setup options that are available and
appropriate.

Each setup parameter is displayed as an

icon on the Microscope Configuration
panel (1) - clicking it reveals the options
(2) and clicking an entry on the list (3)
assigns all of the chosen attributes to the
parameter.

The Stereo Microscope Setup dialog is

reached by clicking on the Setup
Workflow (4) and then on the Microscope
tab (5).

Continued... 107

106 © 2011 Leica Microsystems (Switzerland) Limited

 Stereo Microscope Setup: Hardware Selection:

Before starting the Stereo Microscope Setup, the stand

and camera have to be configured for use with Leica
Application Suite and optionally, a new Hardware
Configuration created.
Go there... 8

LAS User Manual 107

 Stereo Microscope Setup: The Selection Panels:

There are two selection panel styles in the Stereo 3: Clicking on an option in the lower panel assigns the
Microscope Setup: function to the control in the upper panel. In the
illustration, Hardware Key K2 is selected in the upper
Single panel in which all of the options are displayed as a panel and when pressed, it will Open the Iris. It could
list and simply clicking on the highlighted entry selects it, have been assigned any of the options in the lower
and panel to suit the user.

Dual panel in which a range of controls are shown on the 4: Long option or control lists automatically display a
upper panel (A on the illustration) and depending upon the Scroll Button. Click on the button and drag it
selection, a list of options is displayed in the lower panel (B downward to view more list entries.
on the illustration.).
5: Different users will have different preferences for the
The example shows the setup screen for the SmartTouch. way in which controls operate. Each user can store his
The controls - keys, touch screen and Dual Rotary preferences in a Configuration that can be recalled
Actuator are listed in the upper panel with references to and loaded every time he uses the equipment.
the image on the right - and the various functions that can
be applied to the selected control are shown in the lower 6: Configurations are saved and if necessary, deleted
panel. using the buttons.

1: Click a control to select and highlight it. Continued... 109

2: As the mouse moves across the items in the list they

are highlighted in dark brown.

108 © 2011 Leica Microsystems (Switzerland) Limited

 Stereo Microscope Setup: Preferred Options:

The available options for some microscope features are 3: ...all of the additional options - checkbox disabled.
grouped as preferred - those recommended as better
suited to the setup - and a wide range of additional Click on the checkbox to toggle enable/disable.
options that will not be preferred.
Continued... 110
1: A checkbox appears bottom-left of the options
panel...

2: ...toggles between displaying only the preferred

options - checkbox enabled - and...

LAS User Manual 109

 Stereo Microscope Setup: Optics Carrier:

In most cases, the Optics Carrier type is

detected by the software, highlighted on
the database list but cannot be changed.

For microscopes on which assignment is

possible:

1: Click on the Optics Carrier icon on

the configuration panel.

2: The current setting is displayed on

the header.

3: Click to select a new setting from the

list.

Continued... 111

110 © 2011 Leica Microsystems (Switzerland) Limited

 Stereo Microscope Setup: Eyepiece:

Eyepiece selection has a single list of

options that will show either the preferred
(checked) or additional options
(unchecked):

1: Click on the Eyepiece icon on the

configuration panel.

2: Scroll to the required setting and click

to select it.

Continued... 112

LAS User Manual 111

 Stereo Microscope Setup: Tube Selection:

Tube Selection has a single list of options

that will show either the preferred
(checked) or additional options
(unchecked):

1: Click on the Tube icon on the

configuration panel.

2: Scroll to the required setting and click

to select it.

Continued... 113

112 © 2011 Leica Microsystems (Switzerland) Limited

 Stereo Microscope Setup: Camera Adapter:

The Camera Adapter has a single list of

options that will show either the preferred
(checked) or additional options
(unchecked):

1: Click on the Camera Adapter icon on

the configuration panel.

2: Scroll to the required setting and click

to select it.

Continued... 114

LAS User Manual 113

 Stereo Microscope Setup: Main Objective:

The Main Objective has a single list of

options that will show either the preferred
(checked) or additional options
(unchecked):

1: Click on the Main Objective icon on

the configuration panel.

2: Scroll to the required setting and click

to select it.

Continued... 115

114 © 2011 Leica Microsystems (Switzerland) Limited

 Stereo Microscope Setup: Objective Revolver:

Leica Application Suite can read settings

from the microscope and will detect if a
turret is fitted. If there is, then all
objectives have to be selected and
assigned:

1: Click on the Main Objective icon on

the configuration panel.

2 & 3: The upper panel lists the

objectives to be selected. Click on an
entry to select it and...

4: ...click to assign the objective type

from the lower panel. Repeat for the
other objective.

Users with coded objective revolvers will

need to manually rotate the revolver to
both positions before selecting the
assignment.

5: The objective magnification is

displayed on the Zoom panel in the
Acquire Workflow.

Continued... 116

LAS User Manual 115

 Stereo Microscope Setup: Fluo Shutter Mode:

The Fluo setting determines how the fluo

shutter will behave as filters are changed:

1: Click on the Fluo icon on the

configuration panel.

2: Click to select the required behaviour.

Continued... 117

116 © 2011 Leica Microsystems (Switzerland) Limited

 Stereo Microscope Setup: UMC Configuration:

The button and wheel functions on the

Universal Manual Control (UMC) can be
changed to suit individual users and
saved as configurations that can be
recalled whenever a user operates the
microscope.

1: Click on the UMC icon on the

configuration panel. The icon appears
only if a UMC is connected.

2: On the upper panel click to select the

control - button or wheel - that is to be
assigned a function. The image of
the UMC (3) is captioned with the
control references. Functions
applicable to the selected control are
listed on the lower panel.

4: Scroll through and click to select the

function to assign to the control.

Continued... 118

LAS User Manual 117

 Stereo Microscope Setup: UMC Configuration: Continued:

To save the UMC setup as a

Configuration that can be retrieved and
loaded at any time:

1: Click on the small arrows to the right

of the Configuration header.

2: From the drop-down list, click to

select User Defined.

3: Click on the Save button and the

Save UMC Configuration dialog
appears.

4: Click inside the UMC Configuration

text box and type a unique name for
the new Configuration.

5: Click the Save button.

6: The new Configuration name appears

in the Configuration list and can be
loaded at any time simply by clicking
to select it.

7: Delete a Configuration by clicking to

select it from the drop-down list and
then clicking the Delete button. This
action cannot be undone.

Two Default settings - Right-handed and

Left-handed users - that revert to the
factory settings, can be loaded by clicking
on the entry on the drop-down list.

Continued... 119

118 © 2011 Leica Microsystems (Switzerland) Limited

 Stereo Microscope Setup: Smart Touch:

The Hardware Keys, Double Rotary

Actuator, Toggle Button and Touch
Screen functions on the SmartTouch
control can be changed to suit individual
users and saved as configurations that
can be recalled whenever a user operates
the microscope.

1: Click on the SmartTouch icon on the

configuration panel. The icon appears
only if a SmartTouch is connected.

2: On the upper panel click to select the

control - Hardware Key, Rotary
Actuator, Toggle Button or Touch
Screen icon - that is to be assigned a
function. The image of a SmartTouch
(3) is captioned with the control
references. Functions applicable to
the selected control are listed on the
lower panel.

4: Scroll through and click to select the

function to assign to the control.

Continued... 120

LAS User Manual 119

 Stereo Microscope Setup: SmartTouch: Continued:

To save the SmartTouch setup as a

Configuration that can be retrieved and
loaded at any time:

1: Click on the small arrows to the right

of the Configuration header.

2: From the drop-down list, click to

select User Defined.

3: Click on the Save button and the

Save Smart Touch Configuration
dialog appears.

4: Click inside the Smart Touch text box

and type a unique name for the new
Configuration.

5: Click the Save button.

6: The new Configuration name appears

in the Configuration list and can be
loaded at any time simply by clicking
to select it.

7: Delete a Configuration by clicking to

select it from the drop-down list and
then clicking the Delete button. This
action cannot be undone.

Clicking to select the Default option from

the drop-down list, will revert to the factory
settings.

Continued... 121

120 © 2011 Leica Microsystems (Switzerland) Limited

 Stereo Microscope Setup: Zoom Display:
The items displayed in the Zoom Display
window on the front of the microscope, can
be changed to suit the user and then saved
as a configuration that can be recalled
whenever the user operates the
microscope.

1: Click on the Zoom Display icon on the

configuration panel.

2: On the upper panel click to select the

display element that is to be assigned
a feature. The image of the Zoom
Display window (3) is captioned with
the element references.

4: Scroll through the list in the lower

panel and click to select the feature to
assign to the element.

Continued... 122

LAS User Manual 121

 Stereo Microscope Setup: Zoom Display: Continued:

To save the Zoom Display settings as a

Configuration that can be retrieved and
loaded at any time:

1: Click on the small arrows to the right

of the Configuration header.

2: From the drop-down list, click to

select User Defined.

3: Click on the Save button and the

Save Zoom Display Configuration
dialog appears.

4: Click inside the Zoom Display text

box and type a unique name for the
new Configuration.

5: Click the Save button.

6: The new Configuration name appears

in the Configuration list and can be
loaded at any time simply by clicking
to select it.

7: Delete a Configuration by clicking to

select it from the drop-down list and
then clicking the Delete button. This
action cannot be undone.

Clicking to select the Default option from

the drop-down list, will revert to the factory
settings for the Zoom Display.

Continued... 123

122 © 2011 Leica Microsystems (Switzerland) Limited

 Stereo Microscope Setup: Selecting Accessories:

Accessories available for the current

microscope are shown in a single list.
When an accessory is fitted:

1: Click on the Accessory icon on the

configuration panel. The icon is
always available whether an
accessory is fitted or not.

2: From the list, click to select the fitted

accessory and enable its function and
features.

Continued... 124

LAS User Manual 123

 Stereo Microscope Setup: Footswitch:

The Footswitch actions can be configured

to suit the user and then saved as a
configuration that can be recalled
whenever the user operates the
microscope.

1: Click on the Footswitch icon on the

configuration panel.

2: On the upper panel click to select the

pedal that is to be assigned a feature.
The image in the Footswitch window
(3) is captioned with the pedal 1 (P1)
and pedal 2 (P2).

4: Scroll through the list in the lower

panel and click to select the feature
to assign to the selected pedal.

Continued... 125

124 © 2011 Leica Microsystems (Switzerland) Limited

 Stereo Microscope Setup: UMC Configuration: Continued:

To save the Footswitch setup as a

Configuration that can be retrieved and loaded
at any time:

1: Click on the small arrows to the right of

the Configuration header and...

2: ...click to select User Defined.

3: Click on the Save button and the Save

Footswitch Configuration dialog appears.

4: Click inside the Footswitch Configuration

text box and type a unique name for the
new Configuration.

5: Click the Save button.

6: The new Configuration name appears in

the Configuration list and can be loaded at
any time simply by clicking to select it.

7: Delete a Configuration by clicking to

select it from the drop-down list and then
clicking the Delete button. This action
cannot be undone.

Continued... 126

LAS User Manual 125

 Stereo Microscope Setup: TLRC Control:

The Transmitted Light Reflectivity Control

(TLRC) can be enabled and disabled on
the Configuration tab of the TL CCIC
panel:

1: Click on the Zoom Intensity Control

icon on the Configuration panel.

2: On the TL CCIC panel Configuration

tab...

3: ...click the TL CCIC entry.

4: Toggle between enabled and

disabled by clicking on the required
option on the Database panel.

5: If the control has not been calibrated,

the warning appears.

Continued... 127

126 © 2011 Leica Microsystems (Switzerland) Limited

 Stereo Microscope Setup: TLRC Calibration:

1: Click on the Fine Tuning tab. 5: ...until the Calibration successful message appears...

2: Follow the list of setup instructions on screen. 6: ...and the Calibrated check box is enabled.

3: Click the Illumination Calibration button. Return to the previous page (Go there... 126 ) to enable the
TL CCIC.
4: The calibration process begins with the results from
each step displayed...

LAS User Manual 127

 DM Microscope Setup

The DM Series Microscope Setup has

been arranged so that configuring the
microscope is both fast and simple.

Each setup feature is displayed on the

Components panel - clicking an icon will
take the user directly to the appropriate
setup dialogs. The feature icons displayed
will depend upon the microscope model
and fitted accessories.

Generally, all of the setup options for each

feature are listed in a database and all
that is required is a double-click on the
entry to assign it to the selected feature.

1: Click on the Setup Workflow.

2: Click on the Microscope tab.

3: Click on a feature icon as described

on the following pages.

Continued... 129

C-mount selection... 148

128 © 2011 Leica Microsystems (Switzerland) Limited

 DM Microscope Setup: Navigating the Dialogs:

Many of the dialogs have features designed to help user 4: Some dialogs have drop-down menus and, depending
navigation. For example: on the selection the Database listings will change. To
reveal the drop-down options click on the small arrows
1: Click on the border around either the top or bottom to the right of the menu header and...
panels and drag it up or down to re-size it. This can be
used to display more Database items. 5: ...click to select the required option.

2: To change the width of the Database columns and so 6: To remove a selected Database item from the
reveal more of the item descriptions, click on the Microscope Values list, right-click on the item and left-
vertical bar between two columns and drag to the left click on the pop-up Delete Item button.
or right.
Continued... 130
3: By default the Database items are listed in ascending
order - lowest numeric value at the top of the list.
Change to descending order by double-clicking on the
column header.

LAS User Manual 129

 DM Microscope Setup: Microscope Data and Links:

To view the microscope data:

1: Click on the microscope icon on the Microscope > 4: Click on the Fine Tuning tab to display all of the
Components panel. The microscope stand type is setup features in a single list. The setup can be
retrieved from the Hardware Configuration and achieved from here or by going to the required
displayed beneath the icon. section:

IL Turret Selection: Go there... 131 .

Mag Changer Setup: Go there... 132

DIC Turret Setup: Go there... 133

Nosepiece Setup and Fine Tuning: Go there... 134

Z-Drive Setup: Go there... 152

2: Click on the Configuration tab and... Stage Selection & Initialisation: Go there... 150

3: ...the data is displayed. Port Selection: Go there... 146

Camera Selection: Go there.. 147 . 147

Condenser Selection: Go there... 154

Function Key Assignment: Go there... 155

130 © 2011 Leica Microsystems (Switzerland) Limited

 DM Microscope Setup: IL Turret Selection:

To set up the IL Turret Filter Cubes: 5: The database item details are displayed in the turret
position and...
1: Click on the IL Turret icon on the Microscope >
Components panel. 6: ...if there is another position to be filled, it is
automatically selected.

7: To remove an item from the turret list, right-click the

entry and left-click the Delete Item pop-up button.

NB: At least one position of the fluo turret has to be

learned in as “EMPTY” to perform bright field contrast
(BF) in transmitted light axis. EMP-BF has to be selected
in case the stand is capable of a fully motorized IC
contrast in transmitted light axis. If the microscope is
capable of a partial manual IC contrast the EMP-DIC has
2: The IL Turret dialog appears with the number of turret to be selected.
positions indicated as dark circles on the graphic.
Alternatively in case an A cube is used: select A-TL to
3: Click on the turret position to select it. perform contrast methods of the transmitted light axis
with this fluo cube.
4: Double-click on the required database item to assign it
to the turret position.

LAS User Manual 131

 DM Microscope Setup: Mag Changer Selection:

To set up the Mag Changer: 3: Click on the position to select it.

1: Click on the Magnification Changer icon on the 4: Double-click on the required database item to assign
Microscope > Components panel. it to the Mag Changer.

5: The database item details are displayed in the

changer position and...

6: ... on the header.

No Mag Changer Setup: *

7: If a No Mag Changer setting is needed, right-click the
position entry and left-click the Delete Item pop-up
button. The Microscope Value will appear as 1,1,1
2: The Mag Changer dialog appears. There is a single which indicates No Changer.
position value that assigns the Mag Changer
component.

132 © 2011 Leica Microsystems (Switzerland) Limited

 DM Microscope Setup: DIC Turret Setup:

To set up the DIC (Differential Interference Contrast) Turret 3: Click on the turret position to select it.
components:
4: Double-click on the required database item to assign
1: Click on the DIC Turret icon on the Microscope > it to the turret position.
Components panel.
5: The database item details are displayed in the turret
position and...

6: ...if there is another position to be filled, it is

automatically selected.

7: To remove an item from the turret list, right-click the

entry and left-click the Delete Item pop-up button.

2: The DIC Turret dialog appears with the number of

positions indicated as dark circles on the graphic.

LAS User Manual 133

 DM Microscope Setup: Nosepiece:

Setting up the Nosepiece is a two-step process:

Select and assign an Objective to the

Nosepiece positions.

Fine-tune the Objective setup.

To assign an Objective to a Nosepiece position:

1: Click on the Nosepiece icon on the Control

Panel.

2: Click on the Configuration tab on the

Nosepiece dialog.

Continued... 135

134 © 2011 Leica Microsystems (Switzerland) Limited

 DM Microscope Setup: Assign Objectives:

1: Click on the Nosepiece position. 4: ...if another position remains to be assigned, it is

automatically selected. Repeat Step (3).
2: In the database, double-click on the required
Objective. 5: To remove an Objective from a position, right-click on
the Nosepiece position and left-click on the Delete
3: The Objective details are displayed in the Nosepiece Item pop-up button.
position and...
Continued... 136

LAS User Manual 135

DM Microscope Setup: Objective Fine Tuning:

1: Click on the Nosepiece icon on the Control panel. Parfocality: Go there... 137 .

Objective Combination tag: Go there... 139

Stage and Z-Stepsize: Go there... 140

Auto Stage positition: Go there... 141

Parcentricity enable: Go there... 144

Parcentricity adjustment: Go there... 143

2: Click on the Fine Tuning tab. Nosepiece rotation: Go there.. 145

3: There are individual dialogs for each of the Objective

fine-tuning features:

136 © 2011 Leica Microsystems (Switzerland) Limited

 DM Microscope Setup: Parfocality:

Even with closely matched objective sets, Parfocality - Set Parfocality from DM Setup (Here):
maintaining sharp focus when an objective is changed -
may need to be fine-tuned. A Leica motorised focus axis
must be fitted.

The process involves initially focussing on a specimen

using the objective with the greatest magnification, and
then checking each of the remaining objectives in turn
and, where necessary adjusting the focus to maintain a
clear, sharp image.

A focus factor is created for each objective, stored and

retrieved when the objective is selected to drive the stage
to the precise position to maintain focus.

There are two options for setting the Parfocality: 1: Click on the Nosepiece icon on the Microscope >
Component panel.
From this setup dialog with focus being checked
through the eyepieces - the monitor Viewer is not 2: Click on the Fine Tuning tab and then click to select
available here - and... the Parfocality dialog. Continued... 138

From the Acquire Workflow with the specimen

displayed on the Viewer. Set Parfocality from Acquire Workflow:

For both options the dialogs are the same but are
accessed differently:

3: Click on the Acquire Workflow and then...

4: ...on the Mic 1 tab.

5: Right-click on one of the objective select buttons

and...

6: ...left click to select Adjust Parfocality on the pop-up

menu. Continued... 138

LAS User Manual 137

DM Microscope Setup: Parfocality: Continued:

1: Click on the Parfocality dialog and...

5: ...click OK. The sequence will advance to the next
2: ... the Start button. The Find Focus option is selected. objective.
Navigate the stage to and adjust for the sharpest
focus on the specimen. 6: When all objectives have been fine-tuned the Finish
icon is highlighted.
3: Click OK.
7: Click OK.
4: The objective with the greatest magnification is
automatically chosen first. Focus the specimen and...

138 © 2011 Leica Microsystems (Switzerland) Limited

 DM Microscope Setup: Objective Combination tagging:

Some objectives can be used both in air and also

immersed - in water or oil. These so-called Combi
(Combination) objectives can be tagged so that when
they are selected the user is given a warning that
immersion can be an option.

1: Click on the Objective Icon to select it.

2: Click the check box to enable (ticked) or disable

the Combi tag.

If either a tagged Combi or Immersion only objective is

selected on the Acquire > Mic 1 tab, the button will flash 3: Immersion only objectives are marked with a 'filled'
to warn the user. teardrop icon whereas...

4: ...Combi objectives are marked with an outlined

teardrop.

LAS User Manual 139

 DM Microscope Setup: Stage and Z-Stepsize:

Only available when a Leica Stage Controller is fitted, it

allows a stage speed - X, Y and Z - to be matched to an 1: Click on the Objective Icon to select it.
objective based upon its magnification. Optimising the
stage speed improves efficiency. 2: Click on the small arrows to the right of the Stepsize
menu header and...
Higher stage speeds are used with low magnification
objectives and low stage speeds with high magnification 3: ...click to assign the appropriate Stage speed.
objectives both with the aim of maintaining the best
navigation speed with 'losing' the image region of interest.

There are five step speeds ranging from SC - fast and

coarse - to S0 - slow and fine - any one of which can be
assigned to an objective so that when it is selected the
appropriate stage speed is engaged automatically.

140 © 2011 Leica Microsystems (Switzerland) Limited

 DM Microscope Setup: Automatic Stage Lowering:

To avoid an objective colliding with the specimen, the

Stage can be automatically lowered whenever the
objective is selected. To assign Automatic Stage lowering:

1: Click on the Objective Icon to select it.

2: Click on the check box to enable (ticked) or disable

Stage lowering for that objective.

LAS User Manual 141

 DM Microscope Setup: Parcentricity:

Adjusting Parcentricity aims to maintain the specimen Set Parcentricity from DM Setup (Here):
feature of interest as close to the centre of the field of
view regardless of the objective magnification.

The process is straightforward. A feature on a specimen

is focussed and moved as close as possible to the
centre of the field of view using the stage navigation 1: Click on the Nosepiece icon on the Microscope >
controls. The stage X and Y positions are stored as an Component panel.
'offset factor' for the selected objective.
2: Click on the Fine Tuning tab and then click to select the
The objective is then changed and the stage adjusted so Parcentricity dialog. Continued... 143
that the same feature is again close to the centre and
the X/Y offset stored. The process is repeated for all of
the objectives. Parcentricity improves if the feature Set Parcentricity from Acquire Workflow:
position is finely adjusted for each objective.

In the future when an objective is selected, the offset

factor is automatically applied to the stage drivers to
maintain Parcentricity.

There are two options for adjusting the Parcentricity:

From this setup dialog with position being checked

through the eyepieces - the monitor Viewer is not
available here - and...

From the Acquire Workflow with the specimen feature

displayed on the Viewer.

For both options the dialogs are the same but are 3: Click on the Acquire Workflow and then...
accessed differently:
4: ...on the Mic 1 tab.

5: Right-click on one of the objective select buttons and...

6: ...left click to select Adjust Parcentricity on the pop-up

menu. Continued... 143

142 © 2011 Leica Microsystems (Switzerland) Limited

 DM Microscope Setup: Parcentricity Adjustment: Continued:

1: Click on the Parcentricity dialog and... 5: ...click OK. The sequence will advance to the next
objective.
2: ... the Start button. The Find Focus option is selected.
Navigate the stage to and adjust for the sharpest Re-focus if necessary and again centralise the feature
focus on a recognisable feature on the specimen. in the field. Repeat the process for all of the
objectives.
3: Click OK.
6: When all objectives have been fine-tuned the Finish
4: The objective with the greatest magnification is icon is highlighted.
automatically chosen first. If necessary, focus the
specimen, move the stage so that the feature is in the 7: Click OK.
centre of the field of view and...

LAS User Manual 143

 DM Microscope Setup: Parcentricity Enable:

Automatic Parcentricity adjustment can be turned on and

off on the Enable panel.

1: Click on the Nosepiece icon on the Microscope >

Components panel.

2: Click on the Fine Tuning tab.

3: On the Parcentricity Enable panel, click the check

box to enable (ticked) or disable automatic
Parcentricity.

144 © 2011 Leica Microsystems (Switzerland) Limited

 DM Microscope Setup: Nosepiece Rotation:

There are two options for determining how the Nosepiece Restricted allows the software to prevent the
behaves when an objective is selected: Nosepiece from ever rotating more than 3600 even if
that means taking the longest path between the
1: Click to select the required mode: current and selected objectives.
Full allows the software to determine the shortest path
- clockwise or counter-clockwise between the current
and selected objective, or

LAS User Manual 145

 DM Microscope Setup: Port Selection: Visual Tube:

Select the tube Eyepiece as follows: 3: From the drop-down list, click to select Visual tube.
The list of Eyepiece options appears on the Database
1: Click on the Ports icon on the Microscope > panel.
Components panel.
4: Double-click on the Database entry to select it.

5: The item number appears in the Microscope Values

list with...

6: ...the number and name displayed in the header.

7: Change a selection by right-clicking the value and

then left-clicking the Delete Item pop-up button to
remove the selection. Then proceed as describe to
make a new selection.
2: Click on the small arrows to the right of the Port
Options drop-down menu header.

146 © 2011 Leica Microsystems (Switzerland) Limited

 DM Microscope Setup: Port Selection: Camera Ports:

The number of Camera (DOC) Ports is determined by

the fitted hardware and the configuration selected here
in setup.

There a three possible setups:

1: No Camera Port - the DOC option is missing.

2: A single Camera Port - DOC 1 and...

3: ...dual Camera Port - DOC 1 and DOC 2.

The Camera Port(s) must be configured camera(s) can

be assigned by:

4: Clicking on the Ports icon on the Microscope >

Components panel.

6: Click on the Manual-Tube option.

7: On the Database panel scroll to the required

configuration and double-click to...

8: ...save it as the port setting. Repeat Step (5) and check

that the required number of Camera Ports is available
from the drop-down options.

5: Click on the small arrows to the right of the Port 9: Remove the configuration by right-clicking it in the
Options header. Microscope Values panel and then left-clicking the pop-
up Delete Item button.

LAS User Manual 147

 DM Microscope Setup: C-mounts & Adapters:

With the Camera Ports configuration selected, the Camera 3: ... click to select the Camera Port to which the
Adapter has to be selected for each port. adapter will be assigned - in the example DOC 1.

1: Click on the Ports icon on the Microscope > 4: On the Database, double-click to select the name of
Components panel. the fitted Adapter.

5: The selection appears in the Microscope Values

panel.

6: To remove a selection, right-click on the Microscope

Values entry and left-click the Delete Item pop-up
button.

Repeat the process for the other Camera Port if fitted.

2: Click on the small arrows to the right of the Port

Options header and from the drop-down...

148 © 2011 Leica Microsystems (Switzerland) Limited

 DM Microscope Setup: Camera Selection:

The Camera Ports and Mounts must be set up before the 3: From the list, click to select Camera (DOC 1). This is
camera(s) can be assigned to them: Go there... 147 Camera Port DOC 1 to which a camera is about to be
assigned.
Fit and connect the Camera(s) so that they can be
detected by the software. 4: The Database lists the fitted cameras - for the
majority of systems there will be just one Camera Port
1: Click on the Camera icon on the Microscope > and therefore one Camera type in the list. Double-
Components panel. click on the Database entry and...

5: ...the Camera type is assigned to the DOC 1 port and

appears in the Microscope Values list.

6: The Camera and its serial number are also displayed

on the header.

7: To remove a Camera from the Microscope Value,

right-click on the entry and then left-click the pop-up
Delete Item button.
2: Click on the small arrows to the right of the Camera
Options header. Repeat the process if a second camera is fitted selecting
the DOC 2 port.

LAS User Manual 149

 DM Microscope Setup: Stage Selection & Initialisation:

Stage Selection and Initialisation are both carried out from 3: Move to the fitted stage name and type on the
the same panel. Database and double-click it.

1: Click on the Stage icon on the Microscope > 4: The stage Article Number appears in the value list
Components panel. with...

5: ...the description displayed on the header.

6: To change the selection, right-click on the selection

in the values list and then left-click the Delete Item
pop-up button. Repeat the process described above
to make a new selection.

2: To select the stage type click on the Configuration tab.

150 © 2011 Leica Microsystems (Switzerland) Limited

 DM Microscope Setup: Stage Initialisation:

Stage Selection and Initialisation are both carried out from 2: To initialise the stage, click on the Fine Tuning tab
the same panel. and...

1: Click on the Stage icon on the Microscope > 3: ...click the Initialise button to drive the stage to its
Components panel. For initialisation the stage must be limits and set the starting position to top-left (X = 0/ Y
lowered sufficiently to avoid collision with specimens = 0).
or objectives.
During initialisation the stage can
move very quickly and the drive
mechanism is powerful enough to
severely damage specimens and
objectives. Also ensure that there are
no cables that are likely to become
entangled.

LAS User Manual 151

 DM Microscope Setup: Z Drive Setup and Initialisation:

The Z-Drive dialog is divided into 2 panels - Focus Position 3: Drive immediately to the Lower Limit Switch or...
and Initialisation.
4: ...to the selected Lower Threshold (Please see the
1: Click on the Z-Drive icon on the Microscope > following page: Go there... 153 )
Components panel.
5: The Initialisation Mode (Please see also Stage
Initialisation: Go there... 151 ) can be set to either:

Default: Factory setting for 4mm lowering.

Lower Limit Switch: Drives the stage to its lowest
position.
None: The stage remains in the focus position last
set.

Click the appropriate buttton.

2: Set the preferred initial Handwheel Speed by clicking Continued... 153
either Fine or Coarse. The selection is highlighted.

152 © 2011 Leica Microsystems (Switzerland) Limited

 DM Microscope Setup: Z Drive Setup and Initialisation:

The current Z position can be saved either as a repeatable 3: Right-click on the Slider to reveal the Settings
focus or as the lower threshold for initialisation. and Help options. Click on Settings and...

1: Click on the Z-Drive icon on the Microscope > 4: ...and the Focus Settings dialog appears.
Components panel.
5: Save the current Z position as either the
repeatable Focus or the Lower Threshold by
clicking the appropriate button.

6: Clear either setting by clicking the appropriate

clear button and...

7: ...click OK.

8: It is not possible to save a Focus Position that is

lower than the Lower Threshold - the warning
2: To set the focus or lower threshold position, click and hold message appears.
down the Focus Drive Slider and move it to the required
position. The distance is displayed in the window.

LAS User Manual 153

 DM Microscope Setup: Condenser Selection:

To select a single condenser or assign different 4: Either a single position or the number of turret
condensers to turret positions: positions is listed under Microscope Values. Click on
the position to be assigned.
1: Click on the Condenser icon on the Microscope >
Components panel. 5: From the Database, double-click the type to be
assigned to the position.

6: The selected type is displayed in the chosen position

and...

7: ...the next position (if the turret option is selected) is

automatically highlighted.

8: The remove an assigned condenser type, right-click

on the position and left-click the Delete Item pop-up
button,
2: Click on the small arrows to the right of the Condenser
options drop-down menu and...

3: ...click to select either a single condenser or the turret.

154 © 2011 Leica Microsystems (Switzerland) Limited

 DM Microscope Setup: Function Key Assignment:

The microscope push-button controls can be assigned to 3: The push-button numbers are listed under the
functions that suit the user. Microscope Values. Click a position to select the
push-button.
1: Click on the Function Keys icon on the Microscope >
Components panel. 4: Double-click the function on the Database list to
assign it to the button.

5: The function is displayed on the Microscope Values

list and on the header.

6: The next push-button is automatically selected.

7: Right-click on an entry and then left-click the Delete

Item pop-up button to clear an assignment.
2: The Configuration dialog appears with the push-
buttons displayed as a graphic 'map', each with a
number.

LAS User Manual 155

 The Acquire Workflow:

The Acquire Workflow provides control for the

attached microscope and camera and also
displays a live image on the Viewer.

Images are acquired, stored and displayed in

the Gallery as thumbnails.

Within Acquire all microscope and camera

controls can be set to individual requirements
ranging from objective magnification, contrast
method, exposure, gain and gamma.

Optional Modules such as:

Montage MultiFocus.
Measurements
Image Overlay
MultiStep
MultiTime Timelapse.
Image Analysis
Power Mosaic and
Phase Expert...

...provide more specialised acquisition options

Leica Application Suite has LAS Archive as an

optional module to provide all of the speed and
power of a database for storing images and
data. It is available as:

Archive Basic: Essential Archive tools with

versatile structuring and storage options,
and...

Archive Standard: Taking Archive to the

very highest levels of reporting and display.

156 © 2011 Leica Microsystems (Switzerland) Limited

 The Acquire User Interface:

The illustration is a graphical representation of the LAS display and Acquire user interface showing the principal
features and quick links (Jump to the link by clicking ):

1: Workflows 158 Click the Acquire Workflow to launch the microscope and camera controls.
2: Tabs: Select microscope, camera or specialised acquisition functions.
3: Tools 221 Toolbox when Camera is selected.
4: Control Panels for:
173 Stereo Microscope.
189 DM Microscope
215 FS Microscope
221 Camera
5: Image Data Form 170 Displays and edits selected data for the current image.
6: The Grid 169 Displays data for all of the images in the selected folder.
7: The Gallery 167 Displays thumbnails of all the images in the selected folder.
8: Acquire Button 171 Click to grab and image from the microscope camera.
9: Side Toolbar 159 Working tools for image sizing, printing and deleting as well as switches for the Gallery and
Grid.
10: The Search Controls 410 Available only with LAS Archives.
11: Gallery Navigation Browser 167 Rapidly find thumbnails in the Gallery.
12: Thumbnail Scaler 167 Slider adjusts the size of the thumbnails in the Gallery.
13: Status Bar Displays Hardware Configuration, RGB Intensity, Stage Position and Magnification data.
14: The Image Viewer 166 Display and working area for the current image: Press keyboard F5 to show full
screen.

LAS User Manual 157

Launching Acquire:

1: Click on the Acquire Workflow and...

2: ...click on a tab to select microscope,

camera, the selected sequence module or
Live Measurement.

3: Some control panels are collapsed to avoid

cluttering the working area. Expand them
by...

4: ...clicking on the arrow to the right of the

panel header.

5: Most panels can be moved to any part of

the Viewer and docked by clicking on the
panel header and, holding down the mouse
button, dragging it to the preferred
location.

6: When a panel is moved a 'snap back'

symbol (X) appears on the header. Click it
to return the panel to its usual position on
the tab.

158 © 2011 Leica Microsystems (Switzerland) Limited

 The Side Tool Bar:

This section describes the Side Tool Bar tools that are common to most of the Acquire features although
the tool range may change with a selected feature.
Click on a Tool Bar button for more information:

Scale Bar and Annotations: Run Annotations and Scale Bar without leaving Browse.

Floating Navigator: Click to enable the Floating Navigator and click again to 'park' it.

Panning: Examine areas of images that extend beyond the Viewer edges into the display area.

Zoom in and...

Zoom Out.

Fit the image to the Viewer area.

Display at Original Size: Displays he image at its captured size:

Hide and Reveal the Record Panels.

Hide and Reveal the Viewer.

Hide and Reveal the Data Grid: Only available with LAS Archives.

Hide and Reveal the Thumbnail Gallery.

View the image Record Details: Not available in Acquire.

Select the Form Details to display: Not available in Acquire.

Viewer Options: Select Dual Viewer, Lock the Views and Lock the Pan View.

Copy Output Image: Not available in Acquire.

LAS User Manual 159

 Annotations & Scale Bar Quick Launch:

Clicking the Show Annotations button displays the required function. All of the function tools are available.
Annotations and Scale Bar Quick Launch menu.
The Scale Bar display can be turned on and off by clicking
On the live image Basic Annotation, Extended Annotation the Show Scale Bar / Annotations option. Turned on and
and the Scale Bar setup can be launched without leaving the annotation tools are available.
Acquire by checking (a tick mark is displayed) the

160 © 2011 Leica Microsystems (Switzerland) Limited

 The Floating Navigator:

The Browse Navigator panel can be displayed

and docked on the Acquire Viewer by:

1: Click on the Floating Navigator button on

the Side Tool Bar.

2: The Navigator panel appears and can be

used in the same way as it would be from
the Browse Workflow except the Toolbox is
not available.

3: Move and dock the Navigator by clicking on

the header bar and, holding down the
mouse button, drag the panel to a new
position.

4: Click on the 'snap-back' button (X) to close

the Navigator.

5: Collapse the Navigator without closing it

completely by clicking on the arrow to the
right of the header. Click again to restore
the panel.

LAS User Manual 161

Panning and Zoom:

1: Panning: The Pan tool allows detailed

areas of an image that exceeds the visible
area of the Viewer to be examined. It will
not work if Fit to Viewer is enabled
because all of the image is being
displayed.

On the Pan Window viewer, click and hold

in the outlined rectangle and drag it to the
area to be examined. The selected area is
displayed in the main Viewer.

To move the Pan Window away from the

main Viewer, click and hold the header
bar and drag it to another part of the
screen.

Click the Pan tool to close the Window.

2: Zoom: Click on the (+) to zoom in to the

image or (-) to zoom out. The zoom level
as a percentage, is displayed top right of
the the Viewer border.

If the monitor Magnification Settings have

been set in Preferences , the image
Magnification value appears bottom right
of the Viewer border.

3: Fit to Viewer: Click to fit the image to the

available Viewer area regardless of the
original size of the image. The Image
Scaled to Fit Window message appears
top right of the Viewer border.

4: Display at Original Size: Click to display

the image at its original size. The image
may appear smaller or larger than the
Viewer area. The Image Unscaled
message appears top right of the Viewer
border.

162 © 2011 Leica Microsystems (Switzerland) Limited

 Hide and Reveal:

The various screen areas - Viewer, Gallery, Report and 3: Hide/reveal the Data Grid: The Viewer will expand to
Grid (where applicable), may be revealed or hidden to cover some of the vacated space. The Grid is only
create the best working environment for the user. Some available if an LAS Archive is installed.
tools are toggles – click once to reveal the area, click
again to hide it: 4: Hide/reveal the Thumbnail Gallery: The Gallery is
hidden and the Viewer expands to include the Gallery
1: Hide/reveal the Record panels: The Image Viewer space.
expands to fill the vacant space.

2: Hide/reveal the Viewer: The Record panels expand to

occupy the Viewer width.

LAS User Manual 163

 Dual Viewer:

The Viewer area can be split to show two images 6: To synchronise the panes and enlarge or reduce the
simultaneously. images as the zoom and fit tools are used, click to
enable the Lock View option.
1: Click the Viewer Options button.
7: Enabling Lock Pan will synchronise the images as the
2: Click to enable (tick mark visible to the left) the Dual Pan tool is used. Click the pane to pan and then on
Viewer option. The Viewer will then divide into two the Panning tool. Both images will automatically move
panes. to and display the image segment shown in the Pan
window.
3: The live image currently being viewed will appear in
the left-hand pane. Dual Viewer more information: 79

4: Display an image in the right-hand pane by clicking

the pane and...

5: ...the thumbnail of the required image.

164 © 2011 Leica Microsystems (Switzerland) Limited

 Turn Off Dual Viewer:

To revert to the live image occupying the entire Viewer:

1: Click on the left-hand Viewer pane - the current live

image.

2: Click on the Dual Viewer button.

3: Click to disable Dual Viewer - the tick mark

disappears - and the live image expands to fill the
Viewer.

LAS User Manual 165

 Full Screen Mode & Second Monitor:

Full Screen Mode: Single Monitor:

1: The Viewer area can be expanded to fill

almost the entire screen by either
clicking Options on the main menu
and...

2: ...selecting Full Screen from the drop

down menu, or...

3: Pressing Key F5. Press F5 again to

return to the normal display.

Second Monitor:

The software detects a second monitor and

changes the Options drop-down menu to:

4: ... Use Second Monitor. Click the option

to use both monitors.

The Viewer and image occupy all of the

second monitor whilst the Gallery and
Thumbnails together with the controls
remain on the primary monitor.

The Side Tool Bar buttons are

appropriately shared between the two
monitors.

5: Alternatively, press Key F5 to move

between using both monitors and
returning to single monitor.

166 © 2011 Leica Microsystems (Switzerland) Limited

 The Gallery:

The Gallery is a thumbnail display of images in

the current folder in both Image Explorer or LAS
Archive.

The Gallery can be hidden or revealed using

the Side Tool Bar tools.

1: Clicking on a thumbnail will immediately

display the full-sized image in the Viewer
and the data associated with it in the
Record and Grid (If LAS Archives is
installed).

2: A Slider is automatically displayed for

multiple rows of thumbnails - click and drag
it to scroll the Gallery.

3: The Navigation Bar (bottom right of the

screen) provides a way of moving through
the thumbnails quickly and is especially
useful with large galleries of thumbnails.
Click on the arrows to move a single image
(3) ...

4: ...or go to the extreme ends of the Gallery.

5: The thumbnails can be re-sized by clicking

and dragging the Scaling Slider - slide left
to reduce the thumbnail size and right to
increase it.

6: Move the mouse over a thumbnail to reveal

basic data bout the image.

7: Right-click a thumbnail to show the Context

Menu. Left-click to select an option.

Gallery Docking Position 168 . 168

LAS User Manual 167

 The Gallery: Docking Position:

Available in Acquire and the Process Workflows as well 2: ...from the drop-down menu, left-click to select the
as Browse, the thumbnail Gallery can be 'docked' either Toggle Gallery Position option.
horizontally - along the bottom edge of the Viewer - or
vertically - along the left-hand edge of the Viewer - to suit The action toggles between horizontal and vertical
the user. docking.

1: Right-click on a thumbnail or on the spaces around Scroll bars, if required, are placed automatically.
the thumbnails and...

168 © 2011 Leica Microsystems (Switzerland) Limited

 The Grid:

The Grid displays data for all of the captured images in a 4: Column widths can be changed by clicking and
folder in a tabular structure. The image names are listed dragging the vertical bars that separate the columns.
on the left and the data items as headers across the top.
5: A small arrow is revealed when a header is clicked.
In Acquire captured images are displayed in the right-hand This allows the image data to be sorted – high-to-low
pane when the Dual Viewer is enabled æ: or low-to-high – by successive clicks on it.

1: The Grid is revealed and hidden by clicking on the 6: To make multiple selections prior to cutting or
Side Tool Bar button. This is a toggle – click to reveal, exporting, hold down the keyboard Ctrl key whilst
click again to hide. clicking individual thumbnails.

2: Clicking on an entry in the Grid will immediately Keyboard combination Ctrl + A will select all of the
display that image in the Viewer right-hand pane and image data.
also highlight the thumbnail. Ctrl + C will copy all the selected image data to the
clipboard.
3: Header positions can be changed by clicking and Ctrl + V will paste into another application.
holding the left mouse button on the header to be
moved, dragging it to the new position and releasing
the mouse button.

LAS User Manual 169

The Data Form:

1: The Data Form displays selected data

associated with the image.

All of the data about the camera,

microscope, exposure, creation date and so
on, are actually stored and all can be
displayed, but the Form can be configured
to display only the more pertinent items.

2: The Data Form can be hidden by clicking

the Form button on the Side Tool Bar. This
is a toggle action - click again to reveal the
Form.

170 © 2011 Leica Microsystems (Switzerland) Limited

 Acquire Image Controls:

The are three methods for starting image

capture:

1: Click on the Acquire Image button at the

bottom of the Acquire Workflow.

2: Press the keyboard F3 function key.

3: Click on Options on the Main Menu and

click the Acquire Image option (4).

LAS User Manual 171

Acquire Workflow: Stereo Microscope Operation: SMS Introduction:

Stereo- and Macroscope Systems (SMS) The following motorised/coded microscope devices are currently
is available on Leica Application Suite if an supported by LAS:
automated micro- or macroscope is Schott KL 2500LCD (31 250 200 & 31 250 201)
connected to the computer. Photonic CLS 150XD and CLS 150LS External light sources
(30 111 480 & 30 110 481)
SMS provides control of the micro- or Motorised focus 300mm and 500mm (10 446 176 & 10 447 041)
macroscope from the computer and, if a M 165 C and M 205 C
digital camera is also fitted to the MZ 16A microscope (10 447 103)
microscope will be especially helpful in MZ 16FA microscope (10 447 063)
optimising specimen images. Z 6 APOA microscope (10 446 368)
EZ 4 D
SMS can control: M 205A, M 205FA, M 165FC
MST 51/MST 59 Motor Focus
Motorised focus, 179 TL RCI Internal light source (10 446 352)
UMC, Universal Manual Control (10 447 080)
Motorised zoom, 177 Foot switch (10 447 398)
X and Y stage positioning, 184 X/Y Motorised stage (10 447 305)
Filter wheel, 176 The following functions are automatically detected and can be controlled
Zoom iris aperture, 181 from SMS:
Internal and external light source intensity, On and Off
Zoom objective changer. 178 CCIC shutter Open and Close
Zoom magnification (Objective changer)
Internal light source (TL), 183
Motorised focus position
External light source (IL) and 182 Fine focus position
Filter wheel position
CCIC and Fluorescence shutters. Fluorescence shutter Open and Close
175 Zoom iris aperture.

Five memory locations allow settings to The following manual microscopes and devices have reduced support in
be saved and recalled, precisely SMS:
replicating the microscope setup. MZ 16F (10 447 064)
MZ 16 (10 447102)
Before using SMS, please check that the MZ 12.5 (10 446 370)
motorised focus cable is fitted, that the MZ 9.5 (10 446 272)
security clamp is properly fitted and in a MZ 7.5 (10 446 371)
position to prevent collision with the MZ 6 (10 445 614)
specimen and that all cables have MS 5 (10 445 613 )
sufficient slack to allow the carrier to travel M 125
to the top of the stand. S6D (10 446 297)
S8 APO (10 810 038 )
Z16 APO (10 447 173)
Z6 APO (10 447 174)
Macrofluo
Fluocombi

172 © 2011 Leica Microsystems (Switzerland) Limited

 Stereomicroscope Operation: LAS Start-up:

If, at start-up, LAS detects a motorised

focus, it will ask to initialise the focus
position by displaying the message (1).

2: Click OK to send the carrier to the top

of the stand and automatically
initialise the focus position.

3: Click Cancel to skip the initialisation.

If initialisation is skipped the exact
focus position may not be reliable and
will not be saved with other settings.

4: Click the Acquire Workflow to reveal

the SMS control panels.
If focus was initialised the Focus
Drive position (5) will display the
current focus position.
If focus was NOT initialised, the value
will be '0'.
Previous settings that were saved
may be recalled and loaded by
clicking the appropriate Memory
button (6).

Continued... 174

LAS User Manual 173

Stereomicroscope Operation: The SMS Controls:

Revealing the SMS controls:

The Stereo- and Macroscope controls in
the Leica Application Suite are revealed
by:

1: Click on the Acquire Workflow.

2: Click on the Mic1 tab. Depending

upon the microscope and the
functions available, the controls may
be displayed on additional tabs
named 'n' sequence - Mic2 or Mic3.

The components of the Stereo- and

Macroscopes must be setup in the Setup
Workflow before the Acquire:Mic tabs can
be used.

Continued... 175

174 © 2011 Leica Microsystems (Switzerland) Limited

 Stereomicroscope Operation: Fluorescence Shutter Control

The Shutter button on the fluorescence

panel opens and closes the shutter. Close
the shutter when the microscope is not in
use to protect delicate specimens.

1: Click on the Shutter button. This is a

toggle action, the Shutter opening and
closing on successive clicks.
The Shutter status (Opened or
Closed) is shown on the
Fluorescence panel header bar and
when it is open a red dot appears on
the button.

2: If an Empty filter wheel position is

selected, the Shutter control is not
available(3) and remains closed.

Continued... 176

LAS User Manual 175

Stereomicroscope Operation: Filter Wheel Control:

1: Place the mouse cursor over a Filter

button to reveal its specification (2).
Click the button and the Filter Wheel
will rotate to select the filter.

To change a filter:
3: Manually turn the Filter Wheel to the
required position. Each filter has an
identifying tag on its rim. Carefully
slide out the filter and insert the new
one.

After filters have been changed:

4: Click on the Refresh button. All of the
Filter button labels will clear, the Filter
Wheel will turn and the fitted filters
identified.

5: The button labels will be displayed

with the correct filter data.

Continued... 177

176 © 2011 Leica Microsystems (Switzerland) Limited

 Stereomicroscope Operation: Motorised Zoom Drive:

There are five control options for the

Motorised Zoom:

Drag and drop:

1: Click and hold the Scale Indicator and
drag it to the required zoom position.
The shadow indicator (2) remains in
the starting position until the mouse
button is released.

Click on the Scale:

3: Click on the Scale Bar at the desired
zoom position. The Scale Indicator
will move to the selected position.

Type a value:
4: Click in the Zoom Drive text box and
type the required zoom position.
Values larger or smaller than the
zoom limits will be ignored.

Preset positions:
5: Click on the arrows to the right of the
Zoom Drive text box and from the
drop down list click to select a preset
position (6).

Fine adjustment:
7: Use the mouse wheel (if fitted) to
move the zoom in small steps.

Continued... 178

LAS User Manual 177

Stereomicroscope Operation: Zoom Magnification:

1: If the Objective Changer feature is

fitted and is enabled on the Hardware
Setup panel, High and Low
magnification levels are available.

2: Click to enable High magnification

or…

3: Click the Low magnification option.

4: The Zoom Drive Scale changes to

reflect the magnification level
selected.

See: Installation and Licensing Help for

selecting hardware and the Hardware
Setup panel.

178 © 2011 Leica Microsystems (Switzerland) Limited

 Stereomicroscope Operation: Motorised Focus:

There are three options for driving the

Motorised Focus:

Focus Control:
1: Fine focus (FF) and motor focus (MF)
buttons are available with MF
designated microscopes and Z6APO
(A) and Z16APO(A) macroscopes.

2: Click on fine (F or FF) to move the

focus in small increments.

3: Click on coarse (C or MF) to move

the focus in large increments.

4: Click, hold and drag the Focus

Control up or down. Release the
mouse button when the desired focus
is reached. Generally, start with
coarse (C or MF) selected to get
close to optimum focus and then
select fine (F or FF) to ‘tune’ the focus
position.

Type a position:
5: Click in the Focus Position text box
and press the keyboard Delete key to
clear the existing entry. Type a new
value and press the keyboard Enter
key.
The default units are micrometers
(µm) but to move in millimeters type
‘mm’ after the value.

Mouse Wheel:
4: Click on the Focus Control.

6: Rotate the mouse wheel to move the

motorised focus up or down. The
focus step for each indent of the
mouse wheel depends on the depth
of focus and the zoom magnification.

Continued... 180

LAS User Manual 179

Stereomicroscope Operation: Memory:

Current microscope settings may be

saved in any one of five separate memory
locations represented by a button and
numbered 1 to 5.

1: Place the mouse cursor over a

numbered memory button to reveal
the microscope settings:
Motorised focus position,
Zoom position,
Filter selected,
Iris diaphragm setting,
Internal and external light source
settings and
Motorised stage X and Y values.

Left click on the button to drive the

microscope to the memorised values.

2: Empty memory locations are denoted

by (--) on the button.

To save the current microscope

settings:
3: Select a memory location - either Set
or Clear - and right-click the button.
From the drop down menu, click on
Set Memory. The button will display
the location number to indicate that
the settings have been saved.

4: To clear a memory location, right-

click on the button and from the drop
down menu, click to select Clear
Memory. The button number will be
replaced with the empty (--) symbol.

Continued... 181

180 © 2011 Leica Microsystems (Switzerland) Limited

 Stereomicroscope Operation: Zoom Iris Aperture control:

There are three methods for changing the

Zoom Iris aperture:

1: Click, hold and drag the Slider to the

left to close the iris, and right to open
it. The aperture value is shown as a
percentage (%) of fully open on the
header bar. The minimum value is
limited to 20%.

2: Click on the Slider Bar and the Slider

will track to the selected position.

3: Click in the Iris Setting text box to

select the existing value and press
the keyboard Delete key to clear it:
Type a new aperture value and press
the Enter key on the keyboard. The
maximum value is 100 and the
minimum value 20.

4: Click on the 'closed iris' icon to close

the iris to 20% of full, or...

5: Click on the 'open iris' to fully open

the aperture.

Continued... 182

LAS User Manual 181

Stereomicroscope Operation: External Illumination control:

The CLS 150XD and 150LS external light

sources may be controlled remotely from
SMS:

1: The Brightness Control may be

rotated to increase or decrease
brightness.

2: The light source may be turned on or

off by clicking the Power button. A red
dot indicates that the source is on.

3: Click, hold and drag the red ‘handle’

on the rotary control, clockwise to
increase brightness, or anti-clockwise
to decrease it. The light output value
is displayed on the header bar.

4: Alternatively, click on the outer rim of

the Brightness Control which will
rotate to the selected position.

Continued... 183

182 © 2011 Leica Microsystems (Switzerland) Limited

 Stereomicroscope Operation: Internal Illumination control:

There are three controls for the Internal

Transmitted Light Base:

CT (Colour Temperature) which

controls the lamp average voltage
and therefore its brightness. This also
affects the light colour.

CCIC (Constant Colour Intensity

Control) which acts like a window
blind, reducing the amount of light
reaching the specimen without
affecting the colour.

Shutter is a toggle action button which

stops light reaching the specimen
altogether.

The CT (1) and CCIC (2) controls work in

combination so for some procedures it will
be necessary to adjust the light colour
using the CT control, and then adjust the
brightness with the CCI control.

Both controls are adjusted in the same

way:

1: Click and hold on the red dot on the

periphery of the control. Drag either
clockwise or anticlockwise to the
desired position and release the
mouse button. Or...

2: Click on the outline of the control and

it will rotate to the selected position.
The CT light colour (k) value and the
CCIC opening (%) are displayed on
the panel header.

3: Click on the Shutter to open it as

indicated by the red dot, or…

4: Click again to close it and the red dot

disappears.

LAS User Manual 183

Stereomicroscope Operation: X/Y Stage Control:

The Motorised Stage is represented on

the control panel as a rectangle with rulers
along the top and left side. Actual X and Y
co-ordinates are displayed in two text
boxes, and the stage initial traverse speed
is selected using one of the three buttons:
Fast
Slow
Auto for precise positioning within the
field of view.

The traverse speed changes automatically

as the stage nears the required position.

To move the stage to target:

1: Select the desired speed by clicking
on the appropriate button. Choose
Fast for longer distances.

2: Double-click within the rectangle

approximately on the X/Y position
required.
The Target Marker will move to the
selected position and the stage will
start to track toward it. As the stage
approaches the Target the traverse
speed will drop to Slow and within the
Target, Auto (precise) will be
selected.
Click in either the X or Y text box and
then use the Mouse Wheel (5) to ‘fine
tune’ the position in 2µm increments.

To move the stage interactively:

3: Click and hold within the stage
rectangle. The 4-Way Arrow
appears.

4: Drag in the required direction. The

Arrow changes to reflect the direction.
The stage will follow the mouse
position until the button is released.
Stage traverse speed automatically
slows as the mouse position is
approached.

5: Click in either the X or Y text box and

then use the Mouse Wheel to ‘fine
tune’ the position in 2ìm increments.

Continued... 185

184 © 2011 Leica Microsystems (Switzerland) Limited

 Stereomicroscope Operation: X/Y Stage Control: Continued:

To move the stage to entered co-

ordinates:
To be used when actual X/Y co-ordinate
values are known. Both co-ordinates are
entered in the same way:

1: Click on the X or Y text box and type

a new value.

2: For positions measured in millimetres,

type ‘mm’ after the value otherwise
the units will default to µm.

3: Press the Enter key on the keyboard.

The stage will go to the entered
positions.

4: If necessary ‘fine tune’ the position in

2µm increments with the Mouse
Wheel after clicking in either the X or
Y text boxes.

5: Optional accessories such as the

Joystick, SmartMove or UMC controls
may be used together with the SMS
controls.

LAS User Manual 185

 Stereomicroscope Operation: Wiring Diagrams (1):

Continued...

186 © 2011 Leica Microsystems (Switzerland) Limited

 Stereomicroscope Operation: Wiring Diagrams (2):

LAS User Manual 187

Stereomicroscope Operation: Wiring Diagrams (3):

188 © 2011 Leica Microsystems (Switzerland) Limited

 Acquire Workflow: DM Microscope Operation:

DM Control provides remote control for all

motorized functions of the Leica DM
microscope series.

If a camera is attached to the microscope, the

camera can be controlled at the same time.

DM Control consists of the following control

control windows depending on the connected
microscope:

Contrast Methods Control 190

Fluorescence Control 191
Illumination Control 193
Objective Nosepiece Control 194
Magnification-Changer Control 195
Focusdrive Control 196
Stage Control 200
Motorised Tube Control 197
Autofocus Control (optional module) 199

The Multi-User Package has been designed

especially for DM microscope users. It allows
users to create profiles that store all of the
hardware settings for a specific task, retrieve
them later and automatically re-configure the
microscope. More about the Multi-User
Package: Go there...

LAS User Manual 189

 DM Microscope Operation: Contrast Methods Control:

1: All available contrast methods for

transmitted and incident light axis are
displayed in the control window.

2: The current contrast method is

highlighted on the control. Each contrast
method can be selected by a left mouse
click.

3: Appropriate contrast methods for the

current objective in the light path are
marked with a triangle . If the selected
contrast method is not valid 'Pseudo
Bright Field' is applied.

Light rings, dark stops, DIC prisms, polarizer

(mot.) and analyzer (mot.) are addressed
automatically if necessary. Mechanical
polarizer and analyser have to be inserted
manually.

Note: For Combi-Contrast FLUO-DIC a

manual analyser has to be inserted in the
appropriate slot on the upper left side of the
stand.

190 © 2011 Leica Microsystems (Switzerland) Limited

 DM Microscope Operation: Fluorescence Control:

1: All learned in Fluo-Cubes are listed in the

control window.

2: The current fluo cube in the light path is

highlighted on the control. Fluo-Cubes
can be selected with a left mouse click.

3: The IL-Shutter can be closed to protect

the sample from bleaching.

To learn in a new Fluo-Cube, please go to the

Setup Workflow.

LAS User Manual 191

 DM Microscope Operation: ExMan and IFW:
Motorized Excitation Manager (ExMan):
Allows the balancing of different fluorochrome
intensities. Suitable for the following Leica dual
and triple filter cube systems: G/R; BFP/GFP;
CFP/YFP; B/G/R, C/Y/R

1: For access to the balancing slider select/

activate the appropriate dual or triple cube
in 'Acquire' and use right mouse key to
open the Excitation Manager Control
Window (2).

Note: This function is only available if the

microscope is equipped with the appropriate
fluo axis.

Internal Fast Filterwheel (IFW):

For fast excitation in red, blue, or green.
Suitable for the following Leica dual and triple
filter cube systems: G/R; BFP/GFP; CFP/YFP;
B/G/R, C/Y/R

3: For access select/ activate the appropriate

dual or triple cube in 'Acquire'. The filter
wheel positions are then directly
accessible.

Note:This function is only available if the

microscope is equipped with the appropriate
fluo axis.

192 © 2011 Leica Microsystems (Switzerland) Limited

 DM Microscope Operation: Illumination Control:

1: The current settings for the active light-

axis are displayed in the control.

2: Depending on which light-axis is

activated (TL or IL), light settings can be
modified.

3: Use left mouse button, the mouse wheel

or the cursor buttons to change the values
for light Intensity (Int), Field Diaphragm
(Fld), and Aperture Diaphragm (Ap)
respectively. For precise adjustment of
the light intensity, the Fine mode is
automatically selected.

4: FIM (Fluorescence Intensity Manager) in

FLUO mode: the intensity of the excitation
light can be reduced in 5 steps to protect
the sample from bleaching.

For COMBI mode (FLUO/DIC or FLUO/Phase,

: use the tabs to switch between the control
panel of TL and IL axis.

Mirrorhouse
To change the fluorescence illumination with
the Mirrorhouse, move the mouse into the
Illumination Window of LAS 'Acquire':

Press: Right Mouse Key.

Select: Mirrorhouse
Select: Light Path

Note: Modifications in the light settings are

stored and automatically recalled when the
microscope in turned on.

LAS User Manual 193

 DM Microscope Operation: Objective Nosepiece Control:

1: All learned in objectives are displayed in

the control window.

2: The current objective in the light path is

highlighted on the control.

3: Objectives which are valid for the selected

contrast method are marked with a
triangle.
Immersion objectives are marked with a
black drop.

Objectives which have been learned in as

combi-objectives (module 'Fine tuning')
are marked with a clear drop.

The selected objective blinks if you are changing the Each objective button shows small status icons:
mode from DRY to IMMersion and vice versa. The
stage is lowered and you have to confirm the change Marks an objective, if it is valid for the currently
of mode with an additional mouse-click. selected contrast-method.

4: Parfocality can be adjusted using the context menu of Marks Immersion-Objectives (Oil, Water,
the right mouse button. This will start the Parfocality Glycerine).
wizard (5). It is recommended that the Parfocality of
all listed objectives is adjusted if new objectives are Marks Combi-Objectives (for use in both modes,
learned in. Immersion- and Dry-Mode).

To learn in new objectives please go to the Workflow Starts the Parfocality Wizard
Setup.

194 © 2011 Leica Microsystems (Switzerland) Limited

 DM Microscope Operation: Magnification Changer Control:
1: The current value of Magnification Changer
is highlighted in the control window.

Since the Magnification-Changer is not

motorized remote control is not possible

Note: The values of the Magnification-Changer

are used for the calculation of the total
magnification (see LeicaScreen).

LAS User Manual 195

DM Microscope Operation: Focus Drive Control:
1: The current position of the Z-Focus is
displayed in the control window.

2: Lower threshold and focus position are

displayed as icons in the control window
and can be recalled. Press the appropriate
button until the threshold or focus point is
reached.
The control window uses the following
status icons:

Focus-Position
Lower-Threshold

3: Please use the right mouse-button to get a

context-menu for additional functions:

Advanced Settings, define/clear the

lower threshold and the focus-position

4: Lower threshold and focus position can be

deleted or set in the Advanced Settings.

196 © 2011 Leica Microsystems (Switzerland) Limited

 DM Microscope Operation: Motorised Tube Control:
1: The current beam-ratio is displayed is
highlighted in the control window. The
beam-ratio for a motorized tube can be
selected.

Each button on the control shows a small

status icon:

The light path is set to 100 % visual

exit (eyepieces).

The light path is set to 100 %

camera port.

The light path is set to 50% / 50%.

Note: After turning on the microscope the light

path is automatically set to 100% visual exit
(eyepieces).

LAS User Manual 197

 DM Microscope Operation: Memory Control
Only available for DM6000.

Allows to store the current combination of

objective and contrasting method (e.g. 20x /D).

1: Using the right mouse key ('settings') the

current combination can be stored...

2: ...available combinations recalled from the

memory.

3: Clear the data on the selected memory

position.

198 © 2011 Leica Microsystems (Switzerland) Limited

 DM Microscope Operation: Autofocus Control
This is an optional module and needs to be
licensed before use.

The cameras supported by digital Autofocus are

listed in the document 'Systems Requirements'.

By calculation the digital Autofocus finds a

reasonable focus plane and adjusts the Z-focus
level automatically. For proper functionality
please adjust the Parfocality values for each
objective.

1: Depending on the samples two appropriate

modes can be selected:

2: Search starts from pre-selected focus:

suitable for sample with approximately the
same thickness. Adjustment of Parfocality
is a prerequisite for this mode.

3: Search starts from the current Z-position:

suitable for samples with variable
thickness.

Within each mode the span of search can be

selected:

4: Near: Search of focus position considers

only the near proximity.

5: Global: Range of search is extended.

Note: For correct operation please install the

newest camera driver. It is highly recommended
to adjust the exposure time of the camera to
values < 10 msec to ensure fast response time.

LAS User Manual 199

 DM Microscope Operation: Stage Control:
The Stage Settings panel contains three tabbed
dialogs that allow the user to initialise and
configure the stage working features.

The tabbed panels are:

Initialise: Sets the top-left (X = 0 and Y = 0)

and bottom-right co-ordinates of the stage
limits. It will not drive beyond these co-
ordinates and all of the other positioning
features are measured from them. Go
there. 201 .. 201

Area: Allows the user to setup a defined

working area within the stage limits. The
stage will not be driven beyond the
boundaries of the user Area. Go there. 202 ..

Positions: Provides the Mode - Live and

Stage to Centre - settings that define how
the stage will be driven to specific points;
Controls the colour and visibility of the Help
Lines - guides that show the stage centre;
If Live Positions will be displayed with a
number or a user description and their
colour, and the nominated camera port for
microscopes with several port options. Go
there. 204 ..

Continued... 202

200 © 2011 Leica Microsystems (Switzerland) Limited

 DM Microscope Operation: Stage Settings:
1: The speed of the stage can be modified by
clicking the control buttons 'Fast' or
'Precise'. The Fast option uses the settings
for pitch entered in:

Setup Workflow > Stage > Fine Tuning Tab

The Precise option drives the stage more

slowly to avoid 'overshoot' and maintain
positioning accuracy.

2: The Stage Settings are accessed by right-

clicking the stage area and...

3: ...clicking to select the Settings option on

the context menu.

Initialise the Stage:

4: Click to select the Initialise tab.

Check that the stage is clear of

! specimens and slides. Make sure the
objectives or magnifiers will not collide
with the moving stage and that the
connecting cables have sufficient slack
to allow the stage unrestricted
movement.

5: Click the Initialise button. The stage will

drive to the top-left corner limit switches to
establish X = 0 and Y = 0. It will then travel
to the bottom-right to set X Max and Y Max
travel.

6: Click Close.

Continued... 202

LAS User Manual 201

 DM Microscope Operation: Stage Settings: Area:
1: The Stage Settings are accessed by right-
clicking the stage area and...

2: ...clicking to select the Settings option on

the context menu.

Area:
Within the extreme limits of the stage
movement, an area that (usually) contains the
image, can be user-defined - the stage will not
drive beyond the boundaries of the setup area.
To create a user area:

3: Click to select the Area tab.

4: Click the Clear Area button to clear any

existing user settings.

5: Drive the stage using the stage manual

controls or SmartMove, to the upper-left
corner of the user area. This is usually set
with reference to the image. Click on the
Step 2 > Store button to save the X/Y
position in memory.

6: Repeat the process but this time driving the

stage to the bottom-right corner of the user
area and click Step 3 > Store.

7: Click the Close button.

Continued... 203

Note: The Area boundary co-ordinates are not

stored permanently and are lost when the
microscope is switched off.

202 © 2011 Leica Microsystems (Switzerland) Limited

 DM Microscope Operation: Stage Settings: Positions:
1: The Stage Settings are accessed by right-
clicking the stage area and...

2: ...clicking to select the Settings option on

the context menu.

Positions:
The Positions tab displays the controls for
setting up the Help Lines (Crosshairs) and Live
Position markers as well as selecting the
current mode:

3: Click to select the Positions tab.

4: There are three Mode options selected by

clicking the appropriate button:

Normal: Disables both Live Positions and

Stage To Centre and hides the Help
Lines. This is normal stage operation.

Live Positions: Allows up to 15

individually selected positions to be
marked on the image. The stage can be
driven directly to a selected position. The
Help Lines are displayed if they are
enabled.

Stage To Centre: Drives the stage centre

directly to a point indicated on the image
by a double-left click. If Help Lines are
enabled they are displayed and
positioned above the point.

Live Positions and Stage To Centre are

mutually exclusive - only one can be
enabled and active. They can also be
selected from the context menu (2) by
clicking the required option.

5: The Help Lines - crosshairs that indicate

the centre of the stage - can be turned on
or off by clicking the checkbox. This is a
toggle action with a tick mark displayed
when the lines are enabled and visible.

Continued... 204

LAS User Manual 203

 DM Microscope Operation: Help Lines and Position Markers:
1: The colour of the Help Lines and...

2: ...that of the Live Position markers can be

changed to suit the user by clicking on the
Colour box and...

3: ...from the Select Colour dialog choosing a

colour from the Basic Colours or...

4: ...dragging the 'target' to the required

position on the colour wheel.

5: Adjust the selected colour intensity by

dragging the slider along the graduated bar.

6: Click OK to save the colour.

7: To display an appropriate description next

to a Live Position marker, click to enable
the User-defined Description check box.
This is a toggle action - click again to turn
off the description display.

Continued... 205

204 © 2011 Leica Microsystems (Switzerland) Limited

 DM Microscope Operation: Camera Port Selection:
The active port option is for microscope models
that have more than one port and allows the
user to select the port that has the active
camera attached.

To select an active port:

1: Click on the small arrows to the right of the
Port header.

2: From the drop-down menu click to select

the port that has the active camera
attached.

3: Click the Close button to close the Settings

dialog.

Continued... 206

LAS User Manual 205

 DM Microscope Operation: Live Positions:
The Live Positions feature allows the user to
select up to 15 individual points on the image
storing each either as a set of X/Y co-ordinates
or as a User Description on a Position List. The
Description option is enabled under Stage
Settings (Go there... 204 ). The stage can then be
driven directly to a point by selecting it on the
Position List and clicking the GoTo button.

1: If the Live Positions option is not already

selected on the Stage Settings > Position >
Mode panel,

2: ...right-click inside the stage panel and...

3: ...from the context menu click to select Live

Positions. A tick mark will appear along
side the option.

4: Click on the small arrows to the right of the

Positions header.

5: On the drop-down list, click to select an

empty position. If necessary use the scroll
bars to extend the list. Positions cannot be
overwritten - they have to be cleared first (
Go there... 210 ).

6: The Store button becomes enabled. Do not

click it yet.

Continued... 208

206 © 2011 Leica Microsystems (Switzerland) Limited

 DM Microscope Operation: Live Positions: Continued:
1: If User-defined Description has been
enabled on the Stage Settings dialog (Go
there.. 204 .)...

2: ...move the stage so that the point of

interest on the image is directly beneath
the Help Lines intersection. (The green
square on the illustration is for indication
only and does not appear in use).

3: Click the Store button.

4: The Position Description entry dialog

appears. Click in the text box and type a
unique name for the for the point.

5: Click the Close button and...

6: ...the Point Number appears over the Help

Lines and therefore, the point of interest,
and...

7: ...the Position Description appears in the

Position List.

Continued... 208

Note: The positions are not stored permanently

and are lost when the microscope is switched
off.

LAS User Manual 207

 DM Microscope Operation: Live Positions: Continued:
1: This sequence applies when the User-
defined Description is disabled.
Two options are available:

Navigate to and store a position, and...

Double-click to store a position.

Navigate and Store:

2: Drive the stage to the point of interest so
that it lies directly beneath the Help Lines -
the centre of the stage.

3: Single-click the Store button. The marker is

placed over the Help Line intersection
and...

4: ... the position is stored as co-ordinates on

the List.

Double-click to Store:
5: Double-left click on the point of interest on
the image - no need to drive the stage to
the Help Line intersection. A marker with
the position number is placed over the point
and its co-ordinates are displayed on the
Position List (4). Empty list positions can be
selected and filled automatically by double-
clicking again over other points on the
image.

6: Up to 15 separate points can be selected

on an image with each being stored on the
Position List.

Continued... 209

Note: The positions are not stored permanently

and are lost when the microscope is switched
off.

208 © 2011 Leica Microsystems (Switzerland) Limited

 DM Microscope Operation: Stage To Centre:
Stage to Centre is a convenient method for examining 2: Double-left click a point of interest on the image...
selected points of interest on the specimen quickly. It drives
the stage immediately to a point indicated by a double-left 3: ...and the stage will move immediately to the centre of
click on the image so that it moves to the centre of the field the field of view.
of view:
4: If required, higher magnification can be selected to
1: Right-click inside the stage area and from the context examine the point of interest in greater detail which will
menu click to select and enable the Stage To Centre stay in focus.
option.
Continue to do this whilst Stage To Centre is enabled
to effectively 'browse' the image.

5: If an empty position on the list has been selected, the

Store button becomes available. Click it to store the
position co-ordinates and return to it later using the
GoTo feature.

To change Stage to Centre settings: Go There.. 203 .

Continued... 210

LAS User Manual 209

 DM Microscope Operation: GoTo and Clear:
GoTo:
The stage can be driven directly to any of the
set positions by:

1: Click on the small arrows to the right of the

Position List header.

2: From the drop-down Position List, click to

select the one required.

3: Click the GoTo button.

4: The stage drives to the position locating it

beneath the Help Line intersection - stage
centre.

5: A fast alternative to driving to a set position

is to click one of the Side Tool Bar buttons.
However, these only respond to the first 5
positions.

Clear:
With a position selected from the drop-down
Position List:

6: Click on the Clear button to remove the

position co-ordinates or point name. The
image marker is also removed. A list entry
must be cleared before it can be used for
another position.

210 © 2011 Leica Microsystems (Switzerland) Limited

 DM Microscope Operation: Adjust Stage Pitch:

Stage movement and accurate positioning

relies upon the Leica Application Suite
software 'knowing' just how far the stage will
travel for a single revolution of a lead screw or
gear train. If tiny manufacturing tolerances
make the travel distance slightly larger or
smaller, then precise alignment of individual
montage images can be compromised.

The Adjust Stage Pitch feature allows the

users to manually position the stage at the
ends of a known distance - usually a
calibration slide - and from the measurement
determine exactly how far the stage travels
for a single revolution of the drive. The value
is stored and used for subsequent stage
positioning.

It is a two-step process - first for the X axis

and then for the Y axis. The measurements
are specific to the fitted stage and must be
repeated if it is changed or swapped.

1: Right-click within the graphic stage area

and... A: Section through a stage lead screw. The manufacturer states a
1.000mm screw pitch - the stage will move 1mm for each turn of the
2: ...click to select the Adjust Stage Pitch screw. If the tolerance is ± 0.002mm then for 25 turns the stage may
option from the drop-down menu. have moved between 24.95 and 25.05mm and perhaps not the
expected 25.00mm.
3: The Stage Pitch Calibration Wizard
appears.

Continued... 212

LAS User Manual 211

 DM Microscope Operation: X Axis Calibration:

1: Place a calibration slide on the stage with

the '0' mark to the left and bring it into
sharp focus. Click the Wizard Start button.
A 'sight' mark appears in the field of view.

2: Drive the stage so that the '0' mark outer

edge perfectly aligns with the sight mark
inner edge (A).
To aid alignment, click the Zoom In button
(4) for a magnified view. Change the sight
mark colour to improve contrast by clicking
the Colour... button (5) and selecting a
preferred colour from the dialog.

3: Click the OK button to capture the stage

position.

6: Drive the stage to the extreme right-hand

end of the calibration slide this time
positioning the inner edge of the calibration
mark with the outer edge of the calibration
mark.

7: Click the OK button.

Continued... 213

212 © 2011 Leica Microsystems (Switzerland) Limited

 DM Microscope Operation: X Axis Calculation:

1: Click inside the Original Pitch X text box and type the 3: The 'OK' button has been replaced with the Restart
calibration slide distance - in the example 1.0mm. button ready to calibrate the Y axis.

2: The X axis calibration is calculated and stored. Continued... 214

LAS User Manual 213

 DM Microscope Operation: Y Axis Calibration:

Follow the same procedure to calibrate the Y axis drive 5: Drive the stage to the end of the calibration slide and
train: again, carefully align it with the sight mark. Click OK
(4).
1: Turn the calibration slide through 90º and check for
position and focus. 6: Click inside the Original Pitch Y text box and type the
calibration slide distance - in the example 1.0mm.
2: Click the Restart button.
7: The Y axis calibration is calculated and stored.
3: Drive the stage to the calibration slide '0' position
carefully aligning it with the sight mark. 8: Click Close.

4: Click OK.

214 © 2011 Leica Microsystems (Switzerland) Limited

 Acquire Workflow: FS Microscope Operation:

This module serves as remote control and status display of all motorized functions of the FS C and FS CB .
If a camera is attached to the microscope, it can be controlled simultaneously.
The module can consist of the following control plug-in windows: (depends on the connected hardware (FS C or
FS Comparison Bridge only):
Objective Magnifications 216
Comparison Bridge Control 217
Illumination Intensity (Cold light sources) 218
Magnification Changer 219
Tube and Photo Port 220
X/Y Stage
Focus Drive

LAS User Manual 215

FS Operation: Objective Magnifications

Objective Magnifications:
This window shows the status of the objectives available and teached-in for both the left and right hand
revolver
turret of the FSC. Switching over between left and right takes place by clicking the corresponding button L / R.
The currently used objective magnification is indicated in red. This is a display function only.
The turret of the FSC is coded but not motorized!

216 © 2011 Leica Microsystems (Switzerland) Limited

 FS Operation: Comparison Bridge Control:

Comparison Bridge Control:

This module enables the direct control of the bridge modes as well as the position and width of the dividing line.
It displays the current status for the a.m. functions and indicates whether the bridge is in the calibrated
(LED = green) or Zoom-mode (LED = red). Four direct control keys are available that switch the comparison
bridge into one of the following modes:
L = full left image (right side = not used)
R = full right image (left side = not used)
LR = Split center image. The dividing line will be positioned to its default position and default width previously
teached-in in DM-control. Later adjustments of either line-position and or line-width result in the switch-off of
the LED in the LR-key. This mode is then called “split image” and no longer “split center”.
MIX = Superimposed image of both, the full left and full right image mode.

The rotary button “Pos.” controls the dividing line position that is used to introduce more or less of the left and
right half-image. In its minimum and maximum settings a full left or full right image and all intermediate positions
can be achieved.
The rotary button “Size” controls the width of the dividing line and can be set from a very thin (almost invisible)
line to a full superimposed image by using the full potential of this function knob. This line or superimposed
strip, can be moved across the image or positioned to any location in the image with the “Pos” key.
The LED “Magnification calibration” indicates two conditions of the comparison bridge:
LED = green, the comparison bridge has identical magnifications left and right taking all optics and objectives
into account. The specified accuracy is less than 1 per mille.
LED = red, the comparison bridge can have different magnifications of the right and left imaging paths by as
much as +/-5%.

LAS User Manual 217

FS Operation: Illumination Intensity:

Illumination Intensity of the cold light sources:

This window allows for the control of two independent cold light sources and displays the illumination intensity
in degree Kelvin. The allocation of the light sources (L & R) depends on the connectors on the rear of the FSC.

218 © 2011 Leica Microsystems (Switzerland) Limited

 FS Operation: Magnification Changer:

Magnification Changer:
The Mag. Changer window allows for the direct control of the 1.5x additional magnification factor to be
introduced at any time with any objective magnification. It acts on both, the eyepieces and the photo port.
After clicking on the desired factor (1x or 1.5x) the current status is indicated in red. It will further be correlated
into the total magnification with the auto-calibration function.

LAS User Manual 219

FS Operation: Tube and Photo Port:

Tube & Photo port:

This is a display only function. The beam splitter has a fixed factor of 50%/50%.

220 © 2011 Leica Microsystems (Switzerland) Limited

 Camera:

The Camera panel provides convenient control

over the functions of a Leica DFC digital
camera ranging from colour balance to
histogram black and white levels.

Images can be acquired in a variety of

sizes, colour depths and file formats to
provide even more flexibility. Setting the
sharpening option and shading reference
acquires images of the highest quality so
that further processing requirements are
minimal.

Leica Application Suite also allows a

focusing region to be defined on a live
image so that areas of significance can be
easily identified and focused more rapidly.

All parameters and configurations can be

saved and recalled at a later date.

All Leica DFC cameras are controlled

from the same interface although the
features available do depend on the
camera type.

Reveal the Camera controls by:

1: Click on the Acquire Workflow and...

2: ...if necessary on the Camera tab.

Using the Slider Controls:

3: Drive a slider control to an approximate

position by clicking on the slider track -
the pointer moves there - and then using
the mouse wheel (if fitted) to fine-tune the
control value.

4: Move a slider in small increments by

clicking and dragging on the slider pointer.

5: Once the slider is selected the mouse

wheel (if fitted) can be used to fine-tune
the control value.

LAS User Manual 221

 Quick Links:

Click on a panel to go directly to the topic: The panels to be displayed or concealed can be set up in
Preferences .

222 © 2011 Leica Microsystems (Switzerland) Limited

 Camera Toolbox: Image Controls:

The Camera Toolbox comprises 9 buttons each of which is

the link to an image control or camera feature.

Click a button for more information:

Automatic and Manual Exposure

Automatic White Balance

DFC Twain tools

Camera and Microscope Linking

High Dynamic Range / Averaging enable

Show Under/Over Exposure

Camera Configurations

Shading Configurations

Pre-defined Camera Setups

LAS User Manual 223

 Camera: Fast Track Exposure:

Often a very acceptable image can be

achieved in seconds by making basic settings
to the exposure using Auto Exposure, and
then fine-tuning the result with white balance.

The following applies to a typical brightfield,

colour image.

Make the basic exposure settings:

1: Set the Saturation to 1.75.

2: Set the Gamma to 0.60.

On the Histogram:

3: Set the black level to 0 and…

4: …the white level to 255 by clicking on

and dragging the sliders.

Run Auto Exposure:

5: Click the Auto Exposure button and then

click again to turn off Auto Exposure.

Set the white balance:

6: Click and drag a Region of Interest

around a white area.

7: Select White Balance from the menu…

…and that is it. Done!

224 © 2011 Leica Microsystems (Switzerland) Limited

 Live Image Display Controls:

Allowing closer and more detailed examination

of the live image before deciding to capture it,
Live Image Display Controls also include an
extended Side Tool Bar and Zoom Level
display.

1: Click on the Acquire Workflow .

2: If necessary click on the Camera tab to

reveal the controls.

3: Image information is displayed in the Live

Bar that appears along the top edge of
the Viewer. When Acquire is selected the
size of the Live Image is displayed.

4: On the Side Tool Bar, clicking the (+)

button will zoom into the image.

5: Zoom out by clicking (-).

6: Zoom to Fit displays the image fitted into

the Viewer window. The image is scaled
so the largest dimension will be fitted to
the Viewer.

7: Same Size displays the image at its actual

size - one camera pixel is represented by
one display pixel. For high resolution
images only part will be displayed and for
low resolution it could appear small and
centred in the Viewer.

8: The Gallery thumbnails are hidden or

revealed in a toggle action if images have
been captured to the current folder.

9: Images zoomed beyond the boundaries of

the Viewer are automatically displayed
with scroll bars top and bottom as
necessary. See also Pan Live Image
226

LAS User Manual 225

 Pan Live Image:

The Pan feature has also been included in the

Live Image Display Controls. It works at any
display level that leaves part of the image
concealed beyond the boundaries of the Viewer
. The ‘hidden’ area can be brought into the
Viewer by manipulating the Pan window.

1: Click on the Pan button.

2: The Pan Window appears. Click on the

header bar and drag the window to any
convenient area on the display.

3: A red outline shows the part of the image

currently displayed on the Viewer. Click
inside the outline and drag it to another
location in the window and the Viewer
display will change to reflect the new
position.

Click on the Pan button (1) again to hide

the Pan Window.

226 © 2011 Leica Microsystems (Switzerland) Limited

 Acquire Image controls:

The are three methods for starting image

capture:

1: Click on the Acquire Image button at the

bottom of the Acquire Workflow.

2: Press the keyboard F3 function key.

3: Click on Options on the Main Menu and

click the Acquire Image option (5).

LAS User Manual 227

 Exposure Adjust:

There are two options for adjusting the

exposure:

229 Automatic with some fine-tuning, and.

..

231Manual with a range of precision

controls.

Before starting work on a live image, using

Automatic Exposure is a good option because
combined with Automatic White Balance it
could produce a perfectly acceptable image
very quickly.

228 © 2011 Leica Microsystems (Switzerland) Limited

 Automatic Exposure Tools:

The software uses current light levels to

establish best values for Brightness,
Saturation and Gamma:

Brightness: A measure of how light or how

dark each colour in the image is. It can
swing from solid black to solid white.
Use small increases in brightness to help
differentiate between colours; too much
and detail begins to disappear.

Saturation: Determines the amount of each

colour that is present. At the highest
setting, each colour will be at its most
vibrant - right hand image - and the
colours cannot be more prominent without
combining to make white.
Use Saturation to achieve colour subtlety
in the image.
Reducing Saturation is a convenient way
of turning a colour image into a
monochrome image - essentially just
shades of grey - without losing detail or
becoming a black solid.

Gamma: A value applied to colour levels to

compensate for different ways in which
the image is viewed. Liquid crystal
displays (LCDs) have a specific Gamma
setting, monitors will have another and
printers yet another.
Changes in Gamma are applied
automatically so for example, when an
image is printed the printer software will
make adjustments before the printing.
Very small changes in Gamma can have
dramatic effects; the examples show a
range of 0.35 to 1.50 with the original in
the centre. Use Gamma to achieve a
contrast ‘match’ to the specimen

LAS User Manual 229

 Automatic Exposure and White Balance:

To apply Automatic Exposure:

1: If necessary reveal the Exposure Adjust

panel by clicking on the arrow right of the
header bar.

2: Click on the Automatic Exposure button.

3: Adjust the Brightness, Saturation and

Gamma controls 229 as necessary to
achieve the required image.

Automatic White Balance:

When Automatic White Balance is applied, all

of the neutral tones - white through grey to
black - are adjusted to remove any ‘colour’
content to maintain a clean, well-defined
image.

4: Click on the Automatic White Balance

button. White Balance is applied to the
entire image.

5: If the image is too dark or too light

Automatic White Balance may fail and an
error message displayed. It may be
possible to lighten or darken the image
with the Exposure Adjust controls or
change the lighting conditions at the
microscope.

6: To undo Automatic White Balance, right

click on the White Balance button and…

7: ...left click the Reset White Balance label.

230 © 2011 Leica Microsystems (Switzerland) Limited

 Manual Exposure:

The image controls change when Exposure

Adjust is in manual mode - Brightness is
replaced by Exposure: Saturation and Gamma
remain but another control - Gain - is added.

If necessary, click the Automatic Exposure

button to ensure it is disabled.

Exposure: Controls the time that the camera

sensing elements are exposed to the
specimen. It is sometimes called the
scanning rate.

At the start of the exposure time period,

all of the camera sensing elements are
reset - they have no usable image
information at all. Then they are exposed
to the specimen and each begins to
‘charge’ to a value that numerically
represents the light falling upon it.
Individual elements are designed to
respond to one of the three colours - Red,
Green or Blue (RGB).

At the end of the exposure period, each

element is ‘read’ and its value used in
combination to create a pixel on the
Viewer.

For any image, there will be an optimum

period of exposure for the elements to
reach values that truly represents the
image (B). Too short an exposure and the
elements will not have sufficient time to
reach the proper value - the image will be
dark and muddy, under-exposed (A). Too
long and the image will be ‘washed out’
and lacking detail, over-exposed (C).

The time scale for exposure will depend

upon the camera and may be measured
in either microseconds(µs), or
milliseconds (ms).

LAS User Manual 231

 Manual Exposure: Gain:

Gain: A function for changing the brilliance

of an image without changing the
exposure. The examples shown are:

Gain = 1.0 (A),

Gain = 1.5 (B) and
Gain = 3.0 (C).

High levels of Gain can make an image

look noisy and badly defined, so as a
general rule leave the setting at 1.0 unless
light levels are particularly low.

Start with a Gain value of 1.0 and

gradually increase the value. Too high a
Gain setting will ‘bleach’ the image, cause
a loss of fine detail and may introduce
‘noise’.

Saturation and Gamma information: 229

232 © 2011 Leica Microsystems (Switzerland) Limited

 Input Options: Camera Selection and Reset:

Input settings cover the camera setup. The

settings are established using several of the
Camera control panels and Preferences.

Users can:

233 Select the Active Camera.

234 Choose and load a Pre-defined
Camera Configuration.
235 Save a User Camera Configuration.
236 Load a User Camera Configuration.
237 Access the Twain User Interface.

Select the Active Camera:

Occasionally, more than one camera each

with its own FireWire connection may be fitted
to the computer. They appear as a drop-down
menu with model names and serial numbers.
The current active camera appears at the top
of the menu list.

To select a camera:

1: Click on the arrow to the right of the Input

Options header to reveal the panel.

2: Click on the small arrows to the right of

the Camera drop down and...

3: ...from the drop down menu click to select

the camera required.

4: To restore a temporarily lost camera

connection, click on the Reset Camera
button.

LAS User Manual 233

 Pre-defined Camera Setups:

Pre-defined Camera Setups are settings for the

most common microscope contrast methods
that a user can quickly select and use.

Pre-defined Setups automatically configure

LAS for image exposure and processing
corresponding to the selected technique.

Select the required technique by:

1: Click on the Pre-defined Setups button and

from the drop down list...

2: ...click to select and apply the required

configuration.

3: Automatic Exposure is automatically

launched to allow the user to make
adjustments if necessary.

4: If adjustments are made to a Pre-defined

Setup it can be saved as an update by...

5: ...clicking on the Save As option and from

the sub-menu...

6: ...clicking on the original setup.

7: To restore Setups to their original settings,

click on the Restore Defaults options.

234 © 2011 Leica Microsystems (Switzerland) Limited

 Save and Delete a User Camera Configuration:

With the camera setup complete, the settings

can be saved and used on another occasion
to perfectly replicate current values.

1: Click on the small arrow to the right of the

Input Options header to reveal the panel.

2: Click on the New Configuration button.

3: On the Save Current Configuration dialog,

click inside the text box and type a new,
unique name for the configuration.

4: Click OK.

To delete the current Configuration:

5: Click on the arrows to the right of the of

the Configuration header bar.

6: From the drop down menu click to select

the configuration to be deleted.

7: Click on the Trash Can (Delete) button

and...

8: ...confirm the deletion on the message

panel by clicking Yes.

LAS User Manual 235

 Load a User a Camera Configuration:

Input settings can be saved as a Configuration

and retrieved later to quickly and precisely
replicate at camera setup.

When the Viewer opens, the configuration

defaults to the Last Used settings.

To Select and Load a previously saved User

Camera Configuration:

1: Click on the arrows to the right of the

Configuration window and...

2: ... from the drop down menu click to

select a saved configuration or...

3: Click on the Select camera configuration

button in the toolbox and...

4: ...from the drop down list click to select

and load a saved configuration.

236 © 2011 Leica Microsystems (Switzerland) Limited

 The Twain Interface:

The Twain User Interface displays settings and tools

associated with the active camera. It is the software
underlying the camera and acquisition functions found in
Leica Application Suite - many of the LAS features can be
adjusted or turned on and off here.

Use the Twain Interface to view the camera data and make
basic exposure settings on a single, compact display.

1: Click on the Camera Settings button on the toolbox.

2: The Twain User Interface appears.

For details about Twain and how to use it 238

LAS User Manual 237

 Twain: Continued:

For a detailed description of Twain and how it is

used refer to the Leica DFC Guides as follows:

1: Click on the Windows Start button and

then on All Programs.

2: From the menu choose Leica DFC Twain

and...

3: ...double-click Image Acquisition or Quick

Start Guide. The Release Notes document
contains latest updates too late to include
in the guides.

238 © 2011 Leica Microsystems (Switzerland) Limited

 Image Formats:

On the Image Formats panel the user can:

239 Select a Colour or Grayscale image.

241 Select Live and Captured image
formats.

Also in this section, how to:

242 Choose the Bitdepth.

243 Enable/disable High Sensitivity Binning

If a colour camera is active, both colour and

greyscale (monochrome) options are
available.

1: Click on the arrow to the right of the

Image Formats header to reveal the
panel.

2: Click on the arrows to the right of the

Image Type header bar...

3: ...and from the drop down menu select

the type required.

Live Format:

Live Format determines the quality and

resolution of the displayed image in theViewer.
The active camera will determine the extent of
the format options available.

To ensure that all of the possible options are

available to the user:

4: Right click on the Live Format text box

and...

5: ...left click on the Show All Modes label to

check it.

LAS User Manual 239

 Image Formats: Format Descriptions:

Depending upon the active camera, there will be a range

of formats available for both live and captured images.

Progressive VGA produces the lowest resolution

images but is suitable for very fast exposure rates.

Colour Binning is a process of grouping adjacent

camera element pixels to create 'super’ pixels. Each
group is then used to 'drive’ a single display pixel.
The feature improves overall sensitivity and speed.

Available binning options depend upon the camera

being used; Three formats - 2 x 2, 3 x 3 and 4 x 4 -
are available, each with a HQ (High Quality) option.
The format numbers describe how the pixel values
are grouped: 2 x 2 = 4 pixel group: 3 x 3 = 9 pixel
group and 4 x 4 = 16 pixel group.

As a guide and depending upon the camera model,

the 2 x 2 Colour Binning 1044 x 772 format is a good
choice for most situations.

Full Frame options display each of the camera

element pixels individually. Resolution and quality is
very high especially if a HQ (High Quality) option is
selected.

Progressive Red, Blue or Green use only the value of

the selected colour. The Viewer displays a grayscale
image representing the intensity of the chosen colour.
Even if the Image Type is set to colour, the image will
appear monochrome.

Avoid saving images using the 16-bit format. They will be

slow to expose and process and if captured and saved in
the same format may be unusable in third-party image
processing applications.

240 © 2011 Leica Microsystems (Switzerland) Limited

 Selecting Live and Captured Formats:

Select the Live Image Format:

1: Click on the arrows to the right of the Live

Format header bar and...

2: ... from the menu click to select a format. If

the camera supports a wide range of
formats, small Scrolling Arrows will appear
at the bottom of the drop down list. Click to
scroll down.

Selecting the Captured Format:

Captured Format determines how the image is

finally captured and saved. In many cases, the
Captured Format will be the same as the Live
Format so, providing the image is saved as a
bitmap on a bit-by-bit basis without
compression, when the image is retrieved it will
be identical to the original.

3: Click on the arrows to the right of the

Captured Format header bar.

4: From the drop down menu click on the

option required.
To save an image with 16-bit colour depth,
a HQ (High Quality) option must be
selected.

LAS User Manual 241

 The Bitdepth:

The Bitdepth is a digital value which determines the

colour range and precision of the saved image. A value of
8 bits provides a spectrum of 256 separate colours
whereas 16 bits yields 65536 colours.

Greyscale images are captured and saved as either 8 or

16 bits per pixel. Colour images require three times the
number of bits per pixel to store the three primary colours
- red, green and blue. So, colour images are stored either
as 3 x 8 bits or 3 x 16 bits (High Quality).

The Bitdepth setting has a considerable effect on the disc

file size so generally 8 bit should be chosen unless more
colour subtlety and variation are needed. The 16 bit
option is not always compatible with some third-party
software.

The Bitdepth setting in Preferences:

The Image Format setting made in Preferences > Save

Images sets the image compression and, in some case
the Bitdepth. They override those made here in Camera.

To check or select the Preferences settings:

1: Click on Options on the Main Header and...

2: ...click to select Preferences.

3: On the Preferences dialog, click the Image tab.

4: Click on the arrows to the right of the In this format

header and...

5: ...click to select the Compression type and

associated Bitdepth. Click OK.

242 © 2011 Leica Microsystems (Switzerland) Limited

 High Sensitivity Binning:

If binning is used for Live Format but not for

the Captured Format, when the image is
retrieved it will appear darker than expected.

To maintain binned live image brightness on

the captured image:

1: Click to check (tick mark visible) the High

Sensitivity Binning check box on the
Exposure Adjust panel.

LAS User Manual 243

 Histogram:

The Histogram is a graphical display of the

colour values in the image represented as 256
points ranging from 0 (Black) through to 255
(White).

In this section:

244 Histogram display options.

245 Checking Under- and Over-Exposure.
246 Automatically cropping Under- and
Over-Exposed areas.
247 Setting the Gamma Level
automatically.

The display detail can be set by the user by:

1: Click on the small arrow to the right of the

Histogram header to reveal the Histogram
panel.

2: Right click on the histogram display

window.

3: From the drop down menu, left click on

Histogram Options and...

4: ...from the display options, click on the

option required:

Combined (A): Shows all of the colours

and the average.

Red, Blue or Green (B): Show that colour

level only.

Average (C): Displays the average of all of

the Red, Green and Blue (RGB) values.

Show Gamma Curve (D): Click to display

or hide the Gamma Curve.

244 © 2011 Leica Microsystems (Switzerland) Limited

 Over and Under-Exposure:

The ‘one-click’ Show Under/Over Exposure

feature is a fast indication of those areas of
the image that are not exposed properly - and
probably not adding to the image quality.

1: To check over- and under- exposure, click

on the Show Under/Over Exposed button.

On the Histogram widow two coloured

panels appear at the extreme ends of the
display. The blue panel indicates under-
exposure at the 0 (Black) end, and the red
panel over-exposure at the 255 (White)
end.
The graphics are also applied to the live
image so...

2: ...this image shows blue as under

exposed areas and red as over exposed
areas.

LAS User Manual 245

 Auto Black and Auto White:

Because under- and over exposed areas of

the image do not usually contribute to its
quality, those levels may be ignored without
detriment.

Auto White and Auto Black automatically

‘crops’ either or both under- and over exposed
levels.

1: Right click on the Histogram window.

2: From the drop down menu select the Auto

White or Auto Black option. The selection
becomes checked and active. Click again
to uncheck and disable the option.

3: Beneath the Histogram window, the slider

corresponding to the white or black option
moves to reflect the ignored light levels.

4: To reset the black and white values, right

click on the Histogram window and from
the drop down left click on the Reset
Black/White Points option.

Auto White and -Black are normally switched

off.

246 © 2011 Leica Microsystems (Switzerland) Limited

 Auto Gamma:

The Auto Gamma option sets the gamma level

based upon the active light levels. Under- and
over exposed levels are not included if they
have been ‘cropped’ either manually or
automatically.

1: Right click on the Histogram window.

2: From the drop down menu select the Auto

Gamma option. The selection becomes
checked and active. Select again to
uncheck and disable the option.

3: To reset the gamma value, right click on

the Histogram window and from the drop
down left click on the Reset Gamma
Value option.

Whilst Auto Gamma is enabled, the Gamma

control on the Exposure Adjust panel is
disabled.

Auto Gamma is normally switched off.

LAS User Manual 247

 HDR & Averaging:

Images that exhibit high contrast between the

light and dark areas, have posed an on-going
problem to the microscopist - capture to
maintain the dark detail and the light
information is badly exposed and probably lost.
Expose for the whites and the dark detail is
compromised.

Leica High Dynamic Range (HDR) provides a

fast solution by automatically capturing a
number of images each at a different exposure,
and then combining them digitally into a single
image that balances the contrast range. Detail
across the entire image is retained and clear.

Users can choose between automatic HDR that

handles the processing and saving in a simple,
fast single step, or a manual operation that
allows some fine-tuning before the image is
saved.

Additionally, an Averaging feature is available

that can reduce the amount of noise in an
image. Reducing noise can enhance the final
image either making it look better or making it
more suitable for analysis.

Continued 249

248 © 2011 Leica Microsystems (Switzerland) Limited

 Setup for HDR and Averaging:

Before using HDR or Averaging:

Set the capture details in Preferences .

Select a capture folder in Browse 299 .

Check that required HDR function is

enabled. If not:

1: Right-click on the HDR button.

2: From the drop-down menu left-click to

select either HDR or Averaging.

3: The button caption displays the selected

function - HDR or AVG (Averaging).

Adjust the specimen illumination so that it

is even, reduces high-lights, reaches into
crevices and does not cast shadows.

Use the Fast Track Exposure sequence

224 and aim for a sharp and reasonably
exposed image.

250 Link to Automatic HDR.

251 Link to Manual HDR with Preview.
254 Link to Averaging.

LAS User Manual 249

 Automatic HDR:

The Automatic HDR feature captures the

range of images to memory, creates the HDR
image and saves it to the hard drive in the set
capture folder.

All of the settings are controlled by the

software - users do not have the facility to
select the Sample Contrast or adjust the
Brightness before the HDR image is saved,
but can use the Enhance controls on the
Process Workflow to make adjustments to it
after capture.

Previous settings made using the Preview

function are ignored.

1: If necessary click to disable Automatic

Exposure and...

2: ...click on the HDR button.

3: On the HDR / Averaging control panel,

click to enable (tick mark visible) the
Automatic check box.

4: Click the Acquire Image HDR button or

function key F3.

5: The Acquisition Progress bar appears

whilst the images are being captured to
memory and the HDR image created.

6: The final HDR image is stored in the

capture folder on the hard drive and...

7: ...a thumbnail is created in the Gallery.

250 © 2011 Leica Microsystems (Switzerland) Limited

 Manual HDR:

In manual mode, the Preview facility is used to

optimise the appearance of the image by
interactively adjusting the contrast range and
brightness of the HDR image. These settings
are then used for actual image acquisition
while HDR is in manual mode.

1: If necessary, click to disable Automatic

Exposure and...

2: On the Side Tool Bar turn on Dual Viewer

so that the live image can be compared
with the HDR processed image. The live
image is on the left and the Preview
image on the right.

Users will find it convenient to select the

After Capture Do Nothing option in
Preferences 49 to remain on the
Acquire Workflow after an image is
captured.

3: Click on the HDR button.

4: On the HDR / Averaging control panel,

click to enable (tick mark visible) the
Preview check box.

Continued 252

LAS User Manual 251

 Manual HDR: Sample Contrast:

Sample Contrast allows the user to select the

exposure values either side of a fixed
exposure point.

The additional exposure times are determined

by the Sample Contrast setting - Low, Medium,
High and Very High. Each option applies
different factors as follows:

Low Contrast: -1 | Centre | 1

Medium Contrast: -1.5 | Centre | 1.5
High Contrast: -2 | Centre | 2
Very High Contrast: -3 | -1.5 | Centre | 1.5 | 3

The Very High Contrast option captures 2

additional images either side of the centre
exposure.

Users should experiment to find the best level

for the image.

1: Click on the arrows to the right of the

Sample Contrast header and...

2: ...click to select the required contrast

level.

Continued 253

252 © 2011 Leica Microsystems (Switzerland) Limited

 Manual HDR: Acquiring the Preview:

Continued from the previous page 251

1: Click the Acquire Preview button.

2: The Acquisition Progress bar appears. A

range of images at different exposures is
being created in memory and then
combined to produce...

3: ...the HDR image with balanced contrast

displayed in the Viewer right-hand pane.

4: If necessary finely adjust the image

brightness.

5: To save the image which is currently only

memory, click on the Acquire Image (HDR
) button or the F3 function key.

6: The progress bar appears again and the

HDR image is saved to the capture folder
with...

7: ...a thumbnail in the Gallery.

The HDR settings derived from Preview

remain active for any further captures unless
changes are made to Sample Contrast or
Brightness.

LAS User Manual 253

 Averaging:

Noise can compromise the level of detail in

digital images, particularly in regions where the
amount of light is low or a higher gain setting
has been used. Reducing noise can enhance
the final image either making it look better or
making it more suitable for analysis. Some
techniques to reduce or remove noise soften
the image as well.

The HDR panel includes a technique that

averages multiple exposures to reduce noise.
Averaging can reduce noise without reducing
detail, because it actually increases the Signal-
to-Noise Ratio (SNR) of the image and may
retain detail by increasing the bit-depth of the
image. Images captured using Averaging tend
to take a little longer to acquire.

Image averaging works on the assumption that

the noise in the image is truly random and
fluctuations above and below actual image data
will gradually even out as more and more
images are averaged.

1: If necessary, click to disable Automatic

Exposure.

2: Click the AVG button. To select the AVG

function 249 .

3: On the HDR / Averaging control panel all

of the controls are disabled except for the
Averaging Image Count slider.

Continued 255

254 © 2011 Leica Microsystems (Switzerland) Limited

 Averaging: Continued:

Continued from the previous page 254

1: Click and drag the Average - Image Count

slider to the left (reduce) and the right
(increase) the number of exposures to be
tested.

2: Click on the Acquire Image HDR button or

the F3 function key.

3: The Acquisition Progress bar appears as

the range of exposures is checked.

4: The exposure with the best signal-to-noise

ratio - least noise - is saved to the capture
folder and a thumbnail appears in the
Gallery.

LAS User Manual 255

 Processing:

The Processing panel provides tools to

improve quality and orientation primarily for a
captured image but may be used for live
images as well.

257 Shading: Corrects light level variations

that often occur due to bright spots
caused by the microscope light source
and the optics.

260 Sharpening: Enhances the edges of

indistinct features on the image making is
clearer and crisper.

261 Flip Horizontal and Flip Vertical: Re-

orientates the image, top to bottom or side
to side.

256 © 2011 Leica Microsystems (Switzerland) Limited

 Shading:

Shading is the name given to variations in the

background light level across an image.

The examples show transmitted light through

a microscope. On the left, the light source and
the optics conspire to create a bright spot in
the centre of the image which gradually
becomes less and less bright toward the
edges.

Even ‘illumination’ on live images can be

achieved in software by applying a ‘blank area’
value to the entire image area. The effect is
shown in the right image.

The light source and any of the optical

elements will produce different shading levels,
so each microscope element combination
should have its own shading setting.

1: If Linking > Shading is enabled it takes

precedence and Processing > Shading is
disabled - greyed out on the Processing
panel - and...

2: ...the Apply to Live check box is available

to allow current settings to be applied to
the live image.

LAS User Manual 257

Creating a Shading Reference:

Make sure the image is sharp and well lit.

Move the stage to a clear, unblemished

area of the image – introduce a very small
amount of de-focus if the chosen area is
not completely clear.

Over-exposing slightly can also help to

‘remove’ blemishes.

Use the Show Over/Under Exposed

control (A) on the toolbox and adjust the
Exposure (B) until just a few red pixels
are visible or...

Replace the specimen slide with a plain

slide (and cover slip if used) of the same
type.

1: Expand the Processing panel if necessary

by clicking on the arrows to the right of
the Processing header bar.

2: Click on the New Shading button.

3: Type a unique name for the shading

reference in the text box.

4: Enable the Overwrite if Exists button to

replace an existing shading reference.

5: Click OK.

6: Click on the arrows to the right to the

Shading header and the new shading
reference is included in the drop-down list.

258 © 2011 Leica Microsystems (Switzerland) Limited

 Select and Load a Shading Reference:

There are two methods for selecting and

loading a Shading Reference:

1: Click on the Toolbox Shading button

and...

2: ...click to select and load a reference from

the drop-down list. It will be indicated by a
tick mark to the left.

3: On the Processing panel, click on the

arrows to the right of the Shading header.
The drop-down list of references appears.

4: Click to select and load a reference.

LAS User Manual 259

 Sharpening:

Sharpening is a software function in which the

boundaries between tonal values are
enhanced. Use it to improve the clarity and
crispness of indistinct detail on an image.

The level of enhancement is selectable

between Low and Robust (very high) but too
much sharpening can make the image appear
grainy and speckled. It is a fast process so the
best approach is to start with the Low setting
and work toward Robust step-by-step.

1: Click on the arrows to the right of the

Sharpening header.

2: From the drop down menu select either

Off to turn off sharpening, or the level
required.
Repeat the process choosing another
level if the result is not suitable.

The illustrations show the original image

(A) and Medium sharpening (B).

260 © 2011 Leica Microsystems (Switzerland) Limited

 Flip: Horizontal and Vertical:

These two options - Flip Horizontal and Flip

Vertical - re-orientate the image. Flipping is
often used to emulate the view through the
eyepieces.

To flip the image from side-to-side:

1: Click on the Flip Horizontal check box.

Click again to return it to its original
position.

To flip the image from top-to-bottom:

2: Click on the Flip Vertical check box. Click

again to return it to its original position.

LAS User Manual 261

 Region of Interest:

A Region of Interest (RoI) is created by

drawing a rectangle on an image. The area
within the rectangle - the Region of Interest - is
then the target of several special functions, the
image outside the region being ignored.

Five special functions may be applied to a

Region of Interest:

262 White Balance: Displays all of the

neutral tones as shades of black and
white but only within the Region of
Interest.

265 Spot Exposure: Automatic exposure is

applied to the entire image but using only
the values contained within the RoI.

266Find Focus: Defines a moveable

Region of Interest in which to focus.

264Region of Interest: When the image is

acquired, only the part within the RoI is
captured - the rest is discarded.

267Check Colour: Used to adjust the

image overall colour balance by using a
small region as the reference for both Hue
and Saturation.

There can be a different region for each of the

five functions and with the exception of White
Balance they are stored and can be recalled
by clicking to enable the check boxes on the
Region of Interest control panel. Each RoI has
a different identifying outline colour.

Create a new region for any of the options at

any time to overwrite the existing one.

White Balance has an immediate effect as

soon as it is selected. A new region has to be
created each time it is used.

262 © 2011 Leica Microsystems (Switzerland) Limited

 Creating a Region of Interest:

Create a Region of Interest by:

1: On the image, click on a corner of the

region and...

2: ...holding down the mouse button drag to

the opposite corner. The region is drawn
on the image with small scaling 'handles'
at each corner and mid-way along each
side. It is not necessary to be precise
because the region can be re-sized and
moved later.

3: Release the mouse button and the

contextual menu appears listing the five
RoI options. The new RoI can be either:

White Balance,
Spot Exposure,
Find Focus,
Cropping Area (Region of Interest) or
Check Colour.

Click to select.

If none of the options is chosen the region

will default to the Region of Interest which
represents the capture area.

A separate RoI of differing dimensions

and position can be created for each of
the options. The location and dimensions
are stored.

As the mouse cursor approaches the region,

the scaling handles re-appear:

4: To stretch the region either vertically or

horizontally, click on a handle. The cursor
changes to a double-ended arrow.
Holding down the mouse button drag the
handle to re-size the region.

5: Scale the region proportionally by clicking

on a corner handle - the cursor changes
to a double-ended arrow - and holding
down the button dragging it diagonally.

6: Move the entire region by positioning the

mouse over an edge. The cursor changes
to a hand. Click and holding down the
cursor button drag the region to a new
position.

LAS User Manual 263

 Crop to Region of Interest:

This feature will capture only that part of the

image within the Region of Interest. The rest
of the image is ignored.

1: Click on the small arrow to the right of the

Region of Interest header to expand the
control panel.

2: If necessary, click to enable the Crop to

RoI check box - a tick mark visible. The
Region of Interest appears green on the
image and can be re-sized and positioned
as described on the previous page.

3: Click on the Acquire Image button or

press the keyboard F3 key.

4: Only the area of the image within the RoI

is captured and scaled to suit the Viewer.

A thumbnail is displayed in the Gallery.

Depending upon the After Capture setting
in Preferences the program will move
to another Workflow or remain where it is.

264 © 2011 Leica Microsystems (Switzerland) Limited

 Spot Exposure:

Spot Exposure will automatically adjust the

exposure of the entire image using only the
area with the Region of Interest.

The RoI can be moved around the image to

compare different exposure results.

1: Click the arrow to the right of the Region

of Interest header to expand the control
panel.

2: Click to enable the Spot Exposure check

box. A Region of Interest is drawn with a
magenta outline. Re-size or move the
region as described previously.

3: The Toolbox Auto Exposure button is

automatically enabled and...

4: ...the image exposure is adjusted using

the values in the RoI.

LAS User Manual 265

 Find Focus:

This feature displays the focus precision for

the area of the live image contained within the
Region of Interest.

The region may be moved around the image

to compare different areas with a focus bar
indicating the ‘best’ level achieved.

1: Click the arrow to the right of the Region

of Interest header to expand the control
panel.

2: Click to enable the Find Focus radio

button. A Region of Interest is drawn with
a yellow outline. Re-size or move the
region as described previously.

3: The level bar moves left to right to

indicate the focus precision. The further
the bar is to the right, the better the focus.

4: To check the focus at another point on the

image, click and hold on the yellow outline
(not a ‘handle’), and drag the RoI a new
position.

5: Turn off Find Focus by clicking the Off

button. Click the Find Focus button (2) to
reveal the Region of Interest at its last
size and position.

266 © 2011 Leica Microsystems (Switzerland) Limited

 Check Colour:

The Check Colour function on the Region of

Interest panel is a convenient control for
creating a region that can be used with the
Check Colour feature.

1: Click the arrow to the right of the Region

of Interest header to expand the control
panel.

2: Click to enable the Check Colour check

box and create a Region of Interest as
previously described. A region is drawn
with a blue outline. Re-size or move the
region as necessary.

3: With Check Colour enabled and the

Check Colour feature launched 296 , the
region is represented by...

4: ...a small dot on the colour wheel which

represents the Hue and Saturation levels
inside it.

LAS User Manual 267

 Calibration Settings:

Calibration ensures that measurements

indicated by the software are given in real
world units taking into account the selected
optical magnification of the microscope and
the size of pixels of the digital camera.
Five calibration options are available
depending upon the level of precision
required:

269 None: Calibration not required -

distances are measured in pixels..

270 Calculated Default Configuration:

Based upon the software's 'knowledge' of
the microscope optical components and
the pixel size of the digital camera being
used. This is the quickest way of
establishing a reasonable but
approximate calibration as it does not
make any checks against a calibration
slide.

271 Calculated User Configuration: Using

a calibration slide on the stage the
calibration of a single, greatest
magnification objective is measured and
from this all the other objective/mag
changer combinations are derived. This is
more accurate than simply using the
Configuration Default.

274 Measured Calibration: Again, a

calibration slide placed on the stage is
used, but all objectives in combination
with all Mag changer settings are
measured. A 'Wizard' helps to speed the
process and the result can be a high level
of calibration precision.

279 Automatic Calibration: Speeds the

Measured Calibration process by using
the software to detect and measure the
calibration slide. The user does not have
to make any measurements.

268 © 2011 Leica Microsystems (Switzerland) Limited

 No Calibration:

If calibration is not required then one display

pixel = one camera pixel.

All measurements are reported in pixels:

1: Click on the arrow to the right of the

Calibration Settings header to reveal the
Calibration panel.

2: Click on the arrows to the right of the

calibration Type header and from the drop
down menu…

3: …click on the None option.

LAS User Manual 269

 Calculated Calibration: Default Configuration:

Calculated Calibration with the Default

Configuration uses the microscope optical
data and camera pixel size to create a
calibration value. This is used to return
measurements in the type selected – Microns,
Millimetres etc. It is a very fast and moderately
accurate method of setting the calibration.

1: Click on the arrows to the right of the

Type header.

2: From the drop down menu click to select

the Calculated option.

3: On the Configuration header click to the

arrows to the right and…

4: …click on the Default option.

5: Select the measurement type by clicking

on the arrows to the right of the header.

6: Click to select the measurement units

required.

270 © 2011 Leica Microsystems (Switzerland) Limited

 Calculated Calibration: User Configuration:

User Calibration calculates calibration values

by making an on screen measurement across
a Calibration Slide placed on the microscope
stage.

Only one calibration is taken with a single

objective/mag combination, the resulting value
is than applied automatically to all other
combinations using the microscope optical
data.

The objective with the greatest

magnification must be selected and Mag
changer set at 1.

For Stereo- and Macroscopes ensure that

that the zoom position is selected at one
of the click stops listed in the zoom
magnification values.

Make sure the Calibration Slide is properly

focused and corresponds to the
measurement units selected in Step (2).

1: Click on the arrows to the right of the

Type header and select Calculated from
the list.

2: Click on the arrows to the right of the

measurement type header and from the
drop down list click to select the
Calibration Slide measurement units.

3: Click the New button.

4: On the Calibration Configuration dialog,

click in the text box and type an
appropriate name for the calibration.

5: Click OK.

LAS User Manual 271

User Configuration: Continued:

1 & 2: Click on the red measurement line

that appears in the Viewer and drag it to
sit over the calibration slide.

Click on the ‘handles’ at either end of the

measurement line and drag them so that
they precisely align with selected marks
on the slide.

Greater accuracy is achieved if both of the

line end strokes are aligned either to the
left or right of the marks. In the illustration
the measurement line represents a
distance of 300 microns.

3: Click inside the Actual length of line text

box type the length of the measurement
line – in this case 300 microns.

4: Click Save.

272 © 2011 Leica Microsystems (Switzerland) Limited

 User Configuration: Continued:

1: When the Save button is clicked the value

in the Actual length of line text box may
change to reflect settings made on the
Acquire > Scale Bar panel. This does not
affect the calibration.

2: Calibration settings can be retrieved and

applied by clicking on the arrows to right
of the Configuration header and from the
drop down list…

3: …clicking to select the Configuration

Name.

4: Delete a configuration by selecting it (as

above) and then clicking the Delete
button. On the Delete Configuration dialog
confirm the deletion by clicking Yes.

LAS User Manual 273

 Measured Calibration:

Measured Calibration provides the most

precise results because every objective/Mag
combination is individually calibrated using a
calibration slide. A Wizard automatically steps
through the combinations.

Place a calibration slide on the stage. It is

important that the slide is clean and correctly
focussed.

1: Click on the arrows to the right of the

Type header and...

2: ... from the drop down list click to select

the Measured option.

3: Select the measurement units by clicking

on the arrow to the right of the header
and…

4: …clicking to select the measurement

type. This must correspond with the
calibration slide that is going to be used.

5: Click the New button.

On the Calibration Wizard dialog two options

are available:

275Manual: The user is responsible for

placing a measurement line across a
known distance on a calibration slide..

279 Automatic: The software detects and

measures the Calibration Slide and
automatically applies the calibration
values.

274 © 2011 Leica Microsystems (Switzerland) Limited

 Manual Measured Calibration:

On the Measured Calibration Wizard dialog:

4: …from the drop down menu click to select the 1
1: Click to select the Manual option. option. For the first pass, every objective will be
calibrated using the 1 Mag Changer. Subsequent
2: Click in the Name text box and type an appropriate passes use the next incremental Mag Changer value
name for the calibration. The calibration values will be with all of the objectives.
stored in a separate file using this name.
5: Click on the Start button.
3: Click on the arrows to the right of the Mag Changer
header and…

LAS User Manual 275

 Apply Measurement:

On the Measured Calibration Wizard dialog:

than trying to align exactly with the centre of the mark.
1: The first objective is automatically selected. Press and hold down the Shift key to display the
Magnifier. Drag it over the calibration slide to help with
2: A measurement line is displayed on the Viewer: Click, the alignment.
hold and drag it so that it is positioned over the
calibration slide. 3: Click inside the Actual length text box and type the
Click and drag the small handles at each end of the distance measured – in the example 300 microns.
line so that it extends to the distance required – in this
case 300 microns. Position both the end stokes to 4: Click on the Apply button.
either the left or right of the division marks – easier

276 © 2011 Leica Microsystems (Switzerland) Limited

 Next Measurement:

1: Click on the Next button.

4: Click Apply.
2: The next objective is selected.
Repeat this procedure for each of the objectives.
3: Repeat the measurement line procedure described on
the previous page.

LAS User Manual 277

 Next Mag:

With the last objective in combination with the Mag 1 Repeat the calibration sequence described on the previous
complete, click the Next button. pages.

Move through all of the Mag Changer values in the same

way until all the combinations have been calibrated.
1: Click on the arrows to the right of the Mag Changer
header and… The configuration and calibration settings must now be
saved 285 .
2: …click to select the next Mag Changer value – in the
example 1.5.

3: Click Restart and the first objective will be selected

again.

278 © 2011 Leica Microsystems (Switzerland) Limited

 Automatic Measured Calibration:

Automatic Measured Calibration offers a fast,

precise method using a calibration slide and
allowing the software to detect it and apply the
values. The user is not involved in any
measurements but an active and licensed
option measurement module must be
installed.

The software can accurately and automatically

detect the slide image and calculate the
calibration from a known interval between the
divisions, providing the image is sharp and the
division lines clearly defined.

Place a calibration slide on the stage. Is is

important the slide is sharply focussed with
distinct divisions visible, clean and clear of
debris for the calibration process to be
successful.

Select high resolution for the camera

capture format: 240 .

Choose an objectives greater than x2.5

on a compound microscope if using a
10µm/division calibration slide.

1: Click on the arrows to the right of the

Type header and...

2: ... from the drop down list click to select

the Measured option.

3: Select the measurement units by clicking

on the arrow to the right of the header
and…

4: …clicking to select the measurement

type. This must correspond with the
calibration slide that is going to be used.

5: Click the New button.

LAS User Manual 279

 Setup:

On the Measured Calibration Wizard:

3: Click inside the Slide Interval text box and type the
1: Click to select Automatic. interval value of the calibration slide - that is the
distance between two adjacent divisions in the
2: If necessary, change the measurement units for the selected measurement units. Typically, this will be 10µ
calibration slide by clicking the arrows to the right of m for a compound slide and 100µm for a stereo
the header and selecting from the drop-down list. calibration slide.

The measurement units for the slide and calibration 4: Click on the arrows to the right of the Mag Changer
do not have to match. header and...

The calibration slide does not have to be perfectly 5: ... select 1 from the drop-down list.
horizontal or vertical, but the closer it is then the faster
the detection. 6: Click inside the Name text box and type a unique
name for the calibration configuration.

280 © 2011 Leica Microsystems (Switzerland) Limited

 Calibration Detect:

1: Click the Start button and... 6: If a scale is not detected or does not conform to the
verification parameters, an error message appears.
2: ...the first objective is selected. The message changes to reflect the error.

3: Click th Get Calibration button. 7: Sometimes detection can be improved by re-

focussing. Click on the Show Live button to view the
The software will now try to find a calibration slide and live image and adjust the focus.
verify that it is precise and suitable as a calibration source:
Calibration slide scale verification parameters can be
4: If a calibration scale is found and verified, a coloured changed: 283 .
overlay is applied to the division strokes on the scale,
a new calibration value is automatically calculated Change the calibration scale detected colour: 284
and...

5: ...the Apply button becomes active. Check that the

calibration intervals have been properly detected and
click the Apply button to apply the new calibration
value for the mag/objective combination.

LAS User Manual 281

 Next Mag:

With the calibration complete for the selected

mag/objective combination: 6: Click the Restart button and the process will begin again for the
new mag value.
1: Click the Next button.
7: When all of the mag/objective combinations have been
2: The next objective is selected. calibrated, save the configuration and values: 285

3: Click Get Calibration and the detection A calibration configuration can be edited to adjust individual
process will begin again. combinations: 286 .

4: Click the Apply button if the detection is The Mag/objective combinations together with the calibration
successful. settings can be printed: 287

Continue this way until all of the objectives have

been calibrated and then:

5: Click on the arrows to the right of the Mag

Changer header and from the drop-down
select the next magnification.

282 © 2011 Leica Microsystems (Switzerland) Limited

 Calibration Settings:

The verification parameters check that a

calibration slide image is sufficiently accurate
to be used as a calibration source. For
example, random fibres on an calibration slide
image could be interpreted as part of the scale
and have to be 'filtered' out.

Users can change the settings but should be

aware that significant changes can result in
compromised calibration accuracy. If in doubt
revert to the factory optimised defaults.

1: On the Measured Calibration Wizard click

on the 'spanner' icon to reveal the Auto
Calibration Settings dialog.

2: The Patch size refers to the spread of

pixels leading to a discernible edge. In the
illustration there are several pixels ranging
from white to dark grey before the black
central 'edge' appears. Increasing the
patch size could 'create' spurious edges.
Keep the patch size as small as possible.

3: Angle tolerance determines how much the

angle between two adjacent scale strokes
can vary from the parallel. The software is
looking for a series of parallel strokes at a
consistent 'pitch' or interval.

4: The interval of the scale strokes must be

close to the value entered by the user.
The Variation threshold determines how
much it can be allowed to vary.

5: Outliers are scratches and debris that

may be present on the image and could
be interpreted as part of the scale. The
Outlier acceptance sets the % level at
which the interval mean (a central
'average' for all of the detected intervals)
can vary. Strokes falling below the Outlier
acceptance are removed.

6: If changes are made to the settings click

the Apply button to save them.

7: To restore the factory settings click the

Defaults button.

LAS User Manual 283

 Calibration Colour:

Change the colour of a detected calibration

slide image by:

1: Click on the Colour button on the Slide

Interval panel.

2: From the Windows Colour dialog choose

a colour by clicking a swatch, dragging the
crosshairs on the wheel or typing Red,
Green and Blue (RGB) values.

3: Click OK.

284 © 2011 Leica Microsystems (Switzerland) Limited

 Save Calibrations:

1: When all objective/Mag combination calibrations are 2: If the file already exists confirm overwriting the
complete, click the Save button to store the existing calibration file by clicking the Yes button on
calibrations. the Warning message.

3: Alternatively, click inside the Name text box and type

a new, unique name and click Save again.

LAS User Manual 285

 Edit:

The Edit feature allows selected Mag/objective

combinations to be calibrated without having to go through
the entire Wizard sequence, especially useful if an Extend the measurement line over the Calibration
objective is changed. Slide or allow the software to detect it (Automatic
calibration only).
1: Click on the Edit button and the Measured Calibration
Wizard appears with un-calibrated objective(s) Type the actual value.
highlighted and the Actual length value reset to 1.00.
Click Apply.
To calibrate a Mag/objective combination, use the
procedure already described: Save and exit the calibration Wizard at any time.

Select a Mag Changer value..

Click to select an objective.

286 © 2011 Leica Microsystems (Switzerland) Limited

 Printing Calibration Settings:

A complete list of all objective/mag

combination calibrations can be displayed and
printed by:

1: Click on the Print button. The Calibration

Sheet is displayed on the computer’s
default browser.

Use the browser's print button to make a

hard copy.

LAS User Manual 287

 Linking:

Exposure Linking associates a microscope

setup with a specific camera setup, whereas
Shading Linking associates a microscope
setup with a specific image shading level.

288 Exposure Linking.

291 Shading Linking.

With Exposure Linking enabled, the Leica

Application Suite automatically checks the
major microscope settings against a
previously stored Linking list and, if there is a
match will retrieve and load the camera
settings associated with it.

The Linking list is created by the user - it is not

a preset part of the Leica Application Suite.

The microscope settings checked are:

Objective or zoom level for stereo

microscopes,
Mag changer,
Camera and port,
Filter and...
Contrast method.

A Link may be created for all of these items in

any combination making it a really powerful
tool for precise repetition.

288 © 2011 Leica Microsystems (Switzerland) Limited

 Exposure: Create a Link:

To create an Exposure Link the microscope

must be setup for the image required.

1: Click to enable the Link check box.

2: The Status Display on the header bar will

show RED.

This indicates there is no stored camera

information for the current microscope
settings.

3: The Linking button on theToolbox will be

enabled - white against a black ground.

4: To create a link, click on the Linking

button.

5: The Status Display will turn GREEN to

indicate an established link.

To turn Exposure Linking on or off:

6: Click to disable (no tick mark visible) the

Link check box.

LAS User Manual 289

 Link Update or Delete:

If changes are made to the camera settings -

in the example the Exposure value has been
reduced -

1: ...the Status Display will change colour to

YELLOW.

2: The Linking button also becomes enabled

again. Click it to update the link.

If the Exposure setting is changed but before

the link is updated, the original exposure can
be retrieved by:

3: Right-clicking the Linking icon and then

left- clicking on the Revert tag.

On stereo microscopes an additional warning

may occur - the Status Display on the header
bar changes to BLACK (4) indicating that the
zoom level has changed but is being ignored.
All other settings are correct. Update the link
to include the revised zoom by clicking the
Linking button (2).

Deleting Links:

5: All of the links for the current camera can

be removed by clicking the Delete button
and...

6: ...confirming the deletion.

This is not reversible: Use with care.

290 © 2011 Leica Microsystems (Switzerland) Limited

 Shading:

Shading Linking associates a specific

microscope setup with a specific shading
level.

Shading is the name given to variations in the

background light level across an image. In the
example, the left-hand image represents
transmitted light through a microscope. The
light source and the optics conspire to create a
bright spot in the centre of the image which
gradually becomes less and less bright toward
the edges.

Even ‘illumination’ can be achieved by

electronically applying a blank area value to
the entire image area. The effect is shown in
the right-hand image.

The light source together with any of the

optical elements will produce different shading
levels, so each microscope element
combination should have its own shading
setting.

Shading Linking when enabled, checks the

microscope:

Objective or zoom level (for stereo

microscopes),
Mag changer,
Camera and port and
Contrast method.

...and automatically tries to find and apply a

matching Shading Link. If a match is found the
status icon on the Linking header bar will be
GREEN. Without a match it will be RED and
no shading settings applied.

Shading Links are not supplied as presets -

they have to be created by the user.

LAS User Manual 291

 Shading Snapshot:

To create an immediate Shading Link if one

does not already exist (the icon on the Linking
header bar is RED):

1: Click to enable the Shading Link check

box - tick mark visible.

2: Click the Snapshot button.

3: When the Linked Shading prompt

appears, make sure the microscope and
camera exposure settings are suitable
and then:

Either move the stage to view an empty

field on the specimen, or...

Remove the specimen slide and

replace it with a slide (and cover slip if
necessary) of the same type and
quality.

A very small amount of microscope de-

focus may be helpful to prevent
contaminants affecting the Shading
reference.

4: On the Linked Shading prompt click Yes.

5: A shading link will be created, and when

complete the Configuration Stored
message will appear.

Click OK and...

6: ...the header bar icon becomes GREEN.

292 © 2011 Leica Microsystems (Switzerland) Limited

 Shading Wizard:

The Shading Wizard creates a Shading Link

for each microscope objective in turn starting
with the one selected. Having completed one,
it automatically moves to the next. The Mag
changer, camera and contrast method remain
the same for each link.

Because it can be stopped after completing an

objective, groups rather than all the objectives
can be processed.

For a single objective use Snapshot:

Before starting the Wizard, the specimen

image must be properly exposed and
focused.

Either move the stage to view an empty

field on the specimen, or...

Remove the specimen slide and replace it

with a slide (and cover slip if necessary)
of the same type and quality.

A very small amount of microscope de-

focus may be helpful to prevent
contaminants affecting the Shading
reference.

1: Click to enable the Shading Link check

box - tick mark visible.

2: Click the Wizard button.

3: If shading links have already been created

for the current microscope settings, a
warning will appear.
Click Yes to overwrite the existing link or
No to cancel the Wizard.

LAS User Manual 293

 Shading Wizard: Continued:

If necessary, carefully adjust Exposure and Gain on the

empty field or blank specimen slide to achieve a very small
amount of over-exposure. Refer to Histogram for details of To prevent contamination affecting the shading, a very
how to turn on over/under-exposure indication: 244 small amount of de-focus may be used.

3: Click the Next button.

On the Wizard dialog:
4: The Wizard will create the Shading Link for the current
1: Click on the Start button. objective.

2: Click on the objective to be processed. It becomes When complete, it will automatically select the next
high-lighted. objective (change the objective manually for non-auto
microscopes) and return to Step (2).
The first objective is selected automatically for
motorised microscopes, but can be changed by 5: When a Shading Link for all of the required objectives
clicking on another. has been created, click the Close button.

294 © 2011 Leica Microsystems (Switzerland) Limited

 Shading Link: Delete:

1: Turn Shading Linking on or off by clicking

the Link button.

2: Shading links can be deleted by clicking

on the Delete button and...

3: ...confirming the deletion by clicking the

Yes button.

Use with care: This will delete ALL of the

Shading Links.

4: Both Exposure and Shading Linking can

be enabled together. In this case the
status icons on the Linking header bar will
appear combined.

LAS User Manual 295

 Check Colour:

This facility is useful for adjusting the overall image colour

sometimes necessary to compensate for colour shift due 5: To apply the Hue and Saturation values to the entire
to small variations in the specimen lighting and camera image, click on the 'target' mark and drag it toward the
characteristics. dot.

Use it mainly for colour ‘fine-tuning’ after carrying out a 6: To select another area of the image by clicking on the
White Balance. region outline (not the handles) and drag it to the new
location.
1: Click on the arrow to the right of the Check Colour
header to expand the panel. 7: The Hue and Saturation windows show the values at
the current target location. Fine tune the values by
2: Left click on the image and drag a rectangle around either clicking the up and down arrows to the right of
the required area to create a Region of Interest (RoI). the window or clicking in the widow and typing a new
value.
3: When the mouse button is released a context menu
appears. Click to select the Check Colour option. 8: Hide or reveal the Region of Interest by clicking to
disable the ROI checkbox.
4: A small dot on the Colour Wheel represents the Hue
and Saturation values within the region. 9: Reset any adjustments by clicking the Reset button.

296 © 2011 Leica Microsystems (Switzerland) Limited

 Reticule and Web Sharing:

Reticule and Web Sharing are optional

modules that have to be installed and
licensed. If either or both are installed then a
control panel will appear in Camera.

Click on to display the help:

973 Reticule
984 Web Sharing

LAS User Manual 297

 The Browse Workflow:

Browse uses the fast Image Explorer developed

especially for Leica Application Suite and based upon
the familiar tree and folder structure of Microsoft
Windows. It provides access to stored images and to
their relevant data such as the time of acquisition, the bit
depth and the calibration.

As well as Image Explorer an optional module LAS

Archive can be installed on the same computer. Users
can switch between the two storage methods
seamlessly.

Images - single or collections - captured before LAS

V3.3 can be imported into LAS Archive. More about LAS
Archive 371

Browse features and quick links (Click ):

309 Quick navigation between folders with a single

click.

310 Create new folders without having to leave LAS.

307Set a folder as the capture location and

immediately begin to grab images.

304Naming images automatically with meaningful

names and auto-incremented sequence numbers.

340Microscope and camera data saved with the

image: Users can add their own comments and
observations

338 A scalable Thumbnail Gallery to show all of the

images in a folder and, with a single click select and
display an individual.

303A wide range of tools for image fast storage and

access.

298 © 2011 Leica Microsystems (Switzerland) Limited

 Browse: User Interface:

The illustration is a graphical representation of the LAS display and Browse interface showing the principal features and
quick links (Click ):

1: Workflows 300 Click the Browse Workflow to open the navigator.

2: Toolbox 303 Common tools for both Image Explorer and LAS Archives.
3: Navigator Window 301 Tab shows Folders for Image Explorer or Archive and Search for Archives.
4: Image Data Form 340 Displays and edits selected data for the current image.
5: The Image Viewer 336 Display and working area for the current image: Press keyboard F5 to show full screen.
6: The Grid 344 Displays data for all of the images in the selected folder.
7: The Gallery 338 Displays thumbnails of all the images in the selected folder.
8: Acquire Button 304 Click to grab and image from the microscope camera.
9: Side Toolbar 317 Working tools for image sizing, printing and deleting as well as switches for the Gallery and
Grid.
10: The Search Controls 410 Available only with LAS Archives.
11: Gallery Navigation Browser 338 Rapidly find thumbnails in the Gallery.
12: Thumbnail Scaler 338 Slider adjusts the size of the thumbnails in the Gallery.
13: Status Bar Displays Hardware Configuration, RGB Intensity, Stage Position and Magnification data.

LAS User Manual 299

Browse: Launching Browse:

1: Click on the Browse Workflow and if

necessary…

2: …on the Browse tab to reveal the main

panel.

3: If LAS Archive is installed and has been

previously selected...

4: ...click on the Return button to switch to

Image Explorer.

5: Click on the small arrows to the left of

the folders to expand them.

6: Click on a folder to reveal the contents.

7: The Scrollbars to the right and bottom of

the navigation window are displayed
automatically if necessary. Click and
drag a Scrollbar to move the explorer
tree within the window.

300 © 2011 Leica Microsystems (Switzerland) Limited

 The Navigator Panel: Image Explorer:

The main panel layout and controls changes

depending upon the storage system being
used - either Image Explorer or LAS Archives.

Image Explorer is the default navigator

supplied as part of LAS Core.

LAS Archives are optional modules designed

especially for those users who need a fast
database for image storage. More about LAS
Archives 390

The illustration shows a typical panel with

Image Explorer active. The control tab label
displays the word Folders.

The principle features and quick links (Click

)

302Floating Navigator: Drag and drop the

Navigator to a location that suits the user,
even on to a second monitor..

303Toolbox: Image grabbing and record

creation.

307 Navigation Buttons: Move between

folders and set the capture folder.
Favourites creates a fast navigation list.

310 Create a new Folder.

313 Rename a Folder.

314 Delete a Folder.

325 Image Sequence Move and Copy.

315 Hide Component Images.

LAS User Manual 301

 The Navigator Panel: Floating Navigator:

The Navigator panel can be detached from its

'dock' and parked anywhere on the Viewer -
or even on another monitor - to provide access
to folders and archives immediately from any
workflow or Optional Module without returning
to Browse.

To move the Navigator:

1: On the Browse Workflow, click on the

Navigator header and, holding down the
mouse button drag the panel to the
required docking position. All of the
Navigator features are available in the
new position...

2: ... including the minimise button that will

close the Navigator leaving only the
header on display. Click the minimise
button to reveal the Navigator again.

3: The Show Navigator button works as a

toggle. If the Floating Navigator has been
moved the first click will return it to the
side panel. The second click will move it
back to the user's docking position.

302 © 2011 Leica Microsystems (Switzerland) Limited

 The Toolbox:

The Toolbox buttons are grouped on a small panel at the

top of the Browse tab (1). Click a Toolbox button for more
information.

Acquire Image: click to grab an image: Has the

same function as the Acquire button at the
bottom of the screen.

Acquire in the Current Image: Grabs an image

and overwrites the current selected image.

Create an Empty Record: Click to create a

record without an image – an image can be
captured into it at a later date.

Duplicate Current Record: Duplicates the

current record and image.

LAS User Manual 303

 The Toolbox: Acquire New Image:

1: Click the Acquire New Image button. If

Capture to Fixed Location is enabled in
Preferences, and the folder (indicated with
a red dot) is selected in the Navigator
window, the image will be captured to that
folder.
The Acquire button at the bottom of the
screen performs the same function.

If a folder other than that chosen as a ‘fixed

location’ is selected, the Unable to Show
Image dialog appears. The options are:

2: Show the acquired image and move to

the Fixed Capture Location to display it
or...

3: Don’t show the acquired image but do

save it to the Fixed Capture Location
anyway.

4: To make your choice the future default

action, click to enable the Don’t show this
operation again check box

304 © 2011 Leica Microsystems (Switzerland) Limited

 The Toolbox: Acquire into Current Record & Create Empty Record:

1: Acquire into the Current Record replaces

the selected image with a new one.
Where necessary, the data is updated but
the Image ID and Image Name remain the
same.

2: Create an Empty Record does not

capture an image and stores only
essential microscope data, but it does
give the record and Image ID and an
Image Name.

The image and data may then be

captured later by clicking to select the
empty record and then using the Acquire
into Current Record (above). This
provides a convenient method of 'loading'
images later.

3: The Empty Record is represented as a

Leica Cube in the Gallery thumbnails.

LAS User Manual 305

 The Toolbox: Duplicate Current Record:

Duplicate Current Record is a simple way of

copying an image and its data.

Some data such as notes and descriptions

may then be changed to avoid having to type
in the description again, the image and the
microscope data remain the same.
1: In the Gallery, click on the thumbnail of
the image to be duplicated.

2: In the Toolbox click the Duplicate Current

Record button.

3: The duplicate appears as a thumbnail in

the Gallery with the name Copy of…
followed by the original Image Name.
Change data on the Form as required.

306 © 2011 Leica Microsystems (Switzerland) Limited

 Navigation Buttons:

The Browse Navigation Buttons allow the user to move

back and forth between folders and files, nominate a folder
into which images should be captured, create a child of an
existing folder and to store 'favourite' folder locations for
speedy retrieval.

Click a Navigator button for more information:

Step Back: Click to move to the previous folder

(left arrow) and return (right arrow).

Set Capture Folder: Makes the selected folder

the capture target – grabbed images will be
stored here.

Move to Capture Folder: Returns to the Capture

Folder from anywhere on the tree.

Create a New Folder: Creates a folder as a child

of the currently selected folder.

Refresh the Current Folder: Re-displays the

Gallery and selected image in the Viewer.

Favourites: Stores and links to frequently used

folders and files.

There are a further three controls below the Navigator

window:

2: Rename 313 Allows the user to rename a folder.

3: Delete 314 Deletes the selected folder and all of its

contents.

4: Hide Component Images 315 When enabled displays

only the 'analytical' images in the Gallery.

LAS User Manual 307

 Navigation Buttons: Recent:

The software keeps track of the navigation by the step 3: ...the Recent tab is clicked.
buttons (1) and allows the user to go directly to a folder
without having to make multiple clicks on the step buttons. 4: Return directly to a folder by double-clicking on it in
the list.
The Recent feature is only available in Folders.

2: The list of visited folders is displayed when...

308 © 2011 Leica Microsystems (Switzerland) Limited

 Navigation Buttons: Set and Move to Capture Folder:

Users can nominate a folder into which

captured images will be automatically saved.
The nominated folder can be changed at any
time:

1: Click to select the Capture Folder.

2: Providing that the Capture to fixed folder

location option is enabled in Preferences
47 , clicking the Set Capture Location
button will make the selected folder the
saved image location.

3: The Capture Folder is indicated by the red

dot to the left.

To Move to the Capture Folder:

4: If, having navigated away from the

Capture Folder a user needs to return to
it, clicking the Move to Capture Folder
button will go directly there.

LAS User Manual 309

 Navigation Buttons: Create a New Folder:

A new folder is created as a child of the

selected folder that does not have to be the
Current Capture folder.

1: Click on the folder that is to be the parent

of the new folder.

2: Click on the Create New Folder button.

3: The new folder appears with the default

name New Folder.

4: With the new folder selected and

highlighted, click on the Rename button
and...

5: ...type a new name. Press the Enter key

on the keyboard and the new folder has
the new name.

310 © 2011 Leica Microsystems (Switzerland) Limited

 Navigation Buttons: Favourites:

Users can store links to folders that they often use with 2: Click the Add Favourite button to create a link to the
the Favourites button. They can then return directly to a folder.
listed folder simply by double-clicking its link. This is a very
useful feature for 'trees' with many folders and saves a 3: The list of links can be seen by clicking to Favourites
considerable amount of time. The Favourites feature is tab...
only available in Folders.
4: ...with the newly created link highlighted. Move the
1: Click to select the folder to add to Favourites. cursor over the link to reveal the complete path.

Continued 312

LAS User Manual 311

Navigation Buttons: Favourites Controls:

1: To change the name of a link click the Edit button and 3 & 4: Move a link up or down in the list by clicking to
enter the new name. select the link and then either the Move Up or Move
Down button.
2: Delete a link from the list by clicking to select the link
and then clicking the Delete (Trash Can) button. 5: Return to a chosen folder by double-clicking its link.
The Folders tab opens automatically and the selected
folder is highlighted.

312 © 2011 Leica Microsystems (Switzerland) Limited

 Folders: Rename a Folder:

1: Click on the folder to be re-named.

2: Click on the Rename button or on the F2

function key.

3: Click in the folder text box and type the

new name finishing by pressing the Enter
key on the keyboard.

LAS User Manual 313

 Folders: Delete Folder:

Use with caution: Folder Delete cannot be

undone.

Trying to delete a folder that contains images

results in the Unable to Delete warning (1). All
of the images must be removed by:

2: Click on the thumbnail of the first image

and...

3: ...holding down the keyboard Shift key,

click on the thumbnail of the last image.
All of the thumbnails and images are
selected.

4: Click on the Delete (Trash Can) button on

the Side Tool Bar.

5: Confirm the deletion.

6: Click the Navigator > Delete button.

7: Confirm the deletion and the folder will be

removed.

314 © 2011 Leica Microsystems (Switzerland) Limited

 Folders: Hide Component Images:

The Gallery when displaying sequences (A)

can become very cluttered with the individual
component images - the important results may
be scattered and time-consuming to find.

The component images can be easily hidden

leaving only the 'analytical' images on view in
the Gallery (B) by enabling (tick mark
displayed) the Hide Component Images check
box (1).

The individual images are not deleted, only

hidden and can be revealed by disabling Hide
Component Images by clicking it again.

LAS User Manual 315

 Folders: Sequence Move and Copy:

Image sequences - montage and multistep for

example - can be copied or moved in a single
step using mouse drag and drop:

To move a sequence:

1: Left click on the sequence name and,

holding down the mouse button...

2: ...drag to the target folder and release the

mouse button. The sequence is moved
from the source folder to the target folder.

To copy a sequence:

3: Press and hold down the keyboard Ctrl

key, left-click and also hold down the
mouse button on the sequence to be
copied. Drag the mouse to the target
folder and release the mouse. Release
the Ctrl key.

The sequence is duplicated in the target

folder.

316 © 2011 Leica Microsystems (Switzerland) Limited

 The Side Tool Bar:

This section describes the Side Tool Bar tools that are common to both Image Explorer and LAS Archive.
Click on a Tool Bar button for more information:

Scale Bar and Annotations: Run Annotations and Scale Bar without leaving Browse.

Floating Navigator: Click to enable the Floating Navigator and click again to 'park' it.

Export: Export the current image or image selection to a location of the users choice.

Image Stitching: Combines a set of overlapping images into a single, aligned overview image.

Print: Print the selected image together with headers, footers and a wide range of formatted data.

Create Report: Produces a printable report. Appears only when an archive is being displayed.

Delete the selected Image(s): Deletes the images and their associated data.

Panning: Examine areas of images that extend beyond the Viewer edges into the display area.

Zoom in and...

Zoom Out.

Fit the image to the Viewer area.

Display at Original Size: Displays he image at its captured size:

Hide and Reveal the Record Panels.

Hide and Reveal the Viewer.

Hide and Reveal the Data Grid: Only available with LAS Archives.

Hide and Reveal the Thumbnail Gallery.

View the image Record Details.

Select the Form Details to display: Allows the user to add or remove image details from the Form.

Viewer Options: Select Dual Viewer, Lock the Views and Lock the Pan View.

Save the Output File: Click to save an image of the output file currently displayed in the Dual Viewer.

LAS User Manual 317

 The Side Tool Bar: Annotations & Scale Bar Quick Launch:

Clicking the Show Annotations button displays the (a tick mark is displayed) the required function. All of the
Annotations and Scale Bar Quick Launch menu. function tools are available.

Basic Annotation, Extended Annotation and the Scale Bar Additionally, any annotations applied to the current image
setup can be launched without leaving Browse by as well as the Scale Bar can be displayed.
checking

318 © 2011 Leica Microsystems (Switzerland) Limited

 Image Stitching:

LAS Image Stitching software has been designed to Background colour is user selectable.
create a composite image of a large specimen from a
sequence of individual 'tiles' captured on a manual stage. Smooth blending feature helps to correct uneven
lighting effects or incorrect shading.
LAS Image Stitching is ideal for small to moderate tile
counts. It is fast and flexible and is part of the LAS Core Automatic image scaling to reduce the final image
and so available to all users. size.

Users to set result image names with automatic suffix

Features: increments.

Designed especially for image capture using manual

stages - demanding precision is not required. Using Image Stitching:

Suitable for a wide range of specimens - colour or 320 Tile Capture Guide.
greyscale. 326 Advanced Settings - Blend and Background
Areas of interest can be chosen from a tile sequence Colour.
- there is no need to stitch an entire sequence.
327 Image Reduction Factor.
The number of tiles and image size are not fixed but 325 Select and Stitch all Tiles in a Sequence.
the recommendation is for no more than 50 tiles or an
image no larger 500MB. 330 Stitch Selected Tiles.

LAS User Manual 319

Image Stitching: Tile Capture Guide:

Image and Tile Requirements: This is an image of a watch escapement magnified x5:

Image Stitching works best with images

that have complex, random detail. Avoid
repetitive, grid-like detail.

Tile overlap is essential 321 Detailed

images do not require as much overlap as
images with sparse features.

All tiles must have the same magnification

and be as sharp as possible. The
minimum tile size is 512 x 512 pixels.

Tiles must have the same Capture Format

- jpg etc 322 .
The field of view extends to just one corner...
All tiles must have same Calibration value
323 and must not be rotated with
respect to each other.

Shading correction 324 is not essential

but is recommended to avoid 'striping' in
the result image.

The image can be colour or greyscale.

Continued 321

...which means that at least 9 individual tiles are required to

represent the entire image:

320 © 2011 Leica Microsystems (Switzerland) Limited

 Image Stitching: Image Overlap:

Because the individual tiles are captured using a manual 1: Capture commenced at the top-left corner.
stage, the edges are unlikely to represent precise fits to
those of their neighbours. Therefore, the stitching process 2: The stage was them moved along the X axis to the
compares each tile with all of the others in the selection second tile making sure there was a good overlap
looking for matches in pixel groups that will constitute an with the first tile.
edge and to achieve this there must be a guaranteed
overlap between adjacent tiles. 3: This was repeated for the third tile, again ensuring a
good overlap with the second.
As a guide, users should aim for about 25% tile area
overlap although for images with random, non-repetitive 4: Then the stage was moved along the Y axis to start
detail this can be lower. the second row, checking there was adequate row-to-
row overlap - and so on until the entire image had
The illustration shows by using different colours, how the been captured.
nine tiles overlap on all internal edges.

Users who have stages fitted with a Vernier scale can

make some simple calculations to make sure there is
sufficient overlap and use the scale to position the stage
rather than just relying upon a visual estimate.

LAS User Manual 321

 Image Stitching: Setting Image Format:

The specimen capture settings and made in

both Preferences > Image and Acquire >
Camera.

1: Set the Image Format and compression by

launching Preferences ( ) and clicking the
Image tab.

2: On the Save Images panel, click on the

arrows to the right of the Image Format
header and...

3: ...click to select the required format.

Jpeg 2000 (J2K) is not suitable for Image

! Stitching.

4: Whilst on the Save Images panel, users

may like to give the tiles an appropriate
name by clicking inside the Default Image
Name text box and typing a new name,
and...

5: ...setting the number of zeros to be

displayed in the suffix following the image
name. In the example the individual tiles
would be named: Harris Sample_00, 01,
02...10, 11 and so on.

322 © 2011 Leica Microsystems (Switzerland) Limited

 Image Stitching: Calibration:

Calibration ensures that measurements

displayed by the software are given in 'real
world' - millimetres, inches etc - units taking
into account the selected optical magnification
of the microscope and the size of the camera
pixels.

For initial system calibration users should refer

to:

Acquire > Camera > Calibration 268 .

If system calibration has changed and the tiles

pre-dates the change, they can be updated to
ensure they are all consistent by using Update
Calibration found in:

Options and Preferences > Update

Calibration .

LAS User Manual 323

 Image Stitching: Shading:

Shading is the name given to variations in the

background light level across an image.

In the example, the image on the left shows

the effects of shading - the light source and
the optics conspire to create a bright spot in
the centre of the image which gradually
becomes less and less bright toward the
edges.

A more evenly 'illuminated' image can be

achieved by applying a digital 'correction'
based upon the brighter part of the image.
This is shown on the image on the right.

Because shading is caused by variations in

the optics, each objective has to have the level
of 'correction' applied individually.

1: Shading is described in detail and applied

in Acquire > Camera > Shading 257 .

2: With the Shading facility turned off, the

stitched image has a 'corrugated' look
caused by the dark edges.

Check the Blend option in Advanced Settings

to help minimise the effects of uneven lighting
326 .

324 © 2011 Leica Microsystems (Switzerland) Limited

 Image Stitching: Select and Stitch all Tiles:

Start Image Stitching by selecting the tiles to be

used - in this example all of them:

1: In the Gallery, left click on the first image to

select it and...

2: ...holding down the keyboard Shift key, left-

click the last tile in the collection. All of the
tiles between the two selections are
automatically selected - the thumbnail
frame is coloured brown.

3: On the Side Tool Bar click the Image

Stitching button.

4: The Image Stitching dialog appears

showing the number of tiles that have been
selected.

5: The stitched image is given a name

preferred by the user. Click inside the
Result image name text box and type a
name.

6: Click the Show advanced settings button to

set the Blend, Background Colour and
Reduction Factor: 326

7: The Further Information button 328 when

clicked, provides a reminder of the image
requirements.

Continued 326

LAS User Manual 325

 Image Stitching: Advanced Settings:

1: After Image Stitching is launched and the Advanced 4: ...on the Select Colour dialog choose a background
Settings button is clicked, the dialog expands to reveal colour from the swatches, by dragging the 'target' on
the Blend, Background Colour and Reduction Factor the wheel or typing values for red, green and blue
options: (RGB) in the text boxes.

5: Adjust the colour intensity by clicking and dragging the

Blend images where they overlap: slider.

2: The Blend option smoothes the tonal transition 6: Click OK. The selected colour appears in the box.
between adjacent tile edges. It especially useful if
Shading was not enabled or incorrectly set when the Continued 327
tiles were captured.

Click the check box to enable (tick mark visible) the

Blend option. Click again to turn it off. The default is
enabled.

Result image background colour:

3: If unimportant tiles are not selected 330 and

therefore not included in the stitching, there will be a
gap in the composite image. Users can select a colour
to appear in the gap(s).

Click in the Background colour box and...

326 © 2011 Leica Microsystems (Switzerland) Limited

 Image Stitching: Advanced Settings: Image Reduction Factor:

Some stitched images can be large and unwieldy making 2: ...from the drop-down menu click to select the
them difficult to include in reports or share digitally. The reduction factor required.
Image reduction factor automatically scales the result
image to reduce its size. Larger values: Result in a smaller result image.

1: Click on the small arrows to the right of the Result Auto: The reduction factor is based upon the pixel
image reduction factor and... count of the individual tiles.

None: The result image is not scaled at all.

3: Click on the Advanced Settings button to close the

settings.

Continued 328

LAS User Manual 327

Image Stitching: Further Information:

1: Click the Show guidance... button to... 3: The Image Stitching program provides a range of
warnings to help the user capture and select tiles that
2: ...display a reminder regarding the essential will help ensure the best possible result.
parameters for tile capture.
Continued 329
Click the button again to hide the dialog.

328 © 2011 Leica Microsystems (Switzerland) Limited

 Image Stitching: Start the Stitching:

1: Start the stitching process by clicking the

Stitch Images button.

2: The completed composite image appears

in the Viewer together with...

3: ...a thumbnail in the Gallery. The caption

comprises the users preferred name
together with an automatically
incremented numeric suffix.

LAS User Manual 329

 Image Stitching: Stitch Selected Tiles:

The tile selection prior to stitching does not

have to include the entire sequence - tiles that
represent an area of the image that is not
important or needed can be omitted from the
selection provided the remainder represent a
contiguous area.

However, if the selected tiles do not stitch

properly, include some of the omitted tiles and
run Image Stitching again.

In this example only the main bearing is of

interest and so only those tiles that are part of it
are selected:

1: In the Gallery, left click on the first image to

select it and...

2: ...holding down the keyboard Ctrl key, left-

click the other tiles required individually.
The illustration shows just the tiles that
together make up the bearing.

3: On the Side Tool Bar click the Image

Stitching button.

4: The Image Stitching dialog appears

showing the number of tiles that have been
selected.

5: Click the Stitch Images button.

6: The finished composite image with all of

the tiles stitched together...

7: ...represented by a thumbnail in the Gallery

. The final image can now be used like any
other.

330 © 2011 Leica Microsystems (Switzerland) Limited

 The Side Tool Bar: Panning and Zoom:

1: Panning: The Pan tool allows detailed

areas of an image that exceeds the visible
area of the Viewer to be examined. It will
not work if Fit to Viewer is enabled
because all of the image is being
displayed.

On the Pan Window viewer, click and hold

in the outlined rectangle and drag it to the
area to be examined. The selected area is
displayed in the main Viewer.

To move the Pan Window away from the

main Viewer, click and hold the header
bar and drag it to another part of the
screen.

Click the Pan tool to close the Window.

2: Zoom: Click on the (+) to zoom in to the

image or (-) to zoom out. The zoom level
as a percentage, is displayed top right of
the the Viewer border.

If the monitor Magnification Settings have

been set in Preferences , the image
Magnification value appears bottom right
of the Viewer border.

3: Fit to Viewer: Click to fit the image to the

available Viewer area regardless of the
original size of the image. The Image
Scaled to Fit Window message appears
top right of the Viewer border.

4: Display at Original Size: Click to display

the image at its original size. The image
may appear smaller or larger than the
Viewer area. The Image Unscaled
message appears top right of the Viewer
border.

LAS User Manual 331

 The Side Tool Bar: Hide and Reveal:

The various screen areas - Viewer, Gallery, Report and 3: Hide/reveal the Data Grid: The Viewer will expand to
Grid (where applicable), may be revealed or hidden to cover some of the vacated space. The Grid is only
create the best working environment for the user. Some available if an LAS Archive is installed.
tools are toggles – click once to reveal the area, click
again to hide it: 4: Hide/reveal the Thumbnail Gallery: The Gallery is
hidden and the Viewer expands to include the Gallery
1: Hide/reveal the Record panels: The Image Viewer space.
expands to fill the vacant space.

2: Hide/reveal the Viewer: The Record panels expand to

occupy the Viewer width.

332 © 2011 Leica Microsystems (Switzerland) Limited

 The Side Tool Bar: Delete Images:

Delete a single image by clicking its thumbnail and then 3: Click the Delete (Trash Can) button.
the Delete (Trash Can) button.
4: Confirm that the images are to be deleted and cannot
Alternatively, delete a group of images by: be recovered and the images and their data will be
removed from the folder.
1: Hold down the Crtl key and click on a thumbnail. Do
this for all of the images to be deleted, or...

2: ...hold down the Shift key and click on the first

thumbnail. Do the same for the last thumbnail in the
group to be deleted and all of the images between are
selected.

LAS User Manual 333

 The Side Tool Bar: Dual Viewer:

The Viewer area can be split to show two captured images 5: Display an image in the right-hand pane by clicking
simultaneously. the pane and...

1: Click the Viewer Options button. 6: ...the thumbnail of the required image.

2: Click to enable (tick mark visible to the left) the Dual 7: To synchronise the panes and enlarge or reduce the
Viewer option. The Viewer will then divide into two images as the zoom and fit tools are used, click to
panes. enable the Lock View option.

3: The image currently being viewed will usually appear 8: Enabling Lock Pan will synchronise the images as the
in the left-hand pane. If it does not or the image needs Pan tool is used. Click the pane to pan and then on
to be changed, click on the left-hand pane and... the Panning tool. Both images will automatically move
to and display the image segment shown in the Pan
4: ...click a thumbnail in the Gallery. window.

Dual Viewer more information: 79

334 © 2011 Leica Microsystems (Switzerland) Limited

 The Side Tool Bar: Copy Output Image:

The output image can be copied (duplicated) by:

1: Click the Copy Output Image button.

2: The image is copied to the current folder with the

output image name plus the suffix '_Copy'.

LAS User Manual 335

 Full Screen Mode & Second Monitor:

Full Screen Mode: Single Monitor:

1: The Viewer area can be expanded to fill

almost the entire screen by either cl
icking Options on the main menu and...

2: ...selecting Full Screen from the drop

down menu, or...

3: Pressing Key F5. Press F5 again to

return to the normal display.

Second Monitor:

The software detects a second monitor and

changes the Options drop-down menu to:

4: ... Use Second Monitor. Click the option

to use both monitors.

The Viewer and image occupy all of the

second monitor whilst the Gallery and
Thumbnails together with the controls
remain on the primary monitor.

The Side Tool Bar buttons are

appropriately shared between the two
monitors.

5: Alternatively, press Key F5 to move

between using both monitors and
returning to single monitor.

336 © 2011 Leica Microsystems (Switzerland) Limited

 Image Options:

A wide range of options is available by right-

clicking the image in the Viewer or its
Thumbnail in the Gallery. The options vary
depending upon which item was clicked, the
operating system and the software installed on
the computer.

1: Right-click the Thumbnail for the context

menu of basic options and...

2: ...click it to select.

3: Some options have additional possibilities

displayed as a sub-menu.

4: Right-clicking the image in the Viewer

displays a different context menu.

5: Additional functions are available some of

which will also have sub-menus (6).

LAS User Manual 337

 The Gallery: Image Explorer and LAS Archive:

The Gallery is a thumbnail display of images in

the current folder in both Image Explorer or LAS
Archive.

The Gallery can be hidden or revealed using

the Side Tool Bar tools.

1: Clicking on a thumbnail will immediately

display the full-sized image in the Viewer
and the data associated with it in the
Record and Grid (If LAS Archives is
installed).

2: A Slider is automatically displayed for

multiple rows of thumbnails - click and drag
it to scroll the Gallery.

3: The Navigation Bar (bottom right of the

screen) provides a way of moving through
the thumbnails quickly and is especially
useful with large galleries of thumbnails.
Click on the arrows to move a single image
(3) ...

4: ...or go to the extreme ends of the Gallery.

5: The thumbnails can be re-sized by clicking

and dragging the Scaling Slider - slide left
to reduce the thumbnail size and right to
increase it.

6: Move the mouse over a thumbnail to reveal

basic data bout the image.

7: Right-click a thumbnail to show the Context

Menu. Left-click to select an option.

Gallery Docking Position 339 .

338 © 2011 Leica Microsystems (Switzerland) Limited

 The Gallery: Docking Position:

Available in Acquire and the Process Workflows as well 2: ...from the drop-down menu, left-click to select the
as Browse, the thumbnail Gallery can be 'docked' either Toggle Gallery Position option.
horizontally - along the bottom edge of the Viewer - or
vertically - along the left-hand edge of the Viewer - to suit The action toggles between horizontal and vertical
the user. docking.

1: Right-click on a thumbnail or on the spaces around Scroll bars, if required, are placed automatically.
the thumbnails and...

LAS User Manual 339

The Data Form:

1: The Data Form displays selected data

associated with the image. All of the data
about the camera, microscope, exposure,
creation date and so on, are actually stored
and all can be displayed, but the Form can
be configured to display only the more
pertinent items.

2: The Data Form can be hidden by clicking

the Form button on the Side Tool Bar. This
is a toggle action - click again to reveal the
Form.

Select Visible Fields 341 .

Configure Grid Columns 342

Record Details 343

340 © 2011 Leica Microsystems (Switzerland) Limited

 The Data Form: Select Visible Fields:

To select the data (Fields) to be displayed on

both the Data Form and on the Grid (if an LAS
Archive is installed):

1: Click on the Visible Fields button on the

Side Tool Bar.

2: To display a data field, click in the check

box to the right of the field name. This is a
toggle action - click again to clear the check
box and hide the field.

3: Display all of the data fields on the Data

Form by clicking the Select All button.
Because there will now be too many items
to display simultaneously, a Scroll Bar is
automatically positioned to the right of the
Data Form.

4: The Clear All button will hide all of the data

fields and is useful before making a new
selection range.

5: Click OK to close the Select Visible Fields

dialog. The chosen data fields are
immediately displayed on the Data Form.

LAS User Manual 341

 The Data Form: Configure Grid Columns:

1: The data fields displayed on the Data Form are also 3: Click on the Reset Column Widths button.
displayed on the Grid (if an LAS Archive is installed) -
click the Grid button on the Side Tool Bar to reveal it.
Reset the column order to match the Data Form by:
The order of columns on the Grid can be changed by
clicking on the column heading to be moved, holding 2: Click on the Visible Fields button on the Side Tool
down the mouse button and dragging the column to a Bar.
new location.
4: Click on the Reset Column Order button.
In a similar manner, column widths can be changed by
dragging the column Dividing Bar to the required width. 5: Click OK.

Reset the column widths by:

2: Click on the Visible Fields button on the Side Tool Bar.

342 © 2011 Leica Microsystems (Switzerland) Limited

 The Data Form: Record Details:

To view all of the available data and if LAS

Archive is installed, select them for use in a
report:

1: Click on the View All Details button on the

Side Tool Bar.

The Record Details dialog displays all of the

data fields available for the chosen image
and the values that have been captured for
them. Use the Scroll Bar on the right of the
window to scroll through the list.

2: To use a field in a report, click on the

required item to highlight it and then on the
Copy as Report Field button. The field
description is copied and pasted into the
report template.

Use the keyboard shortcut keys to:

Select all fields: Ctrl + A
Copy all fields: Ctrl + C
Paste copied fields: Ctrl + V

3: Click the Close button to close the dialog.

LAS User Manual 343

 Browse: The Grid:

The Grid displays data for all of the images in a folder in a 5: A small arrow is revealed when a header is clicked.
tabular structure. The image names are listed on the left This allows the image data to be sorted – high-to-low
and the data items as headers across the top. or low-to-high – by successive clicks on it.

1: The Grid is revealed and hidden by clicking on the 6: To make multiple selections prior to deleting or
Side Tool Bar button. This is a toggle – click to reveal, exporting, hold down the keyboard Ctrl key whilst
click again to hide. clicking individual thumbnails.

2: Clicking on an entry in the Grid will immediately Keyboard combination Ctrl + A will select all of the
display that image in the Viewer and also highlight the image data.
thumbnail. Ctrl + C will copy all the selected image data to the
clipboard.
3: Header positions can be changed by clicking and Ctrl + V will paste into another application.
holding the left mouse button on the header to be
moved, dragging it to the new position and releasing 7: The Grid data can be exported to a range of other
the mouse button. applications by right-clicking on the Grid then
navigating to and clicking to select an application.
4: Column widths can be changed by clicking and
dragging the vertical bars that separate the columns.

344 © 2011 Leica Microsystems (Switzerland) Limited

 Browse: Device Settings, Store and Recall:

Providing the Store and Recall module is

registered and enabled, the Store and Recall
Settings panel details the microscope data -
camera and microscope setup - relevant to the
displayed image.

1: Click on the expand/collapse arrow to the

right of the header bar to display the
settings.

2: Click the Recall button and...

3: ...the Recall Confirm dialog appears. Click

Yes to confirm.

If the system has an automated microscope

attached, it will automatically return the
microscope and camera to the settings that
were used to acquire the image.

4: To cancel the restore and return to the

previous settings, click the Undo button.

5: If the image XY and/or Z Stage positions

are required, click to enable (tick mark
visible) the appropriate check box and...

6: ...when the Recall button is clicked the

stage movement warning appears. Check
that the stage and objective are clear of any
possible obstructions and click the No
Obstructions - OK to Recall button and the
stage will move to the original capture
positions.

LAS User Manual 345

 Process Workflow

By using the Process Workflow with its Basic Annotations

and Enhance features, images can be used to convey a Basic Annotations to add simple pointers, labels and
wealth of added information and detail. captions to your image.

Annotations can either be saved with the image so that Enhance has the tools to brighten, adjust saturation,
they can be edited at any time, or when the user is gamma and contrast as well as rotate and crop an
satisfied with the results merged with it so that the data is image.
still visible when the image is exported.

346 © 2011 Leica Microsystems (Switzerland) Limited

 Process: Basic Annotation

Digital images can be annotated with

graphics and text to provide information
or to indicate features of interest.

The annotations are stored as a file with

the image but separate from it and
recalled whenever the image is displayed.
They can be positioned or modified if
required. However, they can also be
merged with the image to produce a
single, integrated image that can be
exported to other applications with the
annotations intact.

The essential tools allow automatic

annotation with an Image Name,
Description, Time and Date simply by
enabling appropriate check boxes. A line
can be drawn at any angle formatted as a
Arrow with a label, a simple Line or to
indicate a point-to-point Distance.

Annotations appear in colour overlaid on

an image regardless of its colour depth -
colour or monochrome.

Where images are larger than the

window and it is possible to scroll the
image, the annotations scroll to retain
their positions. If the image is zoomed or
reduced the annotations are scaled
accordingly but limited to a readable size.

Basic Annotations normally reside on the

Process Workflow but can be invoked
from all of the Workflows (except Setup) Basic Annotation features:
using the Side Tool Bar icon. The Control Panel: Go there... 348

The Annotation tools are designed to be Display and Image Name and Description: Go there... 349
quick and easy to use; For greater power Display the Time and Date: Go there... 350
and functionality the optional module
Extended Annotation module can be Draw a Line with a Caption Label: Go there... 351
added to the suite. Selecting the Font: Go there... 352
Changing the Font and Background colours: Go there... 354
Defaults and Merging: Go there... 355

LAS User Manual 347

Basic Annotation: The Control Panel and Features:

To launch the Basic Annotation Control

Panel:

1: Click on the Basic Annotation icon on

the Tool Side Bar and select the
Basic Annotation option.

2: The Control Panel can be closed and

re-docked in the Process Workflow by
clicking the ‘X’ to the right of the
dialog caption.

3: Collapse the panel if it is obscuring

the Viewer or other controls by
clicking the small arrow to the right of
the dialog caption (13). Click and drag
the Control Panel dialog caption to
move it to another part of the screen.

The Basic Annotation features are:

4: Display Image Name: Go there…
5: Display a Description: Go there…
6: Show the Time and…
7: …the Date: Go there…
8: Draw an Annotation Line: Go there…
9: Set the Font and Colour: Go there…
10: Set and Reset the Defaults: Go
there…
11: Show/Hide the Control Panel: Go
there…
12: Merge: Go there…

Continued… 349

348 © 2011 Leica Microsystems (Switzerland) Limited

 Basic Annotation: Display Image Name and Description:

1: Click to enable the Image Name

check box and the name by which the
image was saved appears bottom left
of the image (6). Click and drag it to
another position if required. Whilst
working on a live image the Image
Name is not yet available but will be
displayed after the image is captured.

2: To display a description on the image,

click to enable the Description check
box. The text is displayed bottom right
of the image (7) but can be moved to
another location by clicking and
dragging.

3: Description text can be typed in the

text box beneath the Description
check box or, for a captured image…

4: …clicking the Form icon to display the

Image Data Form and…

5: …clicking in the Description text box

and typing a description.

Continued… 350

LAS User Manual 349

Basic Annotation: Display Time and Date:

1: Click the Time check box to display the current time in The illustration shows Date and Time together with the
the top right corner of the image. It can be clicked and Image Name and Description dragged to a new location.
dragged to another location. The time format (12 or
24 hour) is determined by the computer settings.
Continued… 351
2: To display the date in the regional format set in the
computer, click to enable the Date check box. If both
Time and Date are enabled they are displayed as a
single line.

350 © 2011 Leica Microsystems (Switzerland) Limited

 Basic Annotation: Draw a Line with a Caption Label:

1: Click the Show check box to enable

line drawing.

2: Click on the Up/Down arrows to the

right of the Size text box to Increase/
Decrease the width of the line. The
size of distance line ends and arrow
heads scales with the line thickness.

3: Click on the Colour button and…

4: …on the Select Colour dialog choose

a new colour from the swatches or
from the colour wheel. Alternatively,
click in the Current Colours text
boxes (7) and type the Red, Green
and Blue values.

5: Adjust the colour by clicking and

dragging the Colour Shade slider.

6: Adjust the colour transparency if

required by clicking and dragging the
slider on the Transparency bar.

8: Click OK. The new colour is shown on

the Colour button.

Continued… 352

LAS User Manual 351

Basic Annotation: Draw a Line: Continued:

1: Choose the Line Style by clicking on

the small arrows to the right of the
Style drop down menu and…

2: …click to select the required style.

The Line Only option draws a plain
line; Distance Line draws a line with
bars at each end; Arrow draws a line
with an arrow head.

A: To draw the line, click and hold down

the left mouse button at the start
point, drag to the end point (B) and
release the mouse button. Whilst
drawing, the line can be dragged in
any direction and at any angle.

3: To add a Caption Label, click in the

Label text box and type the caption. It
can be clicked and dragged to re-
position it on the image.

4: Line Width, Colour, Style and the

Label can be changed at any time
(before merging) by selecting the Edit
option from the Style menu, clicking
on the item to be changed and
altering the settings as detailed
above.

Continued 353 ...

352 © 2011 Leica Microsystems (Switzerland) Limited

 Basic Annotation: Font Selection:

To change the Font settings for all of the

Basic Annotation labels:

1: Click on the Font button.

2: On the Font dialog, select the Font

Type Face, Style and Size as
required and…

3: …click OK.

Continued… 354

LAS User Manual 353

Basic Annotation: Font and Background Colour:

The Font colour and the Label

Background colours are both selected in
the same way:

1: Click on the (Font) Colour button or

…

2: …the Background button.

3: On the Select Colour dialog choose a

new colour from the swatches or from
the colour wheel. Alternatively, click in
the Current Colours text boxes (4)
and type the Red, Green and Blue
values.

5: Adjust the colour value by clicking

and dragging the Colour Shade
slider.

6: If required, adjust the colour

transparency by clicking and dragging
the slider on the Transparency bar.

7: Click OK. The new colour is shown on

the appropriate button.

Continued… 355

354 © 2011 Leica Microsystems (Switzerland) Limited

 Basic Annotation: Defaults and Merging:

1: Clicking the Set button will save all of

the current Basic Annotation settings
for future image captures. However,
any of the settings can be adjusted on
individual images before merging.

2: Click the Reset button to return the

settings to the original installation
defaults.

3: Recall.

4: The Show check box when enabled

displays the Basic Annotation control
panel and activates the tools.

Merging:
When Merging is enabled all of the
Labels, Captions and Lines drawn on the
image will be permanently included as
part of the image and cannot be altered.

5: Merge on a captured image is a ‘one

shot’ button and only merges
annotations with the current image.

6: Merge on live images is a check box

setting and whilst it remains enabled
will merge annotations with all
captured images.

LAS User Manual 355

Process: Enhance

Enhance provides a range of easy to use

yet powerful controls for image
enhancement.

The controls work post-capture to change

Brightness, Contrast, Gamma, Hue,
Saturation and Intensity. The image can
be Flipped, Rotated or Cropped and
saved as a new image or to replace the
original.

To reach the Enhance control Panels:

1: Click on the Process Workflow.

2: Click on the Enhance tab.

3: Some Control Panels are minimised -

expand them by clicking on the small
arrow to the right of the panel.

Continued 357 ...

356 © 2011 Leica Microsystems (Switzerland) Limited

 Enhance: Crop:

Cropping is the process of removing

unwanted parts of the image leaving only
the area of interest.

1: This step is not mandatory but

strongly recommended. Right click on
the image. The Viewer menu
appears.

2: Select Send To and then select a

drive to save a copy of the image -
just in case the cropping or colour
adjustment go wrong, this is a pristine
backup.

3: Click the Fit To Screen icon on the

Side Bar to display the entire image.

4: Click on the Enhance tab.

5: Click on the arrow to the right of the

Crop header to reveal the panel.

6: Click Start on the Crop panel.

7: Click-and-hold on a point on the

image where the top left- hand corner
of the crop mask will be. Drag to the
right and down. The mask appears as
an outlined rectangle. When the area
of interest is enclosed by the mask,
release the mouse button.
The position and size of the mask (in
pixels) is displayed in the Crop panel
as the mask is being drawn.

8: Either click the Finish button if the

mask is satisfactory, or click Cancel
to clear the mask and start again.

Continued... 358

LAS User Manual 357

Enhance: Crop: Continued:

1: To keep the masked area, click Apply

on the Confirm panel or click Discard
to start again.

2: If Apply is clicked the Save Changes

prompt appears:

3: Click Replace to replace the original

image with the cropped area.

4: Click Create Duplicate to keep the

original image intact with the same
image name and also create an new
image of just the cropped area with
the same image name prefixed by
Copy of...

358 © 2011 Leica Microsystems (Switzerland) Limited

 Enhance: Orientation:

The image may be flipped - top to bottom

or side to side and rotated about its
central axis.

1: Click on the arrow to the right of the

Orientation bar to reveal it.

To flip the image top to bottom:

2: Click on the Flip Vertical button.

To flip the image side to side:

3: Click on the Flip Horizontal button.

To rotate image about its central axis:

4: Click on the Rotate Up arrow to rotate
clockwise by one degree increments
or on the Rotate Down arrow to rotate
anti-clockwise by one degree
increments, or...

5: Double-click the Rotate by window to

highlight the existing value. Type the
number of degrees to rotate and
press the Enter key on the keyboard.
The image will rotate to the required
position clockwise.
To rotate anti-clockwise, precede the
number with the negative (-) sign.

6: On the Confirm panel, click Apply to

keep the new orientation or Discard to
start again.

Continued... 360

LAS User Manual 359

Enhance: Orientation: Continued:

1: On the Confirm panel, click Apply to

keep the new orientation or Discard to
start again.

2: If Apply is clicked the Save Enhances

prompt appear. Click Replace to save
the cropped image overwriting the
original.

3: Create Duplicate to save the cropped

image with the original name prefixed
with Copy of... The original image
remains intact. Click Cancel to restore
the image and start again.

Before proceeding to colour adjustment, it

is good practice to make a copy of the re-
orientated image as a backup.

360 © 2011 Leica Microsystems (Switzerland) Limited

 Enhance: Colour:

Two panels provide a range of powerful

tools to adjust and modify the image
colours - Brightness: Contrast: Gamma
(BCG) and Hue: Saturation: Intensity
(HSI). To reveal the panels click on the
arrow to the right of the bar.

While the panels use complex

mathematics to manipulate the image,
ultimately colour is simply a matter of
perception - what our eyes and our brains
‘see’- and if what we see suits our
purpose, then the colour is ‘correct’.

Before adjusting the image colour,

consider how it is going to be presented.
For archiving or electronic transmission,
perhaps very little adjustment will be
necessary; for projection - in a Powerpoint
presentation for example - Saturation may
need to be increased to maintain colour
vibrancy on the screen; for paper printing,
Gamma and Intensity may need
increasing to keep colours ‘pure’ and
clean.

1: Print it to check how it looks and...

2: ...if the image is satisfactory, only

then Apply and keep it.

LAS User Manual 361

Enhance: Brightness:

The image in the Viewer comprises tiny,

individual ‘dots’ called pixels. Each pixel is
a mixture of three primary colours - red,
green and blue (RGB) and each colour is
represented by a value. For 8 bit colour,
the values are in the range 0 to 255.

All of the colour controls manipulate the

values of the three colours to produce a
certain effect.

If all three colour values are set to

‘0’(0:0:0) then black is the result. If they
are all set to ‘255’(255:255:255) white is
the result.

On both the Brightness/ Contrast/ Gamma

and Hue/ Saturation/ Intensity colour
panels, click and hold the slider and move
it to the left to decrease or right to
increase values. The displayed numbers
are not a reflection of the colour byte
values but rather a scale associated with
the parameter being changed.

1: Brightness: increases or decreases

the value of all three colours
simultaneously. The illustrations show
(left to right) the result to the image of
a swing of -300 to +300 with the
middle image representing the
original captured value - ‘0’on the
brightness scale. A maximum
negative value will produce a black
image and a maximum positive value
a white image. See: Process:
Enhance, Contrast and Gamma. Go
there... 363

362 © 2011 Leica Microsystems (Switzerland) Limited

 Enhance: Contrast and Gamma:

1: Contrast increases or decreases the

colour values individually both with
respect to each other and also white
levels. It is a proportional adjustment.
The three illustrations represent a
swing of 1000 in both directions with
the original image in the centre.
The perception of contrast depends
upon the ambient light levels. The
three small squares opposite are
identical in both illustrations and yet
those surrounded by black are
perceived as having a lower contrast -
they look closer to each other in
terms of colour - than those
surrounded by white. If your image is
to be projected in dim lighting
conditions, consider increasing the
contrast to compensate.

2: Gamma is a value applied to colour

levels to compensate for different
ways in which the image is viewed.
Liquid crystal displays (LCDs) have a
specific Gamma setting, cathode ray
tube (CRT) monitors will have another
and printers yet another. Changes in
Gamma are applied automatically so,
when an image is printed for
example, the printer software will
make adjustments before the printing
takes place.
Very small changes in Gamma can
have dramatic effects; the examples
show a range of 0.35 to 1.50 with the
original in the centre. Generally, avoid
altering the Gamma settings unless
really necessary.

Continued... 364

LAS User Manual 363

Enhance: Contrast and Gamma: Continued:

1: All three controls may be applied to

the image simultaneously.
To reset the changed values and start
with the original image, click Discard
on the Confirm panel. To keep the
changes, click Apply.

2: If Apply is clicked the Save Enhances

prompt appear. Click Replace to save
the cropped image overwriting the
original.

3: Create Duplicate to save the cropped

image with the original name prefixed
with Copy of... The original image
remains intact. Click Cancel to restore
the image and start again.

See: Brightness. 362

See: Hue and Saturation 365 .

364 © 2011 Leica Microsystems (Switzerland) Limited

 Enhance: Hue and Saturation:

Hue, Saturation and Intensity control the

actual colours, the amount of colour and
the vibrancy. As each is adjusted, the
lower of two Spectrum Bars (1) changes
as a comparison to the static spectrum
above it.

2: Hue is another word for colour. As the

slider is moved, the colours shift from
the dominant red in the middle
illustration which is the original,
toward green on the left or blue on the
right. The Spectrum Bar shifts to
reflect the change in dominance.
Use Hue to correct any perceived
colour imbalance, especially on
printed images.

3: Saturation determines the amount of

each colour that is present. At the
highest setting, each colour will be at
its most vibrant. The right hand
illustration is the high setting and the
colours cannot be more prominent
without combining to make white.
Use Saturation to make powerful (if a
little bit ‘unnatural’) images.
Reducing Saturation is a convenient
way of turning a colour image into a
monochrome image - essentially just
shades of grey - without losing detail
or becoming a black solid.

LAS User Manual 365

 Enhance: Intensity:

1: Intensity is close to Brightness in the

way it affects the image. It is a
measure of the ‘strength’ of each
colour swinging from solid black to
solid white. Use small increases in
Intensity to help differentiate between
colours; too much and detail begins to
disappear.

All three controls – Hue, Saturation and

Intensity - may be used together to
achieve a desired effect and they may be
combined with Brightness, Contrast and
Gamma to 'fine tune’ an image.

2: Discard, return to the original, or

Apply changes on the Confirm panel.

3: If Apply is clicked the Save Enhances

prompt appear. Click Replace to save
the cropped image overwriting the
original.

4: Create Duplicate to save the cropped

image with the original name prefixed
with Copy of... The original image
remains intact. Click Cancel to restore
the image and start again.

366 © 2011 Leica Microsystems (Switzerland) Limited

 The Analysis Workflow:

The Analysis Workflow appears when the

optional 'Analysis' modules are installed.
In this version of the Leica Application Suite,
the Interactive Measurements or Automatic
Measurements modules may appear here:

Interactive Measurements: Go there... 709

Automatic Measurements: Go there... 781

LAS User Manual 367

 Optional Modules:

The underlying capabilities of the Core functions

can be enhanced with a range of advanced
modules and applications. Each LAS module
provides the flexibility to tailor a system solution
to fulfill individual needs with upgrade options
available for future requirements.

Installing the Demo Licence and Enabling

Optional Modules: Go there... 369

Optional Modules include:

LAS Archive Basic.

LAS Archive Standard
Extended depth of focus
Movie recording
Image measurements
Macro Programming
Image Overlay
Power Mosaic
Image Analysis
Web sharing of images
Grain Expert
Phase Expert
LAS Macro

...and many others, described in this

manual and Help.

Use of the optional LAS Macro capability is

described the LAS Macro Editor help file. Go
there... 368

368 © 2011 Leica Microsystems (Switzerland) Limited

 Optional Modules: Demo Licence and Enabling

Optional Modules, once installed have to be

licensed. Users who intend to apply online for
a licence or those who already have a licence
file or dongle should consult the Installation
Guide for the next steps.

This section is for users who want to install

the 60-day Demo evaluation licence.

The Demo Licence dialog is reached through

the Framework - the part of LAS that loads
first and acts as a 'container' for all of the
other Core features and modules.

The Framework cannot be reached through

Leica Application Suite - if LAS is running it
must be closed down using the normal exit
procedure.

The way in which Framework is launched

depends upon the user rights:

A: Administrators with User Rights OFF:

1: Press and hold down the keyboard Shift To reach the Registration dialogs:
key. With the Framework loaded:
2: Double-left click on the LAS desktop 5: Click on Options and from the drop-down menu...
icon. The Application Suite Framework
loads. 6: ...click to select Registration.The Registration dialog appears.
B: Administrators with User Rights ON:
3: Right-click on the LAS desktop icon.

4: From the drop-down menu left-click on

Run as Administrator. The Application
Suite Framework loads.

Continued... 370

LAS User Manual 369

Installing the Demo Licence: Continued:

1: Click on the Demo tab on the Licence Registration 5: Click the Start All button to simultaneously enable all
dialog. of the modules.

2: Click on the Install button to install the Demo licence. 6 & 7: The Enable All and Disable All work as a 'switch'
pair to simultaneously enable and disable all of the
3: When the licence is installed the Start button modules.
becomes active: Click it to display the Status of
Modules dialog. 8: Click Close.

4: To enable a module, click on the associated Demo 9: The status of all the modules can be displayed by
button. Once a module is enabled its evaluation 'clock' clicking the Show Licence State button and selecting
will start running and cannot be turned off even if the the required option from the drop-down menu.
module is later disabled.

370 © 2011 Leica Microsystems (Switzerland) Limited

 Archives: Basic & Standard

LAS operates in 2 distinct modes depending how the Implementation Overview

user wishes to manage the acquired images. These are: Images are combined with text and numeric data,
microscope information and camera parameters in
individual records of a database that you can tailor to the
LAS Image Explorer and specific needs of the application.
LAS Folder Archive.
The content of a record is defined by use of the Archive
LAS Image Explorer is used when images are stored to Design tool on the Setup Workflow. The Archive
the hard drive in an informal manner. The user can save Designer allows you define hierarchical levels by which
the images anywhere on the hard drive in any folder and data is grouped (e.g. Lab Name >Procedure >Customer
organizes the images as convenient. While this method Name > Experiment >Specimen Number >Result). There
has the benefit of simplicity, there is a chance that over is virtually no limit to the number of different fields or the
time the images will become harder to find. volume of information stored. The original high-resolution
images are held external to the database. New fields can
LAS Folder Archives are used where a more disciplined be added to an existing database or redundant fields
approach to the organization of data is demanded. This removed without difficulty. System information, including
might occur where it is mandatory to save additional user operator name, data and time are added automatically,
defined fields with the images or where several users while all the microscope and camera parameters are
need to add data to images in a systematic manner. This included by default for both Basic and Standard editions.
also means that images are saved in locations where
other users will be able to find them. In this case it is a Continued... 372
distinct advantage to restrict the locations that the images
are saved. We call these ‘Folder Archives’ because the
images are still saved to the Windows file system but
only in predefined locations.

LAS User Manual 371

Archives: Basic & Standard: Data Organisation

LAS keeps the metadata in a file associated with the If you need to export image files directly to external
image and in the same location although this file is media or to a personal file space, you should use the
normally hidden. If you do make changes to these files export tool. This will ensure that a copy of the image is
you do so at your peril! For the purposes of managing made and you will not then be likely to damage or destroy
your disk space, you can choose where the database and vital data in the database. The exported images from a
associated files are kept if you want to Because the sequence will each use the image name as part of the file
archive uses the normal Windows file system, you can name. This ensures that sequence of images will be
use standard back-up tools. readily recognizable by you once exported.

Once the Archive is created, it remains in the location on Summary:

the hard drive that was originally specified. When the In order to achieve a fast, reliable and scalable tool for
Archive is created, the location of the images and microscope data capture and processing, it is important
metadata is also specified. However, the images can to use modern database technology and to enforce some
easily take up many Gb of space and may fill your hard simple rules about how the user interacts with that data.
drive. If this happens, the entire archive can be moved This may impose some limitations on the ways in which
using normal Windows tools to an alternative location. you work with your data and may mean that you have to
This location can be on a remote drive but please be change some of the procedures you operate in your
aware that this will affect the performance of recall for work. In creating LAS Archive, we have taken great
large images and be dependent on the network speed. trouble, however, to provide new features that will allow
you to find new working practices to achieve your
When more than one user accesses the same data, workload and in a way that will allow traceability,
there is inevitably some security needed. This may be a scalability and security for the future.
matter of preventing less experienced users from
accidentally damaging or destroying valuable data. LAS
supports the Windows user log-on so that if you wish you
can create databases with restricted access.

Each image in the database has an image name.

Software in LAS ensures that image names are unique to
a folder by appending an incrementing number where
possible and by providing numbered sequences for the
sequence modules.
Clearly it is vital that you have easy access to the images
that are in the database and LAS provides several
feature to expedite this.

372 © 2011 Leica Microsystems (Switzerland) Limited

 Archives: Basic & Standard: An Introduction to Archiving: Archive Structure

Every single image or collection of

images is associated with a Record. It is
a self-contained entity storing not only the
actual image data but a wealth of
information associated with it. Images
may be:

1: Single images.

2: Image sequences such as time

lapse, size only being constrained by
the hardware and preferences.

3: Multiple images like those from

Montage.

4: Image groups produced as Z-

Stacks…
…or any created and modified within
Leica Application Suite.

Continued... 374

LAS User Manual 373

 Archives: Basic & Standard: An Introduction to Archiving: Archive Structure: Continued:

Depending upon the hardware, each

image has data automatically stored with
it:

1: System Data such as image bit

depth, size and sequence.

2: User Data like image name, creation

date and time.

3: Microscope Data that will allow the

conditions to be quickly replicated –
model name, mag-changer,
nosepiece and so on.

4: Camera Data relating to exposure,

gamma, brightness and all of the
essential settings.

The data is stored in Fields within the

Record. All of the essential Fields
appropriate to the hardware setup have
been pre-defined by Leica – it is only
necessary to click Acquire Image – but
for more versatility users can add their
own Fields and definitions with the LAS
Basic and Standard Editions.

374 © 2011 Leica Microsystems (Switzerland) Limited

 Archives: Basic & Standard: An Introduction to Archiving: The Example Archive

LAS Archive has a pre-defined and pre-

loaded archive called Example Archive,
which is an archive structure only; it does
not contain data. This is what you see
immediately after LAS Archive Basic
Edition is installed and started for the first
time.

Example Archive is a Single-Level

Archive (1). This means that all of the
records are directly associated with
Example Archive – or whatever a
structural copy is named – it cannot have
Level 2 record groups.

Example Archive is ready to use

immediately. It is the fastest way - just a
few minutes - of getting into production,
especially for a single-station user.

LAS User Manual 375

Archives: Basic & Standard: Editions: Basic

There are two editions of LAS Archive

both purchased as optional modules:

Basic Edition: Essential functionality

and tools for LAS Archives.

Standard Edition: Builds upon the

facilities available in the Basic Edition
and additionally allows the creation of
multi-level archives with many field
types in addition to text fields.

Basic Edition:
Provides for 1 or 2-Level archives. This
means that image records can be
associated directly with the Primary
Archive (1) or indirectly by grouping shown
on the illustration as (2) and (3) – Record
Group A and Record Group B.

The Primary Archive is Level 1 and the

Record Groups are Level 2. With Basic
Edition as many Primary Archives as
required can be created, each with a
different name. In turn, they can contain
as many Record Groups as needed also
with unique names. However, Record
Groups can have the same name
providing they reside in different Primary
Archives.

Basic Edition also features:

Microscope and camera data

captured automatically with the
image.

Fast Archive Search with detailed

filtering to locate specific images and
data.

User Field Selection for Form display.

Attaching documents to image in any

format - not just text.

Audio recording inclusion with image.

Add Multiple Text Boxes to the
Archive.

376 © 2011 Leica Microsystems (Switzerland) Limited

 Archives: Basic & Standard: Standard Edition

Optional module Standard Edition Archive

extends the power and flexibility of the
Basic Edition to provide:

Multiple archive levels.

Wide choice of Named Archive Fields

- Memo, Boolean, Numeric, Date and
Keywords.

Form Layout design and control.

Highly Detailed Reports to include

scaled images. Export reports to
Adobe Acrobat (pdf) and Browser
(html) facility.

Increased flexibility and security by its

use of specialised fields - fields into
which only pre-determined data can
be loaded.

LAS User Manual 377

Archives: The Navigator Panel: LAS Archives

The illustration shows a typical Browse

Navigator Panel with an archive selected.
LAS Archives are optional modules that
can run on the same computer as Image
Explorer although not at the same time.
See Selecting the Storage method: Go
there… 371

The panel layout with LAS Archive running

displays the Archive tab (1) and the
Search tab (2). The rapid Search facility
(4) is also available.

LAS Archives has addition function

buttons (3) compared to Image Explorer.

378 © 2011 Leica Microsystems (Switzerland) Limited

 Archives: Select Image Explorer or LAS Archive

The image storage method – Image

Explorer or LAS Archive – is selected on
the Setup Workflow.

1: Click on the Setup Workflow.

2: If necessary, open the Archive panel

by clicking the tab.

3: To select and use Image Explorer,

click the LAS Image Explorer button.
Image Explorer and LAS Archive
cannot run simultaneously so clicking
a button cancels the other. Return
immediately to Browse by…

4: …clicking the Workflow.

5: Select LAS Archive by clicking the

button.

6: The Archive toolbar,…

7: …the Archive List Window and…

8: …the Management controls

immediately become active.

LAS User Manual 379

Archives: Archive: Activate Archive:

1: With Archive selected…

2: …a previously selected archive (if

any) will be highlighted and checked –
a green tick mark to the left – will be
displayed in the Archive List Window.

3: To change to another archive either

double-click it or click it once and then
click the Set As Current (Active)
Archive button (4).

5: The selected archive is activated and

the green tick mark appears to the
left. Return to Browse by clicking on
the Workflow.

380 © 2011 Leica Microsystems (Switzerland) Limited

 Archives: Archive: Rename an Archive:

To rename an existing archive, ensure

that LAS Folder Archive is selected and...

1: Click on the Rename button.

2: On the Rename dialog, click on the

small arrows to the right of the
Archive Name header and from the
list of archives...

3: ...click to select the one to be

renamed.

4: Click in the New Archive Name text

box and type a new name.

5: Click OK.

6: Click OK on the Rename Archive

Confirmed dialog.

LAS User Manual 381

Archives: Archive: Delete an Archive:

Use with caution as a deletion cannot be

reversed.

1: An archive cannot be deleted if it is

currently active.

2: …click on the Delete button.

3: On the Delete Archive dialog click on

the arrows to the right of the Archive
Name header and from the drop
down list...

4: ... click to select the archive name.

5: Type the name of the user that

installed Leica Application Suite in
the User Name text box. Most often
this will be Admin.

6: Click to check the Confirm Deletion

check box.

7: Click OK.

8: On the confirmation dialog click OK

and the archive is deleted.

382 © 2011 Leica Microsystems (Switzerland) Limited

 Archives: Archive: Create a New Archive:

Concise details for creating different types

of archive are described in Optional
Modules > LAS Archive > Basic Edition:
Go there... 409

LAS User Manual 383

Archives: Archive: Convert an Existing Archive:

The speed and efficiency improvements

made by Microsoft in their latest SQL
Database Engine are included from LAS
Archive Version 3.3 to keep it in the
forefront of data processing technology.
So that archives created with software
earlier than V3,3 are not lost, a Convert
Archive tool has been included in Archive
Management.

To convert an archive:

1: Ensure that Folder Archive is selected

and click on the Convert LAS V3.2
Archive button.

2: On the Convert dialog, click on the

arrows to the right of the Archive to
Import header and from the list of
available archives, click to select the
one to import.

3: Click inside the Imported Archive

Name text box and type a new,
appropriate name for the imported
archive.

4: Click on the browse button to the right

of the Root Folder text box and
navigate to the location that the
imported archive will be stored. On
the navigation dialog it is possible to
create a new folder.

5: Click OK. The existing archive

structure is copied - the archive
remains intact - to the new location
with necessary upgrades to the
structure. None of the images or their
associated data will be lost or
corrupted.

384 © 2011 Leica Microsystems (Switzerland) Limited

 Archives: LAS Archive: Main Panel

A complete list of available archives is

displayed by:

1: Clicking on the Setup Workflow.

2: Clicking to select LAS Folder Archive.

3: The list appears in the Archive

window with the currently selected
archive (if any) having a green tick
mark to the left. An archive can be
made active by double-clicking on it.

4: With an archive selected and active in

Setup, when the Browse Workflow is
selected...

5: ...and the Browse tab visible...

6: ...the Archive and Search tabs are

revealed and...

7: ...the active archive displayed in the

Navigator window.

8: If the Gallery is enabled the archive

thumbnails will be displayed and the
first image and its data present on
the Viewer.

Continued... 386

LAS User Manual 385

Archives: LAS Archive: Swapping Modes:

Swapping in Navigator between the

conventional folders of Image Explorer
and LAS Archive is as simple as a mouse
click.

In the illustration, an Image Explorer

folder is currently selected - the Folder
tab (1) is visible and the selected folder is
highlighted.

An archive name is displayed further down

the tree (2) and is indicated by the Leica
Cube icon to the left of the name.

Simply double-clicking on the archive

name will change the storage mode from
Image Explorer to LAS Archive - the
Archive tab (3) appears...

...and opens and loads the images (4).

Go back to browsing in Image Explorer

Folder mode by clicking on the Return
button (5).

386 © 2011 Leica Microsystems (Switzerland) Limited

 Archives: LAS Archive: Renaming an Archive:

Rename an Archive by:

1: Click on the Archive to select it.

2: Click on the Rename button.

3: The Archive Name in the Navigator

window changes to a highlighted
outline. Type the new name and
press Enter on the keyboard.

The Archive now appears under its

new name in the Navigator.

LAS User Manual 387

Archives: LAS Archive: Delete an Archive:

To Delete an Archive:

1: Click on the archive to be deleted.

2: Click on the Delete button.

3: For security reasons, an archive that

contains images and data cannot be
deleted - they have to be removed
first by...

4: ...holding down the keyboard Ctrl key

and...

5: ...clicking on the individual thumbnails

in the Gallery.

6: Click on the Delete (Trash Can)

button on the side tool bar.

Continued... 389

388 © 2011 Leica Microsystems (Switzerland) Limited

 Archives: LAS Archive: Delete an Empty Archive:

Continued from previous page:

1: Confirm that the images and data

(Records) should be deleted by
clicking the Yes button.

2: To delete the empty archive click on

the Delete button and...

3: ...confirm the deletion on the Delete a

Folder dialog by clicking the Yes
button.

LAS User Manual 389

 LAS Archive Basic Edition

With the optional Basic Edition Archive installed, all of the

features and tools needed to produce powerful and Basic Edition Archive also includes features to bring even
versatile Archives are immediately available to create: more sophistication to archiving:

User designed Single Level Archives. Fast Archive Search with detailed filtering to locate
specific images and data.
User designed 2-Level Archives with almost unlimited
Record Groups (Folders). Attaching documents of any format - not just text.

Archives using existing structures either from User Audio recording.

designs or the comprehensive library of templates
supplied as part of the module. Adding Multiple Text Boxes to the archive.

As well as User defined data fields, microscope and

camera data is captured automatically with the image.

Elegant and detailed Form display for every image.

Forms can be configured to display the data that the

User requires and hide everything else.

390 © 2011 Leica Microsystems (Switzerland) Limited

 LAS Archive Basic Edition: Create a New Archive: Introduction:

This section describes setting up both a

simple single level archive and a
comprehensive 2-Level archive.

Three options are available when

creating a new archive:

From New: Starting from a

completely blank structure and
configuring precisely to the user
requirements.

From Template: Creating an archive

based upon a template that can be
either supplied in the Leica Template
Library or one created by the user
previously.

From Archive: Copies the structure

and fields of an existing archive.

Archives are created in the Setup

Workflow:

1: Click on the Setup Workflow and if

necessary...

2: ...click on the Archive tab.

3: On the Storage Method panel click

Folder Archive.

4: The Archive Toolbar and...

5: ...the Archive window together with...

6: ...the Management tools become

active.

Continued... 392

LAS User Manual 391

 LAS Archive Basic Edition: Create a New Archive:

To create a completely new archive that can be structured 3: On the Define Folder Archive panel click in the Name
precisely to the users needs: text box and type a unique name for the new archive.

1: Click on the Create Archive button on the toolbar. The

New Folder Archive dialog appears. Continued... 393

2: On the Source panel click the New button. The three

Source options are mutually exclusive - clicking one
automatically de-selects any other.

392 © 2011 Leica Microsystems (Switzerland) Limited

 LAS Archive Basic Edition: Create a New Archive: Storage Location:

1: Click on the browse button to the right of the Select 3: Click OK. The new storage location appears in the
Location text box. Select Location text box.

2: On the Windows Navigation Browse for Folder

dialog, navigate to the location where the new Continued... 394
archive is to be stored.

LAS User Manual 393

LAS Archive Basic Edition: Save for a Single Level Archive:

1: Click on the OK button. 3: Users that require just a simple archive should just
click the Save button (Bottom right-hand of the
2: The Single Level Archive Structure appears. This Viewer). There is no need to change any of the data
will be the basis for the Record Form for each field names.
captured image. To create a 2-Level Archive: Go
there...
Continued 395 ...To start capturing into a single level
archive.

394 © 2011 Leica Microsystems (Switzerland) Limited

 LAS Archive Basic Edition: Start Capturing Images:

With the new archive saved:

1: The new archive name appears in

the Archive List and is set as current
and active.

2: To start capturing images click on

the Browse Workflow and...

3: ...click on the new archive.

4: Click on the Set as Capture Location

button to ensure that all images are
saved in the new archive and...

5: ...start capturing images either in

Browse or in the Acquire Workflow.

Acquire: Camera setup and image

capture: Go there... 221

LAS User Manual 395

LAS Archive Basic Edition: Create a 2-Level Archive:

1: A new archive called Copper

Impurities has been created as
previously described. This is intended
to be a 2-Level archive which means
that within the archive separate
collections of images will be captured
and stored as Record Groups, each
having a unique name.
There will be three Record Groups in
the archive - Batch #12, Batch #13
and Batch #18 - chosen to reflect the
job in hand. Figure (A) illustrates the
structure.

2: Every image within the archive, no

matter what Record Group it is stored 3: The default reference for the image is Image Name but this can
in will have all of the System Data also be changed by clicking in the User text box and type a new,
Items - capture date, exposure more appropriate name, in this example is is called Sample. The
settings, microscope stand and so on Abc to the right of the text box indicates that the new name can
- saved with it. This data is displayed comprise letters and numbers.
as Fields on the Form associated with
each image. The default name for the 4: The next step is to create the Record Groups. Click on the New
Form is Image Data but can be Level button.
changed to something more
appropriate. Click in the Alias text box
and type a new name. In the example, Continued... 392
the new Form Name is Copper
Impurities Results.

396 © 2011 Leica Microsystems (Switzerland) Limited

 LAS Archive Basic Edition: New 2-Level Archive: Name the New Level:

Continued from the previous page: 3: In this example each batch will be given a reference
1: After the New Level button is clicked a dialog number - #12, #13 and #18 - and that will appear as
appears. This represents the structure of any the first User Field (pre-loaded with the example text
Record Groups added to the archive. Folder Name). Click in the first User Field text box
and type an appropriate name - Batch Number in
2: The three Record Groups that re going to be added the example.
will be called Batch #12, Batch #13 and Batch #18
so in this example the structure will be given the
name Batch. Click in the Alias text box (this is pre- Continued... 398
loaded with the example text New Folder) and type
an appropriate name - Batch in the example.

LAS User Manual 397

 LAS Archive Basic Edition: New 2-Level Archive: Add a Field:

Continued from the previous page:

1: In this project each Record Group is to have an 3: The Source Reference is a vital and required piece
additional field to identify the source of the sample of information so the Required check box is enabled
batch. A new field to contain the Source Reference which means that an image can only be acquired if
is added by clicking the appropriate Fields button - this field is complete.
in this example it is a numeric code so the 123 Almost any number of User Specific Fields can be
(Numbers only allowed in this field) button is clicked. added in this way; This example requires only two
described so...
2: The new field is added to the Record Group with the
selected field type to the right - in this case 123. 4: ...click the Save button.
Click in the Field text box and type an appropriate
name - in this example Source Reference.

398 © 2011 Leica Microsystems (Switzerland) Limited

 LAS Archive Basic Edition: New 2-Level Archive: In Setup and Browse:

1: The new 2-Level archive now

appears in the Setup Workflow >
Archive window and is selected and
ready to be used.

2: In the Browse Workflow it appears on

the Archive tab with...

3: ...the Form Name (Copper Impurities

Results) and ...

4: the Record Group (Batch) both

ghosted in the Viewer. These will only
be displayed if the Hide/Show Form
button on the Side Tool Bar is
enabled (5).

Continued.. 400

LAS User Manual 399

LAS Archive Basic Edition: New 2-Level Archive: Create a Record Group:

1: Click on the archive to select it.

2: Click on the Create New (Batch)

button.

3: A new folder icon appears to

represent the Record Group with the (
Batch) name against it.

4: Rename the new Record Group - in

this example it will be called #12 - by
clicking on the Rename button.

5: Type a new name for the Record

Group and press Enter on the
keyboard.

6: The new name appears on the Form.

Continued... 393

400 © 2011 Leica Microsystems (Switzerland) Limited

 LAS Archive Basic Edition: New 2-Level Archive: Set the Capture Location:

1: Before images can be captured to the

correct Record Group, it has to be set
as the Capture Location. Click on the
Record Group and...

2: ...click on the Set Capture Location

button. A red dot appears to the left of
the Record Group to indicate it is set.

3: Capture images in Browse or in

Acquire.

4: As images are captured the System

Data is displayed on the Form.

5: Because in this example Source

Reference is a required field, it must
be completed before an image can be
saved.

For details of Deleting and Importing an

archive, see Browse: Go there... 421

LAS User Manual 401

 LAS Archive Basic Edition: Copying an Archive Structure:

It is a simple and very fast matter to copy an archive 4: Click on the Browse button to the right of the Select
structure - all of the pre-defined fields but no data - give Location text box and...
it a meaningful name that reflects the tasks in hand and
then start capturing images and data into it. 5: ...on the Browse for Folder dialog, navigate to the
folder (6) in which to save the new archive and...
1: Click on the Setup Workflow.
7: Click OK. The path of the new location appears in
2: Click on the Create Archive tab and the New Folder the Select Location text box.
Archive dialog appears.

3: Click in the Name text box and type a name for the Continued... 404
new archive.

402 © 2011 Leica Microsystems (Switzerland) Limited

 LAS Archive Basic Edition: Copying an Archive Structure: Continued:

1: Click on the From Archive button. 3: ...click to select the required archive

2: Click on the arrows to the right of the From Archive 4: Click OK.
header to reveal a list of archives available for
copying and...
Continued... 403

LAS User Manual 403

LAS Archive Basic Edition: Copying an Archive Structure: Continued:

1: A new archive with a new name appears with all of 3: Click Save to save the new archive.
the fields and their properties based upon the
original archive. 4: It appears in the Archive List, is selected and active
and ready to be used just like its 'parent'.
2: The Field Names can be changed as required by
clicking in the text boxes and typing a new,
appropriate name. The field type cannot be
changed.

404 © 2011 Leica Microsystems (Switzerland) Limited

 LAS Archive Basic Edition: Archives as Templates:

Any archive may be used as a template

for future archive creation, especially
useful if a range of archives have to share
the same corporate or end-user style.

Only the structure and field names of the

source archive are replicated – not the
actual data or images.

1: Click to select the Setup Workflow.

2: Click to select the archive to be used

as a source for the template.

3: Click the Edit button. The archive

fields will appear.

4: Click on the Save as Template

button.

5: On the Windows dialog navigate to

the folder in which to save the
template.
Leica Application Suite has a default
folder in which to store templates at:
C:\Documents and Settings\All
Users\Documents\Leica Application
Suite\Archive Templates
…and it is recommended that this be
used wherever possible.
Give the template a new and unique
name.

6: Click Save and the template will be

created.

Continued... 406

LAS User Manual 405

 LAS Archive Basic Edition: Archives from Templates:

A new archive can be created quickly and easily using 4: Click on the Browse button to the right of the Select
either a template saved by the user or from the range of Location text box and...
pre-configured templates provided by Leica and
designed to suite a wide range of applications and 5: ...on the Browse for Folder dialog...
disciplines.
6: ...select the folder in which the new archive will be
1: Click to select the Setup Workflow. saved.

2: Click the Create New button. 7: Click OK.

3: On the New Folder Archive dialog, click in the Name

text box and type a name for the new archive. Continued... 407

406 © 2011 Leica Microsystems (Switzerland) Limited

 LAS Archive Basic Edition: Archives from Templates: Continued:

1: On the New Folder Archive dialog click the From C:\Documents and Settings\All
Template button. Users\Documents\Leica Application Suite\Archive
Templates
2: Click on the Browse button to the right of the From
Template text box. 4: Click to select the template style required. The
name appears in the File name text box.
3: The pre-configured templates supplied by Leica are
stored in a reserved location: 5: Click Open.

Continued... 408

LAS User Manual 407

LAS Archive Basic Edition: Archives from Templates: Continued:

1: The new archive created from the template appears 3: Click on the Save button to save the new archive.
with all of its pre-defined fields.
4: The new archive appears in the Setup > Archive
2: Field names can be changed by clicking in the Field window,
text box and typing an appropriate name. The Field
Properties can be set or cleared but the Field style
cannot.
The Unique property when set means the value in
the field cannot be the same as any other.
The Required property when enabled means that
the image will not be saved until there is some valid
data entered in the field.
To set/clear (enable/disable) a property click on the
check box to the left.

408 © 2011 Leica Microsystems (Switzerland) Limited

 LAS Archives Basic Edition: The Data Grid:

The Grid displays data for all of the images in a folder in a 4: Clicking on an entry in the Grid will immediately
tabular structure. The image names are listed on the left display that image in the Viewer and also highlight the
and the data items as headers across the top. thumbnail.
Hold down the keyboard Ctrl key whilst clicking to
1: The Grid is revealed and hidden by clicking on the make multiple selections prior to deleting or
Side Tool Bar button. This is a toggle – click to reveal, exporting.
click again to hide. Keyboard combination Ctrl + A will select all of the
image data: Ctrl + C will copy all the selected image
2: The header positions can be changed by clicking and data to the clipboard and Ctrl + V will copy into
holding the left mouse button on the header to be another application.
moved, dragging it to the new position and releasing
the mouse button. 5: A small arrow is revealed when a header is clicked.
This allows the image data to be sorted – high-to-low
3: The column widths can be changed by clicking and or low-to-high – by successive clicks on it.
dragging the vertical bars that separate the columns.
6: The Grid data can be exported to a range of other
applications by right-clicking on the Grid then
navigating to and clicking to select an application.

LAS User Manual 409

LAS Archives Basic Edition: Search:

The Search option provided with optional

Basic and Standard editions, is both fast
and flexible. It is possible to search on all
fields within a record group or on specific
selected fields. An editor is available to
create search configurations which can be
used at any time simply by clicking them.

Rapid Search:
Locating specific items which have known
names or text strings, is achieved quickly
by:

1: Select All Fields in the Search Toolbar

field selector - bottom right of the
Viewer.

2: Type the name or text string into the

Search For window.

3: Click on the Search button.

4: If a match is found the Search tab is

automatically selected showing the
Archives in which the search was
made, and...

5: ...the appropriate record is populated

- if there are more than one that
satisfies the search criteria, the first is
selected.

6: The image Thumbnail(s) appear in

the Gallery.

410 © 2011 Leica Microsystems (Switzerland) Limited

 Search: Create a New Configuration:

A Search Configuration is a file that

contains all of the parameters - such as
fields and search strings - needed to carry
out a fast, repetitive search.

Search Configurations are stored under

unique names and can be retrieved and
used for fast searching, or modified to
reflect changing search requirements - for
example the search fields could be
extended and search strings altered.

Mixing field types in a single configuration

has made LAS Archive Search even more
powerful. Now, up to 10 fields of different
types - text, Boolean, date and so on, -
can be mixed and each setup for different
search criteria. And Boolean limits can be
set to ensure that the search is narrow
and very precise; Use the AND command
to ensure that only those images that
conform to every field search string are
returned; Alternatively, the OR command
will only retrieve images that fulfill at least
one search parameter.

Create a New Search Configuration:

1: Click on the Browse Workflow.

2: Click to select the Browse tab and...

3: ...check that the correct archive is

displayed in the Archive window.

4: Click on the Configuration Editor

launch button on the Search Toolbar.

LAS User Manual 411

Search: Create a New Configuration: Continued:

1: On the Search Configuration Editor dialog, click the 5: The fields to search can be set to All - the entire
New button to create a new configuration. record set - or narrowed to include only specific
groups by clicking on the arrows to the right of the
2: Give the new configuration a unique name by clicking header and...
in the Configuration Name text box and typing.
6: ...clicking to select the group to be searched.
3: Click OK.

4: The new name appears in the Search Configuration

list.

412 © 2011 Leica Microsystems (Switzerland) Limited

 Search: Create a New Configuration: Continued:

All of the available fields are shown on the left pane of the 3: ...the field name appears in the Selected Fields pane.
Search Configuration Editor. To include a field in the Any number or any type of field can be selected.
search:
4: To remove a field from the configuration, click to
1: Click on the field to select it. select the field in the Selected Fields pane and click
on the De-select button.
2: Click on the Select button and...

LAS User Manual 413

 Search: Create a New Configuration: Continued:

The search format for a field depends upon the field type - 3: Now the actual search string has to be entered. Click
Text and Memo fields will be searched for character in the window and type the required string to search
strings, Numeric for numbers - and so on. The appropriate for. In the example 'M60' has been entered so any
format options are automatically displayed for the image with a Sample ID field containing the
selected field and are explained on the following pages characters 'M60' will be returned as fulfilling the
(Go there...). The illustrations show a text field called search criteria. The 'Contains' option means that text
'Sample ID'. such as 'Sample M60' or 'Batch M60 Local' will satisfy
the criteria.
The search format is selected by: The search string appears in the Selected Fields
pane to the right of the Field name.
1: Click on the arrows to the right of the format header.
4: Click the Save button to save the configuration.
2: From the drop down menu click to select the desired
format. In the illustration the 'Contains' format has
been chosen.

414 © 2011 Leica Microsystems (Switzerland) Limited

 Search: Options:

Each field type has a range of search

string options associated with it which are
displayed automatically. The types and
options are:

Text and Memo options: Accepts

numbers and characters in the search
string:
1: Click on the arrows to the right of the
header and click to select from the
drop down.

=: The field data must match exactly the

search string.

Contains: The search string can appear

anywhere within the field data.

Starts with: The field data must begin

with the search string.

Ends with: The field data must end with

the search string.

Between: Two search strings are

entered and each computed as an
ASCII string value - converted to a
number. The field data, also
converted to an ASCII value, must lay
between the two.

Is Null: The field must be empty

(nothing).

LAS User Manual 415

Search: Options: Continued:

Date options:
2: Selected from the drop down with
additional options available if the
down arrow (6) is clicked.

The Date Picker provides a simple

way of moving between Years and
Months (click the arrows (4), and by
clicking on the required day (5).

On: Date in the field corresponds

exactly with the date selected.

Before: The field date must occur

before the selected date.

After: The field date must occur after

the chosen date.

Between: Two date windows (3) open

and a date is selected for both. The
field date must occur between the
two.

Is Null: No date in the field: Nothing.

Within options:
Last Day: During the last 24 hours.
Last Week: During the last 7 days. If it is
10am on Tuesday all records that
have a date and time equal or later
that 10am on the previous Tuesday
will be found.
Last Month: If the current date is 17
March then all records from and
including 17 February of the same
year are found. Leap years are
automatically accommodated but the
time of day is ignored.
Last Year: All images with a date on or
later than the same date during the
past year will be found. Leap years
are accommodated automatically but
the time of day is ignored.

416 © 2011 Leica Microsystems (Switzerland) Limited

 Search: Options: Continued:

Numeric options:
1: Click on the arrows to the right of the
header.

2: Values are entered in the text boxes.

The option chosen will determine if 1
or 2 text boxes open.

=: The field value must match the

search value exactly.

<: The field value must be smaller than

the search value.

>: The field value must be greater than

the search value.

Between: Two values are entered for

the search value limits and the field
value must lay between the two.

Is Null: No value in the field: Nothing,

not even zero (0).

Boolean options:
3: Select either True or False. The field
setting must match.

Multiple Field Search:

4: Up to 10 fields of any type 'mix' of
can be selected for a search and in
this case the Search Combination is
enabled. It is based upon two
Boolean parameters...

5: ...AND determines that all fields must

conform to the search strings to
return an image.

6: OR means that one or more fields

must satisfy the search conditions to
return the image.

LAS User Manual 417

 Search: Configuration Delete and Save:

Delete a Search Configuration: Save a Configuration As...

1: Click on the arrows to the right of the Search 4: Clicking the Save As button will...
Configurations menu and click to select the
configuration to be deleted. 5: ...reveal the Search Configuration dialog. Click in the
text box and type a new name for the configuration.
2: Click on the Delete button.
6: Click OK to save the search configuration.
3: Confirm the deletion. Deleted configurations cannot
be recovered. Revert:
7: The Revert button will set the configuration to the last
one saved. Any changes made since will be lost.

418 © 2011 Leica Microsystems (Switzerland) Limited

 Search: Run:

If a new configuration has just been

created, all that is necessary is to click on
the Search button (5) to run the search.

Search using an existing

Configuration:
To run a search using a previously
created configuration:

1: Click on the Archive tab and...

2: ...check that the required archive is

selected and active.

3: Click on the arrows to the right of the

Search Configurations header on the
Search bar.

4: From the pop-up menu, click to select

the configuration required.

5: Click on the Search button.

LAS User Manual 419

Search: Run: Continued:

1: If a match is found the Search tab is

automatically selected...

2: ...showing the Archives in which the

search was made, and...

3: ...the appropriate record is populated

- if there are more than one that
satisfies the search criteria, the first is
selected.

4: The image Thumbnail(s) appear in

the Gallery.

420 © 2011 Leica Microsystems (Switzerland) Limited

 LAS Archive Basic Edition: Attach:

Any other file, including Audio and Text

can be attached to an archive using the
Attachments panel in the Browse
Workflow. This feature is only available
with Basic and Standard Editions.

Audio, either speech or music is limited to

a running time of 2 minutes, and text files
should be no greater that 32 MB.

1: Click on the arrow to the right of the

Attachments header to reveal the
Audio panel (2) and Text File panel (3)
.

To attach, play and erase an Audio

file:
Audio input can come from either the
computer microphone input or the line
input for an external CD or tape
player.

4: Click on the Record button to start the

recording.

5: Click the Stop button to stop recording

which will automatically stop at 2
minutes duration. The length of the
recording in seconds is displayed
beneath the Stop button.

6: To play the recording click on the Play

button.

7: Halt the playback with the Pause

button and click it again to resume
playback. The recoding Length and
current position as shown on the
panel.

8: Erase the recording by clicking the

Trash button.

LAS User Manual 421

Attach: Attach, View and Detach a File:

1: Click on the Attach button.

2: On the Windows Open dialog,

navigate to the file to be attached,
click to select it and…

3: …click Open. The file is attached to

the database and…

4: …it is listed in the Filename window.

In principle any number of text files
may be attached to a database limited
only by an individual file size of no
more that 32 MB and an overall
database size of 4GB.

5: To detach a file, click on the file to be

deleted in the Filename window and
then on the Detach button.

6: Click on the Launch button to display

the selected file in Windows Notepad
(7).

422 © 2011 Leica Microsystems (Switzerland) Limited

 LAS Archives Standard Edition

The Standard Edition Archive extends the power and

flexibility of the Basic Editions to provide:

Multiple archive levels.

Wider choice of Named Archive Fields - Memo,

Boolean, Numeric, Date.

The Keywords feature establishes preferred field

descriptions.

Highly Detailed Reports to include scaled images.

Export Report to Word, Adobe Acrobat (pdf) and

Browser (html) facility.

LAS User Manual 423

LAS Archive Standard Edition: Additional Fields:

The Standard Archive option provides a

further 4 field types:

1: Numeric: 32-bit floating point

numbers.

2: Memo: Alpha-numeric with over

32,000 characters. Line returns are
added automatically as the text is
entered.

3: Boolean: Yes/No: True/False. On the

Report this appears as a check box
which, when enabled equates to true
(Yes).

4: Date: On the report this is selected

using the Date Picker by...

5: Clicking on the arrow to the right of

the date text box.

6: Using the left and right pointing

arrows to scroll through months and
years, either forward or backward.

7: Clicking to select the date required.

Today's date is circled in red.

To add a new field to an archive:

With the archive active:
Select Edit mode.

Click to select either User or Image

field groups.

Click on the required field button and

the new field will appear.

The name can be altered by clicking

in the field window and typing a new
name.

Click the Save button to save the

amended archive.

424 © 2011 Leica Microsystems (Switzerland) Limited

 Create a Report: Select a Standard Template:

A report contains an image or collection of images

together with associated data extracted from the database
and presented as a document in Microsoft Word, Adobe
Acrobat Pdf or Browser Html format.

Reports are formatted according to predefined templates

that are created in Microsoft Word (2003 or later) to create
report templates.

Although the information and images comes from and is

generated by the Database, the report is stored using
Windows navigation and folders which allows other
applications to use it.

Standard Templates determine how many images may be

displayed on a page. Images and text are automatically
scaled to fit the page.

1: Click on the Create a Report icon.

2: To use a Standard Template supplied with LAS

Archives, click on the arrows to the right of the window
and from the drop-down template list...

3: …click to select the option required.

Continued...

LAS User Manual 425

 Create a Report: Select a User Template:
To select a template created by a user:

1: Click on the Create a Report icon.

2: On the Create Report dialog, click on the browse

button.

3: On the Windows dialog, navigate to the folder in which

the user templates are stored and...

4: ...click to select the template.

Creating a User Template: Go there... 432

426 © 2011 Leica Microsystems (Switzerland) Limited

 Create a Report: Selecting Multiple Images:
If multiple images are to be shown on the
report all must be selected:

1: On the keyboard, press and hold down

the Shift key and click on each of the
thumbnails of the required images or…

2: Press and hold down the Shift key and

click on the image entry on the Grid.

3: The examples opposite show the effects

of choosing a template which will display 4
records and...

4: ...another to display 16 records.

Images and text are automatically scaled to fit

the text box.

LAS User Manual 427

 Create a Report: Image Embedding:
Reports created using Microsoft Word can include images Images can be very large and occasionally – especially in
in two ways: reports that have many images – there is insufficient
machine memory and a Report Generation Error occurs.
Image Embedding: Copies the image into the report so
that it becomes an intrinsic part of it. Embedding can To cure the problem either:
result in very large files but they are always integrated - Reduce the number of images.
text and images cannot be separated.
Acquire smaller images by using a more compressed file
Image Linking: A path is established between the report format and lower resolution.
and the image source and the image is included only
when the report is displayed, printed or exported - by e- Resample the images to a lower resolution.
mail for example. Linking creates small, compact files but
if the image source is moved or changed the link is lost Use a report template that allows the use of LAS high
and the images will not appear. quality thumbnails rather than full-size images.

1: Click to choose Embed Images (check box ticked) or

Link to the Image source (check box not ticked).

428 © 2011 Leica Microsystems (Switzerland) Limited

 Create a Report: Select a File Format:
Select the format in which the file should be created by: If the template is already open in another application -
1: Click on the small arrows to the right of the File Format perhaps in the process of being modified:
header. 5: The Report Generation Error occurs. Close the other
application and click the Create button again.
2: From the drop-down list, click to select the required
format. The Output File already exists:
6: The Confirm Overwrite warning appears. Confirm the
To display the report after it has been created: overwrite or click No and give the file another name.
3: Click to enable (ticked) the Display Report check box.

To create the report:

4: Click the Create button.

LAS User Manual 429

 Create a Report: Unable to Export Images:
Earlier versions of Microsoft Office Word could
not accept 16 bit images; Trying to insert a 16
bit image causes the Unable to Export Images
warning (1).

The user's options are:

Resample the images to 8-bit in LAS.

Re-capture the images with an 8 bit format by

clicking the Image tab in Preferences (2),
clicking on the small arrows to the right of the
Format header (3) and choosing an
appropriate format from the list.

Use a template that allows thumbnails (8-bit

jpeg by default) to be included.

430 © 2011 Leica Microsystems (Switzerland) Limited

 Create a Report: Modify an existing Template:
The existing Standard Templates supplied with LAS can be
modified to suit the user. The page layout - margins and
indents - can be altered, as well as new image data fields
added and fonts changed.

1: On the Create Report dialog select the template to be

changed and...

2: ...click the Modify button.

3: Word is launched and the selected template opened. Users should consider copying a Standard Template,
Refer to the steps shown below in Create a New
Template to make the changes.
! saving it under a different name and making changes to
the copy to preserve the original.

LAS User Manual 431

 Create a New Report Template:
LAS Standard Report Templates can be modified to either
remove fields or include others, but there are occasions
when a new, bespoke report is a more appropriate solution.

Both LAS and Word must be active. The screen images

included here have been created using Word 2010; Earlier
versions may have slightly different dialogs but the process
is the same.

1: Open Word and create a new document.

2: Click Save As and...

3: ...navigate to a folder of choice,...

4: ...give the document an appropriate name and...

5: ...from the Save as type drop-down menu, click to

select Word 97-2003 Template (*.dot). This ensures
compatibility across different versions of Word.

6: Click Save.

Continued... 458

432 © 2011 Leica Microsystems (Switzerland) Limited

 Create a New Report Template: Page Formatting:

1: Click on the Word Page Layout tab and: Select the number of Columns: The new template can
use columns which is useful if there are a number of
Select a Margin: Use standard layout or set up new ones. small images that need to fitted on to a page.

Choose Portrait or Landscape: From the page The Watermark, Page Colour and Border features found
Orientation options. in the Page Background section can be used in a
template.
Select the page Size. Format A4 is widely used but
regions will have their own requirements. 2: When the page format is complete, click the Save
button.

LAS User Manual 433

 Create a New Report Template: Headings and Static Text:

The report template can have as many lines of ‘static’ text In the example a heading and sub-heading have been
- headings, corporate disclaimers, terms of business etc. - added to the template.
as required. These are items that remain unchanged
unless specifically altered before the template is used to Simply type a heading or any other line(s) of text and if
create a report. required, format it with the usual Word facilities. In the
example the font size has been increased to 11 point, the
style changed to bold and the text filled with blue.

434 © 2011 Leica Microsystems (Switzerland) Limited

 Create a New Report Template: Field Structure: TableStart:
The template is a link between Leica
Application Suite and the report itself. Special
template fields - easily recognised by the « and
» characters that enclose them - mark where
image data from LAS is to be inserted into the
report. Any of the LAS data items can be used.

The part of the template that contains the fields

is marked with a top and a bottom 'boundary'
fields; They are called TableStart and TableEnd
respectively - no space between the words.
The TableStart field must appear before any
other fields and data fields cannot occur after
TableEnd.

Like all other fields, TableStart and TableEnd

are enclosed with the « and » markers and are
placed on the template using the Word Field
Insert procedure as follows:

1: On the new template move the cursor to

where the data field is to be inserted.

2: Click on the Word Insert tab

3: Locate the Field Insert feature - it varies

with the Word version - and select it.

Continued... 436

LAS User Manual 435

 Field Structure: Select Mail Merge:
This step sets up the Field Insert feature to use
the Mail Merge option. This setting is retained
whenever Field Insert is selected unless
changed by the user:

1: On the Field dialog, click on the small

arrow to the right of the Categories header.

2: From the drop-down menu click to select

the Mail Merge option.

Continued... 437

436 © 2011 Leica Microsystems (Switzerland) Limited

 Field Structure: Select Merge Field:
When the Word Field dialog appears:
3: Click OK.
1: On the Mail Merge menu, click to select Merge Field.
4: The TableStart field appears at the cursor on the
2: Click inside the Field properties text box and type: template with the markers inserted.

TableStart:RecordData Continued... 438

There are no spaces between the words,

A colon between TableStart and RecordData and
No need to type the « and » markers.

LAS User Manual 437

 Create a New Report Template: Retrieve the Data from LAS:

Again, the Word Field Insert facility is used to create fields 1: On LAS > Browse click on the Field Information
that will display the data imported from an LAS Archive. button on the Side Toolbar.

It is a 2-step process: 2: The Record Details dialog appears which lists all of
the fields available for that archive. In the example
Copy the field name from LAS. the Image Name field is selected to be copied to the
new template in Word.
Create a field on the template and link it to the LAS
Archive field by pasting the LAS field name into it. 3: LAS Field Names are not the same as the captions
displayed on the Record Details dialog. – they can
LAS must be running in the Browse Workflow with the only be retrieved by using the Copy as Report Field
required archive selected. function which copies the real field name to the
Windows Clipboard. From there it is available to
Word.

Continued... 439

438 © 2011 Leica Microsystems (Switzerland) Limited

 Create a New Report Template: Inserting LAS Data Fields:
This step copies the LAS Field Data link from the clipboard Some versions of Word may have a paste button
and places it on the new template: available that can be used instead of the Ctrl+V
combination.
1: Position the cursor on the new template where the filed
data is to appear. The LAS Field Data link appears in the text box.

2: Click to select the Merge Field option on the Field 4: Click OK.
Names menu.
5: The LAS Field Data link complete with markers
3: Click inside the Field name text box and... appears on the new template.

Press and hold down the Ctrl key. Continued... 440

Press the V key

LAS User Manual 439

 Create a New Report Template: Insert a Field Prefix:
A prefix can be added before the Field Data as a descriptor 2: Click inside the text box and type the prefix - in this
that will appear on the display and printout: case the words Image Reference =.

1: Click to check the Field Options > Text to be inserted 3: When the Field Data is inserted into the template the
check box. prefix appears before the data.

Continued... 441

440 © 2011 Leica Microsystems (Switzerland) Limited

 Create a New Report Template: Inserting a Date Field:
The Word Date Field is dynamic - the date automatically
displays the date today when the report is printed or 3: The Date formats menu appears; Click to select the
displayed: format required.

On the Insert > Field dialog: 4: Click OK.

1: Click the small arrow to the right of the Categories 5: The date appears on the template.
header and...
Continued... 442
2: ...from the drop-down menu click to select Date and
time.

LAS User Manual 441

 Create a New Report Template: Add an Image:

Report images are contained with Word Text Boxes. The 3: Using the handles that surround the text box, re-
first step is to create a text box: position and re-shape the text box to contain and limit
the extent of the image. Images are automatically
1: On the Word Insert tab, click to select Text Box. scaled to fit inside the text box.

2: If text box styles are available in the Word version, Continued... 443
click to select Simple Text Box.

442 © 2011 Leica Microsystems (Switzerland) Limited

 Add an Image: Continued:

Next, a field that draws the image is created within the

Text Box. 4: Click OK.

1: Click inside the Text Box and on the Insert > Field 5: The field appears inside the text box.
dialog...
Only a single image text box is required on the new
2: ...click to select MergeField from the Field names list. template even if it is required to show multiple images.
LAS will export all of the image selected in the Gallery,
3: Click inside the Field name text box and type: drawing and placing them in sequence on the report. New
report pages are created automatically as they are
Image:FullSize required.

A colon between Image and FullSize. Continued... 445

No space between Full and Size.
No need to add the « and » markers.

LAS User Manual 443

 Create a New Report Template: Insert a Page Break:
If only a single image is to be displayed on each report 2: Click on the Word Insert tab.
page, a Page Break can be inserted after the fields and
image text box to force a new page for the next image. 3: Click the Page Break option. This will vary with
different versions of Word.
1: After the fields and image text box, click to mark where
the Page Break is to occur. Continued... 445

444 © 2011 Leica Microsystems (Switzerland) Limited

 Create a Report Template: Completing the Template:
The final step in creating a new template is to indicate the There are no spaces between the words,
end of the data. Field Insert is used again: A colon between TableEnd and RecordData and
No need to type the « and » markers.
On the Word Field dialog:
3: Click OK.
1: On the Mail Merge menu, click to select MergeField.
4: The TableEnd field appears at the cursor on the
2: Click inside the Field properties text box and type: template with the markers inserted.

TableEnd:RecordData Finally, save the new template, close Word and test it: Go
there... 453

LAS User Manual 445

LAS Archive Standard Edition: Keywords:

The Keyword feature can create lists of

words and associate them with archives
so that they can be used to populate
selected fields.

This means that spellings – especially for

complex subjects, possibly in another
language – only have to be captured once
and after that can be used indefinitely,
confident that spelling and ‘cases’ are
consistent (1)

So, Keywords are fast and accurate and

can ensure that field searches will be
precise.

Keyword lists can be shared across any

number of archives with any changes to
the list be reflected in all of the archives.
This is because each archive maintains a
link to the Keyword list rather than copying
it.

New lists can be created by typing, by

cutting and pasting words or phrases from
an existing document or by importing a
complete existing list.

446 © 2011 Leica Microsystems (Switzerland) Limited

 Keywords: Create a New Keyword List:

When a new archive is created the

Keyword dialog automatically appears on
screen ready for a new list to be created
or imported. For existing archives:

1: Select the archive and if necessary

make it active current by double-
clicking it or clicking the Set Archive
as Current button.

2: Click on the Edit button. The field

structure and the Keyword dialog will
appear. If there is a Keyword List
already associated with the archive,
its name will be displayed in the
Keyword List window (3) and its
contents in the main window (4).

LAS User Manual 447

Keywords: Typing a Keyword List:

1: On the Keyword dialog click on the

New button.

2: When the New Keyword List dialog

appears, type a name for the new list
and click OK.

3: The name appears in the Keyword

List window and the other controls
become active.

4: Change the name and start again by

clicking the Delete button and
returning to step (1).

Typing the Keyword List:

5: The first word or phrase must be
preceded by clicking the Add a Child
button. The New Keyword dialog
appears.

6: Type the first word or phrase. Words

can be copied from other open
documents – a text editor or internet
browser for example – and pasted
into the New Keyword text box using
the Ctrl+C (Copy) and Ctrl+V (Paste)
keyboard combinations.

Generally, only letters and numbers

are allowed in keywords; If an invalid
character is typed a red (!) appears to
the left of the entry text box and the
flyout prompt itemises the invalid
character.
In the example it is the comma
between Duda and 1923 shown as [,].
Delete the invalid character and…

7: …click OK.

448 © 2011 Leica Microsystems (Switzerland) Limited

 Keywords: Typing a List and Grouping:

1: The first word or phrase appears in the Keyword 4: Clicking on the entry that is to ‘head’ the group.
Editor main window.
5: Click on the Add a Child button and type the word or
2: To add more words click the Add New Word at this phrase to be indented (3).
Level button and when the New Word dialog
appears… 6: The word will appear indented in the main window.
The last word or phrase added becomes the selected
3: …type a word or phrase and click OK. item automatically, so to continue to indent use the
Add New Word at this Level button.
Keyword Grouping: To revert to the original level, click to select the entry
To make long lists of keyword easier to read or to group that ‘headed’ the group and use the Add New Word at
related entries, the list can be indented by: this Level button.
Several indent levels are permissible.

LAS User Manual 449

 Keywords: Editing and Deleting:

To edit a word or phrase: Delete a Keyword:

7: Click to select the word and… 7: Click to select the word and…

8: …click the Edit button. 10: …click the Trashcan (Delete) button.

9: The word appears in the Edit Keyword dialog. Make 11: Confirm (or cancel) the deletion.
the changes and click OK.

450 © 2011 Leica Microsystems (Switzerland) Limited

 Keywords: Create a List with Cut and Paste:

New lists can also be created from

existing documents and imported as
Keywords.
The list is created in a simple test editor
such as Wordpad or Notepad – DO NOT
use Microsoft Word to create the list, but
individual words or phrases can be cut
from Word and pasted into Wordpad for
example.

1: Either type directly into the text editor

or cut and paste from another source.
Avoid punctuation characters
especially commas (Keyword Lists
are comma-delimited), using mainly
letters and numbers.

2: To create an indented entry or block,

insert the Paragraph (sometimes
called Pilcrow) character at the
beginning of the line. These will
create an indent when the list is
imported into the Keyword Editor.
The Paragraph character can be
inserted by holding down the
keyboard Alt key and typing 0182 on
the Numeric Keypad usually situated
to the right of the main keyboard.
The character can be copied (highlight
and use Ctl+C) and then pasted (
Ctrl+V) if there are many entries to
indent.
Do not use tabs or additional spaces
to make an indent.

LAS User Manual 451

Keywords: Cut and Paste: Continued:

3: Save the file as a .txt document either

as Text (ASCII), MS-DOS Text
(ASCII) or Unicode (UTF-8) but not as
Rich Text Format (RTF) which, like
Microsoft Word includes extensive
formatting.

4: Although Keyword Lists can be saved

to any folder on the hard drive, Leica
Application Suite has a default
location:

C:\Documents and Settings\All

Users\Shared
Documents\Applications\Leica
Application Suite\Keyword Lists

…which is recommended.

452 © 2011 Leica Microsystems (Switzerland) Limited

 Keywords: Importing a Keyword List:

With a new Keyword List created in a text editor and 1: Click on the Keyword Editor Import button.
saved as a text (.txt) file, it can now be ‘imported’ into an
archive. The import process actually establishes a link 2: On the Windows dialog, navigate to the folder in
between archive and Keyword List so that any changes which the new Keyword List is stored. The example
made to the list will be reflected in the archive. shows the LAS default folder, Folder Lists. Click to
select the list and…
To make major changes in he list and NOT have the
archive affected, save the text file under a different name 3: …click Open.
and use it as a completely separate list.

LAS User Manual 453

Keywords: Importing a Keyword List: Continued:

4: If there are invalid characters in the List a warning 5: When import succeeds the list appears in the
appears indicating the problem characters and where Keyword Editor main window. Long and wide lists
they are located. The list will have to be corrected and automatically have scroll bars to the right and bottom.
saved with the text editor.

454 © 2011 Leica Microsystems (Switzerland) Limited

 Keywords: Creating a Keyword Field:

Keywords can only be copied to archive

fields that has been designated as
Keyword Fields.

When a Keyword List (or lists) is

associated with an archive, the Keyword
button on the Archive panel becomes
active:

1: Click to select either the Category

(Record Group) or Image panel.
Keywords in the Category will appear
on every image captured within that
group; Keywords associated with the
image will only appear if they are
selected for that image.

2: Click on the Keyword button. A

keyword text box opens on the
selected panel.

3: Click in the Keyword text box and

type a meaningful name for the
Keyword Field.

4: If it important that a keyword always

appears in the field, click to enable
the Required check box. With
Required enabled, a user will be
forced to select a keyword from the
List before an image can be saved.

LAS User Manual 455

Keywords: Creating a Keyword Field: Continued:

5: In cases where several Keyword Lists

are associated with an archive, select
the appropriate list from the drop
down menu by clicking on the arrows
to the right of the Keyword List name
and…

6: …clicking to select the list required.

Several Keyword Fields may be

added to the Record Group or Image
parts of the archive and this, together
with the facility to associate a range
of Keyword Lists with the archive,
makes Keywords a powerful and fast
method of gathering data. And if, for
example, a list comprises the cost
centres within an organisation and is
set as Required, every image without
fail could become the property of a
particular department.

7: Click Save.

456 © 2011 Leica Microsystems (Switzerland) Limited

 Keywords: Using Keyword Lists:

Keywords are copied to keyword fields in the Browse Use this process any time a keyword is required:
Workflow. For Required keywords in the Category
(Record Group), these are selected when the group is 4: Click on the button to the right of the Keyword Field –
created as follows: this could be in Category or in Image. The Select
Keyword dialog appears.
1: Click on the Create a New Category button.
5: Scroll through the Keyword List to the required entry
2: Type a name for the new category. and then click in the check box to the left of it. A tick
mark should appear. Clicking the entry itself is not
3: If the keyword is Required, a red (!) will flash to the sufficient.
right of the Archive Navigator. This means that the
new category cannot be saved until a keyword has 6: Click OK and the keyword will be copied to the field.
been selected.
7: Click on the Save button.

LAS User Manual 457

Keywords: Exporting a Keyword List:

Keyword Lists may be exported to any

folder on the computer or to an external
storage device such as a memory stick.
The Export feature makes a copy of the
Keyword List at the new location; The
original remains intact.

If there are several lists associated with an

archive:

1: Click on the arrows to the left of the

Keyword List on the Keyword Editor
and from the menu…

2: …click to select the List to be

exported.

3: Navigate to the export target folder by

clicking on the browse button to the
right of the Save Keyword List to
window, and…

4: …selecting it on the Windows dialog.

Click Save.

5: Click on the arrows to the right of the

Format text box and from the menu
click to select the character coding –
ASCII is the default and generally
preferred.

6: Click OK.

7: A message indicates that the Export

is complete. Click OK.

458 © 2011 Leica Microsystems (Switzerland) Limited

 Keywords: Deleting a Keyword List:

A Keyword List may be deleted –

disassociated with the archive – at any
time BEFORE a Keyword Field has been
created. After that, clicking the Delete
button will result in the Cannot Delete
message (1).

LAS User Manual 459

LAS Archive: Exchanging images and data between LAS and Leica Image Manager

If you wish to simply transfer and image from an

LAS Archive to a memory stick then is it simples
to use the Export Image facility.

See Export from LAS Archive to File System 461

For those organisations using Leica Image

Manager (IM) as a networked archive store and
organiser, features are provided in LAS that allow
images from workstations running under Leica
Application Suite to be sent to Leica IM (from
version 5 onward) and managed like any other IM
image.

See Sending Images from LAS into Leica IM V5

462

Images stored in an IM archive can also be

transferred into LAS.

See Receiving Images into LAS from Leica IM V5

473

460 © 2011 Leica Microsystems (Switzerland) Limited

 Archives: Export from LAS Archive

The Side Tool Bar is displayed on the right

hand edge of the Image Viewer. The
Export button is situated in the top group.

To Export the image being displayed

using LAS Archive to a selected
destination folder:

1: Click a thumbnail to select and

display the image to export.

2: Click on the Export button.

For detailed help on the Export
procedure look in Functions Widely
Available: Go there...

LAS User Manual 461

Sending Images from LAS into Leica IM V5

To be able to use this feature, the Leica

IM Image Data Read-in module must be
licensed.

Leica Image Manager can store all of the

data fields captured with the image in LAS
providing they are nominated but it cannot
itself display them – they are stored in a
special file associated with the image.
However, when the image is retrieved
from Leica Image Manager by LAS, the
data will be displayed in the Browse and
Process Workflows in the usual way.

Setting up Image Manager to receive

LAS images and data:
Images and data from LAS can be sent to
either an existing or a new archive in
Leica Image Manager. Run Leica Image
Manager and to use an existing archive:

1: Click on File on the main menu and

from the drop down menu…

2: …click to select an archive from the

Recent list or…

3: …click the Open Archive option. This

will reveal the Open Archive dialog.

4: Click to select an archive from the list.

5: Click OK.

Continued… 464

462 © 2011 Leica Microsystems (Switzerland) Limited

 Export to Leica IM V5: Creating a New Archive:

To create a New Archive:

1: Click on Options on the main menu.

2: Click on System Administration and…

3: …click to select New Archive. The

New Archive Wizard will appear.
Follow the instructions.

A new archive need not have a range

of fields – these will be selected later
and the data for them will be supplied
by LAS.

However, a new archive MUST have

a Text field for every level and that
must be chosen as the Identifier (Key)
, as well as at least one Image field at
the lowest level. If either are missing
the archive will not be created.

Continued... 464

LAS User Manual 463

Export to Leica IM V5: Setup AutoSave in Image Manager:

The AutoSave option in Leica Image

Manager must be enabled so that the
incoming images (from LAS) are directed
to the selected archive and all of the data
is correctly stored.

1: Select Options on the main menu…

2: …and click AutoSave.

3: On the AutoSave dialog, click the

right-hand tab that represents the new
data sheet – the label on our example
is Component_Location.

4: The fields existing in the chosen

archive are displayed in the Compose
window.

5: If it is intended that the LAS Image

Names are used to refer to them
within Leica Image Manager, click to
enable Use original filename…

Continued… 465

464 © 2011 Leica Microsystems (Switzerland) Limited

 Export to Leica IM V5: Setup AutoSave: Continued:

The image storage format can be

changed as it is imported into Leica Image
Manager. Jpeg compression is
recommended because images are more
compact and large archives will import
quicker.

To change the image format:

1: Click on the small arrows to the right
of the Image Format header and from
the drop down list…

2: …click to select the required format.

3: The Image Directory (in Image

Manager) will be set to that of the
selected archive but can be changed
by clicking on the browse button to
the right of the Image Directory
header and…

4: …using Windows Browse for Folder

dialog to…

5: …navigate to the required directory.

6: Click OK to close the dialog.

7: Click on the left-hand tab and also…

8: …on the Activate checkbox to enable

AutoSave.

Continued… 466

LAS User Manual 465

Export to Leica IM V5: Link the Import Plug-in to Image Manager:

When Leica Application Suite is installed it

makes available to Leica Image Manager
an Import Plug-in, a small software
application that will ‘instructs’ Leica IM
how to store the data fields that will be
sent from LAS.

The next step is to link Leica Image

Manager to the Plug-in:

1: Click Options on the main menu.

2: From the drop down list, click on File

Preferences and then…

3: …on Plug-in Setup.

4: On the Plug-in dialog, click to select

Leica XML.

Continued… 467

466 © 2011 Leica Microsystems (Switzerland) Limited

 Export to Leica IM V5: Link the Plug-in: Continued:

With the Leica XML Plug-in selected:

1: Click on the Include button.

2: The Leica XML Plug-in is listed in the

Order of Usage window and…

3: …the Plug-in is marked with a pin in

the All Plug-ins window.

4: Check that the Fixed Application

check box is enabled and…

5: …the application being pointed to is

ImagePackageImporter.exe. If it is
not, click on the browse button to the
right of the Fixed Application window
and…

6: …on the Browse (Open) dialog,

navigate to the Leica Application
Suite directory and select the
ImagePackageImporter application
(7).

8: Click Open.

9: Click OK on the Plug-in Setup dialog.

Continued… 468

LAS User Manual 467

Export to Leica IM V5: Select the LAS Fields to be included:

With the Leica XML Plug-in linked to

Image Manager all of the LAS data fields
are available to Leica IM. Now they can be
displayed and selected for inclusion with
the imported images.

1: Click Options on the main menu.

2: From the drop down list click to

select System Administration and…

3: …click to sleect Edit Archive.

4: The Edit Archive dialog appears with

the current Image Manager archive
displayed including the Leica IM
fields: Shown on the illustration are
the minimum that would be allowed –
one Text field as the key (Indicator)
and one Image field.

5: Click on the Add button.

Continued… 469

468 © 2011 Leica Microsystems (Switzerland) Limited

 Export to Leica IM V5: Select the LAS Fields to be included: Continued:

On the Field Properties dialog:

1: Click on the small arrow to the right of
the Type header.

2: From the drop down list click to

select Image Header Data.

3: Click on the arrow to the right of the

File Type and from the drop down…

4: …click to select Leica XML.

5: A list of all the available LAS data

fields now appears in the Field
window. Not all will be appropriate to
the imported images so click to select
only those required. The selected
data will be imported into Image
Manager but will be packaged in a
separate file that is associated with
the image. The fields are not
available to Leica Image Manager but
will be retrieved and displayed by LAS
along with image.

6: Click OK.

Continued… 470

LAS User Manual 469

Export to Leica IM V5: Select the LAS Fields to be included: Finish:

Returning to the Edit Archive dialog:

1: The selected LAS fields have been
added to the list.

2: Click OK to complete the Leica Image

Manager setup.

3: Double-click the desk-top icon to

start Leica Application Suite.

Continued… 471

470 © 2011 Leica Microsystems (Switzerland) Limited

 Export to Leica IM V5: In LAS: Select the Images to Export into Image Manager:

In Leica Application Suite, select the

images to be exported to Leica Image
Manager – it can be a single image or an
entire range.

1: Click to select a thumbnail in the

Gallery or if a range of images is
required click on the first thumbnail to
be included and holding down the
keyboard Shift key, click on the last
thumbnail in the range. All of the
images encompassed by the two
selections are highlighted.

2: Click on the Export button.

3: On the Export Images dialog, click to

select Image Manager and if
required…

4: …click on the Don’t ask again check

box to skip the Export Images dialog
in the future.

5: Click OK and the export begins.

Continued… 472

LAS User Manual 471

Export to Leica IM V5: Done...

Images exported from Leica Application Suite in Leica Image Manager.

472 © 2011 Leica Microsystems (Switzerland) Limited

 Receiving Images into LAS from Leica IM V5:

Existing images, sequences or movie

clips stored in Leica Image Manager 5
(IM V5) prior to and including Leica
Application Suite 3.3, can be easily
imported into an LAS Archive or Folder
created after Version 3.3. The process is
slightly different for archives or folders.

Start LAS and then...

Start Image Manager.

Import into an LAS Archive:

Importing is carried out on a field-by-field
basis so the target (receiving) archive
must have the same structure and field
names as the source otherwise data will
be lost.

1: Click on the Setup Workflow.

2: Select the Archive tab.

3: Click on the Folder Archive button.

4: On the list of available archives click

to select the target archive into which
the images will be imported.

5: Return to the Browse Workflow.

6: Click the target archive and...

7: ...select it as the Capture Archive.

Continued... 474

LAS User Manual 473

 Import From Leica IM V5: Into an LAS Archive: Select the Image in Leica IM V5:

Go to Image Manager and navigate to the

location of the image(s) to be sent to LAS
Archive:

1: Right click on the thumbnail of the

image to be sent to LAS in the Leica
IM5 gallery.

2: From the context menu, click to

select Export to LAS.
If this option is not available on the
menu see the instructions in the
Appendix: Go there... 477

3: The Import to dialog appears with the

target archive in LAS and...

4: ...the destination folder in the

appropriate window. 6: Click OK and the image will be sent to LAS.

5: Click to enable both Include Single

Capture Images and Include Sub- Continued... 475
Folders.

474 © 2011 Leica Microsystems (Switzerland) Limited

 Import From Leica IM V5: Into an LAS Image Explorer Folder:

1: Click on the Browse Workflow.

2: On the Folders tab, navigate to the

target folder in LAS into which the
images from Leica IM5 will be
imported.

Continued... 476

LAS User Manual 475

 Import From Leica IM V5: Into an LAS Image Explorer Folder: Locate the Image in Leica IM V5:

Go to Leica Image Manager and navigate to

the location of the image(s) to be sent to
LAS Archive:

1: Right click on the thumbnail of the

image to be sent to LAS in the Leica
IM5 gallery.

2: From the context menu, click to select

Export to LAS.
If this option is not available on the
menu see the instructions in the
Appendix: Go there... 477

3: The LAS Import dialog appears with the

target archive in LAS listed. Click Yes
to confirm the import and the image will
be imported.

476 © 2011 Leica Microsystems (Switzerland) Limited

 Import From Leica IM V5: Appendix: TheLeica IM V5 Context Menu Setup:

Right clicking on a thumbnail in the Image

Manager gallery, reveals a context menu.
To be able to export images from Leica IM5
into LAS archives or folders, a small
program must be enabled. The program
resides in Leica Application Suite but is
launched from Leica Image Manager 5 by
clicking on an entry in the context menu
called Export to LAS.

To put the option on the context menu:

1: In Leica IM5, click on Archive on the
main menu and...

2: ..from the drop down menu click the

Properties option.

3: On the Archives dialog, click to enable

Assigned application.

4: Click in the Name text box and type

Export to LAS.

5: Click on the browse button to the right

of the Application text box and...

6: ...on the Open dialog navigate to:

C:\Program Files\Leica
Microsystems\Leica Application Suite
V3...

...and click to select

ImagicPackageImporter.exe

7: Click Open.

8: Click OK

The Export to LAS option will now be

available on the Leica IM5 context menu.

LAS User Manual 477

Extended Annotation

Extended Annotations is an optional

module that adds considerable power and
flexibility to the image annotation tasks.

With three line styles, two shapes -

Rectangle, including Square, Ellipse also
including Circle - and an adaptable
Caption Label text tool, Extended
Annotations provides all of the essential
features to produce high quality,
professional annotated images.

A large number of graphic objects can be

created and, if required permanently
merged into an image.

The Object Grid that can be hidden or

revealed, improves productivity by listing
all of the graphic objects and their basic
properties. Clicking on an entry
immediately selects the object on the
image, especially convenient when
dealing with large, complex annotations.

Feature details include - click on a feature

to go there...

Drawing Lines 488 : 488

Drawing Rectangles and Ellipses: 489

Drawing a Caption Label: 490

Inserting an Image: 491

Merging Graphics with an Image: 492

478 © 2011 Leica Microsystems (Switzerland) Limited

 Extended Annotation: The Control Panel:

The Extended Annotation tools are

available on all of the Workflows except
Setup. To launch the Extended
Annotations interface:

1: Click on the Extended Annotations

icon on the Side Tool Bar and click to
select the Extended Annotation
option.

The Toolbox tools are:

2: The Pointer used to select drawn
items.

3: Text tool for creating labels captions.

4: Three Line tools – Plain Line, Arrow

and Distance Line.

5: Rectangle tool.

6: Circle and Ellipse tool.

7: Insert Image tool.

8: Trash Can (Delete) button.

9: Undo and Redo tools.

10: Display the Object Grid, a list of all

of the drawn objects.

11: The Show check box must be

enabled for the Control Panel to be
displayed and the tools activated.

12: Merge incorporates the drawn

objects with the image permanently.

A & B: These areas of the panel change

according to the tool selected.

Continued… 480

LAS User Manual 479

Extended Annotation: The Object Grid:

The graphic objects drawn on an image

are listed on the Object Grid to make
access and editing quick and easy.

1: Graphic objects – rectangles, text and

lines…

2: …are listed on the Object Grid if the

Show Grid check box is enabled.

3: Clicking on a Grid entry immediately

selects the object on the image -
control 'handles' appear around it.

4: The caption text can be edited in the

Label text box or…

5: …the object deleted by clicking on

the Trash Can icon.

Continued… 481

480 © 2011 Leica Microsystems (Switzerland) Limited

 Extended Annotation: Move, Edit and Delete an Object:

A graphics object drawn on an image can

be selected and moved by dragging,
edited or deleted as follows:

1: Click to select the Pointer in the

toolbox.

2: Click on the object to be moved…

3: …eight small ‘handles’ appear around

the periphery.

To Move the Object:

4: Click in the centre of the object – the
cursor changes to a four-quadrant
arrow - and holding down the mouse
button, drag the object to a new
location and release the button.

To Edit the Object…

5: …by stretching it, click on one of the
small ‘handles’ – the cursor changes
to a double-ended arrow - and holding
down the mouse button, drag it to
change the shape.

To Delete the Object:

6: Click on the Trash Can (Delete) icon
in the toolbox.

To Navigate between Actions:

7: Click the left pointing arrow (Undo) to
go back to the previous action - the
right pointing arrow (Redo) becomes
active. Click it to return to the last
action.

Continued… 482

LAS User Manual 481

Extended Annotation: Line Thickness and Colour:

1: Select a Line, Rectangle or Ellipse

tool from the toolbox.

Set the Line Thickness:

2: Increase or decrease the Line
thickness by clicking the Up/Down
arrows to the right of the Width text
box.

Select the Line Colour and

Transparency:
3: Set the line colour by clicking on the
Line button and…

4: On the Select Colour dialog, choose a

colour from the Basic Colours
swatches, from the Colour Wheel by
dragging the ‘target’ or…

5: …by clicking in the Current Colour

text boxes and typing new Red,
Green and Blue values.

6: If necessary, adjust the Shade by

clicking and dragging the slider.

7: Adjust Transparency by clicking and

dragging the slider.

8: The transparency value is displayed

in the Transparency text box. A value
of 255 represents a solid colour and 0
is totally transparent.

9: Click OK.

10: Shapes can have an outline that is

enabled by clicking the Outline check
box. The Outline thickness and colour
is set by following the steps above.

Continued… 483

482 © 2011 Leica Microsystems (Switzerland) Limited

 Extended Annotation: Setting the Fill Colour and Transparency:

Rectangles and Ellipses can be filled with

a colour and its transparency set to solid
(Value 255) or completely transparent
(Value 0) and all of the graduations
between.

1 & 2: Click on either the Rectangle or

Ellipse tool on the toolbox.

3: Set the shape fill colour and

transparency by clicking on the Fill
button.

4: On the Select Colour dialog, choose a

colour either from the Basic Colour
swatches, by dragging the ‘target’ on
the Colour Wheel or…

5: …clicking in the Current Colour text

boxes and typing new values for Red,
Green and Blue.

6: Adjust the Shade by clicking and

dragging the slider on the Shade Bar.

7: Set the transparency by clicking and

dragging the Transparency slider
downward or…

8: …typing a value in the Transparency

text box. Set the value to 0
(Completely transparent) and enable
Outline (9) for an ‘open’ rectangle or
ellipse.

10: Click OK.

Continued… 484

LAS User Manual 483

Extended Annotation: Caption Labels: Selecting the Font Attributes:

Caption Labels are created using the Text

tool. The Font Type Face, Style, Size and
Colour can be selected as well as the
Label Background Colour and
Transparency. The illustration (1) shows a
typical Caption Label drawn directly onto
the image.

To select the Font Type Face, Style and

Size:

2: Click on the Text icon on the toolbox.

3: Click on the Font button and…

4: …on the Font dialog select the Type

Face, Font Style and Size.

5: Click OK.

Continued… 485

484 © 2011 Leica Microsystems (Switzerland) Limited

 Extended Annotation: Text and Background Colours:

The colour of the text and the background

are both changed in the same way. In
Figure (1) the Background Colour
transparency has been set to 0 (Totally
transparent) so that the Caption text
appears directly on the image.

2: Click on the Text icon in the toolbox.

3: Click either the (Font) Colour button

or the Background button.

4: On the Select Colour dialog, choose a

colour either from the Basic Colours
swatches, from the Colour Wheel by
dragging the ‘target’ or…

5: …by clicking in the Current Colour

text boxes and typing new Red,
Green and Blue values.

6: If necessary, adjust the Shade by

clicking and dragging the slider.

7: Adjust Transparency – mainly

applicable to the Background – by
clicking and dragging the slider.

8: The transparency value is displayed

in the Transparency text box. Set the
value to 255 for a solid colour, 0 for a
completely transparent Background
or ant value between for different
transparency levels.

9: Click OK.

Continued… 486

LAS User Manual 485

Extended Annotation: Adding an Outline:

The Text Box in Figure (1) has a red

outline created as follows:

2: Click on the Outline check box to

enable outlining.

3: Set the width of the Outline by clicking

on the small Up/Down arrows to the
right of the Width text box to increase/
decrease the Outline thickness.

4: Change the colour of the Outline by

clicking on the Line button and
following the steps described
previously: Go there… 482

Continued… 487

486 © 2011 Leica Microsystems (Switzerland) Limited

 Extended Annotation: Positioning Text:

There are three options for positioning text

on a Caption Label:

1: Set Left,

2: Set Centre and

3: Set Right.

The text is automatically positioned as the

Label is drawn.

4: Click the Text icon in the toolbox.

5: Click on the small arrows to the right

of the Position header and…

6: …from the drop down list click to

select the text position.

LAS User Manual 487

Extended Annotation: Drawing the Line and adding a Caption Label:

There are three Line styles available in

Extended Annotations:

1: Plain Line.

2: Arrow and

3: Distance Line that displays a distance

in pixels between the end strokes.

4: To draw a line, click on the icon of the

style required in the toolbox.

5: Adjust the width and set the colour:

Go there… 482

6: To include a Caption Label with the

line, click in the Label text box and
type the required text.
Set the Text and Background
properties: Go there… 484

7: Click on the image at the starting

point for the line and, holding the
mouse button down drag in the
required direction to the required
length. Release the mouse button.
The head for the Arrow line appears
at the starting point.
Edit position, length and angle as
described previously: Go there… 481

When lines are selected and dragged to a

new location the Caption Label follows: To
re-position the Caption Label only, select it
using the Point tool, click in its centre and
drag to a new position.

488 © 2011 Leica Microsystems (Switzerland) Limited

 Extended Annotation: Drawing a Rectangle or Ellipse:

There are two shape tools included with

Extended Annotations, the Rectangle
(including Square1) and the Ellipse
(including Circle 2).

1 & 2 Click on either the Rectangle or

Ellipse tool on the toolbox.

3: Set the shape fill colour and

transparency: Go there… 483

If a Caption Label is required:

4: Set the font attributes and colours: Go
there… 484

5: Click in the Label text box and type

the label text.

To draw the Rectangle or Ellipse:

6: Click on the image and holding down
the mouse button, drag diagonally to
create a shape of the required size.
Edit the position and size: Go there…
481

LAS User Manual 489

Extended Annotation: Drawing a Caption Label:

To draw a Caption Label:

1: Click on the Text icon in the toolbox.

2: Set the Font attributes, Font and

Background colours: Go there… 484

3: If an Outline is required set the

thickness by clicking on the Up/Down
arrows to the right of the Width text
box and enable the Outline check
box.
Set the Outline colour: Go there… 482

4: Click inside the Label text box and

type the caption text.

5: Click on the image and whilst holding

down the mouse button, drag
diagonally to create the Caption Label
.

6: When the label shape is correct,

release the mouse button.
Edit the Caption Label size and
position: Go there… 481

490 © 2011 Leica Microsystems (Switzerland) Limited

 Extended Annotation: Insert an Image:

It is quick and easy to insert smaller

images over the main image. Inserted
images should be typically low resolution -
640 x 480 pixels or less - so that they do
not slow loading.

If an outline is required around the

inserted image:
1: Click to enable the Outline check box.

2: Adjust the outline thickness by

clicking on the arrows to the right of
the Width text box.

3: Select the Outline colour: Go there…

482

To insert the image:

4: Click on the Insert Image icon in the
toolbox.

5: Click on the main image and holding

down the mouse button…

6: …drag down and to the right to draw

a box. The inserted image will be
scaled to fit inside the box.

7: Release the mouse button and the

Windows Open dialog appears.

8: Navigate to the folder containing the

image to be inserted and click on its
name. Click Open.

9: The image is inserted into the box.

Edit the insert size and position: Go
there… 481

Continued… 492

LAS User Manual 491

Extended Annotation: Merge:

When Merging is enabled all of the

Caption Labels, Shapes and Lines drawn
on the image will be permanently included
as part of the image and cannot be
altered.

1: Merge on a captured image is a ‘one

shot’ button and only merges
annotations with the current image.
Click to merge.

2: Merge on live images is a check box

setting and whilst it remains enabled
will merge annotations with all
captured images. Click to enable -
click again to disable merging.

492 © 2011 Leica Microsystems (Switzerland) Limited

 MultiTime

The Leica Application Suite MultiTime

optional module is a highly effective
solution for the automatic acquisition of
images over time. The time span and
acquisition may range from many images
per second or just a few delayed by
minutes.

After acquisition they may be visualised,

enhanced and documented. Other LAS
modules can perform analysis and
measurement of the images.

There are 2 distinct components to

MultiTime that are operated
independently:

MultiTime Time-lapse: Images are

acquired with a delay starting at 1 sec
.

MultiTime Movie: Images are

acquired in a compressed image
stream directly to the hard drive as
fast as possible, equivalent to making
a video recording.

If images are to be collected over a long

time, the user must ensure that the
specimen and microscope are unaffected
by changes in temperature, focus position
and the electrical supply. Achieving these
conditions is not the remit of the MultiTime
module.

LAS User Manual 493

MultiTime: Time Lapse:

Time Lapse is an imaging technique that acquires images image data at full frame rates during the operation.
with a predefined delay between each image acquired.
Time Lapse imaging typically has a significant time delay
The images are stored to the hard drive at defined between each image processed. The camera can acquire
intervals and can be recalled individually, in a continuous images at its full frame rate, but only one image is
loop or as an AVI movie file. This delay time is the processed per period. This is because it is not efficient to
distinguishing feature between normal imaging and Time process every single image if only certain images are of
Lapse imaging. interest.

Time Lapse imaging is best suited for continuous image

acquisition over long periods, or where there is no need
for

494 © 2011 Leica Microsystems (Switzerland) Limited

 Time Lapse: Launch Time Lapse and set Capture Archive:

With the MultiTime module installed and

enabled:

1: Click on the Select Acquisition Mode

button.

2: From the menu, click to select

MultiTime. The 'T' symbol will appear
on tabs in both the Acquire and
Browse Workflows.

Select the Fixed Capture Location:

The Time Lapse images will be captured
into the fixed location.

3: Click on the Browse Workflow and...

4: ...if necessary click on the Browse

tab. Click to select the capture
archive and then...

5: ...click on the Set Fixed Location

button.

6: To indicate the fixed capture location

archive, a red dot appears to the left
of it.

LAS User Manual 495

Time Lapse: Image Exposure:

1: Click on the Acquire Workflow tab.

Both exposure and capture format
depend to a great extent on the time
lapse duration between individual
images.

For example, a short time lapse of

say 1 second, will demand a short
exposure time to allow the image to
be captured, processed and written to
disk, for the control files to be created
and written, and for the imaging
elements in the camera to be reset
ready for the next image. If these
functions together take longer than
the time lapse, images will be lost.

For a short time lapse:

Make sure the exposure time is also
short – as a guide 150mS should be a
maximum. Adjust the microscope
light settings if necessary to achieve a
short exposure.
Consider using VGA or 4x4 Binning
for the Captured Format, avoid using
High Quality (HQ) and set the
Captured Bitdepth to 8 bits keeping
disk files small and processing times
low.
Turn off Store and Recall in
Preferences.

See: Store and Recall. 495

Long time lapse:

Whilst the exposure time may not be
as critical, still consider the lower
resolution capture formats. Using high
resolution for multiple images will
rapidly consume disk space.

Always carry out an Auto White

Balance.

Check for a Shading Link or create a

new one.

Regularly backup MultiTime projects

to CD or DVD and free up disk
space.

496 © 2011 Leica Microsystems (Switzerland) Limited

 Time Lapse: Set-up:

The MultiTime module must be installed

and enabled. See: Registration.

1: Click on the Acquisition Mode

selector.

2: From the menu, click on the

MultiTime icon.

3: Click on the Acquire Workflow if it is

not already visible.

4: An additional tab marked ‘T’ will be

displayed. Click on it to reveal the
MultiTime controls.

The main panel – Define Time Sequence

– is further divided into 3 smaller panels:

Start
Sequence and
Test Acquire Time

There is an additional panel – Options –

which can be revealed by:

5: Clicking on the arrows to the right of

the header bar.

LAS User Manual 497

Time Lapse: Set-up Options:

The Start panel provides three start up

options for the MultiTime sequence:

Manual: The Sequence starts as soon

as the Acquire Time Lapse button is
clicked.

At date and time: Starts the capture on

a specific date and at a specified time.

After delay: Waits for a specified time

before starting the capture.

Manual start:
1: Click on the Start: Manual button and
go directly to the Sequence panel to
set up the time lapse.

At date and time start:

2: Click on the At date and time button.

3: Click on the arrow to the right of the

date window and the calendar
appears.

4: Use the left/right arrows to scroll

through the months and years.

5: Click on the day date on which the

sequence will start.

6: The time of day is divided into three

fields – hours: minutes: seconds.
Double-click on a field to select and
then use the up/down arrows to the
right of the window to set the required
value. Click and hold on an arrow for
a fast scroll. The hours field (24 hour
clock) rolls over at a count of 24, and
the minutes and seconds at 60.

498 © 2011 Leica Microsystems (Switzerland) Limited

 Time Lapse: Set-up Options: Continued:

After delay start:

The delay can be from 1 second to 23
hours: 59 minutes: 59 seconds.

7: Click on the After delay button.

6: The time is divided into three fields –

hours: minutes: seconds. Double-click
on a field to select and then use the
up/down arrows to the right of the
window to set the required value.
Click and hold on an arrow for a fast
scroll. The hours field (24 hour clock)
rolls over at a count of 24, and the
minutes and seconds at counts of 60.

LAS User Manual 499

Time Lapse: Sequencing:

1: On the Sequence panel, click the up/

down arrows to the right of Number of
images acquired window to set the
number of images in the sequence.

2: The Interval Between Images (time

lapse) window is divided into three
fields – hours: minutes: seconds.
Double-click on a field to select and
either type a value or use the up/down
arrows to the right of the window to
set the required value. Click and hold
on an arrow for a fast scroll. The
hours field (24 hour clock) rolls over
at a count of 24, and the minutes and
seconds at counts of 60.
Intervals may extend from 1 second
to 23 hours: 59 minutes: 59 seconds.

3: The number of images is multiplied

by the interval and the overall
sequence duration is shown in the
Total time window. In the example, 6
images are required with an interval
of 10 seconds. Including an interval
for capture and data writing for the
6th. image (in case further images
are added to the sequence), this
equates to 60 seconds or 1 minute
(00:01:00). However, if no further
images are added, the final interval
will be ignored.

4: Click on the Add Sequence icon and

the image count and interval appear
in the Sequence window (5).

6: When using Fluorescence

techniques, enable Close shutter
when not acquiring to avoid damage
to the specimen.

See: Testing MultiTime. 501

500 © 2011 Leica Microsystems (Switzerland) Limited

 Time Lapse: Testing:

With the number of images required and

the interval between each capture set, and
the sequence loaded to the Sequence
window, it is easy to check that the
exposure and data capture will fit within
the interval:

1: Click the Test button.

2: The result (in seconds) of exposing

the image and emulating a write to
disk together with the necessary data
files is displayed beneath the Test
button.
In the example, the interval between
images is set to 1 second. The test
capture and write amounted to 0.846
seconds so this sequence would have
succeeded with a small margin to
spare.

3: This test fails because either the

exposure is too long or the captured
image format results in a very large
file – or both!
The test result here is 3.446 seconds,
much longer than the 1 second
interval, so most images are going to
be missed.

See: Image Exposure and Capture

Format. 496

LAS User Manual 501

Time Lapse: Configuration:

To achieve complete flexibility, MultiTime

can be configured with any number of
capture sequences.

1: The Start option is executed first – in

the example ‘After delay’ of 2 hours
has been selected. When the delay
has elapsed each of the sequences
will be executed in turn.

The example sequences are:

2: 10 images with a 30 minute delay
between each.

3: 100 images with a short 1 second

delay between each image, and
finally…

4: ...25 images with 1 minute between.

Each sequence was added to the overall

program by clicking on the ‘Add Sequence
’ icon.

Changing the Sequence order:

To move a sequence up or down the
execution order:
4: Click on the sequence to be moved.

5: Click on the up/down order icons.

Deleting a sequence:
4: Click on the sequence to be deleted.

6: Click on the Trash Can icon.

Starting the MultiTime sequence:

7: To start a sequence or sequence
program, click on the Acquire button.

502 © 2011 Leica Microsystems (Switzerland) Limited

 Time Lapse: Project names:

A sequence or sequence program, may

be saved as a file for future use as
follows:

1: Click on the arrows to the right of the

Options header to reveal the panel.

2: Click in the Configuration text box and

type a name for the sequence.

3: Click Save.

Project name:
When a MultiTime sequence starts, a new
folder is created into which all of the
images and their data files will be loaded.

By default, these folders are given the

name T-MultiTime n where ‘n’ is a
sequential number if other folders of the
same name exist.

The prefix ‘T’ is always present because it

denotes a MultiTime project, but change
the folder name to one of your choice by:

4: Clicking in the Folder Name text box

and delete by backspacing the word
MultiTime.
Type in the new name. In the
example the word VeeGon (the name
of the test) has been typed so the
folder will be named T-VeeGon.

To retrieve an existing configuration:

5: Click on the arrows to the right of the
Configuration text box.

6: From the drop down list, click to

select a configuration.

To delete a configuration:
6: Select a configuration from the list.

7: Click on the Delete button.

LAS User Manual 503

Time Lapse: Capture Display Options:

Three options are available to determine

the state of the Viewer during the capture
sequence:

1: Click the Show Acquire Panel to

remain in the Acquire Workflow and
watch the images in real time. To
momentarily ‘freeze’ the image as it is
captured. But there must be a
sufficient interval between images to
allow the Viewer to refresh.

2: Enable the Show images during

acquire checkbox. This option works
only if the Show Acquire Panel is
selected.

3: Click Show Browse Panel to watch

the sequence on the Browse
Workflow. This has the advantage
that the captured images are
displayed on the Viewer as they are
made and the Gallery (4) is
progressively populated.

Continued... 505

504 © 2011 Leica Microsystems (Switzerland) Limited

 Time Lapse: Capture Display Options: Continued:

Progress, Pause and Stop panel:

During the capture sequence progress is
displayed as:

5: The number of images acquired so

far and the number remaining to be
captured.

6: The current time, Next image acquire

time, sequence time remaining and
estimated time of completion. If there
are several sequences in a project,
the completion time is for the entire
project not just for the current
sequence.

7: Graphical representation of progress,

both as a bar and a percentage.

8: To suspend the sequence, click on

the Pause button. The button label
changes to Resume. Click again to
continue the capture. Note however,
that images may be lost during Pause
.

9: Stop the sequence by clicking the

Stop button. The Confirm Sequence
Termination message appears (10).
Click Yes to stop the sequence
completely or No to continue the
sequence.

LAS User Manual 505

Time Lapse: Replay:

When the capture sequence finishes, the

Browse Workflow opens…

1: …with the name of the sequence in

the View Image Set window.

2: Normally, the MultiTime images are

played continuously, but an interval
may be set and inserted between the
images by clicking on the up/down
arrows to the right of the Delay
window. The delay interval is in
seconds.

3: Start/Pause the sequence by clicking

the Play button.

4: Skip one image forward button.

5: Move to the end of the sequence

button.

6: Skip back one image button.

7: Move to the start of the sequence

button.

506 © 2011 Leica Microsystems (Switzerland) Limited

 Time Lapse: Convert to Movie:

A sequence of images captured as a

MultiTime project may be converted into a
movie file which can then be played on
most media software.

To convert a project into a movie:

1: Click on the arrow to the right of the
Save Time Lapse Image Set as Movie
header to reveal the panel.

2: Click on the arrows to the right of the

Movie File Type text box and from the
list select the movie file format. The .
avi (Audio Video Interleave) format is
the default and the most popular for
video exchange.

3: Set the number of frames per

second. The ‘smoothest’ – least jerky
– movies will have a larger number of
frames per second. The range is 1 to
25. Double click on the Frames text
box and type a value.

4: Click on the Save as Movie File

button and the Creating xxx File
progress panel appears (5). xxx
refers to the file format.

6: Movie files are stored in the same

folder as the MultiTime project and
can be recognised by the file
extension – in this case .avi – and
they also appear in the Gallery as
thumbnails.

7: The resulting movie file can be shown

most popular video software
packages – in this case Windows ®
Media Player.

Preferences > Play Movie: Go there... 51

LAS User Manual 507

MultiTime: Movie:

The Movie module must be installed and enabled before compromising viewing quality with fast exposure to
it is available. See: Registration for details of installation. achieve a frame rate of 20 to 25 frames per second. This
will provide smooth playback.
The Movie module captures images or 'frames’ in real
time as a single, continuous file directly to the computer Movies may be split into a number of chunks or ‘clips’ all
hard drive. Frames are captured as quickly as the stored as separate files. Between the clips, capture is
camera and the computer will allow broadly governed by suspended so time and disk space are not wasted on
the resolution and exposure time. Aim for low resolution periods of specimen inactivity.
without

508 © 2011 Leica Microsystems (Switzerland) Limited

 Movie: Launch Movie and Select the Capture Archive:

1: Click on the Select Acquisition Mode

button and from the menu...

2: ...click to select Movie. The Movie

caption will appear on tabs in Acquire
and Browse Workflows.

Select the Fixed Capture Location:

The Time Lapse images will be captured
into the fixed location.
3: Click on the Browse Workflow and...

4: ...if necessary click on the Browse

tab. Click to select the capture
archive and then...

5: ...click on the Set Fixed Location

button.

6: To indicate the fixed capture location

archive, a red dot appears to the left
of it.

Continued... 510

LAS User Manual 509

Movie: Setup:

6: If it is not already selected, click on

the Acquire Workflow tab. The Movie
tab should now have been added to
the Acquire panels.

7: Click on the Camera tab. The number

of frames that can be captured each
second and therefore, the overall ‘real
time detail’ of the film, depends upon
the Exposure time, the Live Format
and the Camera.

8: Make sure the image quality is good

and the Exposure time is as short as
possible commensurate with a good
image. Carry out an Auto White
Balance if necessary.

9: Consider using 3 x3 Binning or

Progressive VGA for the Live Format
because although High Quality (HQ)
formats may be used they can result
in very large file sizes and very long
exposure times. For smooth playback
aim for 20 to 25 frames per second.

510 © 2011 Leica Microsystems (Switzerland) Limited

 Movie: Select Recording Mode:

1: In the Acquire Workflow click on the

Movie tab.

Two recording options are available:

Continuous results in a single clip

lasting as long as frames can be
captured within the constraints of the
hard disk space available.

Preferences > Movie Settings: Go there...

Define a sequence of clips creates a

single file for each clip. Clip duration
and interval may be set individually.
When the clips are replayed in Browse
they are 'assembled' into a single,
continuous movie.

The movie starts and records for the

duration of the first clip. It then stops until
the first interval has passed. The next clip
then runs for its designated duration,
recording stops and the second interval
starts, and so on until all the clips have
been recorded.

To select the Recording Mode:

2: Click on the Continuous button and
then go to Options.

3: Click on the Define sequence button.

With this option the selected
sequence setup panel appears, then.
go to Defining Clip Sequence.

Continued... 512

LAS User Manual 511

Movie: Define Clip Sequence:

On the Sequence panel:

1: Set the number of clips for the
sequence by clicking the up/down
arrows to the right of the Number of
clips text box.

2: Set the recording time for the clip(s)

by clicking on the appropriate field in
the Recording time window and typing
in a number, or using the up/down
arrows to the right of the window. The
fields are hours: minutes: seconds
(hh:mm:ss). If more than one clip is
required, each will record for the
same time.

3: If there is more than one clip, set the

interval between them by clicking on
the appropriate field in the Interval
window and typing in a number, or
using the up/down arrows to the right
of the window. The fields are hours:
minutes: seconds (hh:mm:ss). Each
clip will be separated by the same
interval.

4: Click on the Add Sequence icon to

load the sequence to the program
window (5).

Almost any number of sequences may be

added to a program. In the example there
are 3 sequences with a total of 5 clips.

6: Remove a sequence by clicking on it

and then on the Delete icon.

7: Change the order by clicking on a

sequence and then on the up/down
icon as appropriate.

512 © 2011 Leica Microsystems (Switzerland) Limited

 Movie: Name and Compression:

To give the movie a name:

1: Click in the Movie name text box and
type an appropriate name.

2: The first time a movie is captured, a

'common’ folder named Movies is
created in the capture directory.

3: Within this common folder, a new

folder is created for every new movie,
with the name specified in the Movie
name text box.

4: The movie ‘.avi’ file(s) together with

several control files, are stored in this
folder.

Select the Compression option:

Compression results in smaller movie files
but can affect image quality. The
compression is set on the Quality slider.
Values up to about 50% on the slider,
have little or no noticeable reduction in
quality: Higher figures represent a greater
level of compression and a smaller file
size, but may result in a noticeable loss of
quality.

Start with a value of 80% and then

experiment by gradually decreasing the to
achieve acceptable quality.

5: Click on the arrows to the right of the

Compression type window and…

6: from the drop down menu select

either None for no compression, or
MJPEG to apply compression.

7: If MJPEG is selected, the Quality

slider appears. This controls the
degree of compression applied to the
images and therefore the display
quality. Click and hold the slider and
drag to the left to decrease the quality
or to the right to increase it.

LAS User Manual 513

Movie: Time Stamp and Saving Configuration:

To display a progress time stamp on

the movie (1):
2: Click on the Font button and from the
Font dialog…

3: …select a font type, style and size

and click OK.

4: Click on the Colour button, select a

font colour from the Select Colour
dialog and click OK.

5: Click on the Include timestamp check

box to enable the time stamp.

Saving the Configuration:

The current settings may be saved with a
unique name and recalled for another
movie at a later date.

To save the current setting:

6: Click in the Configuration text box to
highlight the text, type a unique name
for the current configuration and
press Enter on the keyboard.

7: Click on the Save button which will

save all of the current settings.

514 © 2011 Leica Microsystems (Switzerland) Limited

 Movie: Existing Configuration and Testing:

Previously saved configurations:

These may be retrieved and used again to
load the settings for a new movie.

1: Click on the arrows to right of the

Options bar to reveal the Options
panel.

2: Click on the arrows to the right of the

Configuration text box and from the
drop down list…

3: Select the existing configuration to

use.

Deleting a Configuration:
4: When a configuration is selected the
Delete button becomes active. Click it
to delete the selected configuration.

Testing the Movie length:

With all of the settings made or an existing
configuration selected, check the movie
length by…

5: Clicking the Test button.

After a short delay the Approximate
maximum length (6) and Frames per
second count (7) will appear.
Aim for a frame count of at least 15
frames/second (fps) and preferable
closer to 26 fps.

If the frame count is too low, adjust

the exposure time on the Camera tab,
or increasing the light level at the
microscope, or by selecting a Live
Capture binning option that results in
smaller image sizes.
After making changes, click the Test
button to review the movie length and
frame count.

LAS User Manual 515

Movie: Start Recording and Progress:

To start recording the movie:

1: Click on the Acquire Movie button.

The Recording Progress panel shows the

Current Clip progress (2) with…

3: Time elapsed as hh:mm:ss,

4: Computed Time to completion as hh:

mm:ss and…

5: Real time Completion expected at.

There is also a progress bar (6) which
provides a graphical representation of
time elapsed for the current clip.

7: To stop the current clip and skip to

the next (if there is one), click on the
Pause button.

8: To halt the entire movie click on the

Stop button.

Overall progress is detailed in the upper

panel.

9: Time now is the real time clock.

10: Time elapsed for the entire movie,

all clips so far.

11: Computed Time to completion as

hh:mm:ss.

12: Real time Completion expected at.

There is also a graphical display of
time elapsed so far (13).

When the movie recording is complete, a

sample reference image (14) is placed in
the capture directory with a thumbnail.
Placing the cursor over the thumbnail
reveals the path and name (15).

516 © 2011 Leica Microsystems (Switzerland) Limited

 Movie: Play Controls:

After capture the display will open in the

Browse Workflow (1) in the specified
movie capture folder. The movie will
normally be played in the Viewer, but the
playback application can be changed by
clicking the Play with button (2) which will
launch the program specified in
Preferences: Image Output Settings: Play
Movie with…

The playback controls (3) on the Browse

Viewer are located on the Play Movie
panel. Click the icons:

4: To Start/Stop the movie.

5: Skip a frame forwards.

6: Skip a frame backwards.

7: Skip to the end of the movie.

8: Skip to the start of the movie.

9: Skip to the next or previous clip by

clicking on the left/right arrows to the
right of the Current movie clip
window.

10: The playback speed may be

adjusted in the text box (10). The
default is 100% which is playback
speed = recorded speed. Increase or
decrease the speed by clicking in the
text box and typing a speed (as a %
of the recorded speed), or clicking on
the up/down arrows to the right of the
text box.

LAS User Manual 517

Movie: Play and Frame Capture:

1: Click on the Start/Stop button. The

button icon changes to Pause on a
brown background.
Click the button again to halt the
movie.

2: The Progress Indicator moves from

left to right as a graphical indication of
running time.
Click and hold the slider and then
move it left or right to re-position the
movie playing position.

3: Running Time elapsed as hh:mm:ss

is displayed in the window.

Frame Capture:
Individual movie frames may be captured
and saved for future examination:

4: Click on the Start/Stop button to stop

the movie at the frame required.
Alternatively, use the Progress
Indicator to position the movie at the
frame and then click on the Start/Stop
button.

5: Click on the Save Image button.

6: The frame is stored in the current

capture folder (not in the movie
folders) with the name of the movie,
the name of the capture folder and a
sequential number which is
automatically generated (7). The file
name extension represents the
capture format (.jpg etc).

518 © 2011 Leica Microsystems (Switzerland) Limited

 Movie: Play Existing Movies:

1: Click on the Browse Workflow.

2: If necessary, click on the Browse tab

to reveal the Archive Navigator.

3: Click to select the archive that

contains the movie file to be played.

4: A thumbnail representing the movie

(s) appear in the Gallery.

5: Click on the Browse: Movie tab to

reveal the Play Movie controls.

See: Movie play controls: 517

See: Movie play and frame capture: 518

LAS User Manual 519

MultiFocus

Images obtained from microscopes have

a known and limited depth-of -focus. For
specimens with varying surface height,
when the Z-position of the specimen is
adjusted, different regions of the
specimen appear in focus. By collecting
digital images at these Z-positions, they
can be combined by an image processing
algorithm into one single sharp composite
image that the effectively extends the
depth-of-focus of the image.

Specimens that can benefit from LAS

MultiFocus are predominately those that
are imaged by light reflecting from the
surface such as geological and fossil
specimens, plant and marine biology,
histology and materials such as paper,
electronic components, metallurgy,
surface coatings and fractures

LAS MultiFocus provides fully integrated

software controlling microscopes with
motorised focus, cameras with high
resolution and colour fidelity that are
combined with a computer that
comfortably handles digital images. The
performance and relative economy of
modern computing makes the use of
sophisticated imaging algorithms
practicable for acquiring and processing
Z-Stacks of multiple images.

The principle is simple: An image that

varies in focus, is taken at steps across
the thickness of the specimen. These
‘slices’ are then mathematically combined
to form a MultiFocus Image – all of the
slices blended into a single, uniformly
sharp image.

Almost any number of slices (together

called a Z-Stack) can be captured – the
higher the number, generally the better
the MultiFocus Image – and each may be
saved and examined.

The MultiFocus application is suitable for

both manual and automatic microscopes.

520 © 2011 Leica Microsystems (Switzerland) Limited

 MultiFocus: Enable MultiFocus:

MultiFocus is an optional module which

must be installed and operational before
use

To enable MultiFocus:
1: Click on the Module Select icon.

2: From the drop down menu click on

the MultiFocus (Montage) icon to
enable it. The icon will only be
available if MultiFocus is installed.

3: The ‘Z’ logo appears on a new tab in

the Acquire Workflow and…

4: Also in the Browse Workflow.

LAS User Manual 521

MultiFocus: Select Image Format:

1: On the Acquire Workflow click on the

Camera tab.

2: Click on the arrow to the right of the

Options panel to reveal it.

3: Click on the arrow to the right of the

Captured Format header bar and
from the list of options…

4: Click to select the appropriate format.

Thick specimens may require a large
number of steps so consider a more
'compact’ format that does not
compromise quality but speeds the
capture time, reduces disk space and
makes processing quicker.

522 © 2011 Leica Microsystems (Switzerland) Limited

 MultiFocus: Select or Create Capture Folder:

Check the Preferences settings to ensure

images have the correct settings.
Preferences: Go there...

1: Click on the Browse Workflow and...

2: ...navigate to the folder in which to

capture the images.

3: If a new folder is required click on the

New Folder button and...

4: ...rename the folder appropriately.

5: Click on the Set Capture Location

button to automatically save the
images to the selected folder. Make
sure Capture to Fixed Location
checkbox in Preferences is enabled.

LAS User Manual 523

MultiFocus: Options:

1: Click on the Acquire Workflow to

select it.

2: Click on the Z tab to display the

MultiFocuscontrol panels.

3: If necessary, click on the arrow to the

right of the Options header bar to
reveal the Options panel.

Create MultiFocus after Stack Acquire:

4: Selecting this option – clicking the
checkbox – will automatically create a
MultiFocus Image when the last Z-
slice is acquired. Left un-checked,
the MultiFocus Image can be created
manually on the Viewer.

Save Sub-images:
5: If the MultiFocus Image is created
automatically (See 4 above), it may
not be necessary to save all of the Z-
slices or sub-images. Check this
option to discard the layers. Save
sub-images must be selected if
Create MultiFocus is unchecked
otherwise the prompt (6) appears. If
Save sub-images is checked, the
MultiFocus Image cannot be re-
created again; the entire process will
have to be repeated.

Enter the Stack Name:

7: Click in the Stack Name text box and
type an appropriate name for the Z-
Stack. The letter Z is automatically
prefixed to denote a MultiFocus
group, and a sequential number is
appended every time a new stack is
acquired.

Continued 525

524 © 2011 Leica Microsystems (Switzerland) Limited

 MultiFocus: Options: Align, Manual Focus and Wait Before acquire:

Align Images before combining:

8: Optical variations - predominantly in
stereo microscopes - can cause an
apparent shift in the point of focus
from layer to layer. The shift will be
corrected in software if the Align
Images option is checked so that all
X-Y points of focus are in exact
alignment.

Perform Manual Focus:

9: Check this option if the microscope
does not have a motorised focus and
each image is to be focussed
manually.

Wait Before Acquire:

10: Measured in milli-seconds, this
option inserts a delay between the
Acquire button being clicked and the
acquisition of the first layer. Click in
the text box and type a value.

LAS User Manual 525

MultiFocus: Configuration: New, Retrieve and Delete:

The Configuration facility allows

MultiFocus Z-step settings and options to
be saved, retrieved and used again.

Create a New Configuration:

1: Click in the Configuration text box and
type an appropriate file name for the
MultiFocus session.

2: Click on the Save button. The

settings are saved to disk and the file
name added to existing
Configurations. After the Z-Stack has
been set up, the Configuration will be
saved again to include the new stack
settings.

Retrieve an Existing Configuration:

3: Click on the arrows to the right of the
Configuration text box.

4: From the drop down list click to select

the Configuration required. The Last
Used option will automatically load the
settings used during the last
MultiFocus session.

Delete a Configuration:
5: Click the Delete button to remove the
Configuration displayed in the text
box. The settings cannot be retrieved.
The Last Used option may not be
deleted.

526 © 2011 Leica Microsystems (Switzerland) Limited

 MultiFocus: Define Z-Stack: Automatic Microscopes:

1: If necessary, click on the Acquire: Z

tab to reveal the MultiFocus control
panels.
Defining the stack consists of setting
the first Z position on the specimen,
called the Start, setting the last Z
position called the End, and then
deciding the number of focussed
'slices' or steps to capture between
Start and End.

Start and End may encompass either

the whole specimen from bottom to
top or any part of the specimen. The
positions chosen are displayed on a
graphical ‘stack’ (2) with the Start
position (3) shown as an arrow on the
left hand side, and the End position
as an arrow on the right hand side.
Windows attached to the arrows
display the microscope focus values.

The Z-Range window (5) will show

the numerical difference between the
Start and the End positions.
Z-Position (6) is the current
microscope focus position.
Z-Step (7) represents the distance
between each of the Z-steps and
Steps (8) is the number of images
that will be captured.
The Optimise step size button (9) will
determine the best step size based
upon the microscope and the optics.
Limit buttons (10) will automatically
move the objective to either the Start
or End position.

LAS User Manual 527

MultiFocus: Define Z-Stack Start:

Generally, the Start position is the part of

the specimen closest to the stage. It
provides the opportunity to place the
specimen on a textured background
which, when in sharp focus is a precise
reference for the software. In subsequent
'slices' the background will be out of focus
and so ignored.

1: Click on the Set Start arrow. It will

turn black indicating it is active and
will automatically track the
microscope movements.

2: Adjust the microscope either with its

manual controls or by using the
Application Suite interface, until the
specimen at the Start position is in
focus.

3: The microscope settings are reflected

in the Start window and the Start
arrow moves down the virtual ‘stack’
(4).

5: Click on the Start arrow which will

become red and locked at the
focussed position.

528 © 2011 Leica Microsystems (Switzerland) Limited

 MultiFocus: Define Z-Stack End:

1: Click on the Set End arrow. It will turn

black to indicate it is active and
tracking the microscope movements.

2: Focus on the part of the specimen

that represents the last Z-step using
either the microscopes manual
controls or the Application Suite
interface. The End arrow will move
along the virtual ‘stack’ in response to
the microscope.

3: Click on the End arrow which will

become red again and locked in the
End Z-step position.

Continued... 529

LAS User Manual 529

MultiFocus: Go To Z-Stack Start/End:

The illustrations show two images:

1: The Start Z-layer with part of the
specimen in focus together with a
sharp, textured background, and…

2: The End Z-layer with a different part

of the specimen ‘sharp’ but the
background out of focus.

Using the 'Go To' buttons:

3: The Go To Start button when clicked
will drive the microscope to the Start
position with the value displayed in
the Z-Position window (5) for a
focussing check.

4: The Go To End button drives the

microscope to the End position for a
focussing check. Again, the
microscope value is displayed in the
Z-Position window (5).

6: The Z-Range window shows the

distance between the Start and End
positions.

530 © 2011 Leica Microsystems (Switzerland) Limited

 MultiFocus: Set Z-Stack Steps:

With the Start and End points set, the

distance between the two is calculated
and displayed in the Z-Range and Z-Step
windows.

There are two methods of setting the

number of layers or steps:

1: Click in the Steps text box and type

the number of steps required. The
distance between each is calculated
and shown in the Z-Step text box.

2: Click in the Z-Step text box and type

the distance required between each
step. The range value is divided by
the entered figure and the resulting
number of steps displayed in the
Steps text box.

3: With the Stack Definition complete,

click the Options: Save button to save
the settings.

See: 532 Acquire Z-Stack Images. 532

LAS User Manual 531

MultiFocus: Acquire Z-Stack Images: Automatic Microscopes:

1: Click on the Acquire MultiFocus

button to start the acquisition.

2: At each step position, the microscope

will automatically move to the next
step and focus. A progress message
is displayed.

3: To cancel the sequence, click on the

Cancel button.

4: When all of the images have been

captured, the Browse Workflow opens
with the Show MultiFocus Image Set
control panel displayed (5).

532 © 2011 Leica Microsystems (Switzerland) Limited

 MultiFocus: Define Z-Stack: Manual Microscopes:

1: If necessary, click on the Acquire: Z

tab to reveal the MultiFocus control
panels.

Defining the stack consists of setting

the first Z position on the specimen,
called the Start, setting the last Z
position called the End, and then
deciding the number of 'slices' or
steps to capture between Start and
End.

Start and End may encompass either

the whole specimen from bottom to
top or any part of the specimen. The
positions chosen are displayed on a
graphical ‘stack’ (2) with the Start
position (3) shown as an arrow on the
left hand side, and the End position
as an arrow on the right hand side.
Windows attached to the arrows
display the microscope focus
positions.

The Z-Range window (5) will show

the numerical difference between the
Start and the End positions.

Z-Step (6) represents the distance

between each of the Z-steps and
Steps (7) is the number of images
that will be captured.

LAS User Manual 533

MultiFocus: Set Z-Stack Start Position:

Generally, the Start position is closest to

the stage because the specimen can be
set against a textured background which
will also be in focus. This gives the
software a benchmark for the background
so that the out-of-focus background of
subsequent steps will be ignored.

1: Focus on the start region of the

specimen making a note of the
microscope scale reading.

2: Click on the Set Start arrow which will

turn black, and type in the scale
reading.

3: Click on the Set Start arrow which will

become red again. The virtual ‘stack’
displays a graphical representation of
‘depth’.

4: The Z-Range value reflects the

entered Start position.

See: Set End position: Manual

Microscopes. 535

534 © 2011 Leica Microsystems (Switzerland) Limited

 MultiFocus: Set Z-Stack End Position:

1: Focus on the region of the specimen

that represents the End position.
Make a note of the microscope scale
reading.

2: Click on the Set End arrow which will

turn black, and type the scale reading
in the window.

3: Click on the Set End arrow which will

become red. The virtual ‘stack’ will
display a graphical representation of
the distance between the Start and
End positions.

4: The Z-Range window displays the

difference between the Start and End
microscope readings.

See: Set Start position: Manual

Microscopes. 534

LAS User Manual 535

MultiFocus: Set Z-Stack Steps:

With the Start and End points set, the

distance between the two is calculated
and displayed in the Z-Range and Z-Step
windows.

There are two manual methods of setting

the number of layers or steps:

1: Click in the Steps text box and type

the number of steps required. The
distance between each is calculated
and shown in the Z-Step text box, or...

2: Click on the Z-Step text box and type

the distance required between each
step. The range value is divided by
the entered figure and the resulting
number of steps displayed in the
Steps text box.

Optimise step size:

3: Enable the Optimise step size button
to have the steps calculated
automatically using the microscope
aperture. On stereo microscopes
check that the actual and displayed
iris settings correspond.

4: With the Stack Definition complete,

click the Options: Save button to save
the settings.

See: Acquire Z-Stack images: Manual

Microscopes. 538

536 © 2011 Leica Microsystems (Switzerland) Limited

 MultiFocus: Set Z-Stack Steps: Continued:

The illustrations show two images:

1: The Start Z-layer with part of the
specimen in focus together with a
sharp, textured background, and…

2: The End Z-layer with a different part

of the specimen ‘sharp’ but the
background out of focus.

3: The Z-Range window shows the

distance between the Start and End
Z-layers, and…

4: The number of steps and the ‘size’ of

each step in the Z-Step window.

LAS User Manual 537

MultiFocus: Acquire Z-Stack Images:

1: Click on the Acquire MultiFocus

button to start the acquisition.

2: At each step position, the prompt

appears. Re-focus the microscope
using the Z-Step value, always
focussing in the same direction.

3: Click OK to capture the image. To

cancel the sequence if it is believed
sufficient slices have been captured,
click on the Cancel button. The
number of captured images does not
have to correspond with the
calculated or entered number of
steps.

4: When all of the images have been

captured, the Browse Workflow opens
with the Show MultiFocus Image Set
control panel displayed (5).

538 © 2011 Leica Microsystems (Switzerland) Limited

 MultiFocus: Show Image Set Slide Show:

When all of the MultiFocus images are

captured, the Browse Workflow opens
with the Show MultiFocus Image Set panel
displayed.

The panel provides the controls necessary

to view the entire set as a ‘slide show’, a
single layer or the MultiFocus Image.

1: Set the delay between images in

seconds by:
Either clicking on the Delay Between
text box and typing a number or…
Using the up/down arrows to the right
of the text box.

2: Click on the Stack Images button to

select the images.

3: Click the Play button. It will change to

red with the pause symbol.

4: The current image and total images

are displayed in the Progress Window
and…

5: An indicator moves across the

Progress Bar. Each image is
displayed in turn in the Viewer.
Click the Play button again to ‘freeze’
the current image.

6: Move to individual images in the set

by clicking and holding the Progress
Bar indicator and dragging it left or
right.

7: Skip an image backward or forward

by clicking the Skip buttons.

8: Choose the first or last images by

clicking the Beginning or End buttons.

9: Select and display the MultiFocus

image by clicking the MultiFocus
Image button.

LAS User Manual 539

 Montage Multifocus:

Leica Application Suite Montage Multifocus provides Confidence Map: An image which contains an estimate
advanced, versatile features for producing excellent of the accuracy of the depth map at all points on the
extended depth-of-focus images using the renowned image..
technology of Auto-Montage from Syncroscopy.
Additionally, the image can be viewed in several different
Digital images from a Z-Stack, spread over the focus ways to examine the surface in greater detail:
range of the specimen, are acquired using the same
features provided by LAS Multifocus but because LAS Anaglyph.
Montage provides tunable algorithms is able to create
Stereo Pair
excellent extended depth-of-focus images covering a
wider range of microscopy contrast methods. Colour Relief
3D Model (Optional extra module) which provides a
LAS Multifocus provides only one montage method and is
perspective visualisation of the extended focus image
effective only on narrow range of specimen types. There
for which the user can change the viewpoint so that the
are no facilities for adapting to different situations and no
image give an impression of depth. Note: LAS Montage
means of enhancing result image or alternative means of
is a pre-requisite for the use of LAS 3D Viewer.
visualising the result image.
The extensive Measure Tools include a fast
LAS Montage has many additional capabilities to extend
the imaging conditions that can be used and improve the
Profile display...
quality of the resulting image.
...that maps and measures the contours of a user-drawn
LAS Montage extends Multifocus by adding functions to
path.
provide:
Z-Stack acquisition is the same for both Multifocus and
Depth Map: An image which contains depth information
Montage. To find out more: Go there... 520
for all points on the image,

540 © 2011 Leica Microsystems (Switzerland) Limited

 Montage Multifocus: Launch and Operating Steps:

Launch Montage by:

1: Clicking on the Acquisition Mode button...

2: ...and then on the the Montage icon.

Montage Multifocus OperatingSteps:

Acquire the Z-Stack: Go there.. 520 . 520
Select the Z-Stack: Go there... 542
Select the Z-Stack images to use: Go there...
544
Create the Montage Multifocus image: Go
there... 545
Make Enhancements: Go there... 547
View the Results: Go there... 552
Edit the Results: Go there... 560
Create additional views if needed: Go there...
554
Use the Measure Tools: Go there... 563

LAS User Manual 541

 Montage Multifocus: Select the Z Stack:

1: Click to select the Browse Workflow.

2: Click the Browse tab and...

3: ... select the required Z-Stack on the

Navigator.

4: Click on the Z tab to open the Montage

control panel.

5: If necessary, click to expand the Show

Multifocus Image Set panel.

The Z-Stack images will be loaded into the

gallery together with any associated Multifocus
image and maps.

Clicking on a thumbnail in the Gallery will

display the image.

From this panel click to review the following

images:

Stack Images 543 . 543

Multifocus Image 545
Depth Map 552
Confidence Map 553
Anaglyph 554
Stereo Pair 555
Colour relief 556
3D Model 558
Leica Map, if it is installed, has its own help:
Go there... 571

542 © 2011 Leica Microsystems (Switzerland) Limited

 Montage Multifocus: Stack Images:

1: To view the Z-Stack sequence click on

the Stack Images button. This will display
the first image in the stack and enable the
controls to view the stack.

2: Enter a delay time between images and

then use the control buttons to scan the
images.

3: Click to play the image sequence

continuously with the specified delay time
between each image. Click again to stop.

4: Moves to first and last image in the

sequence.

5: Steps forward or back in the sequence.

6: Click and drag on the red slider to move

forward or back through the sequence.
The display also shows the position in the
sequence while the sequence is played.

LAS User Manual 543

 Montage Multifocus: Source Image Selection:

The stack images are listed in the Source

Images panel from where they can be selected
for inclusion in the Montage.

Click to select the Process Workflow and then

the Z tab.

1: Click on the arrow to the right of the

Source Images header to reveal the
panel.

2: The image sequence can be swapped

between starting with the top of stack or
starting with the bottom of the stack by
clicking the arrows and selecting the
option from the drop-down list.

3: Clicking on an image in the list will select

it and display it in the Image Viewer.

Clicking again toggles between IGNORE

and Include in Multifocus. Any image that
is ignored will not be used in the
generation of the Multifocus image.

4: If the Align Images before combining box

is checked on the Montage Multifocus
panel (Go there... 545 ) then relative
alignment data will be shown here.

544 © 2011 Leica Microsystems (Switzerland) Limited

 Montage Multifocus: Create Multifocus Image:

If the source images are not exactly

registered with each other, having been
captured with a stereo microscope or
from scanned photographs, this should
be corrected with the Align Source
Images function before proceeding with
the Montage operation.

1: Click on the Process Workflow.

2: Click on the Z tab.

3: On the Montage Multifocus panel,

click to enable (tick mark visible)
Align source images first.

4: Choose the Montage Method that is

closest to the contrast method used when
the Z-Stack was created by clicking on the
arrows to the right of the Method header
and selecting from the drop-down list:

Reflected light: Four degrees of smoothing -

none, minimum, medium or maximum ...

Transmitted light: Methods 1, 2 and 3.

Fluorescence: Minimum and maximum

intensity.

5: Select the Background to reduce the effect

the background has on the Multifocus
image.

Continued... 546

LAS User Manual 545

Montage Multifocus: Patch, Preview and Create:

1: The Patch Size refers to the spread of

pixels used to calculate the montage. A
recommended range is 8 to 20: Too small
and and the detail will be good but noise
could be a problem. Too large and whilst
the depth maps will be smoother there
could be less detail.

Adjust the Patch Size by clicking the up/

down arrows to increase/decrease it.

2: Click to enable the Enhance after Create

check box to automatically apply any
settings made on the Enhance dialogs (
Go there.. 547 . 547 ). This is a convenient
way of creating a final Multifocus image
but enhancements can always be made
after the image is created.

3: To preview a result, click to enable the

Preview check box and...

4: ...click on the arrows to the right of the

header and choose the image to be
previewed from the drop-down list.

5: The Preview is automatically generated

and the area can be enlarged or reduced
by clicking and dragging the corner arrow.

Click the STOP button to halt the preview

generation.

Click and drag inside the Preview window

to move the area of interest.

To close the Preview window, click to

disable the Preview check box (2).

6: Click the Create Multifocus Image button.

546 © 2011 Leica Microsystems (Switzerland) Limited

 Montage Multifocus: Enhance:

The Montage and Depth Map images may be

processed prior to publication. Smoothing the
Depth Map can be used to remove artefacts
that may appear in regions that lack in focus
detail. Sharpening the Montage image may
improve the images appearance.

The Montage Enhance panel is divided into

three main control areas:

Background Confidence (A)..

Enhance Multifocus Image (B) and

Enhance Depth Map Image (C).

Additionally, there is a Preview facility and the

Enhance Images button that applies the
enhancements.

Continued... 548

LAS User Manual 547

 Enhance: Background Confidence:

Note - background is now removed using the

Background selection from the Montage
MultiFocus panel. This information is retained
only for reference.

Please set control 1 to 0.

Adjusting the Background Confidence value

will define a new Background Mask layer
which can be applied to both the Multifocus
and Depth Map images.

The Background Mask can be derived from

any or all of the layers and is simply the parts
of the images that remains after all of the
desirable image information has been
extracted.

1: The threshold for the Background

Confidence can be set to a value between
0 and 100% by clicking inside the text box
and typing a value or using the up/down
arrows.

Any pixel with a confidence value in the

range 0 to Background Confidence% will
be regarded as a background pixel.

2: The boundary between the Background

Mask layer and the rest of the image may
be smoothed depending on the value of
the smoothing factor. Click on the arrows
to the right of the header and from the
drop-down list select from:

Hard edges.

Soft Edges(Min)

Soft Edges(Med)

Soft Edges(Max)

Continued... 549

548 © 2011 Leica Microsystems (Switzerland) Limited

 Enhance: Multifocus Image:

Background:
Apply various effects to the Montage image,
but only within the region where a pixel has a
confidence value in the range 0 - Background
Confidence% - the Background Mask.

1: Click on the arrows to the right of the

header and from the drop-down list click
to select:

No Effect: No effect on Montage image..

Background: Over the Background Mask

region the background layer is copied to
the Montage image

Smooth: (3 levels): Over the Background

Mask region a smoothing filter is applied
to the Montage image

Flood Black, Grey or White: Over the

Background Mask region, the Montage
image is set to a solid colour

Darken: Over the Background mask

region, the intensity of the Montage image
is reduced

Whole Image:
Sharpen or smooth the parts of the Multifocus
image where the confidence is higher than
the Background Confidence%.

2: Click on the arrows to the right of the

header and from the drop-down list click
to select:

No Effect: No effect on Montage image..

Sharpen (3 levels): or...

Smooth (3 levels).

Continued... 550

LAS User Manual 549

 Enhance: Depth Map Image:

Background:
Apply various effects to the Depth Map image,
but only within the filter region where a pixel
has a confidence value in the range 0 -
Background Confidence% - the Background
mask.

1: Click on the arrows to the right of the

header and from the drop-down list click
to select:

No Effect: No effect on Depth Map image.

.

Smooth: (3 levels): Over the Background

mask region a smoothing filter is applied
to the Depth Map image

Whole Image:
Sharpen or smooth the parts of the Depth Map
image where the confidence is higher than
the Background Confidence%.

2: Click on the arrows to the right of the

header and from the drop-down list click
to select:

No Effect: No effect on Depth Map image

or..

Smooth (3 levels).

Continued... 551

550 © 2011 Leica Microsystems (Switzerland) Limited

 Enhance: Preview:

To preview the Montage Enhance settings:

1: Click to enable the Preview check box.

2: Select the image type to preview by

clicking on the arrows to the right of the
header and choosing from the drop-down
list. The Preview Viewer opens.
To move the area of interest within the
Viewer, click and drag the image.

3: When the Enhance settings are

acceptable, click on the Enhance Images
button to apply them to the images.

LAS User Manual 551

 Montage Multifocus: Depth Map:

A Depth Map image is generated as part

of the Montage process.

The Depth Map is a record of which

source image provided the in-focus
region for the montage image at each
pixel location, expressed as a grey level.

The Depth Map image is the same size

as the source images used to generate
it, but is always monochrome. The grey
levels depend on the particular Montage
Method used to generate the depth map.

The Depth Map image is displayed in the depth

map image window and is saved in the dataset
file itself, but can also be exported to an image
file if required.

552 © 2011 Leica Microsystems (Switzerland) Limited

 Montage Multifocus: Confidence Map:

A Confidence Map is also generated for

each montage operation. It is an
estimate of accuracy of the depth map
image at each pixel location, expressed
as a grey level.

White represents high confidence and

therefore an accurate depth value. Black
represents low confidence and an
uncertain depth value.

Low confidence will often represent

areas of low contrast where several
planes will appear to be in focus.

The Confidence image is the same size as the

source images used to generate it, but is
always monochrome. Values are expressed as
percentages, ranging from 0% (no confidence
that depth map is correct for that pixel) to 100%
(complete confidence).

LAS User Manual 553

Montage Multifocus: Anaglyph:

The Montage Anaglyph operation

generates the anaglyph image, which
when viewed through suitable red and
green/blue lenses offers a 3-dimensional
monochrome view of the montage
image, with depth information
synthesized from the depth map.

It is the same size as the Montage

image used to generate it, but always
comprises 3 RGB planes.

The anaglyph image will be saved as

one of the result image files.

1: Select Method from drop-down list.

Choose Red/Green or Red/Blue to match
the glasses to be used. Choose Red/Cyan
for better printed results - the effect
depends on the Montage image.

2: Check Colourise to add a hint of the

original Montage image colour back to the
Anaglyph image - the final effect depends
on the Montage image.

Colourise is not available with Red/Cyan

option because all 3 colours are effectively
already used.

3: Set the amount of Separation - the

maximum amount, in pixels, by which left
and right-eye images are displaced to
provide the stereo effect. Lower values are
recommended in most cases.
Click and drag the slider or if the mouse
has a wheel for scrolling, click on the slider
and use the wheel to adjust the parameter
value.

4: Check Preview to see the effect of the

settings in the Preview window

5: Click Create Anaglyph to create see the

finished Anaglyph image in the Viewer.

554 © 2011 Leica Microsystems (Switzerland) Limited

 Montage Multifocus: Stereo Pair:

The Montage Stereo Pair operation

generates the Stereo Pair image, which
when viewed correctly offers a 3-
dimensional colour view of the montage
image - with depth information
synthesized from the depth map.

The stereo pair image comprises two

similar images, in colour or greyscale
depending on the source image, each
the same size and pixel depth as the
Montage image used to generate it.

The Stereo Pair image will be saved as

one of the result image files.

1: Select Source from drop down box. The

stereo pair may be generated from:

A monochrome representation of the

Montage image,.
From the original colour Montage image - if
the source images is colour, or
From the colour relief image if that has
been generated.

2: Select Layout. For normal use, select Left/

Right. However, some people find it easier
to view a stereo pair with the images
flipped Right/Left

3: Set the amount of Separation - the

maximum amount, in pixels, by which left
and right-eye images are displaced to
provide the stereo effect. Lower values are
recommended in most cases.
Hint: If your mouse has a wheel for
scrolling, click on the slider and use the
wheel to adjust the parameter value.

4: Click Create Stereo Pair.

LAS User Manual 555

 Montage Multifocus: Colour Relief:

Montage Colour Relief generates the

colour relief image that offers a view of
the Montage image colour-coded with
depth information from the Depth Map.

The Colour Relief image is the same

size as the Montage image used to
generate it, but always comprises 3
RGB planes.

Experiment with the controls to find a

representation which works well with
your particular images:

Images of surfaces consisting mainly of

highlight detail tend to look good with mode set
to Light Features and Saturation slightly left of
centre.
Images of biological samples with dark fibres
tend to look good with mode set to Invert.

A pair of red/green or red/cyan spectacles as

used for anaglyphs, may help to interpret colour
as depth.

The Colour Relief image will be saved as one of

the result image files.

Continued... 557

556 © 2011 Leica Microsystems (Switzerland) Limited

 Colour Relief: Methods:

1: Select Method from drop down box.

Experiment with this control to find which
mode works best with the images:

Light Features applies the colour effect

mostly to the lighter parts of the
montage image.

Dark Features applies the colour effect

mostly to the darker parts of the
montage image.

Invert applies the colour effect to the

darker parts of the montage image and
darkens the background.

2: Click on Apply Confidence to use the

confidence image to suppress unfocused
parts of the montage image.

3: Select the Colours scheme in which to

plot the colour relief image.

4: Adjust the Saturation. Saturation varies

from purely monochrome intensity (left-
most position) to full-coloured depth map
with no intensity information (right-most
position).

Click and drag the slider or if the mouse

has a wheel for scrolling, click on the
slider and use the wheel to adjust the
parameter value.

5: Check Preview to see the effect of your

settings in the Preview window

6: Click Create Colour Relief to see the

finished image in the Viewer.

LAS User Manual 557

Montage Multifocus: 3D Model:

The 3D Model is an Optional Module

that must be licensed and installed. If it
is not, the button will not appear in the
tool list.

The module is launched from the

Browse Workflow:

1: Click on the Browse Workflow.

2: Click to select the Z Tab.

3: On the tool list, click the 3D Model

button.

Continued... 559

558 © 2011 Leica Microsystems (Switzerland) Limited

 Montage Multifocus: 3D Model: Continued:

The 3D Model is a 3D surface view of the

image based on the Depth Map information
and can be viewed from different angles by
clicking on the model and dragging the mouse.

The model can be created in three different

Display Modes:

A: Solid Colour: The model is displayed as a

surface relief with a single colour. The
colour can be set by clicking on the Set
button..

B: Multifocus Image: The model is displayed

as a surface relief overlayed with the
Multifocus image.

C: Colour Scale: The model is displayed as

a surface relief overlayed with the colour
relief image showing the depth map
contours.

1: Select the display mode from the drop

down box.

The brightness of the image can be

adjusted using the Ambient light slider and
the illumination direction with the
Directional light slider.

2: With the Solid Colour mode selected, the

colour can be changed by clicking the Set
button and choosing a colour from the
dialog.

3: Set Rendering to Speed or Accuracy to

change the response time of the model
display in Colour Scale mode.

4: The image can be 'stretched' along the Z

axis by clicking and dragging the Z Axis
Scale slider.

5: Click the Create Image button.

LAS User Manual 559

 Montage Edit:

The Montage Edit tool is used to copy and

blend pixels from one image - the source, - to
another - the target. It is best described using
a hypothetical example.

Image (A) is the resulting Montage from the Z

Stack of a bolt. Most of the image is sharp with
the exception of the serial number (circled)
which is important to the job but out of focus
and indecipherable. This will be the target of
some edited pixels.

Image (B) is Z Stack image number (10) and

whilst it is mainly out of focus, the serial
number is sharp and readable. This will be the
source of the edited pixels.

(C) Montage Edit allows the user to extract a

section of a selected stack image and 'blend' it
into the final montage.
Now the Montage is sharp and the serial
number from Z Stack image (10) has been
included and maintains its sharpness.

The Dual Viewer facility allows the user to view

both the final montage and the edit source
side-by-side so that the result can be seen
immediately and editing is an easy and fast
operation.

The Depth Map can also be the target of the

edit.

Continued... 561

560 © 2011 Leica Microsystems (Switzerland) Limited

 Montage Edit: Dual Viewer:

Both the source and target of the edited pixels can be

viewed side-by-side by using the Dual Viewer facility: 4: Click in the right-hand pane.

1: Click on Viewer Options icon on the Side Tool Bar. 5: Click on the source of the pixels - in this case
From the drop-down list click to select Dual Viewer. Image010.jpg. The image appears in the right-hand
The Viewer separates into two panes. pane.

2: Click in the left-hand pane. Continued... 562

3: In the Gallery, click on the target thumbnail - in this

case the Montage Multifocus image but it could be
the Depth Map. The image appears in the left-hand
pane.

LAS User Manual 561

 Montage Edit: Select the Edit Area:

1: Click the Edit Mode check box to enable

editing.

2: Click inside the Select Layer text box and

type the source image number - in the
example image 10.
Alternatively, click inside the text box and
then click on the image thumbnail which
will automatically transfer the image
number.

3: Region Select Style allows the easy

selection of an area using the mouse. The
selected area is outlined with a red line.

4: There are 4 possibilities for the Region

Edge Style:
The left-most selection uses a hard edge
with no blending of the edited area and
the background image.
Each of the other 3 edge styles add
progressively heavier blurring of the
compound image and the selected layer,
while still remaining within the selected
outline. Click to select.

5: Define the source edit area:

Create a rectangular area by clicking at

each corner..

Create a freehand area by clicking and

dragging to draw the edit area.

Double-click to close the shape and

define the edit area.

If a mistake is made while defining an outline,

a right mouse click can be used to delete the
last point. This can be repeated until the line is
entirely removed.
To quickly remove an entire line, uncheck the
Edit Mode checkbox.
The Undo / Redo panel can be used if the
results of an edit operation are not as desired.

The edited target is automatically saved. To

revert to the original Montage Multifocus
image create a new image (Go there... 545 )
that is not influenced by the edit.

562 © 2011 Leica Microsystems (Switzerland) Limited

 Montage Multifocus: Measure Tools Position Readout:

Measure Tools allows the user to read depth

information from a point or line on a Montage
Multifocus Image and display a profile of the
surface as a graph on a chart. Measurements
may be absolute or relative to a reference
point set by the user.

This mode is controlled by several checkboxes

on the panel. The panel is inactive if no depth
map is available for the Multifocus image. (The
depth map is created when Montage
Multifocus makes a new Multifocus image.)

1: Check to enable the Position Readout

check box to give a continuous readout of
the current mouse position as the mouse
is moved over the image.

2: The X and Y positions are given in

calibrated units relative to the top left
corner of the image.

The Z position is taken from interpreting

the depth map and is displayed in
microns.

Distance is measured from the top left

corner of the image or from a Reference
Point placed on the image. Distance
follows a path constrained to the surface
and prescribed by the Depth Map. The
Distance along the surface will be longer
than the straight-line distance between the
start and end point.
It is best practice to use calibrated images for Distance
3: Set a Reference Point on the image by measurement. A nominal one micron per pixel is used as a default
clicking the Set button and... for un-calibrated images.

4: ...clicking on the image where the Continued... 564

Reference is to be placed.

5: Click the Clear button to remove a

Reference Point.

LAS User Manual 563

 Measurement Tools: Profile Display:

Two check boxes control the Profile display:

1: Enable Profile (with Edit disabled) to

draw...

2: ... a straight line on the image with the

mouse. Each new line will replace any
previous one.

3: The Profile display will show the graph of

the surface section. Use the controls top-
right to display the Profile full screen or to
close it.

4: Click to enable Edit to interact with an

existing line or move a reference point.
The line can be moved by clicking and
dragging it or rotated by clicking a
dragging an end point.

A Reference Point can also be

repositioned. The Profile graph changes
accordingly.

Placing a Reference Point on the image

sets a zero level for Z values and the
display will change to include a Zero line
with heights both above and below it.

Continued... 565

564 © 2011 Leica Microsystems (Switzerland) Limited

 Measure Tools: Change Line Colour:

Change the Line and Reference Point colour by: 2: From the Select Colour dialog click to select a basic
colour, drag the target to the required colour on the
1: Click on the Line Mode colour button. circle or enter values in the Current Colours text
boxes.

Continued... 566

LAS User Manual 565

 Profile Panel Context Menu:

The Profile Panel has a context menu selected with a To zoom a portion of the graph, left-click and drag a
right-click. rectangle around the area of interest.

The Profile can be copied to the clipboard,. Select Un-Zoom to remove the zoomed area.

Saved to a named file and... To Pan a zoomed chart hold down the left Shift key,
click and drag the mouse.
Printed.
Save Profile As exports the profile to a text file that can
Selecting Show Point Values provides a read-out of the be imported into Microsoft Excel.
graphs co-ordinates when the cursor is placed near the
graph.

566 © 2011 Leica Microsystems (Switzerland) Limited

 Montage Methods Appendix: Essentials & Reflected Light:

The purpose of the Montage Methods is to create a single the same X/Y location. Nine source images comprise the
in-focus image by combining parts of multiple images Z-Stack in the illustration with the single pixel location
taken at different Z positions. Each individual image may represented bottom right of each.
contributes some focussed areas to the final image.
On comparison with neighbouring pixels in the same
The multiple images are acquired in order of Z position image, a numerical measure of ‘sharpness of focus’ is
ideally at steps in the Z position that are separated by less calculated for that pixel within each source image.
than the depth of focus of the objective being used. This A graph of focus against depth (Source Image Index) is
set of images is called an image- or Z- 'stack' and the then plotted for that pixel location in all source images.
individual images are referred to as slices.
The focus comparison and the algorithms described in the
Consider a single pixel from each source image (slice) at following illustrations are repeated for every pixel X/Y
location within the series of source images.

Reflected Light: No Smoothing: Fixed: The resulting Montage image is generated by pasting in
Selects the single source image plane which is in best the single pixel value from the selected source image at
focus at each pixel location. that location.

The resulting Depth Map - a list of each in-focus pixel with Reflected Light No Smoothing is the quickest algorithm,
its source slice and X/Y co-ordinates - is generated with so is useful for determining optimal parameter values
whole integer values referring to a single source image before performing a Reflected Light with Smoothing or
index for each location and the depth map image will be Transmitted Light montage operation. The Depth Maps
seen to have a ‘layered’ appearance. produced are not suitable for 3D viewing or
measurement.

Note - this method is used by the MultiFocus module.

Depth Map image value (3).

Montage Image value copied from single source image
(3).

LAS User Manual 567

 Montage Methods: Reflected Light with Smoothing:

Reflected Light: Smoothing: Blended: The Depth Map image will be seen to have a ‘smooth’
Takes into account the effects of adjacent in-focus planes appearance.
at any one pixel location, and attempts to predict the point
of maximum focus more accurately than the no smoothing The Montage image is generated by interpolating between
method. adjacent source images according to the fractional part of
the depth map value.
The Depth Map is generated with fractional floating-point
values which shows slopes within the sample more Reflected Light with Smoothing gives good results with
accurately. many types of specimen.

Depth Map image value 2.7.

Montage Image value calculated from relative proportions
of source images (2) and (3).

Blended Variations Min, Med & Max: three values. This tends to squeeze the Depth Map a bit
Blended (Min): The best focus for each pixel is compared more enthusiastically and should cope better when the
with its focus in the two immediately adjacent source sample has a continuous gradient but focus has
images and an interpolation is performed between those been moved too far between successive source images.
three values. This is expected to give the best results for
most cases Blended (Max): This is like Med above but then applies a
gentle smoothing to the Depth Map only. This should
Blended (Med): The best focus for each pixel is biased probably be viewed as a last resort to pull out acceptable-
towards whichever of the immediately adjacent source looking results when the other two have failed.
images has stronger focus in a ratio calculated using all

568 © 2011 Leica Microsystems (Switzerland) Limited

 Montage Methods: Transmitted Light:

Transmitted Light: Method 1: Weighted: Transmitted Light: Method 1 depth is particularly effective
Takes into account multiple in-focus planes at any one on transparent biological samples, where more than one
pixel location. plane may contain focussed detail; The result is a
combination of all focused planes, rather than a choice
The Depth Map is generated with fractional floating-point between them.
values but may contain inaccurate depth values which do
not correspond to an optimally-focussed source image. The result is expected to be a ‘soft’ focus not sharp.

The Montage image is generated by calculating relative

proportions of all source images.

Depth Map image value of 3.6 is not accurate.

Montage Image calculated from relative proportions

of all source images.

Transmitted Light: Method 2: Weighted Exponentially: The resulting Montage image is generated by calculating
Takes into account multiple in-focus planes at any one relative proportions of all source images.
pixel location, but biased more strongly towards the best
focus. Particularly effective on biological samples with intrusive
background.
The Depth Map is similar to that generated by the weighted
depth algorithm, although it will usually be more accurate.

Depth Map image value of 5.4 is not accurate.

Montage Image calculated from relative proportions of

all source images.

LAS User Manual 569

 Montage Methods: Transmitted Light:

Transmitted Light: Method 3: Compound Weighted:

Takes into account multiple in-focus planes at any one The resulting Montage image is generated by calculating
pixel location, but biased more strongly towards the best relative proportions of all source images.
focus.
Particularly effective on biological samples with intrusive
The Depth Map is similar to that generated by the background.
weighted depth algorithm, although it will usually be more
accurate.

Depth Map image value of 5.4 is not accurate.

Montage Image calculated from relative proportions of

all source images.

Fluorescence Min and Max Intensity: Projection: Place this value in the result image and repeat for all X/Y
A single pixel from each source image at the same X/Y locations.
locations is used as in all other cases.
Often used on fluorescent images where light is being
Determine the maximum or minimum value that is found. generated from various Z positions in the specimen.

Minimum

Maximum

570 © 2011 Leica Microsystems (Switzerland) Limited

 Leica Map

Leica Map is a powerful and sophisticated metrological installed and licensed for Leica Map to work properly. The
application for analysing complex surface profiles and Montage stack must be captured using a well-maintained
structures. motorised microscope stand because the Z-positions must
be accurate. Refer to the Montage help: Go there... 540
It works in conjunction with the LAS Montage module to
produce comprehensive reports using a wide range of The following pages show how to export data to and
tools for both surface data manipulation and display. The launch Leica Map, and also provides a brief illustration of
reports can be customised to suite corporate or project how it can be used. It has its own extensive help that can
styles and guidelines. be launched from the Main Menu or used contextually by
pressing key F1.
The working data is sourced from a Montage image
sequence that has a processed montage image and depth Leica Map is a very flexible and adaptable program but
map. This is the only data 'route' so Montage must be outcomes are always dependent upon the suitability of the
original specimen and the users interpretation of the
results.

LAS User Manual 571

 Leica Map: Installation and dongle

Leica Map is a Windows application not an

optional module that runs independently from
LAS.and is installed from the LAS DVD. The
dongle provided must be fitted to a USB slot on
the computer whilst the program is running.

Leica Map can be used initially in demo mode

with full functionality for a limited period. It is
recommended that it is not installed until you
are ready to start an evaluation.

However after the demo period expires, a

license must be purchased for continued
operation. A Leica Map dongle (separate from
the LAS dongle) will be .supplied to enable the
license. It is recommended that the Leica Map
dongle is attached to a USB port on the
computer before it is booted to make sure of
validation before Leica Map is launched.

Leica Map can be launched directly from the

desktop icon once installed for use with images
directly from the computer hard drive.

572 © 2011 Leica Microsystems (Switzerland) Limited

 Leica Map : Select the File Format

Leica Map must be configured to accept the

Montage file formats and layer structure.

To select the file format:

1: Click on Options on the main menu and...

2: ...from the drop-down menu click to

select General Preferences. Alternatively,
press F7 on the keyboard.

3: On the Preferences panel, click the File

Formats tab and...

4: ...the Select Formats button.

5: The list of file formats appears. Scroll

down the list until the Leica Formats entry
appears and click to enable the check
box.

6: Click OK.

Continued... 574

LAS User Manual 573

 Leica Map : Select the Layer Type

Select the Layer type as follows:

1: Click on Options on the main menu and...

2: ...from the drop-down menu click to

select General Preferences. Alternatively,
press F7 on the keyboard.

3: On the Preferences panel click the

Multilayer tab.

4: Click to select the Load each layer as a

separate studiable radio button.

5: Click OK.

574 © 2011 Leica Microsystems (Switzerland) Limited

 Leica Map: Montage Results

Users that have just captured a Z-Stack

sequence in Acquire and have automatically
come to the Browse Workflow because of the
'After Capture' settings in Preferences, can
skip this page.

For a previously captured Z-Stack sequence:

1: The Montage module must be active. If it

is not click on the Selector button and...

2: ...from the Acquisition Mode list click to

select Montage.

3: Click on the Browse Workflow and...

4: ...on the Browse tab.

5: In the Navigator locate the folder

containing the Montage Z-Stack sequence
and open it.

6: In the Z-Stack sequence there must be a

Depth Map and...

7: ...a Montage extended focus image.

Continued...

LAS User Manual 575

 Leica Map: Launch Leica Map

To launch the Leica Map application from

Leica Application Suite with Montage active:

1: Still in the Browse Workflow, click on the

Z tab.

2: Click on the Leica Map button. This will

only be available if Leica Map is installed
and the dongle is present and validated.

3: Leica Map is launched and the flash

screen appears.

If Leica Map is already active it is detected

and the Montage data transferred to it
automatically.

Continued...

576 © 2011 Leica Microsystems (Switzerland) Limited

 Leica Map: The Template Option

Before image analysis begins, the user has the template 3: ...navigating to the folder in which the templates are
option: stored. The location for the default template is:

Use an existing template - either the default provided C:\Users\Public\Documents\Leica Application

with Leica or one created by the user, or... Suite\Leica Map Templates\default.mnt.

A basic report which can have specific analyses added 4: Click the Export button.
to suit the project.
Any previously saved Leica Map report with the .mnt
Using a Template: extension stored in any location can be used as a
1: To use a template for the report, click to enable the template. However, but some Montage images may not be
Template check box - a tick mark appears. appropriate to the Operators and Studies used in the
template and in this case Leica Map will automatically
2: Select a template by clicking the browse button and... close.

Template examples are shown on the following pages.

LAS User Manual 577

 Leica Map: The Default Template

The default 3D Parameters template (1) creates a report Height distribution and Skewness), Functional Parameters
that contains much of the data prescribed in the ISO (Surface Bearing and Extreme Height) and Features (Peak
25178 Surface Texture Standards. Density etc) derived from segmentation.

A Topography pseudo colour image (2) is accompanied by Also included but not shown on the illustration, are
a Height Parameters table (3) containing data such as Waviness and Roughness pseudo colour images.
Mean Height of the Surface, Maximum Height of the
Peaks, Valleys and the entire Surface, and the Root Mean Pages can be added to the report together with a wide
Square Height. Spatial Parameters include Texture Aspect range of specific data and images. Operators and Studies:
Ratio and Texture Direction. Go there... 580

The Abbot-Firestone Curve (4) is represented as well as a

table (5) showing Height Parameters (such as Kurtosis

578 © 2011 Leica Microsystems (Switzerland) Limited

 Leica Map: No Template

1: If the Template option is cleared the browse button is The user can then add tables, analyses and images -
no longer available and a basic report is created. called Studiables - appropriate to the project using
Operators and Studies: 580 Go there... 580
2: Click Export and only essential images such as the
Topography... More pages can be added and the various components
moved around with borders, text, captions, company
3: ...and Intensity layers are included on an otherwise logos, names and report headers added as required. The
blank report sheet. resulting document can then be used as a template for
other, similar projects.
4: The report example shown here also includes the
original Montage image.

LAS User Manual 579

 Leica Map: Operators

Leica Map performs surface analysis by

applying Studies and Operators to the data
being analysed (known as a Studiable), in this
case Depth Map and Montage Images.

Studies are graphical representations of a

studiable such as a 3D view or an analysis
performed on a studiable such as a step
height measurement.

Operators are mathematical transformations

used to modify a studiable by adjusting
settings in an associated dialog box.
Examples would be to level a surface or to
extract the waviness by filtering.

Many Operators are interactive providing

controls to adjust for example, filter structures,
thresholds and edge detection.

1: With an image on the report selected, on

the Main Menu click the Operators option.

2: The Operator dialog appears. The

controls and output displays differ with
the Operator chosen. Change the
parameters as needed and...

3: ...to include the results in the report click

the OK button. Clicking the Cancel button
will close the dialog and not include the
results in the report.

Continued...

580 © 2011 Leica Microsystems (Switzerland) Limited

 Leica Map: Studies
Studies provide different methods for
representing the surface data.

1: On th Main Menu click the Studies option.

2: From the drop-down, click to select the

required study type.

3: If necessary, a new page is added

automatically to the report and the study
graphic displayed. 3D View and Distance
Measurement are illustrated.

Some studies have built-in interactivity. For

example, on a corner of the 3D View click
and hold down the mouse button whilst
moving the mouse will rotate and tilt the
image. Similarly, the dotted lines on the
Distance Measurement can be dragged to
measure a specific distance or angle.

4: Right-click a study to reveal and change its

parameters.

5: Display features such as axis colours and

graduation, image tinting and slicing for
both Operators and Studies can be
changed using the mini-toolbars available
when an image is selected.

An additional set of tools can change

border colour and thickness and alter the
image sequence if they are overlapping.

Illustrations and Reserves, both available

from the Main Menu can be used to add
captions, images and and a range of
graphical features to enhance and
customise a report.

Leica Map has a dedicated help file accessed

through the help option on the Main Menu or
contextually by pressing the F1 key when the
program is running.

LAS User Manual 581

 Image Overlay
Image Overlay is an LAS module that enables
the acquisition of Channel images with a LAS
compatible microscope and the creation of a
composite image from a sequence of these
images. The Channels are typically Fluorescent
channels but can also be any other available
contrast method.

Sequences of channels may be defined and for

each channel the microscope and camera
settings set and saved. Once a sequence is
acquired a coloured overlay image may be
made using a variety of methods.

If using a colour camera the Image Type should

be set to greyscale.

See: Acquire: Camera: Image Formats: 239

Fully motorised microscopes will automatically

select filters, but users of manual machines
may still use Image Overlay simply by physically
turning the filter turret when prompted.

Between 2 and 8 images are captured during an

Overlay sequence. The subject and
magnification are the same for all, but individual
images use a different filter and exposure to
optimise contrast for a specific part of the
image.

Additional enhancement and ‘separation’ is

achieved with pseudo colour – a digital staining
technique – all controlled within Leica
Application Suite.

The final step is to bring all of the images

together in a single combined overlay in which
individual parts can still be easily identified to
illustrate their place in the ‘whole’.

582 © 2011 Leica Microsystems (Switzerland) Limited

 Image Overlay: Enable Image Overlay:

The Image Overlay module must be

installed and enabled; If it is not the icon
will not appear on the Acquisition menu
(2).

To start Image Overlay:

1: Click on the Select Acquisition icon
and from the menu…

2: Click to select Image Overlay. After it

is selected an additional tab marked
with the Lambda ( λ ) symbol (3)
appears in both the Browse and
Acquire Workflows.

LAS User Manual 583

Image Overlay: Setup Tools:

1: Select the Acquire Workflow by

clicking on its tab.

2: Click on the Image Overlay tab

marked with the Lambda (λ) symbol
to reveal the Channel Setup panel
comprising…

3: The Setup Tools and…

4: The Channel Dialog.

Using the Setup Tools:

5: Camera Live ‘freeze’ halts the camera
activity leaving the latest image on the
Viewer. Click again to resume taking
live images. Use the freeze button in
combination with…

6: Capture Single Image which will save

the current image to the capture
folder. The filename is created
automatically and a thumbnail of the
image is displayed in the Working
Gallery.

Continued... 585
See: Channel Dialog 587

584 © 2011 Leica Microsystems (Switzerland) Limited

 Image Overlay: Setup Tools: Continued:

7: The Next Filter button moves the

focus of interest to the next available
filter on the Channel dialog. The focus
‘rolls over’ from bottom to top
automatically.

8: Click the Move Up button to move the

selected channel up in the filter
sequence.

9: Click the Move Down button to move

the selected channel down in the filter
sequence.

10: To save the channel dialog

sequence, click the Create New
Sequence button.

See: Channel Dialog 587

LAS User Manual 585

Image Overlay: Channel Dialog:

The Dialog is a list of the filters available

on the microscope with details of:

1: The filter position on the turret.

Initially, this will also represent the
image capture sequence in automatic
mode but may be changed.

2: The filter name.

3: The contrast method. Right click on

the entry to reveal a menu with
options based upon the filter type:
Edit Description to change the
description:
Refresh to update the filter
information:
Reset Pseudo Colour reverts to the
filter's default value. This is only
available on monochrome cameras.
Extended Filter reveals an additional
menu to determine how the filter
works with the internal filter wheel.

4: Image exposure time. Initially, this

value will be the same for all filters but
may be changed individually to
achieve the best results.

5: The Gain value. Again, these will be

set to the same common value but
may be altered for each filter.

6: To the left of each filter entry is a

check box. When this is enabled
(ticked) the filter will be actively used
in the sequence and, with its
associated image becomes a
Channel.

Continued 586 ...

586 © 2011 Leica Microsystems (Switzerland) Limited

 Image Overlay: Channel Dialog: Continued:

Select a Channel by:

7: Clicking anywhere on a filter entry.

8: Clicking to enable the checkbox.

At least two and up to the total filters
available may be chosen for a
complete Channel sequence.

Changing sequence order:

9: Select an enabled Channel by clicking
the Next Filter button.

10: Click either the Move Up or Move

Down buttons to change the order in
which images will be captured.

Saving a Channel sequence:

When all of the required filters have been
selected:

11: Click on the Create new sequence

button to save the Channel sequence.

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Image Overlay: Adjust Channel Exposure:

Each Channel may have Exposure,

Gamma and Gain values set individually
to achieve the best image.

1: Check that the Camera is in live

mode. The message on the top bar of
the Viewer indicates the camera
state. If it reads Paused then click
the freeze button to return to live. The
button icon should have a small
square in the centre, not an arrow.

2: Click on the Next Filter button to

move to the Channel required.
Motorised microscopes will
automatically turn the filter turret to
the correct position, but non-
motorised machines will have to be
set manually. The filtered image will
appear in the Viewer.

3: If necessary, alter the Exposure time

by clicking and holding the Exposure
slider and drag it left, to decrease
exposure, or right to increase it.

Continued 589 ...

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 Image Overlay: Adjust Channel Exposure: Continued:

4: The Gamma value is normally set to

best suit the viewing medium – in this
case a colour monitor. Small changes
can have a dramatic effect on colour
density but in some cases can help to
improve contrast. Click and hold the
Gamma slider and drag left to reduce
the Gamma value or right to increase
it.

5: Gain will brighten or darken the image

without affecting the Exposure time.
Try to keep Gain to low values to
avoid introducing 'noise' into the
images. Click and hold the Gain slider
and drag it left to darken the image or
right to lighten it.

LAS User Manual 589

 Image Overlay: Snapshot and Working Gallery:
The Capture Single Image function provides a simple ‘
snapshot’ method of checking images from individual 2: Click on the Capture Single Image button.
Channels after making changes to the Exposure settings.
3: The 'snapped' image appears in the Gallery.
If necessary, click on the Channel to snap. Wait for the
Viewer to settle and then: 4: Remove snapshots from the Gallery by clicking on the
thumbnail to select it and then pressing the keyboard
1: Click on the Camera Live button to halt the Camera. Delete key.
Check the Viewer top bar: The message should read
Paused.

590 © 2011 Leica Microsystems (Switzerland) Limited

 Image Overlay: Auto Create Sequence Record:

Changes made to any of the Channel

settings are immediately reflected on the
live image in the Viewer and also appear
in the appropriate columns in the Channel
dialog.

Always check that the correct filters are in

the specified positions on the filter turret –
some microscopes can do this
automatically, but most require a manual
check.

Final settings are made on the Options

panel:

1: Click on the arrows to the right of the

Options bar to reveal the panel.

2: If the Always create new overlay

folder check box is enabled, every
time an Overlay sequence is acquired
a new folder within the current
capture archive will be created.
Situations, in which a substantial
throughput is expected, will benefit
from this time-saving option.

3: If Auto create overlay is selected, the

text in the Default Sequence Name is
used as part of the folder name.

Continued 592 ...

LAS User Manual 591

Image Overlay: Auto Create Sequence Record: Continued:

4: The folder name comprises the

Default Sequence Name and a
sequential number. Creation and
naming are fully automatic.

5: Save the configuration, including the

Channel sequence by clicking on the
Save button and…

6: Entering an appropriate name for the

configuration and…

7: Clicking the OK button.

8: The new name appears in the

Configuration window and the settings
may be used again for future
overlays.

See: Selecting an existing Configuration.

596

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 Image Overlay: Auto Create Overlay:
For speed and efficiency, select: Adjusting Channels before creating:
1: Auto create overlay by clicking the check box and… 1: Uncheck the Auto create overlay check box for a
preview before committing. Each of the images may
2: Click on the Always create new overlay folder check then be examined and adjusted for colour balance and
box. brightness before creating the overlay.

3: Click on the Acquire Overlay button.

These options will create a new folder within the capture

folder with the Default Sequence Name as part of its name.
The images for each of the filter channels (4) will be saved
inside the folder and they will be automatically combined to
produce the Overlay (5).

LAS User Manual 593

Image Overlay: Auto Overlay Without Auto Folder:

1: Click on the Auto create overlay

check box to enable it.

2: Disable Always create new overlay

folder by clicking the check box until
the tick symbol disappears.

3: Click on the Acquire Overlay button.

The Channel image location dialog
appears (4).

There are 3 options on the dialog:

Create a new sequence folder (5) will
create a new folder within the capture
folder and save the images to it – the
equivalent of checking the Always
create new overlay folder check box.

Empty the current sequence folder (6)

will delete all of the images inside the
current folder and replace them with
the new acquisition.

Add to the current sequence folder (7)

is a powerful option that allows any
number of channel images and
subsequent overlays to be stored in
the same folder. Each acquisition is
given its own sequential number to
differentiate between the sets.

Select the required option by clicking

the button next to it and clicking on
the OK button (8).

9: The entire acquisition sequence may

be stopped by clicking the Cancel

594 © 2011 Leica Microsystems (Switzerland) Limited

 Image Overlay: Auto Overlay With Delete Channels:
Enabling the Delete channels after overlay
creation option, removes the individual
Channel images after the overlay has been
created automatically. This is an excellent ‘
housekeeping’ arrangement for very high
throughput where there is no requirement to
add pseudo colour to the images.

1: Check the Auto create overlay check

box and…

2: ...the Delete channels after Overlay

creation check box.

3: Click on the Acquire Overlay button.

The Channel images are deleted after
the overlay is created and saved in the
current Capture Folder – not in a newly
created folder or the last newly created
folder.

LAS User Manual 595

Image Overlay: Recall Configurations and Manual Acquire:

Previously saved configurations stored in

the current capture folder can be re-called
by clicking on the arrows to the right of
the Configuration window (1) and then
choosing from the list (2). Changes may
then be made to the exposures and to the
Options with the altered settings saved as
a new configuration (3).

Manual Acquire:
Primarily designed for non-motorised
microscopes, the Manual Acquire facility
prompts for the filter turret to be moved
manually to the correct position before the
image is captured.

4: Click to enable the Manual acquire

option.

5: Any or all of the other options may be

selected in Manual Acquire mode.

6: Click on the Acquire Overlay button.

7: When the Manual Sequence Prompt

appears, turn the turret to the filter
position specified on the prompt
and…

8: Click Continue. The image will be

acquired and appear as a thumbnail
in the working Gallery. The prompting
process repeats for all of the selected
filters.

9: To end a capture sequence

prematurely, click on the Finish
button.

596 © 2011 Leica Microsystems (Switzerland) Limited

 Image Overlay: Creating and Viewing Overlays:

When the Channel images have been

captured…

1: The Browse Workflow opens with…

2: ...the Image Overlay or a preview

selected. Thumbnails of the captured
channel images together with the
composite overlay if auto-create were
selected, will be displayed in the
Gallery. The Viewer shows the last
image captured or the overlay.

There are three panels on the Image

Overlay tab:

3: The General panel on which the

Overlay Method determines the
manner in which the channel images
are combined to create the overlay.

4: The Image Groups and Overlay

Method settings can be saved as the
ongoing default.

Continued 598 ...

LAS User Manual 597

Image Overlay: Creating and Viewing Overlays: Continued:

The Adjust Appearance panel provides

the controls for:

6: Adding false (pseudo) colour to

monochrome images.

7: Adjusting colour balance with

Histogram controls.

8: Changing the Gamma value of the

channel images and the overlay, and

9: ...varying the ‘mix’ strength or

dominance of a channel image.
Images captured in colour will not
have all of the colour controls
available.

The Overlay Image panel (10) determines

which of the channel images are to be
included in the overlay. At least 2
channels must be selected before an
overlay can be created.

11: The Create Overlay button creates a

new Overlay based upon the selected
settings. Once the channel images
have been captured they may be
altered at will and almost any number
of different overlays created.

598 © 2011 Leica Microsystems (Switzerland) Limited

 Image Overlay: Colour Control:
Monochrome Channel images can have false or
pseudo colour applied to them before they are
combined into a single overlay to improve
contrast and clarity of the final image.

1: The Monochrome Channel image with the

Adjust Appearance panel as it looks when
capture is complete. Had this been a colour
image the Apply False Colour check box
would not be available.
Select another image by clicking on its
thumbnail in the Gallery. It will be displayed
in the Viewer.

2: To apply colour, click on the Apply False

Colour check box to enable it. A basic
colour may be applied to the image based
upon the type of filter used in its capture.

3: Move the Channel Colour slider to the

desired colour.

4: The applied colour is immediately displayed

on the Viewer.

Continued 600 ...

LAS User Manual 599

Image Overlay: Colour Control: Continued:

5: Refine the image by adjusting the

sliders below the Histogram and also...

6: ...the Gamma value.

7: The Mixing Strength slider controls the

predominance the Channel image will
have in the overall result. The range is
from 0 to 200%. The higher the value,
then the greater the ‘presence’ of the
image in the overlay.

8: Test the result by clicking the Preview

check box.

9: All of the channels are combined in a

temporary overlay.

10: If the result is acceptable click on the

Apply button to create an overlay. A
new thumbnail will appear in the
Gallery. In this way any number of
overlays can be created to illustrate
various aspects of the image.

11: To reset the channel images to their

original filter colour values, click on a
thumbnail and then on the Restore To
Default button.

600 © 2011 Leica Microsystems (Switzerland) Limited

 Image Overlay: Selecting Image Group and Overlay Method:
1: To select the method of combining
the Channel images to create the
overlay, click on the arrows to the
right of the Overlay Method text box
and from the drop down list…

2: ...click to select the required method.

In the illustrations, images (3) are the
original, common channel images that
have had false (pseudo) colour
applied.

4: Represents the overlay combining the

three common channel images using
the Maximum option. The highest
pixel value from the same location in
the three common images is used to
create the overlay.

5: The same process using the Addition

option. The pixel value from the same
location in all three channel images
are added together.

6: Shows the result with the Average

option. The pixel values in the same
location in all three channel images
are added together and then
averaged.

7: To save the settings as default, click

the Set as Default button.

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Image Overlay: Fine Tuning:

False colour is applied to a channel image

electronically by association only – the
original remains intact and unchanged.
This means that colour may be ‘removed’
completely or ‘fine tuned’ at any time after
capture and a new overlay created.

Individual colours in overlays cannot be

altered, but the Histogram (1) and the
Gamma (2) controls are available to
lighten or intensify overall effect and in the
process enhance a particular feature.

It is not necessary, nor always desirable to

include all of the channel images in an
overlay. The Overlay Image panel
provides the means to include or exclude
individual channels.

3: A channel is included when the check

box to its left is checked. Click to un-
check and exclude.
At least two channels must be
selected in order to create an overlay.
If only two are still selected the
program will prevent any more
exclusions.
The colour bar to the right of a
channel name indicates the filter
colour used in its capture.

4: Click the Show Overlay button to

select and display the last overlay
created or…

5: Click on the Create Overlay button to

produce a new overlay using the
revised channel images.

602 © 2011 Leica Microsystems (Switzerland) Limited

 MultiStep

This section contains the following topics:

Introduction.
Acquire MultiStep images
Browse MultiStep images

Introduction
This module allows for the creation of
Scan Patterns using Motorised X/Y Stage
control through the Leica Application Suite
interface. The subsequent images may
then be joined together to create one large
image or mosaic.

1: On installation of the MultiStep

module and the subsequent start up of
LAS, the MultiStep module can be
found by selecting the Select
Acquisition Mode button to the left of
the Workflow bar.

2: On selection of the MultiStep mode

icon a new tab will appear in both the
Acquire and Browse Workflow items
labelled S.

LAS User Manual 603

MultiStep: Acquire Images:

On the Acquire Workflow panel an S tab

will be visible if the MultiStep option is
active. Selection of this tab reveals four
control panels:

1: Scan Position

2: Stage Position

3: Define MultiStep Sequence

4: Options

604 © 2011 Leica Microsystems (Switzerland) Limited

 MultiStep: Scan Position Control:

This shows the position of the current field

of view. The user can mark points on the
specimen by driving the stage manually.
The live window will be updated to reflect
the true position on the stage.

1: The Navigation or Soft Joystick can

also be used to navigate to new
positions.

2: Select Scan Method from the drop-

down menu. A position is located
using the microscope (e.g. the corner
of an area of interest) to mark the
initial field in the scan pattern.

3: The Create button is then pressed to

mark this co-ordinate on the graphic.
(See right: Cross hair in red).

4: The Create button then changes to

Expand. When a second coordinate is
added (again by driving the stage to
the desired position and then selecting
the Expand button), a graphic
representation of the scan area is
shown (5).

Continued... 606

LAS User Manual 605

MultiStep: Scan Position Controls: Continued:

6: The Clear button allows the user to

remove the scan pattern and start the
process again.

7: The Shrink button allows the user to

reduce the size of the scan area. This
is achieved by placing the red cross
hair at the position the Scan Pattern is
to shrink to.

8 and 9: The Zoom In and Zoom Out

buttons allow the user to enlarge the
Scan Position display.

10: This display can then be navigated

using the scroll bars that will appear,
or by holding the mouse button down
over the scan position graphic and
panning the position by moving the
mouse. Releasing the mouse stops
the panning.

11: Double clicking on the stage graphic

causes the stage to move to that
position.

606 © 2011 Leica Microsystems (Switzerland) Limited

 MultiStep: Define Sequence:

This is a data/textual representation of the

scan pattern which allows fine tuning of
the scan pattern.

1: Scan Origin is the location of the first

field of view in the pattern and is the
central point of that field.

2: Go To drives the stage to the Scan

Origin.

3: Set takes the current stage position

and sets it as the Scan Origin and the
scan pattern is shifted appropriately.
Scan Definition:

4: Step Size is the distance between one

field and the next, both horizontally
and vertically.

5: If Auto is selected the component

images will be adjoining. De-selecting
Auto allows the user to acquire images
with overlaps or images with spaces
between them.
If the Auto checkbox is selected this
value will equal the dimensions of one
field of view. If the Auto checkbox is
unselected the step size is user
defined.

6: The Pattern Size gives the x and y

dimensions of the rectangular pattern.

7: The Fields boxes show the number of

whole fields of views in the pattern.
Changing the Step Size will change
the Pattern Size as will changing the
number of fields of view.

Continued... 608

LAS User Manual 607

MultiStep: Define Sequence: Autofocus:

9: This option allows the user to turn on

Predictive Focus.

10: Pressing the associated button that

appears when Predictive Focus is
selected displays the Predictive Focus
Setup dialog (11).

Continued... 609

608 © 2011 Leica Microsystems (Switzerland) Limited

 MultiStep: Define Sequence: Continued:

12: Selecting Add creates a new

predictive focus point based on the
current stage position.

13: A prompt panel reminds the user

that the Predictive Focus Setup focus
dialog requires a minimum of three
predictive focus points.

14: The Shutter allows the user to define

whether the shutter is closed during
stage moves and only opened prior to
an acquisition by selecting Close
when not acquiring (15). This is
particularly useful if not essential when
undertaking fluorescence specimens.

LAS User Manual 609

MultiStep: Options: Configuration and Show During Acquisition:

The Configuration section allows the user

to store and recall previously saved
configurations.

1: To save a new configuration based on

the current settings, type a name into
the configuration drop-down and then
select Save (2). This entry defines the
mosaic name and the sub-folder
name if sub-images are saved .

3: To remove a previously saved

configuration, select it from the drop-
down list and then press the Delete
button.

4: The Create Mosaic image if checked

means that a mosaic image will be
created after the last image in the
Scan Pattern has been acquired. This
will be displayed and stored in the
Gallery.

5: If Save sub-images is selected, all

acquired images will be saved as an
image set. If de-selected these
images will not be saved.

Show During Acquisition

6: Mosaic shows the dynamic build up of
the mosaic on the screen as each
image is acquired

7: Single Images shows each acquired

image on the main screen as it is
captured in turn.

Continued... 611

610 © 2011 Leica Microsystems (Switzerland) Limited

 MultiStep: Options: Reduction Factor:

Auto provides a final image at a size

comparable with an individual field image
size.

None provides an image at max size --

size of final image is sum of sizes of
individual images.

8: Values (e.g. x2) indicate an image of

size Max Size / (value* value). Highest
value is capped to give an image
of approx 640 x 480. It displays the
image size and amount of MB storage
that will be used.

NB: Memory requirements can only

be shown accurately if the image is to
be saved as in BMP or TIF formats.
JPGs will show the maximum size for
the image. Disk space requirements
are additional because field images
need disk space too. Even if system
is set up to acquire 16-bit images,
mosaic images will be acquired in 8
bits

9: Selecting the Acquire MultiStep button

will start the Scan Pattern acquisition
process as defined above.

10: A Progress bar will show how the

process is proceeding:

11: Pause: Will halt the acquisition which

can then be restarted.

12: Stop: Will halt the acquisition where

it is resulting in either a partial mosaic
being created and/or a partial set of
images depending upon your settings.

LAS User Manual 611

MultiStep: Browse MultiStep Images: Select Image Set:

Under the MultiStep tab S (1) there are

two control panels:

View Image Set.

MultiStep

View Image Set

This allows the user to select a previously
saved set of MultiStep acquisition
images.

2: The images may then be stepped

through (like a slide show), by using
the video play controls which allow the
user to go to the beginning or end of
the sequence or step through images
one at a time forward or back or play
the sequence at the specified frame
rate.

3: The rate at which images can be

displayed is altered by entering a
value into the Delay box.

4: The red slider in the sequence

visualization can also be grabbed
using the mouse (like using a scroll
bar) and the sequence navigated by
moving this back or forth through the
image sequence.

5: The image being displayed is listed in

the text box.

Continued.. 613 .

612 © 2011 Leica Microsystems (Switzerland) Limited

 MultiStep: Browse MultiStep Images: Browse Controls:

After selecting the sequence in Browse,

the MultiStep panel allows the user to view
the images and create a Mosaic image.

1: MultiStep Image: Selecting this

automatically loads the MultiStep
image from the currently selected
image set in the gallery. The View
Image Set controls will be disabled as
the mosaic image is being displayed.

2: Image Set: Selecting this option loads

the first image from the image set

3: The Create MultiStep Image button

allows the user to re-create the
MultiStep Image from the current
image set.

Continued... 614

LAS User Manual 613

MultiStep: Browse MultiStep Images: Reduction Factor::

1: Auto provides a final image at a size

comparable with an individual field
image size.

None provides an image at max size.

Size of final image is sum of sizes of
individual images

2: Values (e.g. x2) indicate an image of

size Max Size / (value* value). Highest
value is capped to give an image of
approx 640 x 480. It displays the
image size and amount of MB storage
that will be used.

Note: Memory requirements can only be

shown accurately if the image is to be
saved as in BMP or TIF formats. JPGs will
show the maximum size for the image.
Disk space requirements are additional
because field images need disk space
too. Even if system is set up to acquire 16-
bit images, mosaic images will be
acquired in 8 bits.

614 © 2011 Leica Microsystems (Switzerland) Limited

 Power Mosaic and Power Mosaic Plus

LAS Power Mosaic software integrates high-performance Once you have created a scanned mosaic, you can save
specimen scanning into the Leica Application Suite to your specimen’s details as a workspace, or export the
provide an easy-to-use application for creating, viewing, full-resolution mosaic for use with image publishing and
and saving ultra-high resolution mosaic images. LAS analysis packages.
Power Mosaic is using in conjunction with a Leica
microscope, Leica DFC digital camera and a stepping The software is designed to offer a comprehensive range
stage. of facilities with a strong emphasis on versatility and ease
of use. With minimal experience it is possible to produce
With LAS Power Mosaic you can scan a selected area or excellent mosaic images. In particular, the accurate
an entire slide quickly and accurately, and then calibration of the camera and motorised stage is
effortlessly relocate to areas of interest with a simple essential and is made convenient by being performed
click. Additionally, with the LAS Power Mosaic Plus automatically.
software, it is possible to acquire images at multiple
focus positions and view the complete mosaic while Good results depend, however, on the system being well
scrolling through focus. configured and set-up and on the user becoming
reasonably familiar with the available facilities and
The ability to generate high-resolution mosaic images functionality.
provides a powerful and novel method of visualising a
specimen. The specimen overview is an aid to The principal purpose of this manual is to provide
understanding the relationships between microscopic practical guidance on configuration, set-up and use of
features and overall structure. LAS Power Mosaic.

LAS User Manual 615

Power Mosaic: Features:

Navigate specimens from LAS Scan Patterns

Power Mosaic integrates seamlessly within LAS. Rectangle, Circular, Annular, Cross (+ and x),
Random
Power Mosaic images can be acquired and
reviewed. Overlap of tiles allows smooth merging of joins.

Images from LAS Power Mosaic can be saved and Correction for camera rotation is performed
measured in LAS.
Save / load scan patterns
Specimen movement is performed by a stepper
stage.
Continued 617 ...
A Leica DFC camera acquires images.

Power Mosaic Acquisition

Uses triggered image capture for very fast

continuous scan and acquire on suitable cameras

Standard scan using step and acquire for low light

applications.

Image streaming for mosaic sizes limited only by

free disk space.

Additional scans can be added to the initial scan to

include all parts of a specimen.

616 © 2011 Leica Microsystems (Switzerland) Limited

 Power Mosaic: Features: Continued:

Export/Import
Point and click relocation of stage to indicated
Save and load mosaic scan pattern and Power position.
Mosaic environment as a ‘workspace’.
Real-time graphic display of current stage position
Select workspace for recall from LAS. on specimen map.
Load last used workspace on start up. Zoom and pan over entire mosaic.
Images exported to tif, bmp or jpg.
Continued... 618
Export mosaic as full resolution bitmap.

Export mosaic as reduced resolution bitmap.

Export user-selected region of interest.

Export image to current LAS capture folder.

Specimen Map
Switch between display of stage overview and
pattern view.

Drag to create pattern scan or enter exact details.

LAS User Manual 617

Power Mosaic: Features: Continued:

Calibration Automatic Focus

Automatically measures calibration and updates Multi-point predictive focus for continuous focus
scan patterns according to selected magnification tracking.
on microscope using a correlation-based
calibration procedure. Predictive focus setup by combined auto scan with
user review.
Camera rotation and sage skew is measured to
allow convenient adjustment. Autofocus can be used for specimens that are not
flat.
Small rotation of camera is compensated for in the
mosaic creation. Predictive and Autofocus can be combined.

Automatic stage orthogonality correction. LAS Power Mosaic Plus

This is an extended version of Power Mosaic and
Microscope Automation
includes all the features of Power Mosaic and in
Oasis XY stage and Z focus control drive board is addition:
used.
Z-Scan optional at each field.
Software joystick for stage and focus movement.
Definition of Z-Scan positions, step number and
Compatible with Leica Microsystems LAS step width.
configured microscopes controlling focus, turret,
condenser, and lamp control as available Extended focus imager can be created.

Stage and focus speed defined per objective Mosaic image can be reviewed while sweeping
linked to LAS. through all focus positions.

Navigator for multiple scan patterns

618 © 2011 Leica Microsystems (Switzerland) Limited

 Power Mosaic: Operating Steps:

The following procedure lists the steps required to 3: Initialise the Stage and Focussing:
obtain high-quality mosaic images. Once the system is This checks the stage travel limits, finishing at the
correctly aligned and adjusted, some of the steps will centre point. The small green 'target' in the stage
not be necessary again. area indicates current position. The 'hatched'
It is assumed that the system is already installed border around the periphery indicates the 'soft'
according to the installation procedure detailed in the limit switch area.
release notes.
Lower the condenser and check that stage will not
1: Prepare the specimen: collide with objectives.
Check the following:
Check/adjust the stage speed: See: Stage
The specimen is as flat as possible to minimise Initialisation. 634
the need for focus changes.
Initialise/adjust the focus limits: See Focus
The slide surface is clean. Initialisation. 635

The specimen is firmly fixed to the stage - loose See Autofocus Setup: 636
slides are a common problem.
Continued.. 620
2: Setup the microscope:
To gain experience with Power Mosaic, start with
an x5 objective as this magnification probably
does not require focus compensation during the
scan. This makes it easier to check that the
scanning is working correctly.

Ensure the condenser is in focus and apertures

are set for Koehler illumination. With an automatic
microscope this should be checked for all of the
objectives expected to be used.

LAS User Manual 619

Power Mosaic: Operating Steps: Continued:

4: Check the camera and adjust the exposure: Set Exposure to approximately 100 µs.
On a clear field of view, debris in the optical path will
be seen as fixed dark regions on the live image On the Histogram display, click to enable ‘Show
when the specimen is moved. If cleaning is Under/Over Exposure’.
necessary, a qualified technician should perform it.
Adjust the lamp voltage until red flecks appear on
Check that there is no visible debris in the optical the image white highlights. Note: On a DM
path. microscope, fine lamp voltage control is achieved
by pressing both stand lamp buttons together.
For a colour camera Saturation = 1.75: Gamma = Expect a high lamp voltage and very bright image.
0.6: Gain = 1: Make fine tuning adjustments
around these values to achieve the required If there is insufficient light, increase the camera
image. gain by small amounts to give the correct
exposure. Check that this does not increase
For a mono camera: Saturation = 1.75: Gamma = significantly image noise.
1.0: Gain = 1.
WARNING: Switch 100% light to the camera. DO
Ensure that the black and white levels on the NOT look at the specimen through the eyepieces.
histogram are reset.
Adjusting the camera for Standard scan
See: Input Options: 631 Standard scan is used when the conditions for
Turboscan are not met.
Turboscan and Standard scan require different
adjustments: Select the camera image format to suit the
specimen detail required. Choose the lowest
Adjusting the camera for Turboscan: resolution without compromising image quality to
Turboscan, the fastest scanning method, can be save disk space.
used if:
The camera has progressive scan mode. Set the lamp voltage to give a comfortable image
in the eyepiece.
There is sufficient light to give an exposure of less
than 200 µs and Adjust lamp voltage and camera exposure to until
red flecks appear on a white region of the image.
The specimen is flat enough to use no focus
change or predictive focus. Continued 621 ...

Select an exposure mode with Progressive scan.

620 © 2011 Leica Microsystems (Switzerland) Limited

 Power Mosaic: Operating Steps: Continued:

5: Set shading 7: Create and perform a test scan

Shading correction has to be set in Power Mosaic. Select the Split Screen view. The 'crosshair' shows
LAS settings are not used. the current stage position.

Shading correction must be repeated every time Move the stage so that the specimen can be seen
the objective is changed. in the live image window.

Move the specimen so a clear region without Select the New Pattern tool, click close to the
artefacts is visible over the whole image. stage 'crosshair' and drag a small scan region.
Click Acquire Power Mosaic.
Set the shading correction: See: Shading: 640
Use Zoom and Pan to check that the mosaic is
6: Calibration for each objective formed correctly. See: Create Pattern Grid: 657
LAS Power Mosaic derives its calibration from the
stage movement to accurately align the tile edges. 8: Extend the scan pattern to the required region
Calibration must be carried out for every objective To include parts of the specimen not included in the
on first use. test scan:
A warning message appears if an objective has not
been calibrated. Click on the Create/Expand tool.
Calibration tests that the values returned are
reasonable: If they are not a warning is displayed. Click on the stage in an area outside the boundary
Camera rotation must be less than 0.1°. Conditions of the test scan and drag to include the required
can alter over time and systems are susceptible to parts of the specimen. See: Create Pattern Grid:
dirt, heat and vibration. Periodically check the 657
calibration to ensure that it remains accurate.
Continued 622 ...
Check microscopes with a manual turret, that the
selected objective matches that selected on the
Acquire:Mic1 tab. The same applies to the Mag
Changer if it is fitted.

Set calibration: See: Calibration: 638

If necessary adjust the camera rotation: See:

Camera rotation: 639

LAS User Manual 621

Power Mosaic: Operating Steps: Continued:

9: Select the Focus Method: Using Predictive focus:

During scanning, two focussing methods are Select the focus points manually or use the
available: automatic Grid.

Predictive Focus is best suited to uniform Select and focus each point manually or choose
specimens and low magnification - up to x10. 'Autofocus on all points'
Focussing is performed either manually or
automatically on a number of points across the Run Predictive focus: See: Predictive Focus: 652
specimen to create a table of values. Points which
have not been pre-focussed use the table to Using Autofocus:
predict a focus position without going through the Set the repeat pattern for fields to be focussed.
time-consuming process of an Autofocus on every
field. Enable/disable field skipping on focus failure. See:
Focus Methods: 651
Autofocus should be used for irregular specimens
at any magnification. Focussing is carried out at
regular intervals across the specimen with options
to set the repetitiveness.

Both Predictive and Autofocus may be used in

combination to benefit from the speed of
Predictive and the precision of Autofocus.

622 © 2011 Leica Microsystems (Switzerland) Limited

 Power Mosaic: Starting

Power Mosaic: Loading the module:

1: Click on the Select Acquisition button
to reveal the available modules. If a
module is already loaded and running
the button icon will differ from the
illustration.

2: The Power Mosaic icon will be

present only if it is installed. However,
it will also need to be enabled:
See: Installing, Configuration and
Licensing: Registration Information.
Click on the icon.

3: Click on the Acquire Workflow tab.

4: When Power Mosaic is loaded and

running the PM tab will be present
with the main panels displayed.

5: An additional panel – the On-screen

Joystick – is available by clicking the
Show Joystick Tools button.
Click and hold on the Joystick header
to drag and dock it on any part of the
screen.

LAS User Manual 623

 Power Mosaic: User Interface Map:

The User Interface is shown in the illustration with the 4: Scan viewing area with on-screen Joystick shown.
Acquire Workflow selected and the Power Mosaic tab
active. 5: Scan Grid Pattern.

1: Workflow tabs. 6: Live Image Field: Split screen view is selected.

2: Control Panels and function tabs. 7: Tool Bar. The tools are explained on the following
page.
3: Acquire scan button.

Continued... 625

624 © 2011 Leica Microsystems (Switzerland) Limited

 Power Mosaic: User Interface Tools:

The Toolbar is located on the right-hand

side of the screen. The tools descriptions
are:

1: Live Screen: The entire screen is

devoted to the live image.

2: Split Screen: Divided between viewer

part of live screen.

3: Grid Pattern View: Entire screen is

devoted to the scan grid.

4: No Tool selected.

5: Stage Area: displayed on Viewer.

6: Display Grid Pattern.

7: Hide/Display Grid Pattern.

8: Zoom In/Zoom Out.

9: Pan.

10: Create a New Scan Grid.

11: Extend/Contract existing scan grid.

12: Go to Stage point.

13: Move Scan Pattern.

14: Clear scanned Tiles.

Continued... 626

LAS User Manual 625

Power Mosaic: User Interface Views:

There are three viewing options available,

each selected by clicking the appropriate
button.

1: View Stage Map: fills the viewing area

with…

2: …the scan pattern grid and scanned

tiles scaled to fit.

3: Split View: displays the pattern grid,

and scanned tiles on the left (5) and a
live image on the right (4).

6: Live Image: fills the viewer with a live

image.

626 © 2011 Leica Microsystems (Switzerland) Limited

 Power Mosaic: On-screen Joystick:

With the On-Screen Joystick visible, it can

be customised to suit individual
preferences.

1: Right click on the Joystick to reveal

the Speed and Properties menu.

2: Click to select the speed at which the

stage will be driven during when the
joystick is moved.
Three options are available: Normal,
Fast or Slow.
Actual speed will depend upon the
stage type so ‘select and test’ to find
the most appropriate.

3: Click on Properties to reveal the

Properties Dialog. Three tabs provide:

Speed: Similar to (2) above but on this

tab X and Y speeds may be selected
individually and the travel direction
can be inverted.

Z-Axis: Allows the Mouse Wheel (if

fitted) to act as a focussing control
and sets the focus step size. To
change it, click on the Step size
window, press the Delete key to clear
the existing value and type a new
value in µms.

Limit Alarm: There is a small indicator at

each of the joystick quadrants and
also top and bottom of the focus
slider. Normally they are green when
the stage and the focus are within
limits. If the Show Limit Alarm LEDs
box is checked, the indicators will turn
red as the travel limits are
approached.

Nudge size: Each joystick quadrant also

has a small arrow displayed. Clicking
an arrow ‘nudges’ the stage in that
direction. It is a very useful facility for
precise positioning. Set the nudge
value in µms by clicking the window,
deleting the existing value and typing
a new one.

Continued... 628

LAS User Manual 627

Power Mosaic: On-screen Joystick: Continued:

Color: The area around the joystick

defaults to pale grey (silver) but may
be changed by clicking to select a
required color. Various color sets are
available.

Speeds (Calibration): The three speed

options may be calibrated individually
by clicking and dragging the
appropriate slider. Again, actual
speeds will vary with the stage type:
Change and test to achieve the best
settings

628 © 2011 Leica Microsystems (Switzerland) Limited

 Power Mosaic: Imaging Setup:

If Power Mosaic Turboscan is going to be

used, the exposure times must be very
short – typically less than 200µs
(microseconds). This requires high light
levels making some contrast methods
unsuitable. If the required exposure times
cannot be achieved, choose Standard
scan instead for which light levels and
exposure times are not critical.

1: Click on the appropriate Mic(roscope)

tab and…

2: …select the required Objective,

Magnification and…

3: …Contrast method.

4: Set the Aperture and...

5: …Intensity to suit specimen and scan

type.

Often, a good image can be achieved

quickly by setting basic exposure
parameters, using Auto Exposure and
then fine-tuning the result with white
balance. This is especially suitable for
Standard scan - Turboscan will probably
require closer attention to exposure:

6: Click on the Camera tab to reveal the

exposure controls.

7: Set the Saturation to 1.75 by clicking

on the slider and dragging it – to the
left to reduce the value and to the
right to increase it.

8: Set the Gamma to 0.6 (1.0 for

greyscale).

9: Gain should be as low as possible –

start with a value of 1.

See: Acquire: Camera for more detailed

information.

Continued... 630

LAS User Manual 629

Power Mosaic: Imaging Setup: Continued:

The image must be properly focussed.

On the Histogram:
1: Set the Black level to 0 and…

2: …the White level to 255 by clicking

and dragging the sliders.

3: Enable Under/Over Exposure by

clicking the check box.

Run Auto Exposure:

4: Click on the Auto Exposure icon and
then click again to turn Auto Exposure
off.

Manual Exposure setting for

Turboscan
5: Adjust the Exposure slider to give a
reading of about 100µs. Gradually
increase the light intensity (see
previous page) until intermittent red
flecking indicating slight over-
exposure, appears.

Set the white balance:

6: Click and drag a Region of Interest
around a white area.

7: Select White Balance from the menu.

Fine tuning:
If necessary, fine tune the image with the
Saturation and Gain controls.

630 © 2011 Leica Microsystems (Switzerland) Limited

 Power Mosaic: Input Options:

Selecting the Input Options:

1: Click on the Camera tab.

2: If the Input Options panel is

concealed click on the arrow to the
right of the header to reveal it.

The panel provides options for:

3: Using a pre-saved Input Options
Configuration or creating a new one.

4: Selecting the Image Type – colour or

greyscale.

5: Setting the Colour Depth.

6: Determining the Captured Format –

how the image will be saved and
stored on disc and…

7: Selecting the Live Format – how the

images will be displayed on the
Viewer.

Selecting the Camera:

If Turboscan is going to be used, only
designated high speed, progressive scan
cameras with a trigger facility can be
selected.

Choosing a Configuration:
If an Input Options configuration has been
previously saved, it can be recalled from
the Configuration menu.

8: Click on the arrows to the right of the

Configuration header.

9: From the drop down menu, click to

select the saved configuration. Each
will have a unique name. All of the
saved settings will be loaded and the
remainder of the Input Options may
be skipped.
A newly created configuration can
also be saved using the Configuration
menu and selecting the ‘Save Current
’ option.

See: Acquire:Camera:Input Options for

detailed procedures.

Continued... 632

LAS User Manual 631

Power Mosaic: Input Options: Continued:

Image Type:
Colour cameras may be used in either
colour or monochrome (Greyscale) mode.

1: Click on the arrows to the right of the

Image Type header and from the drop
down menu…

2: …select either Colour or Greyscale.

If Turboscan is being used with a
Progressive Capture Format, the
Image Type will automatically revert
to Greyscale.

Captured Colour Depth:

3: Click on the arrows to the right of the
Captured Colour Depth header and
select 8 bit. Avoid the 16 bit option if it
is available. It will greatly increase
stored image size, slow the scan and
may not be compatible with other
image processing software.

Continued... 633

632 © 2011 Leica Microsystems (Switzerland) Limited

 Power Mosaic: Input Options: Continued:

Captured Format:
Power Mosaic has been designed to
capture high quality images so avoid using
the low resolution options. For Turboscan,
select one of the Progressive options (but
not VGA):

4: Click on the arrows to the right of the

Captured Format header and from the
drop down menu…

5: …choose the highest possible

resolution. The ‘binning’ options will
save disk space but are unlikely to
improve scan speed. Use the
navigation arrows to reveal other
options.

Live Format:
Live format is not used during Power
Mosaic, but for the sake of completeness
keep the Live Format the same as the
Captured Format.

6: Click on the arrows to the right of the

Live Format header and from the drop
down menu…

7: Select the same format as the

Captured Format.

See: Acquire: Camera for more detailed

information: 241

LAS User Manual 633

Power Mosaic: Stage Initialisation:

The Stage must be initialised the first time

an objective is selected and again if the
stage stalls for any reason.

Initialising determines the limits of travel in

both the X and Y directions and ‘matches’
the stage speed to the objective.

Warning: Ensure that there are no

obstructions before initialising – turn to an
empty turret position and lower the sub-
stage condenser.

To compensate for different stage types,

ages, slackness and stiffness in the
mechanisms as well as various imaging
setups, the stage speed can be de-rated
from its maximum. Lowering stage speed
also helps prevent ‘stalling’ which results
in a loss of initialisation values.

1: Click on the Initialise and Set Speed

button.

2: On the Automation Settings dialog,

set the Drive Speed slider to a high
value - start at 90%.

3: Click the Initialise Stage button.

4: Click Yes on the Confirm message.

If the stage stalls, reduce the speed
and re-initialise.

5: After initialisation completes

successfully, reduce the speed by
10% and click on the Apply button.

634 © 2011 Leica Microsystems (Switzerland) Limited

 Power Mosaic: Focus Initialisation:

The speed of the focus drive mechanism

can be adjusted below its maximum to
allow for possible overrun and minimise
shaking.

Additionally, the limits between which the

focus mechanism can operate are set to
prevent the specimen colliding with the
objective (Upper) and allow sufficient
clearance for the specimen to be
accessed (Lower).

Select Live Image and focus.

1: Click on the Initialise and Set Speed
button.

2: On the Automation Settings dialog,

adjust the Focus Speed by moving
the slider. Determining the actual
value will depend upon the
microscope and the imaging setup
and may require some trials.

3: Click on the Initialise Focus button.

4: Set the Upper Focus Limit (relative to

the current position) by clicking on the
text box and entering a new value.
For example, to limit the focus
position to 100um above the current
position, type in ‘100’.

5: Set the Lower Focus Limit in the

same way. Positive numbers will be
automatically converted to a negative
value so no need to type the leading
sign.

6: Click OK.

7: Click Apply to apply the settings and

Close to save them.

8: If Apply is not clicked, the Settings

Not Changed message appears.
Click No to apply the new settings

Go to Autofocus Setup: 636

LAS User Manual 635

Power Mosaic: Auto Focus Setup:

Auto Focus initialisation sets the travel

range of the focussing mechanism and
also the threshold value for focussing ‘
success’.

Select the split screen display option.

1: Click on the Auto Focus tab.

2: The current stage position on the Z

Axis is shown by a vertical green line.

3: A blue horizontal line represents the

Focus Curve.

4: Using the on-screen Joystick,

SmartMove or stage controls,
navigate in the live image pane to a
part of the specimen that contains
some detail, avoiding repetitive or
symmetrical patterns. It may be
helpful in ‘proving’ the Auto Focus
effectiveness to de-focus the
specimen.

5: The Range value (in microns)

determines the travel limits of the
focussing mechanism. It reflects the
thickness of the specimen. Click in
the Range text box and enter a value
– the mechanism will drive upward
from the current stage Z position by
50% of the value, and downward by
an identical distance. Between these
limits, Auto Focus will achieve the
best focus possible.

6: Find a field on the image that can be

sharply focussed and click the
Calibrate button. This will measure
any drive backlash and store the
value to be used with the current
objective.

7: Click the Apply button. ALWAYS click

Apply when changing values.

8: Click on the Auto Focus button.

Continued... 637

636 © 2011 Leica Microsystems (Switzerland) Limited

 Power Mosaic: Auto Focus Setup: Continued:

1: If there is sufficient detail and contrast

in the specimen area chosen, the
curve will look similar to this centred
about the new stage Z Axis position,
and…

2: …the image should be sharp. The

Score bar will extend across to the
right.

3: The Threshold value represents a

difference (%) of the lightest to
darkest pixels in the image. For Auto
Focus to ‘succeed’ the calculated
value must be at or above the
Threshold. Below the Threshold and
there probably was not sufficient
detail to guarantee accuracy.

4: Set the Threshold by clicking in the

text box, typing a new value and
clicking the Set Threshold button.

5: Click Apply. The Threshold line will be

drawn to reflect the new value.
After changing Threshold repeat Auto
Focus to ensure that the value is not
so high it prevents focussing.

6: Click Close.

Note: In the unlikely event that the Z focus

shows at the peak but the Score indicates
something less than peak, gearing
backlash could have caused the
discrepancy. In this situation change the Z
scan direction to 'Up' (bottom to top of
specimen). Please refer to the Oasis
manual.

LAS User Manual 637

Power Mosaic: Calibration:

Input options and lighting levels must be

set and the specimen in place.

The Calibration process is automatic: It

will determine the ‘pixel/micron’ value for
the objective being used. As objective
magnification increases so the actual field
of view on the specimen decreases.
Calibration correlates the field dimensions
(microns) to the number of camera
elements (pixels) required to capture it. It
must be carried out at the start of a
session, and whenever the optics are
changed.

Calibration also checks for camera

rotation – the angle of rotation from the
Stage X and Y axes.

1: Select the Live Screen view.

2: Click on the Set Calibration button.

When the Calibration Dialog
appears…

3: …a small rectangle is displayed on

the live image. Use the on-screen
joystick or stage controls to navigate
to a well defined and detailed region
of the image lying within the
rectangle. Avoid uniform patterns.

4: Click on the Calibrate button.

5: If there is insufficient detail in the

selected region a warning will appear.
Repeat the process from Step (3).

Continued... 639

638 © 2011 Leica Microsystems (Switzerland) Limited

 Power Mosaic: Camera Rotation:

The values for camera rotation returned

by the Calibration, relate to the stage X
and Y axes. If the rotation is excessive a
warning will be displayed (2).

Rotation for both axes should not exceed

0.10 degrees (1).

Check that the camera mount is secure.

Loosen slightly the camera clamp (3) so

that the camera may be rotated.

Rotate it by very small amounts and

repeat the calibration until the rotation
angle is within limits.

Tighten the clamp.

Once set correctly and providing the

camera and stage do not change, rotation
should seldom require adjustment.

LAS User Manual 639

Power Mosaic: Shading:

The Shading Reference electronically

corrects any light level fall-off toward the
edges of the image that is often inherent
in optical systems.

The exposure, light intensity, required

objective and mag level must all be
properly set with the specimen in place. If
there is a change in objective, the Shading
Reference must be carried out again.

Select Live Screen view:

1: Navigate to a portion of the image
that is free of detail with a clear
background.

2: If necessary, click on the Optics

Setting header to reveal the panel. If
a Shading Reference has not been
carried out, the check box will appear
greyed out.

3: Click the Set Shading button and…

4: …on the dialog click the Set button.

5: When complete, the Set Shading box

(7) will be checked and shading will
be enabled.
If required, it can be disabled by
clicking the check box.

6: Click the Close button on the dialog.

640 © 2011 Leica Microsystems (Switzerland) Limited

 Power Mosaic: Load Configuration:

A Scan Pattern configuration may be

saved for future recall as a fast and
accurate way of loading settings.

To retrieve and automatically load a saved

configuration:

1: If necessary, click on the arrow to the

right of the Scan Pattern panel to
reveal it.

2: Click on the Load button.

3: On the Open dialog, click to select the

configuration required – the selected
file name appears in the window (4).
Scan Pattern configuration files have
the file extension .pat.

5: Click on the Open button.

Only the Scan Pattern settings are
loaded; Optics and Z-Stack settings
must be adjusted for each session.

6: The Folder defaults to the selection

made either in Preferences or Browse
. Use Windows navigation to open an
alternative folder.

See: Preferences:Save in Directory:

See: Browse:Select Default Directory.

LAS User Manual 641

Power Mosaic: Select Scan Pattern:

There are two scan pattern options which

govern the stage tracking as the image
tiles are collected – bi-directional and uni-
directional.

The bi-directional pattern scans left to

right, moves down to the next row,
reverses and scans right to left.

The uni-directional pattern scans left to

right, returns to the left, moves to the next
row and scans from left to right again.
(Depending upon the specimen shape the
scan direction may be from top to bottom).

1: Choose bi-directional for speed.

2: Choose uni-directional for greater

accuracy.

The selected check box is coloured red.

Select Scan Speed:

Turboscan is the fastest way to capture a
image, but it does require a very fast
exposure time - 200µs (micro seconds) or
less and a progressive scan camera with
trigger facility. During Turboscan, the
stage moves continuously – it does not
stop to make an exposure – which is why
a short exposure is needed.

During Standard scan the stage halts at

every tile position so the exposure time is
immaterial. Standard scan can also
operate with non-progressive cameras
and does not require a trigger facility.

3: Choose Turboscan for speed but only

if the exposure time is less than
200µs. Trying to scan with a higher
exposure time will display the warning
message (4).

5: Click to select Standard scan speed

for exposure times greater than
200µs and non-progressive cameras.

642 © 2011 Leica Microsystems (Switzerland) Limited

 Power Mosaic: Auto Step Size:

The Step Size represents the distance

between adjacent tiles in µm (micro-
meters). Normally, Auto Step Size is
enabled to allow mosaic creation, and the
program calculates the number of tiles
required to cover the image on the basis
that they will all abut adjacent tiles (2).
Turning off Auto Step Size allows the step
to be adjusted so that tiles may be
overlapped or spaced apart (8).

1: Enable Auto Step Size. The check

box will be coloured red with a tick.
Auto Step Size must be enabled for a
mosaic to be created.

2: The tiles are butting and arranged to

cover the image. In these examples
the Cross Scan Pattern has been
used for clarity.

3: Click to disable Auto Step Size. The

check box becomes grey.

4: Click the Details button.

5: The Details Dialog appears with…

6:…the tile X and Y co-ordinates

enabled for editing. To change the
step size, click on either the X or Y
text box and type a new value.

7: Click Apply. If necessary, re-enter the

step values to get the required
spacing or overlap (8).

LAS User Manual 643

Power Mosaic: Advanced Options:

The Advanced options allow individual

image tiles to be saved temporarily
(Buffered) to a nominated folder and, if
they are saved, set the size of the
thumbnails in that folder.

Saving tiles has the advantage of

identifying and keeping relevant parts of
the image, and discarding less important
parts so saving valuable disc space.

It is important that there is sufficient disk

space to accommodate all of the images
otherwise the scan will stop. The
recommended approach is to have a
partitioned section of the disk – D:
PMTemp for example – in which to buffer
the images. Make sure that computer
privileges extend to the partition.

Thumbnail size is also important in that

they are initially stored in RAM for fast
access. If the thumbnails are too large,
volatile RAM becomes clogged and the
speed advantage is lost.

1: Click on the Advanced button.

2: On the dialog click to enable or

disable tile buffering.

3: If tile buffering is required, click on

the Browse button to reveal the
Browse for Folder dialog.

4: Click to select a folder or…

5: …create a new folder. Use Windows

navigation to reach other levels or
directories.

6: Click OK.

7: If necessary, use the arrows to the

right of the Thumbnail Reduction
Factor text box. The larger the
number the greater the thumbnail
detail – and also the amount of disc
space occupied.

8: Click Apply and…

9:…click Close.

644 © 2011 Leica Microsystems (Switzerland) Limited

 Power Mosaic: Advanced Options: Scan Direction:

The stage Scan Direction may be either

side-to-side or front-to-back. The options
are:
Automatic: Allow the software to
determine the best direction,

Horizontal: Select side-to-side, or

Vertical: Select front-to-back.

Generally, Automatic is the best option to

ensure the most efficient scan, especially
when Turboscan is being used.

1: Click on the arrows to the right of the

Direction header to reveal the options
drop down menu.

2: Click to select the required direction.

Although both specimens 3 and 4

illustrated are the same, a vertical scan
on Figure (4) would be best because
there are fewer direction changes.

LAS User Manual 645

Power Mosaic: Rectangular Scan Pattern:

Select Pattern Type:

There is a wide range of scan patterns
available designed to provide total
flexibility and efficiency in the capture and
storage of images.

Each pattern type can be configured to

best suit the task in hand. On the following
illustrations, each small rectangle
represents a separate tile.

1: Click on the arrows to the right of the

Pattern Type header.

2: From the drop down menu select the

pattern required.

3: Click on the Details button. This

opens the appropriate pattern dialog
(4).

5: For the Rectangle Pattern (7), the X

and Y tile counts are available for
editing. Click in the X or Y text box
and enter a new value.

6: Click on the Apply button and the new

configuration is applied to the image.
Repeat steps (5 and 6) until the
pattern is suitable.

For clarity, the tile layouts on the following

pages are shown neatly abutting each
other. However, if the camera rotation is
not perfect the tiles may appear at an
angle and possibly overlapping (8). The
software has been designed to
accommodate these variations.

See: Create the Pattern Grid: 657

646 © 2011 Leica Microsystems (Switzerland) Limited

 Power Mosaic: Circular Scan Pattern:

The following pages illustrate the Pattern

Types available and the options for each.

Circular Pattern Type:

The options are:

Full Coverage and

Inside only.

1: Click on the Diameter text box and

enter a value for the circle diameter in
millimetres. The circle does not have
to cover the entire image.

2: Click on the arrows to the right of the

Coverage text box and…

3:…select either Full Coverage (4) or

Inside only (5) from the drop down
menu.
Full Coverage covers the entire
circle with overlap where necessary.
Inside only places tiles within the
circle.

6: Click Apply to apply the values to the

image. Repeat from Step (1) if
necessary to adjust the pattern.
Click Close to save the pattern and
exit.

LAS User Manual 647

Power Mosaic: Cross & CrossX Scan Patterns:

Cross and CrossX Pattern Types:

The options are:

X and Y tiles for the Cross pattern:

Tile count only for the CrossX pattern:

1: For the Cross pattern:

2 Click on the X or Y text boxes and

enter a value. The X value represents
the horizontal tile count and the Y
value the vertical count.

3: Click Apply to apply the settings.

Repeat from Step (2) to change the
pattern.
Click Close to save and exit.

4: For the CrossX pattern which is a

regular X:

5: Click on the X text box to enter a new

total tiles values.

6: Click Apply to apply the setting.

Repeat from Step (5) to change the
pattern.
Click Close to save and exit.

648 © 2011 Leica Microsystems (Switzerland) Limited

 Power Mosaic: Random Scan Pattern:

Random and Random without overlap

patterns:
The options are:

Random the number of tiles some of

which may overlap:

Random without Overlap the number of

tiles none of which will overlap:

1: Random pattern creates a specified

number of tiles randomly inside a
specified boundary. Some of the tiles
may overlap.

2: Click on the Target Width and Target

Height text boxes and enter values for
the scan area boundary. The scan
area does not have to cover the entire
image.
Click on the Total Fields (tiles) text
box and enter a value for the number
of tiles.

3: Click Apply to apply the settings.

Repeat from Step (2) the change the
values.
Click on the Close button to save and
exit.

4: Random Pattern without Overlap:

This option is the same as Random
except that none of the tiles will
overlap.

LAS User Manual 649

Power Mosaic: Annular Scan Pattern:

Annular:
Options are:

Full Coverage and

Inside only.

This option automatically populates an

area defined between two concentric
circles with a calculated number of tiles.

1: Click on the Diameter text box and

enter a value for the outer (larger)
circle in millimetres. The circle does
not have to cover the entire image.

2: Click on the Inner Diameter circle text

box and type a value for the smaller
circle (shown coloured on the
illustrations for clarity). It must be
smaller than the outer diameter
otherwise the settings will be ignored.

3: Click on the arrows to the right of

Coverage text box and from the drop
down menu…

4: …click to select either Full Coverage

or Inner only.

5: Inner only places the tile within the

outer circle avoiding inner circle.

6: Full Coverage ensures the entire

outer circle is covered without
including the area of the smaller
circle.

7: Click Apply to apply the settings.

Repeat from Step (1) to change the
values.
Click on the Close button to save and
exit.

650 © 2011 Leica Microsystems (Switzerland) Limited

 Power Mosaic: Focus Method:

Three automatic focus methods are

available:

Predictive Focus: which uses a set of

known focus points to interpolate
(predict) any unknown points.
Predictive Focus is fast with good
results but should be used only when
the specimen is either flat or has a
uniform, slope across focus points.
Use objectives up to 10x or for very
flat specimens up to 20x.

Autofocus: uses contrast differences in

groups of adjacent pixels to establish
sharpness. Because it is a continuous
process of focussing and checking,
Autofocus is slower than Predictive
but yields extremely good results.
especially over irregular specimens.

Predictive and Autofocus: in

combination uses the speed of
Predictive and the quality of
Autofocus to achieve very good
results, quickly. Use for specimens
that are predominantly uniform but
also have local irregularities.
Predictive will come close to focus
and the Auto will fine tune it.

Both Predictive and Autofocus methods

and the combination are selected by
enabling the check boxes (1).

When a method is enabled its setup

button becomes active (2). Setup is
explained on the following pages.

For perfectly flat specimens both methods

can be turned off.

Continued... 652

LAS User Manual 651

Power Mosaic: Predictive Focus:

The Predictive Focus function, focuses on

a number of predetermined points on the
image and creates a ‘table’ of their focus
values. The focus position for any other
point can be predicted by interpolating the
table values.

The greater the number of pre-determined

points then the greater the accuracy of the
prediction and better the overall focus.
This is especially true for very specimens
that have irregular focus.

1: Click on the Focus Method: Predictive

checkbox to enable it.

2: Click on the Setup button to display…

3: …the Predictive Focus dialog.

4: Three pre-determined points are

automatically placed on the image.
Their positions are displayed in the X/
Y columns on the dialog.
Normally, the set points will be
coloured red to indicate that they
require focussing. If they are green
and have a value in the Z column
denoting that they are already
focussed, possibly as the result of a
previous scan, the points should be
focussed again.

Continued... 654

652 © 2011 Leica Microsystems (Switzerland) Limited

 Power Mosaic: Predictive Focus: Continued:

The Grid feature automatically creates a

regular pattern of points across the image.
The number of points can be established
by typing a value.

1: Click on the Grid text box to high light

the existing value. Press the keyboard
delete key to remove the value. Type
a new value.
The value represents the number of
points that will be created along the
top row of the grid. On the second
row the number of points is reduced
by one. The next row is increased by
1 and so on across the entire grid. In
the illustration, a value of 3 has been
typed.

2: Click on the Grid button and the new

points will be created and listed on
the dialog.
Points sitting on completely clear
areas of the image (3) will result in a
failed Autofocus because there is no
detail to focus on. To avoid a failure,
either delete the points (See: Delete a
focus set point) 654 or adjust the
Focus Threshold Setting upward to
include values closer to white (clear).
However, setting the threshold to a
very high value can affect the
precision of Autofocus.See:
Initialisation: Focus 635

Continued... 660

LAS User Manual 653

Power Mosaic: Predictive Focus: Continued:

Reset Focus Point values:

1: To clear all focus point values, click
on the Reset button. The values will
clear and ‘???’ will be displayed in the
Z column of the dialog.

Delete a Focus Point:

2: To delete a focus point click on the
point on the dialog list.

3: Click the Delete button. The list entry

and the point disappear.

To create a New Point:

4: Click on the Move Stage To Point
button.

5: Select a new point on the image and

click. The point appears on the image
coloured red to indicate that it
requires focussing.

6: Click on the Add button.

Delete all Points:

7: All of the points may be deleted by
clicking on the Delete All button. The
dialog list will clear and the points will
disappear from the image.

Update:
Having added or deleted points, update
the list and displayed points by…

8: …clicking on the Update button.

Continued... 660

654 © 2011 Leica Microsystems (Switzerland) Limited

 Power Mosaic: Predictive Focus: Continued:

For multiple focus points:

1: The illustration shows a range of pre-

determined points that have been
added manually to comprehensively
cover the specimen.

2: Click on the Autofocus on all Points

button, or for a single point, click on
the Autofocus button.
The program will cycle through each
of the set points automatically
focussing. As each is completed the
point turns green and the value
appears in the Z column of the dialog.

3: Check ‘failed’ points (circled for

clarity) by…

4: …selecting the Move Stage To button

and then on the focus point. Manually
check the point for detail. Either
delete the point or adjust the Focus
Threshold Setting, reset the points
and repeat the process.

LAS User Manual 655

Power Mosaic: Autofocus:

Setup Autofocus Skipping:

To speed up the scan especially on
specimens that are more-or-less uniform,
automatic focussing can be skipped on
some tiles. The number of tiles to skip is
set up with Autofocus.

It is also possible to resume focussing on

a tile immediately following a focus ‘failure’
rather than skipping tiles, to maintain
focus integrity.

Autofocus failure generally occurs when

there is insufficient detail in the specimen.

1: Click to enable the Autofocus check

box.

2: Click on the Setup button and the

Setup Autofocus dialog appears.

3: To enable tile skipping click on the

Autofocus every… check box.

4: Set the number of tiles to skip by

either clicking on the text box,
deleting the existing value and typing
another, or clicking on the increase/
decrease arrows to the right of the
text box.

5: To Retry on Failure, click the check

box to enable.

6: Click on the Apply button and…

7: Click on the Close button.

Go to Create the Pattern Grid... 657

656 © 2011 Leica Microsystems (Switzerland) Limited

 Power Mosaic: Create Pattern Grid:

This step draws a pattern grid

representing the scan tiles over the image.
Power Mosaic will use this grid as a
scanning ‘map’. Because the specimen
has yet to be scanned, its precise position
on the stage is unknown so the first grid
drawing locates it.

1: Click on the Split View button and

navigate to an appropriate part of the
specimen using the on-screen
joystick.

2: Click on the Pattern View button.

3: Click on the arrows to the right of the

Pattern Type header and from the
menu...

4: …select by clicking the Rectangular

option. A different pattern may be
selected later.

5: Click on the Draw Pattern tool and…

6: …positioning the cursor close to the

stage marker (small, green
'crosshair'), click and hold...

7: …and drag diagonally to the right. An

arbitrary grid pattern should appear. If
it does not, …

8: …click on the Show Grid Lines button

to reveal it.

Continued... 658

LAS User Manual 657

Power Mosaic: Create Pattern Grid: Continued:

9: Click on the Clear Tiles button to

clear any previous images.

10: Confirm clearing the images on the

warning message by clicking Yes.

11: Click on the Acquire Power Mosaic

button. The specimen will be scanned
with the tiles filled in sequence.
The first drawing usually encloses
only part of the image as shown in the
illustration. If this is the case go to the
next page.

9: If none of the specimen is found click

on the Clear Tiles button and repeat
the process starting the drawing at a
different location.

Continued... 659

658 © 2011 Leica Microsystems (Switzerland) Limited

 Power Mosaic: Create Pattern Grid: Continued:

With the pattern grid enclosing only part of

the specimen, it can be extended so that
the specimen is fully covered.

1: Click on the Create/Expand button.

This feature allows the existing grid to
be expanded.
The Create/Expand button may also
be used to reduce the pattern grid if it
is too large.

2: Click on the top left corner of the

existing pattern grid, hold and drag
diagonally to the right. A dotted
rectangle follows the cursor to
indicate the extent of the grid.
Release the mouse button when the
specimen is considered to be
completely enclosed (3).

4: Click on the Clear Tiles button and

confirm the clear on the warning
message.

5: Re-scan by clicking the Acquire

Power Mosaic button.

6: To re-position the scanned image

within the grid, click on the Move
Scan Pattern button, click and hold on
the scanned image and drag it to
reposition.

7: The Magnifier may help in the

repositioning - click on the tool and
use the mouse buttons to enlarge (left
button) or reduce (right button) the
view.

Continued... 660

LAS User Manual 659

Power Mosaic: Create Pattern Grid: Continued:

Having re-positioned the image within the

pattern grid, it is possible that there are
too many tiles. Excess tiles may be
removed as columns or rows only by:

1: Click on the Details button and on the

dialog, change the fields (tiles) to
remove any excess.

2: Click the Apply button and…

3: …click the Close button.

4: Select the Move Scan Pattern button

and re-position the grid to make sure
the fit over the specimen is
acceptable.

5: If an alternative Pattern Type is

required, click on the arrows to the
right of the Pattern Type header and
from the drop down menu click to
select the required pattern.

It may be necessary to re-adjust the

pattern grid-to-specimen fit using the
Move Scan Pattern button.

Go to Save Configuration... 661

660 © 2011 Leica Microsystems (Switzerland) Limited

 Power Mosaic: Save Configuration:

With the Power Mosaic configuration

complete, it is possible to save the
settings for instant use in the future.

To save the current settings:

1: On the Scan Pattern panel click the
Save As button.

2: On the Save As dialog…

3: …type a file name and…

4: …click the Save button. Files are

automatically saved with the .pat
extension.
The Save As dialog defaults to the
folder selected in Preferences or
Browse.

See: Preferences:Save in Folder:

See: Browse:Select Default Folder:

5: The Save As folder may be changed

on the Save As dialog using Windows
navigation.

LAS User Manual 661

Power Mosaic: Scan:

Before starting a Power Mosaic scan, the

thumbnails and tiles from previous scans,
if not required, should be deleted. This is
not mandatory – keeping previous
information can be a vital part of an
ongoing session in which the specimen is
in several parts for instance.

Thumbnails of each tile are stored in RAM

to make access fast and immediate. They
are used to ‘paint’ the mosaic on the
Viewer. Keeping thumbnail size as small
as possible (See: Advanced Options:
Thumbnail Reduction Factor 644 ) will help
prevent filling the RAM too quickly.

The tiles are stored initially in a temporary

file on the hard drive. The individual
images are much larger and used to paint
the mosaic at greater resolution to
preserve detail.

Clearing previous scans removes the

thumbnails from RAM and the tiles from
the temporary file. If the previous scan is
left intact, new thumbnails and tiles will be
added to the existing with sequential
image numbers.

1: Click on the Clear Tiles button to

clear the previous scan. This is only
the previous scan and does not
remove any prior to that.

2: The Clear Mosaic message appears.

Click Yes to continue.

3: Click on the Acquire Power Mosaic

button.

4: If Turboscan is selected and the

exposure speed is too high, a dialog
appears giving the option to use
Standard scan instead or to cancel.

662 © 2011 Leica Microsystems (Switzerland) Limited

 Power Mosaic Plus: Z-Stacks:

If Power Mosaic Plus is installed and

enabled, the Z-Stack option is available.
For every tile position on the XY plane,
Power Mosaic Plus creates a stack of tiles
in the third dimension or Z plane.

The number of tiles in the stack and the

distance between them can be set to
reflect the specimen.

Once captured the individual stack tiles

can be processed in several ways and
eventually into a single composite image
that represents the best focus across the
entire specimen regardless of thickness.

Z-Stacks can occupy very large amounts

of disc space especially if all of the tiles
are saved, so ensure that a liberal amount
of space is available. The best
arrangement for image storage is to have
a partitioned area on the hard drive (Drive
D: for example) separate from other
programs and data, or a completely
independent drive.

1: Click on the arrows to the right of the

Z-Stacks panel to reveal it.

2: Click the Enable Z-Stack check box

to enable scanning.

Continued... 664

LAS User Manual 663

Power Mosaic: Z-Stacks: Continued:

3: To select the required processing

method, click on the arrows to the
right of the Method header and from
the drop down menu…

4: Select the method:

Z-Stack will save all of the captured

stack tiles for further, individual
viewing.

Best Focus chooses the tile with the

sharpest image from each stack and
discards the rest.

Z-Stack + Best Focus combines both

options, retaining all of the stack tiles
but also selecting the best from each
stack which it places at the lowest
level.

Extended Focus examines all of the tiles

from a stack, choosing the ‘best’ pixel
from each at a given location. These
are then combined into a single tile
and the rest discarded.

Maximum and Minimum Projections

emulate Extended Focus to create
composite images based upon the
darkest and lightest pixels in each
pixel column in each stack tile.

Continued... 665

664 © 2011 Leica Microsystems (Switzerland) Limited

 Power Mosaic: Z-Stacks: Continued:

The Z-Range – the focussing distance

based upon objective depth of focus - in
microns that will be travelled – Step Size
and Number of Steps, are inter-related;
Change one value and the others will be
calculated and changed automatically.
Actual settings will depend upon the
specimen.

1, 2 and 3: Use the up/down arrows to

the right of each text box to enter a
value. For large values, click on the
appropriate text box and type a value.

4: The Set Step Size check box if

enabled will automatically load a step
distance based upon the depth of
focus and adjust the Z-Range and
Number of Steps accordingly.
Changing the Step Size manually will
turn off Set Step Size.

5: Click on the Clear Tiles button to

discard any previous scans. The
Clear Mosaic warning will appear…

6:…click Yes to clear.

7: Click the Acquire Power Mosaic

button.
Z-Stacks are only available in
Standard Scan mode. If Turboscan
has been selected it will be ignored

Continued... 666

LAS User Manual 665

Power Mosaic: Z-Stacks: Continued:

When the scan is complete the entire

image is displayed with the tile grid
overlaid (1).

Two methods – Z-Stack and Z-Stack +

Best Focus – have a slider (2) associated
with them which allows each stack step to
be viewed individually, a powerful image
analysis tool.

The slider scale represents the number of

steps or multiples of steps. Click, hold and
drag the slider to display the step results
in turn.

666 © 2011 Leica Microsystems (Switzerland) Limited

 Power Mosaic Plus: Pattern Navigator:

Pattern Navigator is a module that is part

of and licensed by Power Mosaic Plus. It
allows users to create Scan Patterns
randomly in any position across the stage
and then scan them either all together or
at selected sites. The great advantage of
this facility is that only areas of interest are
captured substantially reducing both
processing time and required disk space.

Since patterns can be saved and restored,

tasks that routinely use the same
specimen layout on a slide (say), benefit
from the speed that having a pre-defined
scanning template can offer. For example,
illustration (A) shows a standard 75 x 25
microscope slide (enlarged for clarity)
divided into 12 specimen ‘wells’. The wells
are not spread evenly across the slide –
there is a larger area at the top than there
is at the bottom - nor are they of uniform
size. To complicate matters even further,
only 6 of the specimens are usually
scanned – they are marked with a red
border in the illustration. It is a simple task
for Pattern Navigator to locate the
required wells, create a scan pattern them
and to store that as a template for future
processing.

Continued... 668

LAS User Manual 667

Pattern Navigator: Workspaces:

Pattern Navigator allows the user to divide

the viewable stage into distinct areas
called Workspaces. The number and
position of the Workspaces is determined
by the users for ease of working. The
entire stage could be classed as a single
Workspace (A); It could be divided into
several Workspaces of differing areas (B)
or a group of uniform Workspaces like 4
slides in a row for example (C). The only
restriction is that Workspaces must not
overlap.

Each Workspace encloses its own group

of Scan Patterns (D). An individual pattern
can be re-sized and moved within its
Workspace providing it does not overlap
with another. Scan Pattern shapes –
rectangular, square, circular, cross etc –
can be mixed within a Workspace to
achieve the most efficient and economical
image capturing.

Because a Workspace acts as a container

for its Scan Patterns, it can be moved to
any location on the stage and its patterns
will move precisely with it.

Workspaces can also be copied, so

having set up a group of patterns the
entire Workspace can be duplicated and
‘pasted’ to other parts of the stage. This is
especially useful for repetitive specimens
– four identical slides in a row for
example.

By default the Workspace outline and fill is

turned off. To turn it on and change its
properties: Go there... 696

668 © 2011 Leica Microsystems (Switzerland) Limited

 Pattern Navigator: Creating and Mapping the Workspace:

Capturing specimen mosaics using

Pattern Navigator comprises three simple
steps:

1: Creating and mapping the Workspace

.

2: Plotting the individual Scan Patterns

within the Workspace.

3: Scanning using the Scan Patterns.

Creating and Mapping the Workspace:

Illustration (A) shows the sample
microscope slide described earlier.

To map the locations of the required

specimens, a rapid scan is made using a
scan pattern that covers the entire
Workspace – in this case the slide area
(B).

Using this as a guide, individual Scan

Patterns are created and placed (just by
dragging and dropping) over the
specimens. The overall scan pattern is
then removed (C).

Continued... 670

LAS User Manual 669

Pattern Navigator: Set the Exposure for TurboScan:

To map the Workspace only low resolution

images are required because they are to
be used only as a guide to the specimen
location. Low resolution capture formats
also reduce the scanning time.

1: In the Acquire Workflow click on the

Camera tab.

2: Click on the arrow to the right of the

Image Formats panel.

3: Click on the arrows to the right of the

image type header and…

4: …select Colour or Greyscale to suit

the image.

5: Click on the Mic tab and...

6: ..on the arrow to the right of the

Objectives header.

7: Select the lowest magnification

objective available.

8: Adjust the Exposure and…

9: …the Gain to achieve an acceptable

image with an exposure time no
greater than 200µs, necessary to run
Power Mosaic in TurboScan mode.

10: It may be necessary to adjust the

Aperture and lamp Intensity (11) on
the Microscope tab.

Continued... 671

670 © 2011 Leica Microsystems (Switzerland) Limited

 Pattern Navigator: Stage Initialisation, Calibration and Shading:

1: On the Acquire Workflow, click on

the PM (Power Mosaic) tab.

2: Click on the small arrow to the right of

the Optics Settings header to reveal
the panel.

Stage Initialisation:
3: Click on the Initialise and Set Speed
button: Follow the Initialisation
procedure: Go there…

Objective Calibration:
4: Click on the Set Calibration button:
Follow the Calibration procedure: Go
there…

Shading:
5: Click on the Set Shading button and
follow the Shading procedures: Go
there…

Continued... 672

LAS User Manual 671

Pattern Navigator: Scan Setup:

On the PM tab:
1: Click on the small arrow to the right of
the Scan Pattern header to reveal the
panel.

2: Click on the arrows to the right of the

Pattern Type menu and from the drop
down list click to select Rectangular
pattern.

3: Click to enable the TurboScan button.

4: Click to enable the Auto Step Size

check box.

5: Click to enable bi-directional

scanning, the fastest option.

6: Disable both Focus Method options.

Continued... 673

672 © 2011 Leica Microsystems (Switzerland) Limited

 Pattern Navigator: User Interface Setup:

On the Side Tool Bar (1) click to select: 6: The Stage Viewable area shown as a hatched
2: …the Stage and Live Image option, outline.

3: …the Stage View, 7: A Workspace with Scan Pattern which at first use will
be arbitrary sizes and positions. Subsequently, the
4: …and Show Scan Pattern. last used Workspace will be displayed.

With this arrangement: 8: The Stage Marker which is the current stage position.
5: The Live Image appears top right of the Viewer.

Continued... 674

LAS User Manual 673

Pattern Navigator: Launch the Navigator:

On the PM (Power Mosaic) tab:

1: click the small arrow to the right of
the Power Mosaic Plus header to
reveal the panel.

2: On the Pattern Navigator panel, click

to enable Pattern Navigator and…

3: …click the Show Pattern Navigator

button. The Navigator dialog appears.

4: On the dialog, the current displayed

Workspace is listed together with the
Scan Pattern(s) associated with it.
Both Workspace and Pattern names
can be changed to suit the user, so
the words may differ but the layout will
be the same.

5: The Enable Navigator Mode check

box should be enabled.

Change Workspace and Scan Pattern

Names: Go there... 689

Continued... 675

674 © 2011 Leica Microsystems (Switzerland) Limited

 Pattern Navigator: Re-sizing the Workspace:

In this example, the specimens are

mounted on a standard 25 x 75mm
microscope slide so the Workspace will be
re-sized to the same dimensions.

1: Right click on the Workspace name.

2: The Workspace dialog appears.

3: Click on the Properties option.

4: On the Workspace Properties dialog,

click on the Vertical Slide button to
automatically re-size the Workspace
to 25 x 75mm orientated vertically.

5: Click Apply and…

6: …click OK.

Continued 676 ...

LAS User Manual 675

Pattern Navigator: Locate the Specimen Top Left-hand Edge:

1: The Workspace has been re-sized to

represent a standard 25 x 75
microscope slide.

The next step positions the Workspace

over the specimen slide with their top –
left-hand corners aligned:

2: Using either the on-screen Joystick or

Leica SmartMove (if fitted),

3: …drive the Stage to the top left-hand

corner of the specimen slide.

4: The Stage Marker moves to reflect

the current stage position.

Continued... 677

676 © 2011 Leica Microsystems (Switzerland) Limited

 Pattern Navigator: Align Workspace and Specimen Slide:

1: The Stage Marker has moved to the

current Stage position which is now
the top left-hand corner of the
microscope specimen slide.

2: On the Navigator dialog, click the Set

Group to Stage XY button.

3: The Workspace and Scan Pattern

immediately move to align with the
Stage Marker and therefore, the
specimen slide.

Continued... 678

LAS User Manual 677

Pattern Navigator: Creating the Scan Pattern:

Create a Scan Pattern covering the entire

Workspace – in this example the size and
shape of a microscope slide.

1: Click on the Create New Pattern

button.

2: Click on the top left-hand corner of

the Workspace and…

3: …holding the mouse button down,

drag to the bottom right-hand corner
and release the button.

4: Delete the original (small) Scan

Pattern by, on the Navigator dialog,
clicking the pattern name in the list.

5: Click on the Delete Pattern(s) button.

Only the full-sized Scan Pattern now
remains.

Continued… 679

678 © 2011 Leica Microsystems (Switzerland) Limited

 Pattern Navigator: Scanning:

Only the Workspaces and Scan Patterns

that are selected are scanned – the check
box alongside has to be enabled:

1: Click to enable the check boxes to

the left of the Workspace and Scan
Pattern.

2: Start the scan of the entire specimen

slide by clicking on the Acquire Power
Mosaic button.

All of the specimens appear positioned

against the Scan Pattern that will be used
to precisely locate individual patterns
ready for high quality image captures.

Continued… 680

LAS User Manual 679

Pattern Navigator: Creating Individual Scan Patterns:

Each specimen is now given its own

individual Scan Pattern and it is these that
will be used to capture the final, high
quality images.

1: Click on the Create New Pattern

button.

2: Click on the large Scan Pattern above

and to the left of the specimen.
Holding down the mouse button drag
down and to the right to create a
pattern that fully encloses the
specimen.

3: Release the mouse button.

4: If necessary, click on the Move

Pattern button and then on the new
pattern. With the mouse button held
down drag the Scan Pattern to the
correct position.

Repeat the process for all of the other

specimen areas. As the patterns are
created they are added to the Workspace
list.

Continued… 681

680 © 2011 Leica Microsystems (Switzerland) Limited

 Pattern Navigator: Remove Guide Pattern:

1: All of the required individual Scan

Patterns are in place. The next task is
to remove the large guide Scan
Pattern.

2: Click on the large Scan Pattern on

the Navigator dialog to select it.
Selected Scan Patterns are
highlighted on the Workspace.

3: Click on the Delete Pattern(s) button.

4: The large guide pattern is removed

leaving just the individual patterns.

Continued… 682

LAS User Manual 681

Pattern Navigator: Save and Load a Workspace Group:

At this point it is advisable to save the

Workspace and its Scan Patterns.

1: On the Navigator dialog, click on the

Save Group button.

2: The Windows Save As dialog

appears.

3: The default folder for saved

Workspaces is OIGroups but this can
be changed to suit the user simply by
navigating to an existing folder or
creating a new one.

4: Type a unique name for the

Workspace group.

5: Click Save.

Loading a Saved Workspace Group:

To re-load a Workspace and its Scan
Patterns:

6: Click on the Navigator Load button.

7: On the Open dialog…

8: …click on the required file with the .

oig extension – not on the folder of
the same name.

9: Click Open and the Workspace will

be loaded and displayed.

Continued… 683

682 © 2011 Leica Microsystems (Switzerland) Limited

 Pattern Navigator: Final Scan:

Before scanning with the individual Scan

Patterns:

1: On the Camera tab, select a higher

quality Captured Format.

2: On the PM tab, select the Scan

Pattern panel and decide upon
TurboScan (high speed but lower
precision) or Standard Scan (slower
but higher precision). The higher
quality image formats will often
preclude using TurboScan due to the
high capture time.

3: Decide upon bi-directional (faster but

lower precision) or uni-directional
scanning (slower but better
precision).

4: Select the Focus Method. This may

require re-calibrating the focus.

5: Select the specimens to scan by

enabling the check box to the left of
each pattern.

6: Click on the Clear Tiles button to

remove the previous scans and...

7: ...confirm the deletion.

8: Then start scanning…

LAS User Manual 683

Pattern Navigator: Features and Functions:

This section describes the wide range of

features available in Pattern Navigator.

On the Navigator dialog with a Scan

Pattern selected:

1: Clicking Move to Middle, drives the

stage to the centre of the selected
Scan Pattern.

2: Set Focal Plane sets the Z axis to the

centre of the current Scan Pattern.

3: Save Camera Settings: Not required.

4: Enable the check box to hide all of

the Scan Patterns except the one
selected.

5: Enable the check box to have Scan

Patterns de-selected (check box to
the left of the pattern name is cleared)
after they have been scanned. The
check box will have to be enabled
before the pattern can be scanned
again.

6: When Overwrite is enabled, any

scanned images already present will
be overwritten with a new image from
the current scan.

Continued… 685

684 © 2011 Leica Microsystems (Switzerland) Limited

 Features: Add and Delete Workspaces and Patterns:

To add a Workspace:
1: Click on a Workspace name on the
Navigator list. This is the Workspace
that will be copied and added. Click
on the Add Workspace button. A
duplicate of the selected Workspace
appears with a new entry in the list. At
least one Scan Pattern is created with
a new Workspace.
Reposition the copy by selecting the
Drag and Drop button (3) on the Side
Toolbar, clicking on the Workspace
copy and dragging it to a new
location. A Workspace can be
positioned precisely by using the
Workspace Properties dialog: Go
there…

Delete a Workspace:
2: Click on the Workspace name to be
deleted on the Navigator list. Click on
the Delete Workspace(s) button. On
the Delete Confirm dialog (4) select
either Delete the Selected Workspace
or Delete All but the Selected
Workspace. Click OK and the
Workspace(s) will be removed
permanently.

Continued… 686

LAS User Manual 685

Features: Add, Delete and Copy Patterns:

To Add a Scan Pattern:

1: Click on the pattern name on the
Navigator list to be duplicated and
added. Click the Add Pattern button
and a copy of the pattern will appear.
Use the Drag and Drop tool (3) to re-
position it.

Delete a Scan Pattern:

2: Click on the pattern name to be
deleted on the Navigator list. Click on
the Delete Pattern(s) button. On the
Delete Confirm dialog (4) select
either Delete the Selected Pattern or
Delete All but the Selected Pattern.
Click OK and the patterns(s) will be
removed permanently.

Copy a Scan Pattern:

5: Click on the Copy Pattern button to
copy the currently selected Scan
Pattern to the 'cursor' and click inside
the Workspace to place a copy.
Repeat for as many patterns required.

Continued... 687

686 © 2011 Leica Microsystems (Switzerland) Limited

 Features: Replicate Workspaces:

Workspace replication makes multiple

copies of the selected Workspace,
including any attached Scan Patterns, and
places them in rows and columns
determined by the user. The original
Workspace is removed.

1: Click on the Workspace name on the

Navigator list that is to be replicated.
And click on the Replicated Selected
Workspace button.

2: The Replicate dialog appears. Set the

number of Rows and Columns
required and their spacing by clicking
in the appropriate window and typing
a value.

3: Click Create. A warning appears

advising that the original will be
deleted and replaced by a new group
each Workspace of which will be a
faithful copy of the original.
The Row and Column settings can be
adjusted and the Create button
clicked again without closing the
dialog.

Continued… 688

LAS User Manual 687

Features: Replicate Scan Patterns:

Scan Pattern replication, makes multiple

copies of the selected pattern and
arranges them precisely in a matrix either
created by the user or based upon the
range of templates supplied with the
Navigator.

1: Click on the Pattern name on the

Navigator list and click the Replicate
Selected Pattern button.

2: On the Replicate Pattern dialog, enter

the number of Rows and Columns as
well as the spacing between, by
clicking inside the appropriate text
box and typing a new value.

3: A range of templates for standard

Well Plate configurations are provided
and a template can be selected by
clicking to enable the check box to its
left.

4: If the current Workspace is too small

to accommodate the selected
template, it is automatically re-sized.

5: For user-defined column and row

layouts click to enable the Other
option and, if necessary, enable the
automatic re-sizing facility (4).

6: Click the Create button. The selected

pattern is replicated on its Workspace
. The Row, Column and spacing
values can be adjusted and the
Create button clicked again without
closing the dialog.

Continued… 689

688 © 2011 Leica Microsystems (Switzerland) Limited

 Features: Workspace Alternate Menu:

Right-clicking on a Workspace name on

the Navigator, reveals an alternate menu.
Click to select the high-lighted option:

Set Origin to Stage X/Y: Moves the

entire selected Workspace and patterns
to the current stage position. The Stage
Marker is centred on the ‘first’ Scan
Pattern.

Rename: Type a new name for the

Workspace. It must be unique.

Clear Tiles: Clears all of the mosaic

filed images form all of the patterns on
the Workspace.

Delete: Deletes the selected

Workspace and its patterns.

Re-sequence Patterns: Re-numbers the

pattern sequence starting at ‘1’.

Select All Patterns: Selects and high

lights all of the patterns on the
Workspace.

Deselect All Patterns: Deselects all

selected Scan Patterns.

Continued… 690

LAS User Manual 689

Features: Workspace Alternate Menu: Continued:

Right click on a Workspace name on the

Navigator list and from the drop down
menu:

1: Click on the Properties option.

2: The Workspace (top left corner) can

be positioned precisely by setting the
Stage-X and Y co-ordinates and
clicking the Apply button (7).

3: Set Position to Current Stage XY

when clicked will move the
Workspace to the current stage
position.

4: The Workspace can be re-sized by

clicking in the Height and Width text
boxes and typing a new value. Click
the Apply button (7).

5: Three pre-set templates – Full Stage,

Horizontal and Vertical Slides – are
provided. Click on an option to
automatically size the Workspace.

6: The default name for Scan Patterns is

‘Pattern’ followed by a sequential
number as they are created. To
change the default name, click in the
Default Pattern Name text box and
type a new name. To retrospectively
rename all existing names, click to
enable the Rename All check box.

7: Click Apply and…

8: …click OK.

Continued… 691

690 © 2011 Leica Microsystems (Switzerland) Limited

 Features: Scan Pattern Alternate Menu:

Right clicking on a Scan Pattern name on

the Navigator, reveals an extended menu
of options. Click to select a menu item:

Set Origin to Stage XY: Moves the

Scan Pattern and its Workspace to the
current stage position. The Stage
Marker is located at the centre of the
Scan Pattern.

Scan: Starts a scan for the selected

pattern only.

Move To..: Selects the Drag and Drop

tool. Click on the Scan Pattern and drag
it to the required location.

Lock: Locks the Scan Pattern at its

current position and a small padlock
icon appears to the left of its name on
the Navigator. Click Lock again to
unlock it.

Clear Tiles: Removes any mosaic fields

in the selected pattern only.

Rename: Type a unique name for the

pattern.

Delete: Deletes the selected Scan

Pattern only.

Continued… 692

LAS User Manual 691

Features: Scan Pattern Alternate Menu: Properties:

To display the Scan Pattern properties:

1: Click on the arrow to the right of the
Scan Pattern header to reveal the
panel.

2: click on the Details button. The Scan

dialog appears – in this case
Rectangular layout.

3 & 4: Change the number of Columns

(X) and Rows (Y) by clicking inside
the appropriate text box and typing a
value.

5: Click Apply and the selected pattern

will change to reflect the new values.

6: A Scan Pattern Type– in this example

the Rectangular layout has been used
– can be changed to better suit the
specimen shape and optimise the
number of images required. Click on
the arrows to the right of the Pattern
Type header and from the drop down
list…

7: …click to select the required type.

692 © 2011 Leica Microsystems (Switzerland) Limited

 Pattern Navigator: Options after Scan:

When the scan is complete, there are

several options that can be invoked
automatically – such as saving the
mosaic.

1: Click on the Navigator Options tab to

reveal the options. Check to enable
the required options:

Save Workspace: Automatically saves

the current Workspace with the word ‘
Workspace’ as the file name followed
by a sequential number – Workspace-
05 for example.

Save Thumbnails to Folder: Saves a low

resolution thumbnail of each scan tile
within the Workspace folder to be
used in the Gallery.

Clear Tiles After Save Workspace/

Mosaic: Clears the scan tiles from all
Scan Patterns. Especially important
where machine memory is at a
premium. Use a greater Display
Thumbnail Reduction Factor to
reduce memory usage. Go there... 694

LAS User Manual 693

Pattern Navigator: Map Properties:

There are a large number of options that

can be used to tailor Pattern Navigator
exactly to a user’s needs. The major
options are explained here. The Map
Properties dialog is reached by:

1: Click on the Interactive Mouse button

on the Side Tool Bar and then right-
click anywhere on the stage display
area. The Map Properties dialog
appears.

2: If necessary, click on the Map tab to

reveal the main functions.

3: The View panel determines the

current Navigator view:

Stage: The viewable Stage area with

the Workspaces and Scan Pattern(s).

Pattern: The selected pattern is

displayed enlarged.

Preview: Not required.

Movement Trace: Displays the trace

path over the Scan Pattern (4).

Annotations: Turns On and Off Pattern

Name and Field Numbers if they are
enabled.

Preview Outline: Not required.

Continued… 695

694 © 2011 Leica Microsystems (Switzerland) Limited

 Pattern Navigator: Map Properties: Continued:

In the Pattern Display Panel:

1: Click on the arrow to the right of

Background Style menu to reveal how
the Scan Pattern Fields should be
highlighted and filled. Click a menu
item to select it.

2: The Scan Pattern border style can be

changed by clicking on the arrow to
the right of the Border Style menu
and clicking to select an option.
Rectangle (3) is the ‘normal’ rectangle
configuration and…

4: …Circle configures the pattern as a

series of overlapping, circular fields.

Field Outlines: Displays an outline

around each field.

Pattern Names: Displays the Pattern

Name beneath the pattern (4).

Field Numbers: Displays a sequential

number against each field (4).

Scan Progress: Displays a progress bar

during scanning.

Use FOV for Tiles: The Field of View

determines the field dimensions.

Continued… 696

LAS User Manual 695

Pattern Navigator: Map Properties: Continued:

1: Click on the Colour tab to reveal the

colour options.

2: On the left-hand panel, click to select

the item to have a colour change. Use
the slider on the right to display all of
the items.

3: Choose a colour from the right-hand

display. The first colour entry has the
option to select a user colour via the
Windows Colour dialog.

4: Click on the Cursor tab to modify the

Cursor – stage marker.

5: Adjust the values to construct the

cursor of choice – even an elliptical
outline (6).

The Workspace can be displayed or

hidden and its shape and colour changed
by:

7: Click on the Workspace tab.

8: Display or hide the Workspace by

checking or clearing the check box.

9: Click on the small arrow to the right of

the Background Style menu to list the
options - display in selected colour
(See Step 2 above) or display the
outline only.

10: The shape of the Workspace is set

by clicking the small arrow to the right
of the Border Style menu and
selecting from Rectangular or Circular
.

Continued... 697

696 © 2011 Leica Microsystems (Switzerland) Limited

 Pattern Navigator: Advanced Options:

Some of the main Pattern Navigator

properties can be accessed through the
LAS Power Mosaic panels as follows:

1: Click on the small arrow to the right of

the Scan Pattern header.

2: Click on the Advanced button.

On the Advanced Settings dialog:

3: Set the scan tiles capture folder by
enabling the Buffer Tiles to Folder
check box,…

4: …click on the browse button and, on

the Windows dialog, navigate to and
select the destination folder.

5: Set the Thumbnail Reduction Factor –

a larger number results in more
memory space-saving – by clicking
on the Up/Down arrows to the right of
the text box.

6: Display/Hide the Scan Pattern Names

by checking or clearing the check
box.

7: Display/Hide the Scan Pattern Field

Numbers by checking or clearing the
check box.

8: Enable/Disable the stage travel Trace

by checking or clearing the check
box.

9: Click Apply to load the settings and

close the dialog.

LAS User Manual 697

Power Mosaic: Browse Save & Display:

After a scan (or scans), the tiles are saved

in a temporary file on the hard drive and
the thumbnails in RAM. Both may be
saved to a nominated folder - the Capture
Folder - on the hard drive.

Select the Capture Folder:

1: Click on the Browse Workflow tab.

2: Click on the archive folder in which to

save the scans.

3: Click on the Set Fixed Capture

Location button.
A red dot appears to the left of the
selected folder.

Continued... 699

698 © 2011 Leica Microsystems (Switzerland) Limited

 Power Mosaic: Save Tiles:

There are two parts to saving the scanned

images – saving the collection of
individual tiles which at this point are still
being stored in a temporary file, and
thumbnails of the tiles that are in volatile
RAM, - and then saving the tiles as a
composite image or mosaic.

1: Click on the PM (Power Mosaic) tab.

2: Click on the Save Workspace

Location button. A Workspace is a
detailed description of the display and
image settings saved as a file with the
extension .sws within the Workspace
Location.
Additionally, another folder to contain
the individual tiles is created within
the Workspace Location together
with Thumbnail images of each tile.

3: On the Browse For Folder dialog...

4: ...navigate to the folder in which to

save the Workspace and...

5: ...click OK.

6: Click on the Save Workspace As

button and on the Save As dialog...

7: ...type a unique name for the

Workspace and click Save (8).
A Saving mosaic images progress
message appears and the tiles are
saved at the format and resolution
selected on Camera:Input Options.

LAS User Manual 699

Power Mosaic: Save Mosaic:

The composite image or mosaic file is

saved directly in the Capture Folder with
several associated control files. Because
mosaic images can be very large in terms
of disk space occupied – several
gigabytes is not unusual – there is a
facility to reduce the size of the saved
mosaic. This is called the reduction factor.

1: Click on the arrow to the right of the

Image Handling header to reveal the
panel.

2: Using the arrows to the right of the

Reduction Factor text box, increase
or decrease the Reduction Factor.
The illustrations show the
considerable difference in image size
with factors of 1 and 4.

3: Click in the Mosaic Image Name text

box and type a name for the mosaic.
The image will be stored with this
name in the Capture Folder.

Continued... 701

700 © 2011 Leica Microsystems (Switzerland) Limited

 Power Mosaic: Save Mosaic: Continued:

It is not necessary to save the entire

mosaic – very often only one part is of real
interest and the remainder can be
discarded. It is possible to ‘crop’ the image
to a selected region and save that alone.
The individual tiles are not affected – they
remain intact.

To save a selected part of the mosaic:

1: Click on the Saved Marked Region
check box to enable it.

2: Click on the edge of the region to be

saved, hold and drag diagonally to the
right to encompass the region.
Release the mouse button. For clarity,
the illustration shows a red dotted line
– on screen it is black.

3: Save the mosaic by clicking Send

Mosaic to Capture Folder.

4: The mosaic can also be saved to the

Windows Clipboard for loading into
another application. Click on the
Copy Mosaic to Clipboard button.

Go to Retrieving the Mosaic... 702

LAS User Manual 701

Power Mosaic: Retrieving the Mosaic:

When a mosaic is retrieved and re-

displayed, it is the Workspace and
therefore the tiles that are targeted with a
new full size mosaic created on the
viewer. This could be saved again as a
mosaic either as a selected region or at a
different size.

To retrieve a mosaic:
1: Click on the arrows to the right of the
Select Workspace header.

2: From the drop down list, click to

select a Workspace.

3: A progress panel (Loading Mosaic

mages) appears and the tiles are
displayed as a mosaic on the viewer.

4: Detail of the mosaic can be displayed

by clicking the Details button and…

5: …the Workspace Details panel

appears specifying the number of tiles
and image area.

To delete a Workspace:
6: Click on the Delete button to delete
the selected Workspace (Steps 1 and
2 above). Deleted Workspaces
cannot be retrieved.

702 © 2011 Leica Microsystems (Switzerland) Limited

 Power Mosaic: Leica ISO Pro Stage:

Although the Leica ISO Pro stage is driven

directly by Leica Application Suite, it still
requires an Oasis Controller Board to
report the X and Y co-ordinates.

Connection between the stage and the

computer is by a 15 pin (not the usual 9
pin) 'D' style DIN plug (1).

Selecting the Stage:

LAS needs to 'know' that an ISO Pro
stage is to be used and this is setup using
the Oasis Configuration Wizard. The
illustrations are from a typical Windows
XP operating system - Windows Vista is
very similar.

2: Click on the Start button and...

3: ...on All Programs.

4: From the pop up menu, click to select

the Oasis Configuration Wizard.

Continued... 704

LAS User Manual 703

Power Mosaic: Heerbrugg ISO Pro Stage: Model Selection:

1: When the Oasis Configuration Wizard

opens, click on the System
Configuration Wizards button.

2: Click Next on the introduction dialog.

3: In the X/Y list box, use the scroll bar

on the right to find the Leica Stage
entry. Click to select it.

4: In the Model list box, click to select

the Leica ISO Pro stage. Click Next.

Continued... 705

704 © 2011 Leica Microsystems (Switzerland) Limited

 Power Mosaic: Heerbrugg ISO Pro Stage: Setup:

1: For the Acceleration Ramp tables

click the No, do not re-calculate radio
button. LAS automatically handles the
motor power.

2: Most of the details of the data

exchange between Oasis and LAS
can be setup automatically so click to
select Yes (Recommended).

Make sure the stage is clear of any

obstructions and click the Finish button to
exit the wizard.

Continued...

LAS User Manual 705

Power Mosaic: Heerbrugg ISO Pro Stage: Settings:

On the Acquire Workflow:

1: ...if necessary click the PM (Power

Mosaic) tab to reveal the Optics
Settings panel. If the PM tab is not
visible the Power Mosaic module has
not been loaded: See Starting: Go
there... 623

2: Click on the Initialise and Set Speed

button.

3: On the Automation Settings dialog,

click on the Oasis Properties button.

4: When the Oasis Properties dialog

opens, click on the Stage tab.

Continued... 707

706 © 2011 Leica Microsystems (Switzerland) Limited

 Power Mosaic: Heerbrugg ISO Pro Stage: Settings (Continued):

On the Oasis Properties > Stage dialog:

1: Check that Encoders Fitted is set to

Yes. If it is not the cable connection
may be loose or not fitted, or the
wrong stage has been selected.
Repeat the Oasis Configuration
process: Go there... 703

2: Click to enable both of the Encoder X

and Y Axes check boxes.

3: Click Apply.

4: Click OK.

5: The ISO Pro stage will perform well in

both bi- and uni-direction modes, but
for scanning precision the uni-
direction mode is recommended.

6: Again, for accuracy and precision opt

for the Standard scan speed in most
situations.

Continued... 708

LAS User Manual 707

Power Mosaic: Heerbrugg ISO Pro Stage: Zoom, Focus & Calibration Procedure:

There are three essential steps to

successful imaging using the ISO Pro
stage with both motorised (Select the Mic1
tab: 1) and manual stands:

Zoom: (2)

Focus: (3) and

Calibrate: (4) by clicking on the Set

Calibration button and then on the
Calibrate button (5) on the Zoom
Calibration dialog. For more details:
Go there... 638

708 © 2011 Leica Microsystems (Switzerland) Limited

 Live & Interactive Measurements

Optional modules Live Measurements and Interactive Display or Hide parameters to suit the image.
Measurements bring all of the flexibility and precision of
Leica Application Suite to image measurement tasks. Live Comments to display alongside and enhance the
Measurements is optimised for working on the live image parameters.
before it is captured, and Interactive Measurements is
aimed at captured and stored images. Both modules are Configurations to save Classes and Settings together in
installed and licensed separately. a file that can be retrieved and applied to images at any
time.
Because many of the procedures - measuring distance,
area and so on - are common to both modules, Leica Templates to be used as moveable 'overlays' to
have created identical toolboxes and similar features that benchmark important positions and locations on live
will make moving from live to captured images quick and images.
seamless.
Display and analysis of selected measurements in the
The toolbox contains a wide range of precision tools correct corporate style with Microsoft Excel.
featuring: Snap to edge nearest to mouse cursor for improved
consistency and speed in identifying measurement
Configuration for line thickness, colour, label colour -
details on stored images.
including a transparency control - position and type font.
Classes to reflect similarities with colour-coding make
analysis much easier.
Alternative Parameters so, for example a circle can be
labeled with diameter, radius, area or circumference
whichever is more appropriate.

LAS User Manual 709

 Live & Interactive Measurements: The User Interface:

The main areas of the user interfaces:

1: Menu Bar: For Options and Help.

2: The Workflows: Live Measurements is on the Acquire Workflow and Interactive is on Analysis.
3: The Measure/Interactive Tab: Click to reveal all of the tools and features.
4: The Measurement Toolbox: Click to select measurement type.
5: The Control Tools: Commands for actions such as New Measurements, Redo and Undo.
6: Function Panels: Reveal Properties, Classes, Configuration, Report and Template controls.
7: The Acquire button: Click to capture an image and data.
8: Side Tool Bar: Hide and reveal the Grid (for Results) and Thumbnail Gallery.
9: Gallery Zoom: Move the slider to resize the thumbnails.
10: The Viewer: Displays the live or captured image.
11: Results Grid: Configure for Details or Summary of measurements.
12: Thumbnail Gallery.
13: The Image Status Display.
14: Status Bar.

710 © 2011 Leica Microsystems (Switzerland) Limited

 Live & Interactive Measurements: Feature Checkout:

Direct links to all of the Live and

Interactive Measurement features and
tools:

Launching the Modules: 714

Measurement Tools and the

Toolbox: 715

Control Tools: 726

Tool Properties: 729

Changing Colour: 730

Font Properties: 731

Creating Labels: 732

Classes: 733

Configurations: 737

Live Measurement Templates: 740

Interactive Measurements: Measure:

744
Selecting Results: 748

Create a Report: 750

Merging Images and Measurements:

758

Appendix: Parameter Descriptions:

759
Appendix: Keyboard Shortcuts: 760

LAS User Manual 711

Live & Interactive Measurements: Calibration and Preferences:

To maintain ongoing measurement

precision, calibration should be carried out
at regular intervals. The Calibration
functions can be found in the Acquire
Workflow in the Camera section. Go
there... 268

Measurement Precision:
The actual precision of LAS
measurements is governed by the
software and extends to 13 places after
the decimal point. To display that
precision would only clutter and confuse
the images and so the number of integers
displayed after the decimal point can be
set by the user to aid clarity. Reducing the
display places does not affect the
calculation precision* which is still carried
out internally using all 13 places.

To set the number of display places after

the decimal point:

1: On the main header click on Options

...

2: ...and from the drop down click to

select Preferences.

3: On the Preferences dialog, if

necessary, click on the Image tab.

4: On the Measurements Display panel,

in the Decimal Places text box, either
click in the box and type a value or
use the up/down arrows to the right of
the text box to increase/decrease the
value.

5: Click OK to save the value. Any

measurements being displayed will be
updated immediately.

*Precision in this context refers to the

number of decimal places saved with the
image and should not be confused with
'accuracy' that depends upon screen and
image resolution, calibration and several
other factors.

712 © 2011 Leica Microsystems (Switzerland) Limited

 Live & Interactive Measurements: Live Measurements: Fast Track:

Live Measurements Fast Track is a simple checklist of Add a comment if needed: Go there... 732
steps to take to get into precise measuring quickly.
Select the items to display on the Display Labels: Go
Measurements are made on a live camera image in there... 732
the Acquire Workflow - no need to capture the image
first. If required, save the settings as a Configuration: Go
there... 737
Carry out a calibration before making
measurements: Go there... 268 Start measuring...

Make the image as large as possible - close the Points to Note:

Gallery and Grid if they are open. Live Measurements cannot be used with sequence
images created with modules such as MultiTime or
Select an image Live Format that provides good Movie. If a sequence module is selected the Measure
resolution with a fast refresh rate: Go there... 241 tab is not available.

Determine if image measurements can be grouped If a Stereo microscope zoom is being used, the
together to make reporting and analysis quick and measurement drawings will change size but their
convenient. Set up Classes on the Measure tab to positions will be incorrect because of an image shift
reflect the groups: Go there... 733 due to the focus change. Use the AX-Carrier option to
correct.
If Classes are not being used, set up the Line,
Background and Font properties for initial Microsoft Excel does not have to be installed on the
measurements. They can be altered later if computer to capture results, but the Excel Viewer at
necessary. Go there... 729 least will be needed to view them, and a complete
Excel installation required if the template is to be
changed.

LAS User Manual 713

 Live & Interactive Measurements: Launching the Modules:

The Toolbox for both Interactive and Live Measurements The Live Measurement Toolbox resides on the Acquire
is the same and all of the tools work identically. The only Workflow:
obvious difference is the Merge control that works
differently on live and captured images. 4: Click on the Acquire Workflow.

The Interactive Measurements Toolbox is reached by: 5: Click on the Measure tab to reveal the Toolbox and
main panel.
1: Click on the Analysis Workflow.
6: The Merge feature is a check box which, when
2: If necessary, click on the Interactive tab to reveal the enabled merges measurements as the live image is
main panel and Toolbox. captured. After merging the measurements cannot
be changed.
3: The Merge button is a ‘one-shot’ function that merges Merging: Go there…
the measurements with the current captured image.
Once merged, the measurements cannot be altered:
Merging: Go there… Continued…

714 © 2011 Leica Microsystems (Switzerland) Limited

 Live & Interactive Measurements: Measurement Tools:

Each measurement tool - the top group in

the Toolbox - is selected by clicking the
appropriate button. Most tools have a
range of parameters than can be selected
by right-clicking the tool and choosing the
parameter from the menu (6). This will be
the parameter that is displayed on the
label. For example, Circle tools measure
Diameter, Radius, Area and
Circumference and any one may be
chosen to be displayed with the
appropriate result.

Some tools also have several modes. For

example, the Line tool can be used to
click and drag a line or click at the two
ends with a line automatically drawn
between, or short line in which the line is
split across narrow features. Where
various modes are available they appear
at the top of the menu (6) with the
parameters beneath (7).

1: The Selection tool. Used to select

and alter existing measurements or
de-select the other tools. Go there...
717

2: Distance Line. Measures the

distance in a straight line between 2
points. Also the line Angle and the
Width and Height of an imaginary
enclosing box - Bounding Box - from
the horizontal. Go there... 718

3: Segment Line: Measures the

distance around the Periphery of an
irregular shape, a Bounding Box
Height and Width and also the
distance between the two End
Points. Go there... 719

4: Area Tool: Measures the Area of an

enclosed figure as well as the
Periphery, individual Segments or
Bounding Box Height and Width. Go
there... 719

5: Angle Tool. Measures the Angle

between two drawn lines and
Bounding Box Height and Width. Go
there... 720

Continued... 716

LAS User Manual 715

Live & Interactive Measurements: Measurement Tools: Continued:

1: Circle and Ellipse: Draws a Circle

using either a straight line
representing the Diameter or Radius
or alternatively, drawn around Three
Points marking the Circumference.
Also draws an Ellipse. Go there... 720

2: Count: Counts each item clicked and

displays the count sequence number.
Go there... 723

3: Multiple Distance Line: Establishes

a base line or datum and then
measures distances from the line to
individual points as offsets or creates
lines parallel to the datum with the
distance between displayed. Go
there... 723

4: Cross: Measures both the Major and

Minor Axes of an object. Go there...
724

5: Triangle and Rectangle: Create

triangular or rectangular areas and
measure sides, enclosed area and
angles. Go there...

6: Hide Labels: Sometimes, especially

when making measurements that are
close together or very small, the
properties label can be obtrusive.
Hide the label temporarily by pressing
and holding down the AltGr key. The
label re-appears when the AltGr is
released.

716 © 2011 Leica Microsystems (Switzerland) Limited

 Live & Interactive Measurements: Selection Tool:

1: The Selection tool. Does not actually

draw but is used to select
measurements already made on the
image ready to edit or move them.
Click on the Selection Tool button and
then on the measurement, the end-
points of which will appear as small ‘
boxes’ or handles to indicate it is
selected (2).

Adjust a Measurement: Click and

drag on a handle.

Delete: Press the Delete key or click

on the Trash Can (3) to remove a
selected measurement.

Undo/Redo Actions: Use the Undo

button to restore the last deletion.

Re-position: Click and drag on a

measurement line (not a handle) to
re-position it. The Label follows.

Re-position Label: Click and drag on

a Label to re-position it independently
of the measurement.

Multiple Selection: Click and drag on

the image (not a measurement) to
draw an enclosing box or 'marquee'
around any number of measurements
to simultaneously select them.

Continued... 718

LAS User Manual 717

Live & Interactive Measurements: Distance Line Tool:

1: Distance Line. The distance in a

straight line between 2 points.

Option Draw: Click and hold down at

the start point.
Drag to the end point and release the
mouse button.

Option Two Point: Click at the starting

point, move the cursor to the end
point and click again.

Option Short Line: For situations in

which the feature to be measured is
so small a continuous line would be
difficult to read and draw. Click on
one side of the feature and holding
down the mouse button drag to the
opposite side of the feature. Can be
dragged to any angle (3).

Measurements Options: Length,

Angle and Bounding Box Width and
Height (2).

Continued... 719

718 © 2011 Leica Microsystems (Switzerland) Limited

 Live & Interactive Measurements: Segment and Area Tools:

1: Segment Line: To measure the

distance around the periphery of an
irregular shape.
Click and release to start (3).
Click to end the segment and start
another.
Repeat around the periphery.
Right click to end. The sum of all the
segments is displayed.
Alternatively, click, hold and drag for a
continuous line that tracks the mouse
path.

Measurement Options: Length,

Bounding Box Width and Height, the
distance between the two End Points
(2).

4: Area Tool: Measures the area of an

enclosed figure.
Click and release to begin (6).
Move the cursor to the next vertex
and click.
Repeat for all sides and back at the
start point.
Right click to end.
Alternatively, click, hold and drag for a
continuous line that tracks the mouse
path.

Measurement Options: Area,

Perimeter, Bounding Box Width and
Height (5).

Note: If the drawn shape pulsates

when complete, some points on the
outline cross. These must be
removed to give a correct area
measurement.

Continued... 720

LAS User Manual 719

Live & Interactive Measurements: Angle Tool:

Angle Tool. Measures the angle

between two drawn lines.
For Baseline (1) and Apex (2) angles:
Click to set the start point of the first
'leg'.
Move the cursor to the end-point of
the first line and click.
Move the cursor to the end-point of
the second 'leg' and click.
Right click to end.
Baseline represents the angle
between a horizontal or 'base' line
and an extension (1).
Apex is the angle between the two
legs irrespective of the inclination (2).

A Four Point Angle (3) is that between

two lines that are not connected at a
point. Furthermore, the lines
projected intersection could be
beyond the limits of the image.
Click on the starting point of the first
line.
Move the cursor to the end of the first
line and click again. The line is drawn
(3).
Click on the starting point of the
second line.
Move the cursor to the end of the
second line and click again. The line
is drawn (4).
Dotted lines are drawn between the
ends of the lines.

If the lines are nearly parallel an

imaginary point of intersection will be
beyond the limits of the image or even
the Viewer, but the software can
accommodate this situation and
return an accurate angle.
If the lines are parallel then the angle
will be 0°.

Measurement Options: Angle,

Bounding Box Width and Height.

Continued... 721

720 © 2011 Leica Microsystems (Switzerland) Limited

 Live & Interactive Measurements: Circle and Ellipse Tools:

Circle: Draws a Circle or an Ellipse.

Using Three Points: Click on a point
on the image (1)
Click on a second point on the image
(2).
Click on a third point (3) and drag the
handle to describe the diameter
required.

Using Diameter: Click and hold down

on the first edge-point of the circle (4).
Drag to the second edge point
representing the diameter and
release the mouse button.

Using Radius (5): Click on a point on

the image representing the centre of
the circle.
Release the mouse button.

Drawing an Ellipse): Click on a point

on the image (6).
Click on a second point (7)
representing either the major or minor
axis of the Ellipse and still holding
down the mouse button drag to
describe the other axis.
Click to complete.

Continued... 722

LAS User Manual 721

Live & Interactive Measurements: Dual Circle Tool:

Dual Circle: Draws 2 circles using

either the three-point, radius or
diameter methods and provides a
range of measurements.
Using the method preciously
described, draw a circle around the
first feature (1) and the around the
second feature (2).

Options: The Area, Width, Height etc

parameters refer to the first circle.
The parameters with the '2' suffix (for
example Radius2) refer to the second
circle. Distance Between Centres is
the distance between the centres of
the first and second circles. Closest
Approch is the smallest distance
between the two perimeters. The
Centre X and Centre Y dimensions
are the X (Horizontal) and Y
(Verttical) distances from the top/left
corner of the image.

Continued... 723

722 © 2011 Leica Microsystems (Switzerland) Limited

 Live & Interactive Measurements: Count and Multiple Distance Tools:

1: Count: Counts each item clicked and

displays the count sequence:

Click on each of the items to be

counted (2). The count value
automatically increments and a small
‘target’ is drawn over the clicked
point. Individual ‘target’ and labels
can be moved for clarity.

3: Multiple Distance Line: Using a

single datum line and then measuring
the distance of objects from that line:
either by individual offset lines or
additional lines parallel to the datum.

Multiple Distance Line (4): Click on

the image to establish the datum line
starting position (5).
Click again to anchor it.
Move the cursor to the distance to be
measured left or right, top or bottom
of the datum.
Click on the end point of the distance
measurement.
Repeat the process for subsequent
lines ending with a right click (6).

Parallel Distance Lines: Click on the

image to establish the datum line.
Click again to anchor it:
Move the cursor away from the datum
line to a position to be measured and
click to draw the parallel line.
Repeat as required ending with a right
click.

Measurement Options: Length, either

as the length of an offset line or the
distance between the datum and a
parallel line.

Continued... 724

LAS User Manual 723

Live & Interactive Measurements: Cross Tool:

1: Cross: This tool measures both the

major and minor axes of an object by
drawing a cross over it. The Cross
may be drawn at any angle.

Major Axis Option: Measures the

major (longest) axis.

Minor Axis Option: Measures the

minor (shortest) axis: Click on an
edge of the area to be measured (2),
and holding the mouse button down
drag to the opposite side (3).
The selected dimension is displayed.
Drag the Cross at an angle if
necessary.

Measurement Options: Angle

measures the skew angle from the
horizontal, Bounding Box Width and
Height.

To adjust axis lengths, click on the

Selection tool and then on the Cross
to reveal handles for re-sizing and
rotating. Click, hold and drag in the
centre to re-position.

Continued... 725

724 © 2011 Leica Microsystems (Switzerland) Limited

 Live & Interactive Measurements: Triangle and Rectangle Tools:

Triangle and Rectangle: Draws an

enclosed 3 or 4-sided polygon.
Click on the starting point of the
polygon.
Drag to the required side dimension
and click again.
Drag to create the required
dimensions.

To modify the polygon:

Click to choose the Select tool.
Click on the triangle to select it. Small
handles appear at the vertices.
Click on handle and drag to change
the dimensions.

Measurement Options: Perimeter, Area,

Width at the greatest horizontal
dimension, Height at the greatest
vertical dimension. Centre X and Y
the horizontal and vertical distances
from the top/left corner of the image
to the centre of the polygon.

LAS User Manual 725

Live & Interactive Measurements: Control Tools:

The lower set of tools are utilities used to

start new measurements, make multiple
deletes and move back and forth between
actions.

1: Start a new set of measurements:

Deletes all existing measurements –
confirm the deletion – ready to start a
new set of measurements. Because
this action also clears the Undo/Redo
history, it cannot be reversed.

2: Delete the selected measurements:

Click the icon to delete the existing
and selected measurements.
Measurements can be selected
individually from the Grid or on the
image, as a group by holding down
the CTRL key and clicking them on
the Grid or image, and by using the
Select All tool (3).

3: Select all measurements: Selects all

measurements on the image.

4: Hide/Reveal all measurements:

This is a toggle action – click once to 6: Undo the last action and Redo the last action after an Undo.
hide all of the measurements and Individual measurements or points can be deleted or restored using
click again to reveal them. the Undo and Redo button. Hover the cursor over the button to
determine the action that will occur.
5: Hide/Reveal selected objects: Use
this tool to hide or reveal a group of 7: Toggle Cursor Colour: Each click on the button will switch the
selected objects. Objects can be cursor between black and white. Right click on the button to set the
selected as a group by holding down cursor to a user-defined colour: Go there… 730
the CTRL key on the keyboard whilst
clicking the objects on the Grid or the 8: Snap to Edge: The Snap tool helps the user to make fast, precise
image, or by using the Selection tool, measurements by automatically 'snapping' to a close, well
clicking on the image and drawing a contrasted edge: Go there... 728
'box' or marquee around the
measurements to be selected.

726 © 2011 Leica Microsystems (Switzerland) Limited

 Live & Interactive Measurements: Control Zoom and Side Bar Tools:

The Zoom feature allows parts of the

image to be enlarged to achieve even
higher levels of measurement accuracy.

1: Position the cursor close to the area

of interest.

2: Press and hold down the Shift key. A

Zoom window opens and will follow
the cursor around the image.
Release the Shift to key to close the
Zoom window.
The zoom level within the Zoom
window can be increased or
decreased by turning th mouse wheel.

3: The overall image zoom can be

increased or decreased by holding
down the keyboard Control (Ctrl)
and...

4: ...turning the mouse wheel.

5: The Results Grid can be revealed or

hidden by clicking the button on the
side tool bar to the right of the Viewer.
This is a toggle action - click once
and the Grid is revealed, click again
and it is hidden.

6: The Thumbnail Gallery can also be

hidden and revealed by clicking the
button on the side tool bar. Again, this
is a toggle action button.

LAS User Manual 727

Live & Interactive Measurements: The Snap Tool: Interactive only:

The Snap tool (not available with Live

Measurements) allows the measuring
tools - Distance, Circle, Parallel, Angle
and Cross - to precisely 'snap' to adjacent
sharp edges on the image.

1: Click on the Snap button to enable

snapping.

2: Click to select the required

measurement tool. The advancing
end-point of the tool displays a small
'X' in the Snap chosen colour.
Carry out the measurement using the
techniques described earlier, but as
the measurement line is extended,
the advancing end-point will detect
and snap to an edge as it get close.

3: Change the Snap properties by right-

clicking on the Snap tool and then
left-clicking the pop-up 'Set Options'
button.

4: On the Options dialog, augment the

advancing end-pont with an easy-to-
detect Patch Box enclosing the Snap
cursor.

5: Increase or decrease the size

(maximum 50) of the Patch Box by
clicking the up/down arrows. The box
size determines the area that is
'searched' for a sharp edge.
If the box is too small edges may be
difficult to detect; Too large and
several edges may prove to be snap
targets. Adjust the Patch Box size to
suit the image complexity.

6: Change the Patch Box, Predicted

Line and 'X' cursor colour by clicking
on the Colour button and from the
Colour Dialog selecting a new colour.

7: Enable the Predicted Edge Line by 8: Click the OK button.

clicking the check box. A line appears
within the Patch Box and it rotates to 9: Use the Zoom feature (hold down the Shift key) to enlarge the
indicate the direction of the nearest image area within the Patch Box and aid precision.
edge. Bring the Patch Box close to a
circle and the Edge Line appears as a 10: Hold down the keyboard 'S' key to temporarily disable snapping
tangent, faithfully following the curve and release the key to restore it. This overrides the Snap feature
of the circle. allowing the user to select and mark the target point.

728 © 2011 Leica Microsystems (Switzerland) Limited

 Live & Interactive Measurements: Properties: Line Thickness and Colour:

The Properties Panel provides all the

facilities for tailoring the appearance of
measurements to suit the user.

Change Line Thickness and Colour, the

Font, its Style, Size, Colour and
Background. Select the Parameters to be
displayed on the labels and add
Comments to them.

Set the Line Thickness and Colour:

1: Expand the Properties panel by
clicking on the arrow to the right of
the header.

2: Change the Line Thickness by

clicking on the Up/Down (Increase/
Decrease) arrows to the right of the
Width text box.

3: Change the Line Colour by clicking on

the Colour button and on the Select
Colour dialog choosing a new colour:
Go there… 730

4: Change the Fill for closed shapes by

clicking on the Background button
and on the Select Colour dialog,
choosing a new colour: Go there…
730

Continued… 731

LAS User Manual 729

Live & Interactive Measurements: Properties: Change a Colour:

The colour of lines, text and fills can be

changed to user preferences. The
procedure is the same for Line, Font,
Background or Class colours.

1: On the Select Colour dialog select a

new colour by…

2: …clicking on a swatch, …

3: …clicking and dragging the small

‘target’ on the Colour Wheel, or …

4: ….by clicking in the red, green and

blue text boxes and typing a value in
the range 0 to 255.

5: If necessary, adjust the shade by

clicking and dragging the slider on
the Shade Bar.

6: Colour transparency can be set by

clicking and dragging the slider on
the Transparency Bar or…

7: …clicking in the Transparency text

box and typing a value in the range
255 for a solid colour to 0 for
complete transparency which, when
used with an outline will display an
‘open’ shape with no apparent fill. The
circle on the right in the diagram has
a transparency setting of 103.

8: Click OK.

730 © 2011 Leica Microsystems (Switzerland) Limited

 Live & Interactive Measurements: Font Properties:

Change the Font Type Face, Style, Size,

Colour and Background.

Select the Font, Style and Size:

1: Expand the Properties panel by
clicking on the small arrow to the right
of the panel header.

2: Click on the Font button.

3: On the Font dialog, select the Font

Type Face,…

4: …the Font Style – bold, italic etc,

and…

5: …the Size in points.

6: Click OK.

Change the Font colour:

7: Click on the Colour button and on the
Select Colour dialog choose a new
colour: Go there… 730

Change the Font Background colour:

8: Click on the Background button and
on the Select Colour dialog choose a
new colour: Go there… 730

LAS User Manual 731

Live & Interactive Measurements: Comments and Label Parameters:

1: Click on the small arrow to the right of

the Properties header to reveal the
Properties panel.

Adding Comments:
Comments are often a useful addition to a
measurement and can be added by:

2: Click in the Comments text box and

type a comment or note.

Displaying Label Parameters:

In the Display Labels window click to
select an entry so that it will displayed as
part of the object label. A tick mark
appears to the left of the selection. Click
again to hide the property – the tick mark
disappears.

2: Comment: A comment or note

entered in the Comment text box.

3: Number: Is a sequential
measurement number.

4: Parameter: Shows the tool value –

Length, Area, Angle etc.

5: Value: The actual measurement.

6: Unit: The measurement units chosen

on Camera > Calibration – mm for
example. Go there… 268

7: Class: The Class Name if one has

been selected. About Classes: Go
there... 731

732 © 2011 Leica Microsystems (Switzerland) Limited

 Live & Interactive Measurements: Classes:

Classes represent a quick and simple

method of grouping items to distinguish
them from other parts of the image.
Classes are saved globally but are
grouped and saved in configurations. A
configuration can be attached to an image
and that same configuration used for all of
the images in an experiment.

Apart from visual colour coding on the

image, classes also group measurements
together on result reports making reading
and analysis far easier. Any or all of the
measurement tools can be used within a
class.

This example shows part of an electronic

printed circuit board:

1: The required class is selected by

clicking it. The class name also
appears in the Classes panel header.

2: The Area tool has been used to

highlight the small diodes. The outline
colour, thickness and the area fill can
be defined for a class as well as the
font style, size and colour. On the
illustration sequential numbering has
been enabled and appears as labels
alongside each component, one of
many description options that can be
turned on or off and saved with the
image.

3: In this illustration all of the 5 classes

have been used to highlight and
colour code different components.

4: A combination of Class Name and

Measurement Number can quickly
identify a component on a Results
Report, another integral part of the
Live and Interactive Measurement
modules.

Continued… 734

LAS User Manual 733

Live & Interactive Measurements: Create a New Class:

If necessary, click on the arrow to the right

of the Classes header to reveal the
Classes panel.

1: Click on the New Class button and…

2: …on the Create New Class dialog

type a name for the class.

3: The line colour for the class can be

set by clicking the Line Colour window
and on the Select Colour dialog
choosing a new colour: Go there…
730

4: Click OK.

5: The new class name must be unique.

If it is not a warning appears and the
name is not accepted unless it is
removed from the Master Class List:
Go there...

6: The new Class Name appears in the

Classes window with its selected
colour shown to the left.

Continued… 735

734 © 2011 Leica Microsystems (Switzerland) Limited

 Live & Interactive Measurements: Edit and Delete a Class:

Edit Class:
To edit the Class Name and/or Line
Colour:

1: Click to select the Class to be edited.

2: Click on the Edit button.

3: On the Edit Selected Class dialog,

click to select the existing name and
type a new name or click in the Line
Colour window and choose a new
colour from the Select Color dialog:
Go there... 730

4: Click the OK button.

5: The edited Class appears in the

Classes window.

Continued....

LAS User Manual 735

Live & Interactive Measurements: Attach/Detach/Delete a Class:

All class descriptions are stored in a single

file called the Master Class File. Classes
may be retrieved from the file (Attach) to
serve in new projects and they can be
moved back into the file if they are no
longer required (Detach). A class can be
shared among any number of projects.

Attach/Detach a Class:
1: Click on the 'paper clip' Attach/Detach
button.

2: On the Attach/Detach dialog all of the

classes in the Master File are listed in
the left-hand pane and those in the
current project in the right-hand pane.
Attach a class by clicking on it in the
left-hand pane and then clicking the
right arrow.

3: Attach all classes by clicking on the

right double-arrow.

4: Detach a class by clicking to select it

in the right-hand pane and then
clicking the left arrow.

5: Detach all classes by clicking the left

double-arrow.

6: Click the OK button.

Delete a Class:
7: Delete a class from the Master File by
clicking the 'Trash Can' Delete button.

8: The Choose Class Action dialog

allows the user either to simply
detach the class and move it back
into the Master File or...

9: ...removing it from the Master File

completely.

736 © 2011 Leica Microsystems (Switzerland) Limited

 Live & Interactive Measurements: Configurations:

The current Classes and properties can

be saved as a Configuration file
associated with the image, and retrieved
to be used at a later date.

1: Each Configuration has a unique

name and can be accessed by
clicking on the arrow to the right of
the Current Configuration window and
from the drop down list…

2: …clicking to select the required

configuration.
Two configurations are provided with
Leica Application Suite – Default,
which are ‘factory’ settings that can
be subsequently changed in
Preferences, and Last Used which, as
its name implies, re-loads the
configuration that was last being used
and had been saved - that could be
the Default.

3: To save the current settings as a new

configuration, click on the New button
and…

4: …type a unique name for the new

configuration.

5: Click OK to save the setting.

6: The new configuration appears in the

drop down list.

Continued… 738

LAS User Manual 737

Live & Interactive Measurements: Edit and Delete a Configuration:

To Edit a Configuration Name:

1: Select the configuration to be
changed from the drop down list.

2: Click on the Edit Configuration button.

3: On the Edit Configuration dialog,

change the name by clicking in the
text box and typing a new name
and…

4: …clicking OK.

To Delete a Configuration:
5: Select the configuration to be deleted
from the drop down list.

6: Click the Delete (Trash Can) button.

7: Confirm the deletion and the

Configuration will be removed
permanently. The operation cannot be
reversed.

Continued… 739

738 © 2011 Leica Microsystems (Switzerland) Limited

 Live & Interactive Measurements: Attach and Detach Classes:

All classes are saved in a Master Class

File that makes re-use in different
configurations simple and fast.

1: Select the configuration to which

classes will be attached or detached
from the drop down list.

2: Click on the Attach/Detach 'paper clip'

button.

3: The dialog shows all available

classes from the Master Class file in
the left hand pane (4) and all of the
classes (if any) currently attached to
the configuration in the right hand
pane (5).

To attach a Class to the Configuration:

6: Click the Class entry and...

7: ...click on the Attach Selected button.

The Class appears in the right hand
window (8).

To Detach a Class from the

Configuration:
8: Click the Class to be detached in the
right-hand pane.

9: Click on the Detach Selected button.

The Class is removed from the right-
hand pane and appears again in the
left-hand pane.

Attach/Detach All Classes:

10: Click on the Attach All button to
attach all of the classes to the
configuration.

11: Click on the Detach All button to

detach all of the classes in the
configuration.
Click OK.

LAS User Manual 739

Live & Interactive Measurements: Live Measurements Templates: Create:

A 'live' Template is a measurement or a

collection of measurements used as a ‘
standard’ that can be overlaid on a live
image to check for conformity.

The illustration (1) represents a ‘perfect’

product and boundary lines have been
drawn around three components to map
their precise relationship to each other.
These measurements are saved as a
Template, retrieved at any time and
placed over a live image to check that it
conforms to tolerances for example.
Templates are saved as discreet files and
are not associated with a specific image.

Having created the measurements:

2: If necessary, click the arrows to the
right of the Template header to reveal
the panel.

3: Click on the Create New Template

button and…

4: …on the dialog give the Template a

unique name.

5: Click OK.

Continued…

740 © 2011 Leica Microsystems (Switzerland) Limited

 Live & Interactive Measurements: Retrieve, Apply and Remove a Template Overlay:

To Retrieve and Apply a Template:

1: Click on the arrows to the right of the
Template drop down and click to
select a template.

2: Click on the Template Apply button

and the template will appear as an
overlay to the live image.
Alternatively, double click on the entry
name.

Remove a Live Image Overlay:

3: Remove an overlay by clicking the
Delete (Trash Can) button and on the
Confirm Delete dialog clicking Yes.

Continued... 742

LAS User Manual 741

Live & Interactive Measurements: Export a Template:

The Template Export facility allows a

template to be saved in a location other
than the default Measurement Templates
folder.

To export a template to a selected folder,

ensure that the template to be exported is
highlighted in the Templates Window:

1: Click on the Export button.

2: On the Browse for Folder dialog,

navigate to the folder to which the
template will be exported.

3: There is a default folder created when

LAS is installed called Measurement
Templates, and is the recommended
folder for all templates.

4: If required, create a new folder by

clicking the Make New Folder button.

5: Click OK.

6: The Export Template Report confirms

that the template has exported
successfully (or not).

Continued... 743

742 © 2011 Leica Microsystems (Switzerland) Limited

 Live & Interactive Measurements: Import a Template:

Templates conforming to the LAS

configuration (.tem file extension) can be
imported from any folder.

1: Click on the Import Template button.

2: On the Open dialog...

3: ...navigate to the source folder and

click to select the template.

4: Click on the Open button.

5: The template is imported and

success noted on the Import
Template Report..

6: ...with the template appearing in the

Templates window.

Although a template is a separate file, a

copy of it as an overlay can be saved with
a captured image by merging it into the
image: Go there…

LAS User Manual 743

Live & Interactive Measurements: Interactive Measurements: Measurements:

Measurements provides a simple method

of creating a range of measurement
overlays for a single, captured image.

1: Click on the arrow to the right of the

Measurements header to reveal the
panel.

2: The Default entry represents the

original image.

3: Make a measurement on the image.

4: Click on the New Measurement

button and…

5: …on the Create New Measurement

dialog type a unique name for the
measurement.

6: Click OK.

7: The new measurement name

appears in the window.

8: Make another measurement – or

delete the first and then make
another. Click on the New
Measurement button (4).

9: Type a name for the new

measurement and…

10: …click OK.

Continued… 745

744 © 2011 Leica Microsystems (Switzerland) Limited

 Live & Interactive Measurements: Interactive Measurements: Measurements: Continued:

1: Continue to add or change

measurements giving each version a
new name.

2: retrieve a measurement by clicking

on its name in the window and then…

3: …clicking on the Load button and…

4: …the measurement is retrieved and

placed in position over the image.

5: Edit a measurement by adding, re-

positioning or deleting measurements
and then clicking the Save button.

6: Delete a measurement by clicking on

its name to select it and then on the
Trash Can (Delete) button.

LAS User Manual 745

Live & Interactive Measurements: Select Results using Details:

The Select Results function displays an

analysis of all the measurements taken on
an image and presents them as:

Details: Each measurement is shown

with all its parameters (or those that are
selected in the Configuration), or

Summary: The measurements are

treated as a complete set of data
tabulated statistically.

Both options present the information as a

grid arranged below the image. The
information can be later edited, deleted or
attached to the image.

If necessary, click on the small arrow to

the right of the Select Results panel.

Grid Setup for Details:

1: Click to enable the Details button.

2: Click the Configure button.

3: On the Configure Details Grid

dialog...

4: ...click the Show All button to enable

all of the parameters or...

5: ...click the Hide All button to disable

all of the parameters.

6: Individual parameters can then be

enabled or disabled.

7: Click OK.

8: Those parameters that have been

enabled will appear as a heading on
the Grid with the appropriate results
beneath. If a chosen parameter is not
applicable to a measurement - for
*
When changes are made to the Grid either in Details or Summary, the
instance, Line Length is not a
parameter of a 3-Point Circle - then warning ! flashes. Save the new Grid Layout as a configuration: Go
the value is reported as a dash (-). there... 748

Continued... 747

746 © 2011 Leica Microsystems (Switzerland) Limited

 Live & Interactive Measurements: Select Results using Summary:

To display the results using the Summary

information:

1: Click to enable the Summary button.

2: Click the Configure button.

3: On the Configure Summary Grid

dialog...

4: ...click the Show All button to enable

all of the parameters or...

5: ...click the Hide All button to disable

all of the parameters.

6: Individual parameters can then be

enabled or disabled.

7: Click OK.

8: Those parameters that have been

enabled will appear as a heading on
the grid with the appropriate results
beneath. If a chosen parameter is not
applicable to a measurement - for
instance, Line Length is not a
parameter of a 3-Point Circle - then
the value is reported as a dash (-).

9: Hide/Reveal the Results Grid by

clicking the Grid button on the Side
Tool Bar.

LAS User Manual 747

Live & Interactive Measurements: Select a Class & Save the Grid:

The results can be configured further by

selecting if only a specified class is to be
displayed or alternatively no classes or all
classes.

To select the Class configuration:

1: Click on the arrows to the right of the
Class header and…

2: …from the drop down list click to

select None, All Classes or an
individual Class.

Save the Grid Configuration:

To save the Grid settings as a
Configuration that can be retrieved and
used later:

3: Click on the New Grid Configuration

button.

4: Click inside the Name text box and

type a unique name for the
configuration.

5: Click OK.

6: To see the new configuration in the

list, click the small arrow to the right
of the Current Grid Layout header to
reveal the list.

Continued… 746

748 © 2011 Leica Microsystems (Switzerland) Limited

 Live & Interactive Measurements: Grid Configuration Edit, Delete & Save:

Edit a Results Grid Configuration:

1: Click on the Edit button.

2: On the Edit Configuration panel, click

inside the Name text box and type a
new, unique name for the
configuration.

3: Click OK.

Delete a Results Grid Configuration:

4: Click on the 'Trash Can' Delete
button.

5: Confirm or abort the the deletion. If a

configuration is deleted it cannot be
retrieved.

Save the Results Grid Configuration

with the Image:
6: Click on the Save button. This will
attach a copy of the Results Grid
Configuration to the image so that
when it is opened again in
Measurements the configuration will
be loaded automatically.

LAS User Manual 749

Live & Interactive Measurements: Create Report:

A report collects the measurement

information and stores it in a readable
format – either to be displayed in Microsoft
Excel or as a Comma Separated Value
(CSV) file. CSV files are stored as plain
text and are very compact, making them
ideal as e-mail attachments. They can
also be used in a wide range of text
processing applications.

The parameters that are exported reflect

those chosen to display in Select Results.
Go there... 748

There are a few differences between Live

and Interactive Measurements detailed on
the following pages.

1: A typical Live Measurement image

with a range of measurement tools
being used.

2: Click on the arrows to the right of the

Create Report header to reveal the
panel.

Live Measurements Panel:

3, 4 and 5: Three Field text boxes –
Specimen, Description and
Observations – are provided as
optional headers for the
measurement file. They do not have
to contain information but to add text
to them simply click in a text box and
type.

Interactive Measurements Panel:

6, 7 and 8: The Field text boxes are the
same as Live Measurements and if
they were completed when the image
was captured they will be displayed
automatically.

Continued… 751

750 © 2011 Leica Microsystems (Switzerland) Limited

 Live & Interactive Measurements: Create an Excel Report:

1: Click on the arrows to the right of the

Format header to reveal the output
options.

2: Select Excel from the drop down

menu.

3: The report content can be Details –

the actual measurements, classes
and results, or…

4: …Summary that is a detailed analysis

of the measurements.

...or both Details and Summary

combined in a single Excel book.

5: To add the report data to an existing

Excel spreadsheet as a separate
page, click the Append button. This
arrangement allows results to be
analysed individually or as a group.

Click the Export button (8).

Interactive Measurements Options:

6: The report can be attached as a file
to the image record by enabling the
Attach to Record check box.

7: Enable the Original Image check box

to include the original image in the
report and…

8: …select Result Image to include the

image together with the overlaid
measurements.

9: Click the Export button.

Continued... 752

LAS User Manual 751

Live & Interactive Measurements: Create an Excel Report: Continued:

1: On the Export dialog, click the browse

button and…

2: …navigate to a folder on the Select a

Report File dialog, …

3: …give the file a name – the proper

file extension (.xls) is automatically
added and...

4: …click the Open button.

5: To use a template to lay out the

report, click to enable the Template
check box and…

6: …navigate to the Template folder. On

first use this is automatic for Leica
Excel Templates that have been
specially created for fast, efficient
reporting and can be easily modified
to suit users’ precise requirements. If
a template is not used images are not
exported.

7: To display the file on screen after the

report is complete, click to enable the
Display Report button.

8: Click Export and the file will be

written. If the Display Report check
box is enabled the appropriate
application (if installed) will be
launched with the information
displayed in it.

Continued… 753

752 © 2011 Leica Microsystems (Switzerland) Limited

 Live & Interactive Measurements: Create an Excel Report Examples:

1: The Excel Front Page with Summary: The image 2: The Statistics sheet. If Summary is not included then
appears only with Interactive Measurements.. the lower part of the sheet - the statistics - will be
blank. See Appendix for explanations of the
parameters: Go there... 759

LAS User Manual 753

Live & Interactive Measurements: Create an Excel Report Examples: Continued:

1: An Excel Details sheet lists all of the measurements 2: The Excel Images sheet. The images only appear
and chosen parameters. with Interactive Measurements- on the left the image
without measurements and on the right with the
measurements overlaid.

754 © 2011 Leica Microsystems (Switzerland) Limited

 Live & Interactive Measurements: Create a CSV Report:

1: Click on the arrows to the right of the

Format header and select CSV from
the options.

2: The Content selection differs from

the Excel options in that Details or
Summary can be selected but not
both. Click on the content required.

Interactive Measurements Option:

3: Interactive Measurements has the
option to attach the CSV report to the
image record. Click to enable the
Attach to Record check box.

4: Click the Export button.

5: On the Export dialog, click on the

browse button and…

6: …on the Select a Report File dialog

navigate to a folder in which to store
the report.

7: Give the report a name and…

8: …click Open.

9: Click Export.

Continued… 756

LAS User Manual 755

Live & Interactive Measurements: Create a CSV Report: Example:

1: Example of a report based on a Comma Separated

Values (CSV) output. CSV output is a single page,
very compact report ideal for data transmission. Here
the data is displayed in a Microsoft Excel format.

756 © 2011 Leica Microsystems (Switzerland) Limited

 Live & Interactive Measurements: Retrieve a Report:

To 756 retrieve a report, when using the

optional modules LAS Archive, previously
attached to an image:

1: Click on the Browse Workflow and...

2: ...open the image archive.

3: Click on the Gallery thumbnail to

select and display the image.

4: Expand the Attachments panel by

clicking on the small arrow to the
right.

5: Attached reports are listed in the File

drop-down. Click to select the
required report and...

6: ...click the Open button to start

Microsoft Excel and display the report
spreadsheet.

LAS User Manual 757

Live & Interactive Measurements: Merging:

Merging is the process of combining

measurements with an image so that they
become a single entity.

Measurements merged with an image

cannot be edited and altered.

The Merge control differs in appearance

and function between Live and Interactive
Measurements.

Live Measurements Merge:

1: Clicking to enable the Merge check
box will merge measurements with
images as they are captured whilst it
is enabled.

2: Click on the Acquire Image button.

Interactive Measurements Merge:

3: The Interactive Merge is a ‘one shot’
button – click it to merge
measurements with the captured
image.

758 © 2011 Leica Microsystems (Switzerland) Limited

 Live & Interactive Measurements: Appendix: Parameters:

Total: Is the total for all tools beneath that heading. For Standard Error: Uses the convention Standard
example, Total Line Length is the total measurement Deviation/Square root(n) where ‘n’ is the number of
for all the tools - Distance and Line - that have length measurements made or Total Count.
as a parameter.
Confidence Interval Lower: The lower range of
Mean: Represents the Total divided by the number of parameter values within which 95% of parameter
measurements made (Total/Total Count). values are likely to fall. It is based upon the
assumption that the values are normally distributed
Mode: The most frequently occurring parameter value with a mean of the same value as Mean.
from measurements using this parameter. If multiple
modes exist, that with the lowest value is displayed. Confidence Interval Upper: The upper range of
parameter values within which 95% of parameter
Median: The actual mid-value in a list of values. For values are likely to fall. It is based upon the
example 676 is the Median of 214, 676 and 1031. For assumption that the values are normally distributed
an even numbered list of values, the values either with a mean of the same value as Mean.
side of the mid-point are averaged.
Total Count: The number of measurements made with
Maximum and Minimum: The largest and smallest this parameter.
measurements made regardless of the tool used.
Total Image Area: Is the area (selected units^2) of the
Standard Deviation: A measure of the spread of a entire image.
parameter values from the measurements using that
parameter. It is based upon a random sample taken
from the values.

LAS User Manual 759

Live & Interactive Measurements: Keyboard Shortcuts:

When the Results Grid is selected, a

range of Shortcut Key combinations are
available.

For all Shortcuts, press and hold down

the Control (Ctrl) key and at the same
time press the auxiliary key:

Ctrl + A: Selects all of the

measurements.

Ctrl + D: Deselects the measurements.

Ctrl + Del(Delete): Deletes all of the

selected measurements.

Ctrl + C: Copies any selected text to the

Windows clipboard. It can then be
pasted to another application by using
Ctrl + V.

For a complete list of all the LAS

Keyboard Shortcuts: Go there... 22

760 © 2011 Leica Microsystems (Switzerland) Limited

 Appendix: Tool Parameters: Line, Segment Line and Area Tools:

All distances are calculated from the

centre of the enclosing line:

Line:
1: Length: Overall length of the drawn
line measured to the centre of the line
ending strokes.

2 & 3: Width and Height: The horizontal

and vertical measurements of a
bounding box enclosing the line.

4: Angle: The angle swept between the

line and the horizontal.

Segment Line:
5: Length: The total length of all the
segments added together.

6 & 7: Width and Height: The

dimensions of a bounding box
enclosing the figure.

8: End Distance: The distance between

the two end points of the figure.

Area:
9: Area: The enclosed space returned in
the selected measurement squared
(for example mm²).

10: Perimeter: The distance around the

figure.

11 & 12: Width and Height: The

horizontal and vertical measurements
of a bounding box enclosing the
figure.

Continued... 762

LAS User Manual 761

Appendix: Tool Parameters: Circle and Ellipse:

Circle:
All distances are calculated from the
centre of the enclosing line:

1: Diameter: Distance across the circle

in any direction.

2: Perimeter: The distance around the

circle.

3: Radius: The distance from the centre

of the circle to the centre of the
enclosing line.

4: Area: The enclosed space returned in

the selected measurement squared
(for example mm²).

5: Width: Distance across the circle

horizontally.

6: Height: Distance across the circle

vertically.

Width and Height are the same as

Diameter but are retained because the
circle is part of the ellipse class that will
have definite Width and Height
dimensions

Ellipse:
7: Angle: Angle between the horizontal
and the Major Axis.

8 & 9: Width and Height: Horizontal and

vertical sides of a bounding box
enclosing the Ellipse.

10: Major Axis: The greatest dimension.

11: Minor Axis: The smallest dimension.

Continued... 763

762 © 2011 Leica Microsystems (Switzerland) Limited

 Appendix: Tool Parameters: Dual Circle:

All distances are calculated from the

centre of the enclosing line. Parameters
without the '2' suffix refer to the first circle
drawn and those with the '2' suffix to the
second circle.

1: Diameter: Distance across the circle

in any direction.
Width and Height are the same as
Diameter but are retained because
the circle is part of the ellipse class
that will have definite Width and
Height dimensions

2: Radius: The distance from the centre

of the circle to the centre of the
enclosing line.

3: Perimeter: The distance around the

circle.

4: Area: The enclosed space returned in

the selected measurement squared
(for example mm²).

5: Distance Between Centres: Distance

between the centres of the first circle
and the second circle.

6: Closest Approach: Smallest distance

between the perimeters of the two
circles.

7: CentreX and CentreY: The distances

between the centre of the circle and
the left edge of the image (CentreX)
and that from the centre of the circle
to the top of the image (CentreY).

Continued... 764

LAS User Manual 763

Appendix: Tool Parameters: Angle, Parallel Line and Cross Tools:

Baseline Angle:
1: Angle: The angle described with
reference to the horizontal.

2 & 3: Width and Height: The horizontal

and vertical measurements of a
bounding box enclosing the drawn
angle.

Apex Angle:
4: Angle: The uppermost angle
described between the two drawn
'legs'.

Four Point Angle:

5: Angle: The angle described at the
intersection of two extended lines.

Multiple Distance Line:

6: Length: The distance between the
Datum line and the selected feature.

Parallel Distance Line:

7: Length: The distance between the
Datum line and the selected parallel
line.

Cross:
8: Major Axis: The length of the longest
arm.

9: Minor Axis: The length of the shortest

arm.

10: Angle: The angle at the intersection

of the horizontal and the axis of the
Cross.

11 & 12: Width and Height: The

horizontal and vertical measurements
of a bounding box enclosing the
Cross at the intersection of the axis
and the line ending.

Continued... 765

764 © 2011 Leica Microsystems (Switzerland) Limited

 Appendix: Tool Parameters: Rectangle and Triangle Tools:

All distances are calculated from the

centre of the enclosing line:

Rectangle:
1: Perimeter: The distance around the
Rectangle.

2: Area: The enclosed space returned in

the selected measurement squared
(for example mm²).

3 & 4: Width and Height: Horizontal and

vertical dimensions of the Rectangle.

5: CentreX and CentreY: The distances

horizontally (X) and vertically (Y) from
the edge of the image to the centre of
the Rectangle.

Triangle:
6: Area: The enclosed space returned in
the selected measurement squared
(for example mm²).

7: Perimeter: The distance resulting in

the sum of the sides.

8: Side Length: The length of the side

commencing with the longest (Side
Length) and finishing with the shortest
(Side Length 3).

9: Angle: The angle described between

two sides commencing with the
smallest (Angle) and finishing with the
largest (Angle 3).

LAS User Manual 765

 Image Analysis

Leica Application Suite (LAS) Image Analysis is an Create histograms to display the distribution of sizes and
optional software module for image processing and shapes.
analysis in quantitative microscopy. LAS Image Analysis
allows the user to: Store and display images in a gallery.

Measure size, shape, position, orientation, intensity Export data to Excel to create user defined reports.
parameters for individual features (eg cells, fibres,
The combination of digital camera and microscope control
nodules, particulates, pores etc).
makes LAS Image Analysis the optimum application for
Measure the area percent, total area, perimeter and automatic measurement in a diverse range of imaging
other parameters that are summed for the entire Field tasks including:
of View (eg bone sections, reflective minerals, tissue
Analysis of the size distribution of porosity.
sections etc.)
Characterisation of the shape of a population of features.
Analyse multiple images and accumulates data for
them. Measurement of particulate dimensions
Show a list of selected parameters for all features Counting powders from pharmaceutical preparations
measured.
Fibre cross-section dimension and shape
Calculate a range of statistics.

Optional LAS Macro capability is described the LAS

Macro Editor help file. Go there.. 912

766 © 2011 Leica Microsystems (Switzerland) Limited

 The User Interface:

Image Analysis requires a number of

steps to be performed in a defined order.
This process starts with the acquisition
of the initial image of the specimen,
through to the detection of the features,
and finally to the presentation and
interpretation of results. These steps
are:

Image Acquisition.
Feature Detection
Measurement
Results and Interpretation

LAS Image Analysis is designed to make

each of these stages as simple as
possible. The Image Analysis Sequence
guides you through the acquisition,
detection and measurement set-up
steps and once established, these
settings can be used repetitively.

LAS User Manual 767

 The User Interface: Overview:

The User Interface conveniently groups tools, controls, 6: The Viewer can be split to simultaneously show both
images, results and image thumbnails so that using LAS original and processed images with Scroll Bars to aid
Image analysis is both fast and intuitive. navigation around enlarged views.

1: The Analysis Workflow that is the launch area for 7: The split Viewer can be locked so that both displays
LAS Image Analysis optional module. respond together to navigation and setup - in this
case a Measure Frame created on the original...
2: The Automatic Tab reveals all of the Image analysis
controls. 8: ... is duplicated automatically on the processed
image.
3: Automatic Processing Sequence tool bar.
9: The Side Tool Bar.
4: The Main Processing Menu - click to select a specific
component and reveal... 10: The Grid displays measurement results.

5: Specific Component Control Panels. 11: The Gallery displays thumbnails of the images.

768 © 2011 Leica Microsystems (Switzerland) Limited

 The User Interface: Workflow Panels:

LAS Image Analysis is part of the Analysis

Workflow:

1: Click on the Analysis Workflow and...

2: ...click on the Automatic tab.

3: Select one of the main components

by clicking on it in the main
Processing Sequences menu. A
green tick mark appears to the left
and the selection is highlighted. The
colour bars against each option
indicate the colour chosen (on the
appropriate panel) to represent the
option processing.

4: Processing progress is indicated by

the rotating 'star' and the Progress
Bar.

5: Some of the control panels are

collapsed and can be expanded by...

6: ...clicking on the arrows to the right of

the header.

7: Most Control Panels can be detached

from the 'parking' area on the
Automatic tab and moved by clicking,
holding and dragging the header, to
any other part of the screen.

8: Restore a detached Control Panel by

clicking on the 'X' to the right of the
panel header.

9: Many panel feature drop-down menus

that are revealed by clicking on the
small arrows to the right of the menu
header and then clicking on the list to
select an item.

10: Some panels have Tool Bars; Hover

the cursor above a tool button to
display a description of the tool.

LAS User Manual 769

The User Interface: Side Toolbar:

The Image Analysis module is located on

the Analysis Workflow.

1: Click on the Analysis Workflow and…

2: …click on the Automatic tab.

3: The Side Toolbar is situated on the

right-hand side of the Viewer with the
buttons grouped according to
function:

Viewer primary controls (4) pan image,

zoom in and out, fit the image to the full
Viewer area and display the image at the
actual size.

Image source and data controls (5) hide

or reveal the Data Form, the image, the
Data Grid and the Gallery. Additionally,
they allow the Data Form to be configured
so that only information needed by the
user is displayed.

Image Analysis controls (6) comprises 2

buttons - the upper expands to reveal a
list of options (7) that can be turned on
and off by clicking the entry.

Single/Dual Viewer splits the Viewer to

show the original and processed image
side by side; Original/Result displays the
previous processed image; Original/
Processed toggles the original and
processed images at full Viewer size;
Show Background Image turns the
background image off leaving only the
processed image; Show Binary Mask
displays processed image areas only and
shows the Binary Mask. Additionally, the
split Viewer display can be locked to
ensure that both images zoom together
and show the Measure Frame on both.

Continued 771 ...

770 © 2011 Leica Microsystems (Switzerland) Limited

 Side Toolbar: Primary and Image Source Controls:

The Side Toolbar: Click on a link for

details:

Viewer primary controls:

1: Pan the Image: Go there...

2: Zoom in and zoom out: Go there...

3: Fit the image to Viewer: Go there...

4: Display the Image at actual size:

Go there...

Image source and data controls:

5: Display Data Form: Go there...

6: Hide and reveal Image: Go there...

7: Hide and Reveal the Data Grid: Only

available if an LAS Archive is
installed: Go there...

8: Hide and reveal the thumbnail

Gallery: Go there...

9: Display the Archive Record details:

Go there...

10: Select the Record Fields to be

displayed: Go there...

Continued… 772

LAS User Manual 771

Side Toolbar: Image Analysis Tools:

The lower group of buttons relates

specifically to Automatic Measurement
operations.

1: Click on the Viewer options button.

When an option is enabled a tick
mark appears to the left of the
selection. Click again to disable the
option.

2: Single or Split Screen toggle. When

enabled the Viewer is split into 2
panes with the original image to the
left and the processed image on the
right. This is a toggle action button –
click again to revert to a single image
in the Viewer and use the Original/
Processed button (Item 4 below) to
select either the original image or the
processed image.

3: Clicking the Original/Result from

Previous Step button displays the
processed image from the previous
operation in the left-hand screen
area. This is a toggle button so click
again to return to the work. Use
Previous Step as a fast means of
comparing processing steps.

4: The Original/Processed Image button

toggles between the original image
and the processed image when the
Viewer is set to display full screen
single image (not split).

5: Show Background Image is a toggle

that hides or reveals the processed
underlying image to show only those
areas that have been modified. The
illustrations show an image that has
been passed through a Binary filter
with processed pixels coloured
yellow. Turning off the background
image displays only the mask pixels.
Click the button again to reveal the
background image.

Continued... 773

772 © 2011 Leica Microsystems (Switzerland) Limited

 Side Toolbar: Image Analysis Tools: Continued:

1: Click on the Viewer Options button to

reveal the options list.

2: The illustrations show an original

image – pink, more-or-less circular
areas containing dark blue features –
with the Threshold Adjust set to
process mainly dark blue. Enabling
the Show Binary Mask button shows
the processed areas – those falling
within the threshold values - as bright
red pixels (the colour is selectable).
Use the Show Binary Mask button to
quickly show the effects of binary
editing.

3: If the Zoom control is used to enlarge

the images beyond the Viewer area,
scroll bars are automatically displayed
around them. The original and
processed images can be scrolled
independently or, if the Lock Image
button is enabled, moved together
from any of the scroll bars. When the
Lock is enabled both images are
automatically synchronised.

4: Saves the results image including all

masks currently displayed. A
thumbnail appears in the Gallery and
the image can be used at a later date
for documentation.

Continued 774 ...

LAS User Manual 773

Side Toolbar: Gallery Thumbnail Sizing and Scrolling:

The size of the thumbnails in the Gallery

can be altered with the slider (1) – slide
left for smaller, right for larger.

The Gallery depth can be changed by

clicking on the horizontal bar that
separates it from the Viewer and dragging
it - up to increase the Gallery area and
down to reduce it.

Scroll through the rows of thumbnails by

clicking and dragging the scroll button (2)
or clicking on the arrows (3) to the top and
bottom of the scroll bar.

774 © 2011 Leica Microsystems (Switzerland) Limited

 The User Interface: Results: Feature and Field:

1: The Measurement Type determines whether only the

selected Features are measured, or the entire Field 3: Field Details and Statistics on two tabs or...
or both Features and Field. Generally, measuring
only the features is faster. 4: ...both Field and Feature Details and Statistics
displayed on four tabs.
2: Depending upon the Measurement Type chosen, the
results displayed in the Grid will have two tabs -
Feature Details and Feature Statistics or... Continued... 776

LAS User Manual 775

The User Interface: Results Grid:

1: Measurement results can be displayed on screen by 4: These can be further filtered by selecting those
clicking to enable the Grid View. which are to be displayed on the Grid by clicking on
the Configure tool that...
2: Click on an entry in the Grid to reveal the feature on
the image the label of which will be shown in a 5: ...displays the Select Feature Details dialog. Click to
contrasting colour. In the same way, clicking on a enable a check box to the right of the parameter to
feature on the Binary Output Image will highlight the be displayed. Click OK.
results for that feature in the Grid. If a sequence is
being measured, the appropriate image will be 6: Most headings when clicked will display a small
automatically displayed. Hold down the keyboard Ctrl arrow to the right. Clicking on the arrow will sort the
key whilst clicking on rows to make multiple results low-to-high or high-to-low.
selections. Hold down the Shift key to make a
sequential selection. Windows control keys can also be used to copy data
from the Grid.
3: The Grid headings are determined by the range of Ctrl + C: Copy selected items to the clipboard.
results chosen in Select Results – either a Ctrl + A: Select all the grid data. Use Ctrl C to copy it.
Predefined range or All Measurements.
Select Results to display on the Grid: Go there… 856

776 © 2011 Leica Microsystems (Switzerland) Limited

 Getting Started:

This section describes three approaches to using LAS

Image Analysis each depending upon the type of image Fast Track Automatic: For clear, evenly illuminated
being processed and the features that need to be images with defined features. No image processing
measured. needed - just detect features, measure and display
results.
Image Analysis works on images that have already been
captured in Browse or Acquire or that have been imported Fast Track with User Interaction: Images need
- Image Analysis itself does not have the ability to capture some work to define and isolate the features of
original images. They can be individual images or part of interest. Minimal image processing but some
a sequence captured with another module such as artefacts need to be removed.
MultiStep.
Advanced Measurements: Images are highly
The detail to be processed must be different in either detailed and less well defined, perhaps due to a
contrast or colour from the background and should be lack of contrast or a focus problem. A range of the
evenly illuminated. Shading correction should have been processing tools need to be used to pin point the
used. features to produce accurate results.

Continued... 778

LAS User Manual 777

Fast Track: Automatic:

Use Fast Track Image Analysis for images with:

Well defined features,
Evenly illuminated with good contrast.
'Clean' backgrounds free from artefacts.

Turn off all Image Processing: Go there... 798

Select No Filter options to turn off Image Processing,
Binary Image Processing and Binary Image Editing

Select Images to Measure: Go there... 782

Images should have good contrast.
Be properly focussed
Have features that are separate and well defined.

Adjust Threshold: Go there... 814

Select the features to be measured.
Separates features from background
Output is a Binary Image ready for measurement.

Measure Frame: Go there... 851

Use Typical to define the area enclosing the
features to measure.

Select Results: Go there... 856

Choose either Pre-determined or All Parameters.
Select the Measurement Type.

Measurements: Go there... 860

Set Parameter Filters.
Choose Name/Unit display options.
Select Font and Style and make measurements.

Histogram and Results: Go there... 870

Show results as Histogram.
View details in the Grid.

Display: Add Reference Data: Go there... 776

Add user-defined project and test information.

Create Report: Go there... 885

Create, save and display Report.
Analyse results.

Continue with Fast Track User Interaction: 779

778 © 2011 Leica Microsystems (Switzerland) Limited

 Fast Track: User Interaction:

Fast Track with User Interaction is ideal for images that have:
Well defined features that may be touching or overlapping.

Turn off Image Processing and Binary Image Processing:

Select the No Filter options: Go there... 798

Select Images to Measure: Go there... 782

Images should have reasonable contrast.
Properly focussed and evenly illuminated.
Features can be touching and overlapping but defined.

Adjust Threshold: Go there... 814

Selects the features to be measured.
Separates features from background.
Output is a Binary Image ready for measurement.

Binary Image Editing: Go there... 838

Remove and Create features.
Draw shapes
Select features to be included or ignored

Measure Frame: Go there... 851

Use Typical to define the area enclosing the,
features to measure.

Select Results: Go there... 856

Choose either Pre-determined or All Parameters.
Select the Measurement Type.

Measurements: Go there... 860

Set Parameter Filters.
Choose Name/Unit display options.
Select Font and Style and make measurements.

Histogram and Results: Go there... 870

Show results as Histogram.
View details in the Grid.

Add Reference Data: Go there... 776

Add user-defined data and notes that will be included
in the report.

Create Report: Go there... 885

Create, save and display Report.
Analyse results.

Continue with Advanced Measurements: 780

LAS User Manual 779

Advanced Measurements:

Detailed images with many features, noisy backgrounds and unwanted

artefacts, require the addition of the powerful Greyscale Image and
Binary Image Processing tools to separate and identify even the most
obscure features.

Select Images to Measure: Go there... 782

Many details overlapping and touching.
Focussing and illumination could be better.
Many and varied artefacts and noise.

Image Processing Pre-Filter: Go there... 787

Improve definition of wanted areas and features.
Start to close holes and fissures.

Adjust Threshold: Go there... 814

Select the features to be measured.
Separates features from background.
Output is a Binary Image ready for measurement.

Binary Image Processing Pre-Filter: Go there... 825

Adjusts the Binary Output Image to remove unwanted features.
Close holes and separate features.

Binary Image Editing: Go there... 838

Remove and create features.
Draw shapes.
Select features to be included or ignored.

Measure Frame: Go there... 851

User Define the area enclosing the features to measure.

Select Results: Go there... 856

Choose either Pre-determined or All Parameters.
Select the Measurement Type.

Measurements: Go there... 860

Set Parameter Filters.
Choose Name/Unit display options.
Select Font and Style and make measurements.

Histogram and Results: Go there... 870

Show results as Histogram.
View details in the Grid.

Add Reference Data: Go there... 776

Add user-defined data that will be included in the report.

Create Report: Go there... 885

Create, save and display Report.
Analyse results.

780 © 2011 Leica Microsystems (Switzerland) Limited

 Main Steps:

This section describes the major components and tools of

the LAS Image Analysis module. Generally, they are used Measure Frame: Create a frame to contain the
in the menu sequence, but it is possible to move back and features to be measured: Go there... 851 .
forth between components without loosing settings.
Select Results: Select and configure the results to
Select Images: Choose the image folders, groups display in the Grid and report: Go there... 856
and sequences: Image Analysis can be used to
process single images and sequences - the Progress Measurements: Count and measure the selected
Star next to the Progress Bar only appears when a features: Go there... 860
sequence is being processed. Go there... 782 .
Histogram: Display the measurement results
Greyscale Processing: Improve and clarify features to graphically: Go there... 860
measure: Go there... 787
Reference Data: Information to be displayed on
Adjust Threshold: Isolate features and create binary the Report: Go there... 883
output image prior to measurement: Go there... 814
Create Report: Configure, display and print the
Binary Processing: Fill holes and fissures in features: Report data and images: Go there... 885
Go there... 825

LAS User Manual 781

Select Images: Choose the Images to be Processed:

LAS Image Analysis is automated which

means that if measurements are to be
made on a series of similar images only
one need be processed and measured.
The processing steps are 'remembered'
by the program and then applied
automatically to the remaining images -
often without any further work by the user.

1: Select LAS Image Analysis by clicking

on the Analysis Workflow...

2: ...and if necessary clicking on the

Automatic tab.

The folder and images to be used in

Image Analysis must be selected in
Browse - either with Image Explorer or
LAS Archives (if installed).

The image selected in Browse is

displayed in the Image Analysis Viewer
and thumbnails in the Gallery.

To change the selected folder: 783 Go

there... 783

Continued… 784

782 © 2011 Leica Microsystems (Switzerland) Limited

 Select Images: Select Archive Folder:

If the required images reside in an LAS

Archive it must be selected and made
active. This is done on the Setup
Workflow as follows:

1: Click on the Setup Workflow tab.

2: Click on the Archive tab to reveal the

control panels.

3: To select LAS Archives, click on the

Folder Archive button.

4: Double click the required archive

entry to make it active.

Continued… 784

LAS User Manual 783

Select Images: Loading Sequences & Random Images:

If an image sequence or collection of

individually selected images is going to be
processed automatically, they have to be
added to the Sequence List.

1: Click on the arrow to the right of the

Sequence header to reveal the
Sequence List panel.

Loading Sequences:
2: If the images to be processed form
part of a sequence, click on a
thumbnail and…

3: …click the Append button. The

program will recognise that the image
is part of a sequence and…

4: …the Append Sequence button will

become active. Click it and...

5: ...all of the sequence images will be

loaded to the Sequence List
automatically.

Loading Random Selections:

2: To selected images at random for
processing, click on the first
thumbnail and with the CTRL key (6)
on the keyboard held down, click on
the thumbnails of the other images to
be included.

3: Click the Append button and all of the

image names will appear in the list.

7: To sort the images in numerical

order, click to enable the Sort check
box.

Creating a Process Sequence: Go there…

896

Continued… 785

784 © 2011 Leica Microsystems (Switzerland) Limited

 Select Images: Removing Images & Accumulating Results:

Remove an Image from the Sequence

List:
1: Individual images can be removed
from the measurement list (but not
the Archive) by clicking to select the
image and then…

2: …clicking the Remove button.

Remove all Images from the List:

3: Remove all of the images from the
measurement list by clicking the
Reset button.

Accumulate Results:
Enabling Accumulate Results will produce
a continuous list, the results for each
image following the previous. This option
allows comparisons across the entire
image collection.

In Select Results (Go there...) 856 enable

display Image Name to easily locate the
results for individual images.

Disable Accumulate Results to have the

results associated only with individual 4: To enable Accumulate Results, click on the Options button and...
images - use this option if there are a
large number of images in the sequence 5: ...from the Process Settings menu, click to enable the Accumulate
to speed up processing. Clicking on a Results check box.
Gallery thumbnail will display the image
and its results in the Grid.
Continued... 786

LAS User Manual 785

Select Images: Export Report, Show Histogram & Refresh:

1: Click on the Options button. From

the Process Settings menu...

Export Report:
2: ...click to enable the Export Report
check box and the results of the
Processing Sequence will be loaded
automatically to the chosen Report
option (Excel or CSV) when
processing is complete.

Show Histogram:
3: During the Processing Sequence the
Histogram for each image can be
displayed if the Show Histogram
check box is enabled.

Refresh Display:
4: Enable the Refresh Display check
box to refresh the display between
each image analysis.

Refresh Results:
5: The Grid results display will be
updated for every image if the
Refresh Results check box is
enabled.

786 © 2011 Leica Microsystems (Switzerland) Limited

 Greyscale Processing: Principles:

The Greyscale Processing component of

LAS Image Analysis provides selected
tools to enhance feature recognition in
both colour and monochrome images.

Skip Greyscale principles: 797

Greyscale Processing Principles:

At the heart of most image processing is a
concept called a Structuring Element,
shown in its simplest form in Illustration
(A). It comprises a matrix of cells, each of
which will be a binary value – either 1 or 0.
The centre (red) cell is called the Origin.

In Illustration (B):
1: The original Greyscale Input Image
with each pixel represented as a
value in the range 0 to 255 - black to
white.

2: The Output Image which will be

created using values determined by
the Input Image and the Structuring
Element.

3: The Structuring Element is used as

an 'overlay' with the Origin (red) Cell
positioned over a pixel on the Input
Image. This is also coloured red and
called the Input Pixel.
Those pixels either side of the Input
Pixel are called Neighbours, shaded
pink. The value of the Neighbour
pixels are used to create...

4: ...a new Output Pixel in the Output

Image.

Continued... 788

LAS User Manual 787

Greyscale Processing: Principles: Continued:

A Neighbour pixel is only tested if the

corresponding cell in the Structuring
Element is set (=1). In the example both
cells in the Structuring Element are set so
the Neighbour pixels either side of the
Input Pixel will be evaluated.

Brightening detail: Dilation:

In this process, the Neighbour pixel with
the greatest value determines the value of
the Output pixel. Since, in a greyscale
image the lighter pixels have the greatest
values (closer to white at 255), the effect
is to increase the area of lighter detail and
reduce that of darker detail.

In the Illustration (B) the Structuring

Element Origin (red) is positioned above
an Input Pixel with the value 22. The
Neighbour pixels values are 18 and 32.
The intention is to brighten the image and
so the Neighbour pixel with the greatest
value is used for the new Output Pixel – in
this case 32.

In Illustration (C) the Structuring Element

has moved right to the next Input Pixel.
Now the values in the Neighbour pixels
are 22 and 36 and again the greatest is
used for the Output Pixel.

Darkening detail: Erosion:

To darken the image the same evaluation
process is used but in this process the
Neighbour pixel with the lowest value
(closer to black at 0) is used as the Output
Pixel. Illustrations (D) and (E) show the
process.

All of the greyscale Input Image pixels are

tested until the Output Image is complete.

Continued... 789

788 © 2011 Leica Microsystems (Switzerland) Limited

 Greyscale Processing: Dilation and Erosion:

A simple example:
Illustration (A) is a greyscale image – a
small block of pale grey pixels each with a
value of 120, surrounded by dark grey
pixels with a value of 30.

A 2 x 3 Structuring Element (1) is to be

applied to the image, first as a Dilation
and then as an Erosion.

Dilation uses the Neighbour pixels with

the greatest values so in Illustration (B)
the pale grey block has ‘grown’ adding
pixels to the top and sides.

The process is reversed by Erosion in

Illustration (C). Erosion uses the smallest
values found in the Neighbour pixels so
the pale grey pixels (with a value of 120
closer to white) have been replaced by
dark grey pixels (with a value of 30 closer
to black).

The image shift – the pale grey block has

moved upward by a row of pixels – is due
to the design of the Structuring Element,
and can be prevented by using a more
appropriate matrix.

Continued... 790

LAS User Manual 789

Greyscale Processing: Dilation and Erosion: Continued:

A practical example:
Greyscale image (1) shows a group of 8
black discs on a white background. The
transition between black (0) and white
(255) on an image is seldom that distinct –
very often there will be several grey pixels
around the boundary of a dark feature
giving it a ‘blurred’ appearance. The
blurring may be due to incorrect focussing,
poor lighting or perhaps the feature itself
is fuzzy.

The blurred boundary can make adjacent

features appear to be touching each other
and, if the intention was to count the
features in an image, the discs in Figure
(1) would be treated as 3 features –
groups (A), (B) and (C) - instead of 8.

The boundaries between 2 adjacent discs

may look like the pixels in Illustration (2),
the black pixels of the discs blurring
through shades of grey with white
background pixels appearing at the
extreme edges.

A Dilation (Illustration 3) uses the greatest

Neighbour pixel values to create an
Output Image so at the edges of the
blurring, white (value of 255) would
replace pale grey pixels (value say of
210). The ‘new’ white background pixels
can be seen in the centre.

The amount of Dilation depends upon the

Structuring Element matrix. Illustration (4)
shows the effect of applying a Structuring
Element with a greater number of cells
testing a wider group of Neighbours. Here
several grey pixels have been replaced
with white pixels to create a larger gap
between adjacent discs.

Continued... 792

790 © 2011 Leica Microsystems (Switzerland) Limited

 Greyscale Processing: Erosion then Dilation:

1: The same greyscale image but this

time 8 white discs on a black
background and apparently ‘grouped’
because of edge blurring. The task is
the same – separate the discs without
losing important dimensional data.
Light features against a dark
background are often better
separated by using Erosion followed
by Dilation.

2: The original ‘blurred’ image.

3: After Erosion – Neighbour pixels with

the smallest value (nearer to black at
0) replace those with greater value
(closer to white at 255) – the grey
pixels around the boundaries of the
discs have been replaced with black.

4: After Dilation – Neighbour pixels with

the greatest value (closer to white at
255) replace those with closer to
black (at 0). The effect is to ‘grow’ the
white discs and, with the appropriate
Structuring Element, to closely re-
establish the discs’ diameter without
causing them to touch again.

An Erosion followed by a Dilation is

called an Opening Filter.

Continued... 793

LAS User Manual 791

Greyscale Processing: Dilation then Erosion:

Illustration (5) is the original blurred image

and (6) is the same image with
successive Dilations until all of the grey
pixels have been replaced with white –
only distinct, separate black discs and
white background remain – 8 features
now instead of 3.

However, removing the grey pixels may

have also reduced the actual size of the
discs which could be detrimental if areas
or dimensions are to be measured. An
Erosion will make the black discs grow –
smallest value Neighbour pixels (those
nearest black at 0) replace higher value
pixels (those closest to white 255).
Erosions continue until the black discs are
close but not touching to complete the
transition (7).

Dilation followed by Erosion is called a

Closing Filter.

Continued... 791

792 © 2011 Leica Microsystems (Switzerland) Limited

 Greyscale Processing: Open and Closed Filters:

Open and Closed Filters have a wide

range of applications in image processing.
Noise removal is one most frequently
used.

1: A greyscale image with clumps of

small features in the background
which could be ‘noise’ or just
insignificant parts of the image,
unwanted artefacts that need to be
removed before accurate
measurements can be made.

2: Successive Dilations have not only

separated the black discs but also
removed the ‘noise’ artefacts
completely – simply because they are
small.

3: Erosion enlarges the discs to their

original size but because the ‘noise’
no longer exists there is nothing to
enlarge.

4: Removing small artefacts in Image

Analysis; The original Input Image
and...

5: ...the Output Image with the artefacts

removed and the grey features
restored to their original size.

6: The Image Analysis panel with Image

Processing Pre-Filter tools in Smooth
Black Detail Mode.

Smooth Black Detail Mode: Go there… 800

Continued... 794

LAS User Manual 793

Greyscale Processing: More About Structuring Elements:

On the edge of the Input Image, the first

Neighbour Cell of the Structuring Element
will point to a non-existent pixel beyond
the edge of the image. For greyscale
images (Illustration A) and depending
upon the process – Dilation or Erosion –
these pixels are assumed to have values
of 0 for Dilation and 255 for Erosion. This
arrangement prevents border effects –
fine, dark lines appearing around the
output image.

Almost any layout can be applied to a

Structuring Element; (Illustration B) shows
a common 3 x 3 cell configuration in
which a set cell (=1) is an active
Neighbour used to evaluate the Input
Image pixel values, whereas a cell
cleared (=0) will be ignored. The
Structuring Element matrix – how many
cells it contains – generally reflects the
size and shape of the features being
detected within the Input image. Image
Analysis uses a circular configuration
which expands or contracts depending
upon the size selected by the user (
Illustration C).

Continued... 795

794 © 2011 Leica Microsystems (Switzerland) Limited

 Greyscale Processing: Specialised Filters:

The Structuring Elements using more

complex mathematical principles are used
in a wide range of greyscale filters to
detect and improve the quality of features
of interest in an image. They are used
extensively in Image Analysis and the
following describes just two of them.

Median Noise Removal:

Illustration (A) depicts a Structuring
Element with a 3 x 3 matrix – the Origin
has 8 Neighbour Cells. Only those cells
set (=1) are used in the filter; Those
cleared (=0) are ignored. The filter
extracts the Neighbour pixel information
from the Input Image – in this case 22, 18,
44 and 82 – and calculates a Median
value, that is a value at the mid-point of
the range, not an average.

For an odd-numbered range the middle

value is chosen, but for even-numbered
ranges the two values closest to the
middle are averaged. Our example is
even-numbered (4 values) and so the two
mid values (18 and 44) are averaged (18
+44 = 62/2 = 31) which becomes the value
of the new Output Pixel.

More refined versions can then

manipulate the values in the Input and
Output Images. For example, having
established the Median Output Pixel
value, subtracting it from the Input Pixel
value (20 – 31 = -11) and then adding
back the Input value (-11 + 20 = 9),
darkens the image and improves contrast.

Illustration (B) shows Image Analysis in

action with the Median Noise Removal tool
selected. (1) is the Input Image and (2)
the Output Image.

Noise removal - Median: Go there... 802

Continued… 796

LAS User Manual 795

Greyscale Processing: Specialised Filters: Tophats:

Some of the most useful morphological

filters, Tophats belong to a sub-class of
operations called residues, since they
produce something which is 'left over'
from the original image.

Tophats can be defined as:

Smoothing > Difference > Tophat

The net effect is to enhance whatever was

removed during the smoothing process,
which will usually be the detail of the
image.

The most common Tophats are

constructed by using grey openings and
closings as the smoothing operations. The
Find black detail-Tophat is defined as:

Tophat = Closed Image - Original

Image

The effect of a black Tophat (1) is to 'pick

out' the small dark features in the image.
The closing will remove the dark features
up to a size defined by the cycles used,
replacing them with the grey level of the
lighter regions which surround them, and
the subtraction of the original image will
thus show the grey level difference
between these 'filled in' pixels and their
grey levels in the original image.

2: The original image and...

3: ...after Find black detail Tophat.

796 © 2011 Leica Microsystems (Switzerland) Limited

 Greyscale Processing: Image Processing Pre-Filters:

Experimentation is an inevitable part of

improving image definition and this
manual cannot possibly list methods for
every image type or quality. Instead, the
following pages illustrate examples to
demonstrate some of the powerful
potential in the Image Processing toolkit –
it is up to the user to select and
manipulate the best tool for the job in
hand.

When Greyscale Pre-Filters are applied to

an image the original is not affected at all
– it remains intact on the hard drive.
Applying a filter creates a Processed
Greyscale Output Image in memory which
is then passed on to other functions such
as Adjust Threshold, for additional
processing.
Although the processed Greyscale Output
Image is created in memory it can be
saved to disk and 'exported' to another
application just like any other image:

Click on the Save button on the right side

toolbar to save the processed image. A
thumbnail of it also appears in the Gallery.

Continued… 798

LAS User Manual 797

Greyscale Processing: The Image Processing Panel

To display the Greyscale Image

Processing tool panel:

1: Click on the Image Processing Pre-

Filter option in the main list.

2: The Pre-Filter panel appears.

3: Each ‘pass’ of a structuring element

can be displayed automatically by
clicking the Interactive check box to
enable it.

4: Disable Interactive and use the Apply

button to update the Output Image
when required. This option is
preferable for larger, complex images
for which selecting the filter and
setting the Size and only then
applying the changes is faster.

5: The structuring element matrix is

determined by the Size text box. To
change the value, click in the window
and type a new value or use the Up/
Down arrows to the right of the box.
Small increments are preferable
because there will be a point at which
the image cannot be improved by
making another ‘pass’.

6: To use LAS Macro Runner, click to

enable the check box. Detailed help
for LAS Macro: Go there...

Continued… 799

798 © 2011 Leica Microsystems (Switzerland) Limited

 Greyscale Processing: Pre-Filter Selection

The contrast between features and

background can be improved for most
images by enabling Auto Contrast:

1: Click the checkbox (Tick mark visible)

to enable Auto Contrast.

2: The original greyscale image and…

3: …the same image with Auto Contrast

turned on. The improvement is very
obvious.

4: To select a Pre-Filter, click on the

arrows to the right of the Mode drop
down window and…

5: …click to make a selection from the

list.

Continued... 800

LAS User Manual 799

Greyscale Processing: Pre-Filter Descriptions:

To access the filters:

1: Click on the arrows to the right of the
Mode menu window and…

2: …click to select the filter required.

The option No Filter disables all of the
filters and can be used as a simple
‘reset’.

3: The original Greyscale Input Image.

4: Smooth White Detail brightens the

Output Image by accentuating the
lighter tones. This is an open filter –
Erosion followed by Dilation.

5: Smooth Black Detail darkens the

Output Image by strengthening the
darker tones. This is a close filter –
Dilation followed by Erosion.

Continued… 801

800 © 2011 Leica Microsystems (Switzerland) Limited

 Greyscale Processing: Pre-Filter Descriptions: Continued:

1: The original Greyscale Input Image.

2: Enhance White Detail accentuates

the white – values approaching 255 –
areas of the image.

3: Enhance Black Detail improves the

contrast between the black (values
toward 0) and the lighter areas of the
image.

4: Find White Detail Top Hat detects

and removes lighter detail from the
image depending upon the size of the
Structuring Element.

5: Find Black Detail Top Hat detects and

removes darker detail from the image
depending upon the size of the
Structuring Element.

Continued… 802

LAS User Manual 801

Greyscale Processing: Pre-Filter Descriptions: Continued:

1: The original Greyscale Input Image

and…

2: …after applying the Sharpen Grey

Transitions (Delineate) filter.
Intermediate grey tones are removed
to provide greater contrast between
areas of the feature.

3: The original Greyscale Input Image.

4: The Noise Removal (Median) filter

removes artefacts down to pixel size
from the image.

802 © 2011 Leica Microsystems (Switzerland) Limited

 Greyscale Processing: Example 1:

The aim of this example is to measure the

number of biological cells on an image.
The problem with the original image (A) is
that the cells overlap, have a range of
differing colours, have irregular outlines
with varying contrast and some have
coalesced into indeterminate ‘blobs’.

Skipping a Greyscale Pre-Filter and going

directly to Adjust Threshold (Illustration B)
results in a fuzzy Binary Output Image
with scattered additional artefacts all of
which could be included in the measured
count. Adjustments to intensity might
have improved the image slightly, but it
would be a time-consuming hit-and-miss
process not necessarily resulting in a
better count. Better to use a Pre-Filter to
isolate individual cells allowing each to be
counted with acceptable accuracy.

Because the cell outlines and contrast are

so variable, an alternative solution would
be to count the ‘nuclei’ – they have a
more-or-less consistent colour and size –
and where cells overlap the nucleus tends
to show through and can be counted.

Continued… 804

LAS User Manual 803

 Greyscale Processing: Example 1: Continued:

1: The Enhance Black Detail filter was

chosen to improve the contrast
between the nuclei and the
surrounding cell tissue. Applying a
structuring element of size 12, many
of the intermediate greys have
disappeared, the cell edges have
become more clearly defined and the
nuclei contrast is especially
enhanced.
There are still areas of the image in
which the cells have clumped
together but these may be removed
or more clearly defined during Adjust
Threshold or with Binary Image Edit.

2: The original image and…

3: …the Greyscale Output Image.

Selecting a Pre-Filter 798 : Go there… 798

Continued… 805

804 © 2011 Leica Microsystems (Switzerland) Limited

 Greyscale Processing: Example 1: Continued:

The next step is to apply the Thresholds.

Check that Show Binary (1) and
Accumulate (2) are both enabled.

3: With adjustments to the upper and

lower Intensity Thresholds, the
resulting Binary Output Image
highlighted the nuclei and some
clumps of artefacts all shown in
green the default colour.

4: A first measurement for cell count

yielded 86 but that included the
clumps of artefacts as well. The
feature highlighted in yellow is
currently selected on the Grid view.

Adjust Threshold: Go there... 814

Measurements: Go there... 860
Grid View: Go there... 856

Continued… 806

LAS User Manual 805

 Greyscale Processing: Example 1: Continued:

There are several methods for removing

unwanted features – Reject and Delete
Modes in Binary Edit for example, but in
this example they were excluded by
refining the upper and lower limits of the
Area Measurement tool.

A first pass with a wide difference

between the upper and lower limits
produced an area value for all of the
features, nuclei as well as unwanted
artefacts. From this it could be seen that
the nuclei areas fell between 10 and
50µm2 whereas the unwanted artefacts
were considerably larger.

The Area Parameter limits where then set

to include the nuclei (1) with the result
that all of the unwanted artefacts were
ignored and the count reduced to 80 (2).

In this example a Measure Frame was

used to select just a small part of the
image since the features were evenly
distributed across the entire image and
the smaller Measure Frame would yield a
good mean result. It also speeds some of
the processing time.

Once the correct filtering and thresholds

have been established, the Measure
Frame can be set to Entire Image and a
further measurement made – in this case
it resulted in a total count of 294 cells.

Measurement Tools: Go there... 860

Area Parameter: Go there... 904

806 © 2011 Leica Microsystems (Switzerland) Limited

 Greyscale Processing: Example 2:

In this example a piece of schist, a

crystalline rock, is the original image (1).
The specimen contains some iron ore
which shows up as red-orange areas
scattered randomly across the image.
The test is to determine what percentage
of the sample comprises iron ore.

In such a complex image there are likely

to be many single, unconnected pixels
that are close to the iron ore colour range
for which we will be searching. Leaving
these pixels in place will slow the
measurement process considerably
without beneficially affecting the end
result, so an overall
‘tidy-up’ to remove ‘stray’ pixels and
consolidate larger groupings is a good
starting point.

The Pre-Filter chosen was Smooth White

Detail (2) with a structuring element of
Size 3. Because Smooth White Detail is
an open filter – erosion first and then
dilation – some features disappear
completely which is good for removing
the stray pixels, but it can also start to
reduce the area of the wanted features.
Since this example will measure area, the
filter has to be used sparingly.

(3). The result of the Smooth White Detail

Pre-Filter.

Continued… 808

LAS User Manual 807

Greyscale Processing: Example 2: Continued:

It took only minor alterations to the Adjust

Threshold 814 intensity and saturation to
produce a clean, well-delineated Binary
Output Image (1) which, when compared
with the original (2) proved to have
faithfully pin-pointed the traces of iron ore
and ignored everything else.

However, there were some holes and

fissures in the main accretions which
looked as though they should be included,
but trying to fill them with the Threshold
controls also tended to draw unwanted
artefacts into the Binary Image.

There is an easier way to fill holes…

Adjust Threshold: Go there... 814

Continued 809 ...

808 © 2011 Leica Microsystems (Switzerland) Limited

 Greyscale Processing: Example 2: Continued:

The Combine Detail filter, which is part of

the Binary Processing Pre-Filter collection
825 , can quickly, simply and precisely fill
holes and fissures in a Binary Output
Image (1).

2: The Reconstruct and Fill options were

enabled by clicking the check boxes.

3: The structuring element Size control

was incremented in single steps until
the holes in the major features were
filled without gathering together some
of the more separate features.

4: A first measurement yielded a total

feature (iron ore) area of 152670µm2
which was displayed on the Grid View
and this was...

5: ...within a manually defined Measure

Frame of 2587631µm2 giving a value
of 5.9% iron ore.

To check that the measure frame

represented an average part of the image,
the frame was reset to Entire image and
the area measurement was taken again.
This yielded a feature area of 577912µm2
within an image area of 9513571µm2 -
6.0%.

Binary Processing Pre-Filters: 825 Go

there... 825
Grid View: Go there... 776

LAS User Manual 809

Greyscale Processing: Example 3:

The diamond chips in the original image

(1) produce an acceptable Binary Output
image if passed directly through the Adjust
Threshold, but some fast processing with
a Greyscale Pre-Filter reduces the amount
of fine adjustments made to the Threshold
and can improve measurement precision
considerably.

LAS Image Analysis are so fast that it is

always beneficial to run an image through
several greyscale filters to check for
improvements.

In this example the image was processed

with the Smooth Black Detail Pre-Filter
that removed some of the intermediate
grey values and increased the chip edge
contrast (2).

However, it had to be used carefully since

this was to be a feature count – how many
chips in the image – and because several
overlap the Smooth Black filter could have
blended them together resulting in a single
feature rather than two.

Continued… 811

810 © 2011 Leica Microsystems (Switzerland) Limited

 Greyscale Processing: Example 3: Continued:

1: Diamond chips is a monochrome

image with predominant greys closer
to the black end of the scale, so the
Grey Range Selector was set to
Blacker than and minor adjustments
made to the Histogram sliders.

2: The fast result is a very good Binary

Output Image with small holes (3) in
just two of the chips.

Continued… 812

LAS User Manual 811

Greyscale Processing: Example 3: Continued:

1: In this example the holes in several of

the diamond chips would not have
made a difference to the final feature
count, but since it was necessary to
pass the image though a Binary Pre-
Filter to make sure that there was
good separation where the chips
overlapped, the Combine Detail
with…

2: …the Fill option enabled, was used to

fill the holes with the Size setting at 6.
Again, this was used sparingly
because some of the chips are close
together and could have been
combined.

3: There were 4 chip pairs that were

either overlapping or touching; They
can be clearly seen on the original.

4: With Separation > Cut Touching

enabled…

5: …two steps were sufficient to

separate the overlaps, indicated by
the red lines on the Output Image (6).

7: To use LAS Macro Runner, click to

enable the check box. Detailed help
for LAS Macro: Go there...

Continued… 813

812 © 2011 Leica Microsystems (Switzerland) Limited

 Greyscale Processing: Example 3: Continued:

1: The final step before counting the

diamond chips was to remove the
fragment (bottom right) to prevent it
being counted achieved with…

2: ...Binary Edit Delete Mode. The

fragment was not actually removed
from the Binary Output Image but only
highlighted and ignored in the count.

3: The count shows 18 chips in the

image and shows that the Cut
Touching tool is working well.

Binary Edit Delete Mode: Go there... 849

LAS User Manual 813

Adjust Threshold: Principles:

Adjust Threshold is a quick and simple

method of selecting the features to
measure or edit and ignoring the rest.

The Threshold principle:

Illustration (A) shows the original
greyscale image on the left and its
graphical representation called a
Histogram, on the right. The Histogram
comprises 256 vertical bars each
representing a pixel value. A value of 0
represents black (1) on the left and a
value of 255 represents white (2) on the
right. This image of quartzite does not
have any absolute black areas - the
darkest being close to a value of 25 - very
dark grey. The two Histogram sliders can
be moved along the base line so that any
values to the left of the black slider are
ignored as well as any of those to the right
of the white slider.

In Illustration (B) the black slider has been

moved to position 50 so all pixels with a
value from 0 to 49 have been ignored with
the result that many of the darker features
have been lost.

Illustration (C) shows the same effect but

this time moving the white slider to the
150 position. All pixels with a value
between 151 and 255 are ignored and the
overall effect is a darker image.

Both sliders have been moved in

Illustration (D) - black is at position 50 and
white at position 150 - so only those pixels
in the range 50 to 150 are reproduced in
the image.

The sliders can be moved to isolate a very

narrow range of pixels (Illustration E) to
the extent that just a single pixel value
could be selected.

814 © 2011 Leica Microsystems (Switzerland) Limited

 Adjust Threshold: The Threshold Dialog:

1: The Threshold dialog is opened by

clicking on the Adjust Threshold
option on the main menu.

2: For colour images the dialog with a

Colour Wheel and controls for Hue
(H), Saturation (S) and Intensity (I)
opens, whereas…

3: …for monochrome images the scale

is displayed as a Histogram with only
the Intensity control available.

4: Additionally, the monochrome image

dialog has a Grey Range Selector.
The other buttons and controls are
the same for both image types.

The Interactive Checkbox:

5: With the Interactive checkbox
enabled (showing a tick mark) each
change in the Threshold settings is
automatically reflected in the Output
Image. For simple images this is the
best setting, but more complex
images may take a little longer to
refresh and so disable Interactive and
click on…

6: … the Apply button to update the

Output Image after settings have
been altered.

Continued... 816

LAS User Manual 815

Adjust Threshold: The Threshold Dialog: Continued:

Dual Viewer and Binary enabled:

1: Click on the Viewer Options button on
the Side Tool Bar and from the
context menu, click to enable Dual
Viewer – original Input Image to the
left and adjusted Output Image to the
right of the Viewer.

2: Click to enable the Show Binary

checkbox which will then display the
features selected for measurement in
the right hand side of the Viewer. The
features are shown in the default
colour red or one of the user's
choosing.

3: Checking the Accumulate check box

allows a sequence of selections made
on the original image to be added
together - accumulated - on the
Binary Output Image. The
Accumulate feature: Go there... 819

4: The threshold for images with good

contrast and well-defined features,
can be established automatically by
clicking the Auto button. Fine
adjustment can then be made with
the other controls.

The Adjust Threshold tool produces a

Binary Output Image in computer memory
that is passed on to subsequent filters and
tools for enhancement or measurement.
The original image is unaffected.

Position the cursor over the Binary Output

Image and right click and hold down the
mouse button. This will temporarily hide
the selected features making it easier to
compare with the original and determine if
all of them have been included. Release
the mouse button to restore the Binary
Output Image.

Change the Binary Image Colour: Go

there... 820

Continued… 820

816 © 2011 Leica Microsystems (Switzerland) Limited

 Adjust Threshold: Selecting Features: Drawing a Region of Interest:

There are two methods of selecting

features to be included in measurements:

The Region of Interest can be used for

both colour and monochrome images
and,
The Grey Range Selector used on
monochrome images only..

Both are fast, precise and easy to use.

Region of Interest selection:

Drawing a Region of Interest around a
feature to include a range of greyscale
values is a fast and intuitive. Pixels across
the image with values matching those
inside the drawn region are selected and
coloured on the Binary Output Image.
The process is suitable for both greyscale
monochrome and colour images.

1: The original image, part of an

electronic circuit board. The gold-
plated rings bottom right which have
distinctive colour values are to be
selected for measurement.

2: Use the Zoom In tool from the side

toolbar to pin-point the feature.

3: Click on the feature and drag down

and to the right to create a small
rectangle – the Region of Interest.
Keep the boundaries of the rectangle
within the feature.

4: Release the mouse button and pixels

with values that match those within
the Region will be selected on the
Binary Output Image.

Continued... 818

LAS User Manual 817

Adjust Threshold: Selecting Features: Drawing a Region of Interest: Continued:

LAS Image Analysis recognises colour or

monochrome images and as soon as a
Region of Interest is drawn displays the
appropriate control panel.

Colour Images: Colour and Intensity:

The Colour Wheel (1) can be used to
adjust and experiment with the Hue and
Saturation values. Hue is measured along
the circumference of the Wheel and
Saturation is represented by a position
along the Wheel’s radius.

2: As the cursor moves over the Wheel

both Hue and Saturation are
displayed in a Tooltip box.

3: Click and drag on the Wheel segment

to alter the Hue and Saturation. The
actual values are displayed in the
Colour Range windows (5).

4: Intensity can be changed by clicking

and dragging the Intensity Bar
Threshold pointers up or down.

5: Fine-tune all three parameters with

the Up/Down arrows to the right of
the Colour Range windows. Each pair
of windows represents the span of
values for that parameter H=Hue,
S=Saturation and I=Intensity.

Monochrome Images:
A Histogram (6) displays the greyscale
values for monochrome images.
Adjustments are made with the Histogram
Sliders or with the Grey Range Selector.

The Grey Range Selector: Go there… 821

Continued… 819

818 © 2011 Leica Microsystems (Switzerland) Limited

 Adjust Threshold: Selecting Features: Accumulate Selection:

The Accumulate tool adds the values of

successive drawn regions to extend the
Threshold range and therefore, include
more features.
For example, Illustration (A) shows three
diamond chips that together include
greyscale values from 45 (near to black)
and 187 (near to white).
The first drawn region (1) encompasses
value from 45 to 103 and all features
falling within that range will be selected.
The second drawn region (2) includes
values from 93 to 187 and with
Accumulate enabled this range will be
added to the previous to extend the scope
from 45 to 187.
Disable Accumulate (5) if each Threshold
selection is to be treated as a completely
new operation.

In the examples,
3: ...represents the features selected
after the first region was drawn and...

4: ...after both regions have been drawn

and the range extended.
However, the higher values now
capture some background artefacts
but they can be removed with small
adjustments to the Grey Range
Selector value.

Reset and Undo:

6: The Reset button will clear all
Threshold values whereas…

7: …clicking Undo will remove only the

last pass.

Continued… 821

LAS User Manual 819

Adjust Threshold: Change Binary Image Colour:

The colour of the selected pixels in the

Binary Output Image can be changed to
suit the user.

1: Click on the Colour button. The Select

Colour dialog appears.

2: Choose a colour from the swatches

by clicking on it or…

3: …click and drag the hue sliders or…

4: …click and drag the marker on the

Colour Wheel.

5: Click OK. The new colour appears in

the Colour button.

6: The illustrations show the effect of

changing the binary Colour from red
to green.

820 © 2011 Leica Microsystems (Switzerland) Limited

 Adjust Threshold: Selecting Features: Grey Range Selector:

The Grey Range Selector is a fast and

easy way to select features on a greyscale
monochrome image. LAS Image Analysis
automatically detects monochrome
images and displays the histogram and
Grey Range Selector.
Three user-set grey ‘levels’ are available
as a drop down menu:

Blacker than selects all features at or

below the entered grey value.

Whiter than selects all features at or

above the entered value, and…

Grey automatically sets two values

(Blacker than + 1 and Whiter than –
1) as a centre band.

To set the Threshold range:

1: Click on the small arrows to the right
of the Grey Range menu header
and…

2: …click to select a range. The Grey

Range text boxes reflect the the set
range selected.

3: To enter a value either click inside the

window and type a new value or use
the Up/Down arrows to the right of the
box that are ideal for ‘fine tuning’.

Changes made to the grey values are

automatically reflected in the Blacker than
and Whiter than settings.

Continued… 822

LAS User Manual 821

Adjust Threshold: Selecting Features: Grey Range Selector:: Continued:

1: The entered values are displayed on

the Histogram…

2: …the sliders of which can be clicked

and dragged to adjust the settings.
The selected range is shown as a
colour on the Histogram display.

The Binary Output Images for the three

Grey Range Selector settings are shown
in the illustrations:

3: Blacker than setting.

4: Whiter than setting, and…

5: Grey setting.

822 © 2011 Leica Microsystems (Switzerland) Limited

 Adjust Threshold: Save the Binary Output Image:

1: Save the Binary Output Image at any

time by clicking on the Binary Save
button on the side tool bar. The
Binary Image is saved in the same
folder as the original and…

2: …a thumbnail using the original

name/number plus the suffix ‘AM’ as
a caption. Saving subsequent Binary
Output Images for any of the tools will
replace and overwrite the preceding
binary save.

LAS User Manual 823

Adjust Threshold: Switch Views and Magnifier:

To switch between the Binary Output

Image and the Input Image:

1: On the Binary Output Image click and

hold down the right mouse button.
This will hide the binary image and
reveal the original input (2).

To enlarge parts of either the Input Image

or the Binary Output Image:

3: Hold down the keyboard Shift key and

click on a portion of the image to
enlarge. The magnified area tracks
the mouse movements (4). Release
the Shift key to close the magnifier.

824 © 2011 Leica Microsystems (Switzerland) Limited

 Binary Processing: Principles:

Binary Processing uses the same

techniques as Greyscale Processing for
improving and modifying images except
that instead of working with values ranging
from 0 to 255, only pixel values of 1 and 0
are used.

Usually, the image presented to the Binary

Processing function has passed through
Greyscale Processing and Adjust
Threshold. Illustration (A) shows the
sequence:

1: The Greyscale Input Image

comprising 8 touching disks; the task
is to count the disks and measure
their areas. Performing a Number
(Count) and Area calculation on this
image would yield just 3 features with
diverse areas because some of the
disks are touching.

2: Shows the Output Image after a

Greyscale Processing Erosion – the
disks are no longer touching so a
Number (Count) would be correct at
8. But some diameter has been lost
so the Area calculation would be
understated.

3: The disks after a Greyscale Dilation.

They are still separate and much of
their original diameter has been
restored. A count measurement
would be much more accurate.

Skip Binary Processing Principles: 830

Continued… 826

LAS User Manual 825

Binary Processing: Principles: Continued:

In Illustration (B):
1: The image has passed though Adjust
Threshold and those grey values
falling with the Threshold limits are
coloured red. The highlights on the
disks have not fallen within the
Threshold limits and so remain white.

The Output Image from the Adjust

Threshold has converted a Greyscale
Input with grey value ranging from 0
(black) to 255 (white), to a Binary Output
Image in which those pixels selected are
set to a value of ‘1’ and those not selected
cleared to a value of ‘0’ – hence Binary
because only two values are used.

2: An enlarged disk. The string of binary

digits below the illustration
represents a single row of pixels
where the green line cuts through the
disk and highlight. The selected (red)
areas of the disk are set to ‘1’ and the
unselected highlight (white) areas
cleared to ‘0’.
There are 'holes' in the disks!

3: For a proper Area measurement

those 'holes' need to be selected and
filled. Binary Processing can do that
quickly and efficiently.

Continued... 827

826 © 2011 Leica Microsystems (Switzerland) Limited

 Binary Processing: Principles: Continued:

The Binary Pre-Filters provide the tools for

modifying the Binary Image in the same
manner as Greyscale Pre-Filters with the
difference that values of ‘1’ and ‘0’ only
are being tested.

Binary Dilation:
In Illustration (A):
1: Is the Structuring Element, an
‘electronic overlay’ with cells that are
also set or cleared to binary values. In
the illustration there are three cells all
of which are set = 1.
The centre cell is called the Origin
(coloured red) and it ‘looks’ at a single
pixel (also coloured red) called the
Input Pixel …

2: …in the Binary Input Image.

The process examines the pixels
neighbouring the Input Pixel to
determine whether the corresponding
pixel (3) in the Binary Output Image
(4) should be set or cleared. The
Structuring Element settings
determine if a neighbour is tested
(Cell=1) or ignored (Cell=0).

The process described in this illustration is

called a Dilation – if any of the neighbours
are set (=1) then the Output Pixel is also
set – which is the case in Illustration (A).
A Dilation has the effect of increasing the
number of selected (=1) pixels.

Illustration (B) shows a simplified form of

dilation:
5: The original object.

6: An imaginary 'roller' tracing around

the periphery of the object.

7: The new outline of the object.

Continued… 828

LAS User Manual 827

 Binary Processing: Principles: Continued:

In Illustration (B) the Structuring Element

has moved one pixel to the right. Now the
Input Pixel (red) and its neighbours are all
cleared (=0) and so the pixel in the Binary
Output Image is also cleared (1). The
process continues for every pixel until a
complete and new Binary Output Image
has been created.

Binary Dilation would be a most suitable

pre-filter for filling the highlight ‘holes’ in
the example image (C).

Continued... 829

828 © 2011 Leica Microsystems (Switzerland) Limited

 Binary Processing: Principles: Continued:

Binary Erosion:
1: The Structuring Element.

2: The Binary Input Image with pixels

either set (=1) or cleared (=0).

3: The Binary Output Image. Set pixels

are included in a measurement but
cleared pixels are not.

The Erosion process examines the Input

Pixel’s neighbours and if any one is
cleared (=0) then the Output Pixel is also
cleared. This has the effect of removing
selected pixels from the image.
In Illustration (A) a neighbouring pixel is
cleared (=0) and so the Output Pixel (4) is
also cleared.

The Structuring Element has moved four

pixels to the right in Illustration (B) and
now both of the neighbours are set (=1) so
the Output Pixel (5) is also set.

To determine the value of pixels on the

extreme edges of the Output Image,
Illustration (C), the Input Pixel (coloured
red) is ‘ghosted’ (6) to become a
neighbour. The illustration represents a
Dilation so the Output Pixel is set, but if
this were an Erosion the Output Pixel
would be cleared because the right-hand
neighbour is cleared.

Erosion and Dilation can be used in

combination to achieve specific results.
Dilation followed by an Erosion is called a
closing filter used for filling ‘holes’ in an
image.

Erosion followed by Dilation is called an

opening filter and is often used for
removing small details such as noise and
dust.

LAS User Manual 829

Binary Processing: The Binary Processing Panel:

1: Open the Binary Pre-Filter controls by

clicking on the entry in the main
menu.

2: In the Mode section, three active

filters can be selected as well as No
Filter by clicking on the arrows to the
right of the window and…

3: …clicking to select the required filter.

4: The Options panel contains the

Reconstruct and Fill tools.

5: Separation, which helps to separate

overlapping or touching features has
a single tool – Cut Touching –
enabled with a check box. There is a
Step control associated with this tool.

6: Three tools – Invert, Skeleton and

Edge Remove – are contained in the
Create Result panel to modify the
Input Binary Image in very specific
ways.

7: To use LAS Macro Runner, click to

enable the check box. Detailed help
for LAS Macro: Go there...

Continued... 831

830 © 2011 Leica Microsystems (Switzerland) Limited

 Binary Processing: Change the Output Colour:

1: The Show Binary control displays the

Binary Output Image as it is
modified. Generally, leave Show
Binary enabled.

2: Click the Interactive check box to

automatically start processing every
time a change is made to the filters or
controls. For complex image requiring
a large number of ‘passes’ at different
settings this can be time consuming.
Better to disable Interactive and…

3: …use the Apply button instead. In this

situation changes can be made to
settings and only processed when
the Apply button is clicked.

4: To change the default processing

colour, click on the Colour button
and…

5: …on the Select Colour dialog use the

palette, colour wheel or sliders to
select a new colour.

6: Click OK and the new selected colour

will appear on the Colour button.

LAS User Manual 831

Binary Processing: The Fill Tool:

1: The original image comprising a

group of pollen grains, ..

2: …is passed through a Greyscale Pre-

Filter > Smooth White Detail that
flattens and merges the background
leaving the selected features in
sharper contrast.

3: Using Adjust Threshold selects most

of the pollen grains but leaves some
with holes and fissures. Trying to fill
them with the Threshold Hue and
Intensity began to create noise on the
background, and so…

4: …Binary Processing was used initially

without filters and...

5: …the Fill tool only. Click the Fill check

box to enable it. This filled the holes
well but left some fissures open.

Continued… 833

832 © 2011 Leica Microsystems (Switzerland) Limited

 Binary Processing: Combine Detail and Cut Touching:

The Combine Detail filter:

1: Using the Combine Detail filter and…

2: …gradually increasing the Structuring

Element Size removes the fissures
leaving the pollen grain features well
defined ready for measurements.

The Cut Touching filter:

3: However, two of the pollen grains that
were almost touching have now
coalesced into a single feature and
have to be separated.

4: Enable the Cut Touching tool by

clicking the check box.

5: Gradually increase the Step value

until a red line (6) appears to indicate
that the features have been detected
and separated.

Continued… 834

LAS User Manual 833

Binary Processing: Discard Detail:

The Discard Detail filter:

1: The original image of diamond chips,
…

2: …has had the darker areas enhanced

with the Greyscale Pre-Filter >
Smooth Black Detail.

3: Applying Adjust Threshold has

selected most of the darker areas but
also included some mid-grey tones
that are not required.

4: Using the Binary Pre-Filter > Discard

Detail and gradually increasing the
size of the Structuring Element (5) will
shrink the selected areas, starting
with the lighter tones, until only those
required are highlighted for use in
measurements.

6: The selected feature areas are

highlighted in green, but a first step
with a Size setting of 10 has had little
effect on the Binary Image and so...

Continued… 835

834 © 2011 Leica Microsystems (Switzerland) Limited

 Binary Processing: The Reconstruct Tool:

The Reconstruct tool:

1: ...the Size setting has been increased
to 20 with the result that some of the
lighter tones have been discarded
and now show as red areas. The
green areas are those required.

2: A final setting of 30 has retained only

the very dark areas which was the
aim of using the Discard Detail filter,
and…

3: …a quick Measurements > Number

(Count) check reveals that only the
required features are included.

4: However, a part of the image has

been rejected – possibly because two
diamondchips were overlapping –
and needs to be included without
drawing in any other lighter features.

5: The Options > Reconstruct tool

targets only small, closely related
areas to include as selected.

6: The previously excluded area is now

included and whilst this would not
affect a Number (Count)
measurement, it would be significant
if Area was being measured.
To separate overlapping features
enable the Cut Touching filter. In this
case using Cut Touching with a
setting of 3 resulted in a Number
count of 4 rather than the original 2.

Separate touching features: Go there... 833

Continued... 836

LAS User Manual 835

Binary Processing: Invert and Skeleton Tools:

The Invert tool:

1: The original image – part of a section
through soft tissue…

2: …having been passed through the

Smooth White greyscale filter and the
Threshold filter to select the
predominantly white area of the
image. The selected areas are shown
coloured green.

3: Enabling the Create Result > Invert

checkbox de-selects the previously
selected areas – now displayed in
colour mauve - and selects
everything else. It is essentially a
‘swap’ tool.

The Skeleton tool:

4: Enabling the Skeleton tool erodes
selected features to the point where
only a ‘backbone’, just 1 pixel wide
remains.

5: The original image shows wool fibres.

6: The Skeleton tool applied showing

the ‘backbone’ which traces the main
Binary selection.

Continued... 837

836 © 2011 Leica Microsystems (Switzerland) Limited

 Binary Processing: The Edge Remove Tool:

1: Click to enable the Edge Remove tool

which will remove any features not
wholly within the boundaries of the
image.

2: The original image. The black

features are to be counted but only if
they are within the image.

3: The illustration shows the Binary

Output image after a Threshold
Adjust - the red objects are selected
for measurement.

4: With Edge Remove enabled any

selected objects that are not
completely within the image boundary
- coloured red on the image - are not
included in a measurement.

LAS User Manual 837

Binary Image Editing:

Binary Image Editing is a tool collection

that provides the methods for working
directly on Binary Images to add, remove,
select and de-select features. Facilities
also include drawing and filling shapes as
well as grouping features.

1: On the Image Analysis main menu,

click on the Binary Image Editing
entry.

2: The Binary Image Editing panel

appears. There are two groups of
buttons:

3: Mode and…

4: Tools. Some of the Mode buttons

affect the way that the Tools behave.

5: The Size window controls the line

thickness for some of the Tools used
for drawing.

The Binary Editing program uses three

separate images:
The original Greyscale image which
remains unchanged.

The Binary Input Image which is the

Greyscale image converted to binary
format showing the features of
interest. The factory default colour for
this is light green.

The Binary Output Image which

represents the edited image still in
binary format. The colour for this is
user selectable.

Continued... 839

838 © 2011 Leica Microsystems (Switzerland) Limited

 Binary Image Editing: Settings:

1: Display the edited Binary Output

Image by enabling the Show Binary
check box. Disabled, and the
unedited Binary Input Image as it was
supplied to the editing program is
displayed in the factory default colour
green.

2: Click on the Apply button to apply to

apply Mode changes to the Binary
Output Image.

3: The colour of selected features on the

Binary Output Image can be changed
by clicking the Colour button.

4: On the Select Colour dialog, choose a

colour from the swatches by clicking
on it or…

5: …click and drag the hue sliders or…

6: …click and drag the marker on the

Colour Wheel.

7: Click OK. The new colour appears in

the Colour button.

Continued... 840

LAS User Manual 839

Binary Image Editing: Viewer Layout and Cursor:

Binary Image editing is easier to use if the

Viewer is set to display only the Binary
Image – turn off Split Screen (1).

The Cursor colour can be toggled to either

black or white to suit the image by clicking
on the Cursor Colour button (2).

Binary editing uses 3 images:

The Input Greyscale (original) which
remains unchanged,

An Input Binary created with the

Threshold tool with the selected
features coloured green (factory
setting), and

The Output Binary which is the result of

the editing process. This will be used
with the measurement tools. The colour
of the selected features is user defines.

Continued.. 842 .

840 © 2011 Leica Microsystems (Switzerland) Limited

 Binary Image Editing: The Magnifier:

To enlarge parts of either the Input Image

or the Binary Output Image:

1: Hold down the keyboard Shift key and

click on a portion of the image to
enlarge. The magnified area tracks
the mouse movements (2). Release
the Shift key to close the magnifier.

LAS User Manual 841

Binary Image Editing: Accept Mode:

To isolate a specific area – a Region of

Interest (RoI) - on the image for
measurements:
1: Click on the Accept Mode button.
Four drawing tools are available to
create the RoI...

2: The Freehand tool, the Rectangle,

the Circle and the Ellipse. Click on the
most appropriate tool – a Rectangle is
shown on the illustrations.

3: Click on the top left corner of the

Region to be selected and holding
down the mouse button, drag to the
bottom right corner. The drawn region
is filled with a semi-transparent colour
to make identifying the selected
features easier.

4: To delete the Region and start again,

click on the Undo button.

5: When the drawn region is correct

click on the Apply button

6: Enabling Show Binary will display the

selected features in the chosen binary
mask colour.

Continued... 843

842 © 2011 Leica Microsystems (Switzerland) Limited

 Binary Image Editing: Reject Mode:

Isolate part of the image to prevent

enclosed features from being included in
the measurements as follows:

1: Click on the Reject Mode button.

2: Four drawing shapes are available:

Irregular Fill Area,
Rectangle,
Circle and
Ellipse.

Click on the appropriate shape to select it

and…

3: Click on the image and holding down

the mouse button drag down and to
the right to draw a Region of Interest
the features within which will be
excluded from the measurements.
The Region has a translucent fill.

4: To delete the Region and start again

click on the Undo button.

5: Click the Apply button to set the

rejected Region.

6: Enable the Show Binary check box to

view the rejected features in the
chosen Binary Output image colour.

Continued... 844

LAS User Manual 843

Binary Image Editing: Drawing Mode:

In Drawing Mode, lines, open shapes and

filled shapes can be drawn directly on to
the Binary Output Image.

1: Enable the Show Binary checkbox

otherwise the drawing will not appear.
On the illustration the shapes are
shown in blue for clarity; Normally,
they appear in the chosen Binary
Output Image colour which in this
case would be red.

2: For the Line and Irregular Freehand

tool, set the line thickness by clicking
on the Up/Down arrows to the right of
the Size text box.

3: When a drawing is satisfactory, click

the Apply button.

4: Use the Undo button to delete

individual drawings or the Delete All
(Trash Can) button to remove all
drawings from the image.

5: Click on the Drawing Mode button.

Draw a Straight Line:

6: To draw a straight line click on the
Line button. Click and hold down the
mouse button on the image where the
line is to begin and drag to the line
end point. Release the mouse button.

Continued… 846

844 © 2011 Leica Microsystems (Switzerland) Limited

 Binary Image Editing: Circles and Ellipses:

Draw a Filled Circle:

1: Click on the Circle tool.

2: Click and hold down the mouse

button on the image at a point
representing the edge of the Circle.

3: Drag in any direction. An outline of

the Circle is drawn as the mouse is
dragged. Release the mouse button
when the Circle is complete.

Draw a Filled Ellipse:

4: Click on the Ellipse tool.

5: Click and hold down the mouse

button on the image at a point
representing the edge of the Ellipse.

6: Drag in any direction. An outline of

the Ellipse is drawn as the mouse
moves. Release the mouse button
when the Ellipse is complete.

Continued... 847

LAS User Manual 845

Binary Image Editing: Freehand Shapes:

Irregular Freehand Shape:

1: Click on the Freehand tool button.

2: Click and release the mouse button

on the image at the shape’s starting
point.

3: Move the mouse to the next point on

the shape and click again. A line will
be drawn connecting the two points.

4: Repeat Step (3) until all of the points

around the shape are plotted and at
the final point double-click to
complete it.

Irregular Freehand Shape with Fill:

5: Click on the Freehand Fill tool button.

6: Click and release the mouse button

on the image at the shape’s starting
point.

7: Move the mouse to the next point on

the shape and click again. Line will be
drawn connecting the two points to
create a closed shape which will be
filled with solid colour.

8: Repeat Step (7) until all of the points

around the shape are plotted and at
the final point double-click to
complete it.

Draw a Filled Rectangle:

9: Click to select the Rectangle tool.

10: Click and hold down the mouse

button on the image at the top left-
hand corner of the Rectangle.

11: Drag down and toward the right-

hand bottom corner of the Rectangle.
Release the mouse.

Continued... 845

846 © 2011 Leica Microsystems (Switzerland) Limited

 Binary Image Editing: Erase Mode:

A group of features selected on the Binary

Output Image are ignored during
measurements by:

1: Enable the Show Binary checkbox.

2: Click to select the Erase button.

3: Select either the Outline, Rectangle,

Circle or Ellipse tool and…

4: …click on the Input or Output image

and draw a shape to enclose the
features to be ignored. The features
are displayed in the default colour.

5: Click on the Apply button when the

grouping is correct.

During the measurement process the

selected features will be ignored and not
included in result data.

Continued... 848

LAS User Manual 847

Binary Image Editing: Keep Mode:

Includes only those features individually

selected in the measurement data.

1: Click to enable Show Binary.

2: Click on the Keep button. The Select

tool is automatically selected.

3: Click on individual features (the

illustration cross hairs are shown
emphasised for clarity) that will be
measured.

4: Click the Apply button.

Features not selected will be ignored.

Each selected feature is displayed in the
default colour.

Continued... 849

848 © 2011 Leica Microsystems (Switzerland) Limited

 Binary Image Editing: Delete Mode:

Individual features that will not be included

in the measurements are selected in
Delete Mode as follows:

1: Enable Show Binary by clicking the

check box.

2: Click to select the Delete Mode

button. The Select tool is
automatically enabled.

3: Click on the individual features to be

excluded from the measurements.
Each is displayed in the default Binary
Edit colour green.

4: Click the Apply button.

When a measurement is made the

features selected will be ignored.

LAS User Manual 849

Binary Image Editing: Additional Controls:

1: Undo reverses the last edit action

and Redo repeats the last action.

2: Delete all actions and drawings.

3: Delete the last drawing by selecting it

with the Selection tool and then
clicking the Delete Selected button.

850 © 2011 Leica Microsystems (Switzerland) Limited

 Measure Frame: Principles:

A Measure Frame determines the area of

an image that will be used for analysis and
measurement. It can be the entire image
or more usually a selected, representative
area. In the illustration:

1: Is the edge of the image.

2: A typical Measure Frame outlined in

blue.

3: The Guard Region.

For a selected area, software rules

concerning objects that straddle or touch
the Measure Frame boundary dictate:

Any object that has its bottom right hand

pixel lying within (not on) the Measure
Frame boundary is included in the
analysis even though any of its other
pixels lie in the Guard Region – outside
the Measure Frame boundary.
Field measurements are performed within
the Measure Frame and any pixels
outside the Frame are not included.

4: This object is completely within the

Measure Frame and is included.

5: Although starting outside the Measure

Frame, the bottom right-hand pixel of
this object is inside the Measure
Frame and will be included.

6: The bulk of this object is within the

Measure Frame but because its
bottom right-hand pixel is either
touching or outside the frame
boundary it is not included.

Continued... 852

LAS User Manual 851

Measure Frame: Create and Entire Image Frame:

To Create a Measure Frame:

1: Click on the Measure Frame option in
the Processing Sequence window.

2: There are 4 Measure Frame options

available all accessed by clicking the
arrows to the right of the Frame Type
list box and clicking the required
option from the drop down list.

Entire Image Measure Frame:

3: To include the entire image in the
Measure Frame click to select the
Entire Image option. The frame
boundary coincides with the edges of
the image.

Continued... 852

852 © 2011 Leica Microsystems (Switzerland) Limited

 Measure Frame: Feature Size and Typical Frame:

Frame to Suit the Maximum Feature

Size:
The Adjust to Max Particle option creates
a frame based upon an feature size the
user enters:

1: Swipe the Particle Diameter text box

and type a value in micro-metres.

2: The Guard Region is based on:

100% of the particle diameter at the
top:
50% of the particle diameter on the
sides:
0% at the bottom.

3: To create a Measure Frame based

upon the particle size in a sequence
such as a series of Multistep images,
click to enable the Auto Import Size
from MultiStep check box. The check
box is only available if the sequence
is selected for measurement and
appended.

Typical Measure Frame:

4: The Typical Measure Frame option
will suit a wide variety of microscope
images. Based upon an ‘average’
particle size, it creates a Guard
Region with:
100% of average particle diameter at
the top:
50% of average particle diameter on
the sides:
0% at the bottom.

It is a quick and easy ‘one-click’

operation.

Continued... 853

LAS User Manual 853

Measure Frame: Manually Define Frame:

Manually Defined Measure Frame:

This option allows the user to create a
frame of specific dimensions and position
it anywhere on the image.

1: Click on the arrows to the right of the

Frame Type header and from the
drop down menu click to select
Manually Define.

2 & 3: Swipe the co-ordinate text boxes

and type, or using the up/down
arrows to the right of each box enter a
value - two co-ordinates are required
Top Left X/Y and Bottom Right X/Y –
or…

4: …click and drag the frame handles to

the required size.

5: The Measure Frame can then be

positioned by clicking and dragging
the handle in its centre.

854 © 2011 Leica Microsystems (Switzerland) Limited

 Measure Frame: Change Frame Colour:

The Measure Frame colour can be

changed by:

1: Click on the Colour button.

2: On the Select Colour dialog choose a

new colour by clicking on a pallete
well or...

3: ...clicking and dragging the Hue

sliders or...

4: ...clicking and dragging the target

mark on the Colour Wheel.

5: Click OK. The Colour button changes

to reflect the new colour.

Continued... 852

LAS User Manual 855

Select Results: Select Measurements:

Use Select Results to determine if all of

the parameters are to be measured or just
a predetermined, commonly used group.
This option is particularly useful for
images having many features with which
there would be an appreciable processing
delay to include parameters not really
needed.
Select Results is also used to determine
the results actually shown on the Grid and
Summary.

1: Click on the Select Results entry on

the main menu. The control panel is
divided into two:

2: Measurement Type: The range of

measurements to be displayed and
reported, and…

3: …Feature Measurement: The

measurements that will be made and
available to display and report.

Continued… 857

856 © 2011 Leica Microsystems (Switzerland) Limited

 Select Results: Measurement Type and Feature Measurements:

Measurement Type:
Select the range of measurements to be
displayed by:

1: Click on the arrow to the right of the

Measurement Type header to reveal
the options.

2: Click to select:
Feature: (3) Displays only the
selected feature results.

Field: (4) Displays results for Field

Data. Measurements that describe a
property of a complete Field of View
are called Field Measurements and
the data produced is called Field
Data. Parameters such as Field Area
and Field Perimeter are Field Data. It
is also so-called because it
represents the summed value for all
objects within the Measure Frame
regardless of whether they are
touching or separate This is in 7: Click to select:
contrast to Feature Data that All Parameters: All possible measurements are made but the range
produces a separate value for each can be modified: Go there...
isolated feature or 'blob' that ends
inside the Measure Frame. Predefined Parameters: A selection of the more commonly used
measurements.
Feature & Field: (5) The most
comprehensive results but with
multiple images can result in a very Continued... 858
large display and report.

Feature Measurements:
6: Click on the arrow to the right of the
Feature Measurements header to
reveal the options.

LAS User Manual 857

Select Results: Configure the Grid:

The parameters calculated and then

displayed on the Grid are determined by
the settings selected on the Select Field
Details and Select Feature Statistics.
The dialog depends upon the results tab
currently displayed, so first click on either
Feature or Field tab.

1: Click on the Setup tool to the right of

the results tabs.

2 & 3: Either the Field Details or Feature

Statistics dialog will appear.

4: Click to enable the check box to the

right of the item required to display.

5: Click OK to save the settings.

6: …click on the Show All button to

select all of the items.

7: Clicking on the Hide All button to de-

select all of the items.

8: To display the Results Grid click on

the Side Toolbar Show/Hide Grid
button. This is a toggle – click to
display, click again to hide.

Continued… 859

858 © 2011 Leica Microsystems (Switzerland) Limited

 Select Results: Results Grid Features:

1: Columns that contain results can be

order High-to-Low or Low-to-High by
clicking on the area to the right of the
column title. The small arrow that
appears shows the direction of sort.
This is a very fast option for isolating
features of interest.

2: Column width can be changed by

clicking on the dividing rule and
dragging it to the desired size.

3: Click on an entry in the Grid to

highlight the feature on the Output
Image.

LAS User Manual 859

Measurements:

LAS Image Analysis Measurements uses

the Binary Output Image of all the
selected features and applies a wide
range of fast, precise measurement
algorithms. The scope of the
measurements depends upon the All or
Predefined options in the Select Results
component.
Detailed descriptions of the Measurement
Parameters can be found in the
Appendix: Go there... 905

1: To reveal the control panel click on

the Measurements entry on the main
menu.

2: The Filter toolbar used to create and

modify Parameter limits.

3: The Parameter Filter list window.

4: To automatically apply setting

changes to the output image, click to
enable the Interactive check box.

5: Disable Interactive to allow manual

updates to images via the Apply
button.

6: On the Label panel, Parameter

determines the name that will appear
against a feature - for example, Area.

7: Use the Mode menu to select the

features to label.

8: Click on the Settings button to display

the Label settings dialog (9).

Continued… 863

860 © 2011 Leica Microsystems (Switzerland) Limited

 Measurements: Displaying the Name & Units: Changing the Label Font:

1: On the Label panel, click on the

Settings button to reveal the Label
Settings dialog:

Display the Feature Name:

2: The Parameter – Length, Number,
Diameter etc – can be displayed
alongside the feature and result by
enabling the Name checkbox.

Display the Feature Measurement Unit:

3: Enabling the Unit checkbox will
display the measurement units –
pixels (px), millimetres (mm) – next to
the feature.
Figure (4) shows how the Name and
Units displays appear.
To set the number of digits following
the decimal place on the display,
change the Measurement Display
value on the Preferences > Image
dialog. Go there...

Changing the Label Font:

5: Click on the Font button. The Font
dialog appears.

6: Click to choose a Font…

7: …a Font Style and…

8: …a Font Size (in points).

9: Click OK.

Continued... 862

LAS User Manual 861

Measurements: Label Position and Outline:

Label Position:
1: Click on the Settings button on the
Label panel. The Label Settings
dialog appears.

2: Click on the arrows to the right of the

Position header to reveal the label
position options.

3: Click to select a label display position:

Centroid displays the label in the

centre of the object.

Below-Centre displays the label below

and on the centre line of the object.

Below-Left displays the label below

and to the left of the feature.

Feature Outline:
4: To draw an outline around the
selected features, change the Outline
Width by clicking the Up/Down
(thicker/thinner) arrows to the right of
the Outline text box. A value of '0'
turns off outlining.

5: Outlined features. The Outline colour

is the same as the Label Font colour.

Continued... 864

862 © 2011 Leica Microsystems (Switzerland) Limited

 Measurements: Changing the Label Colour:

The colour of the Labels and results

displayed on the Measurements Binary
Output Image can be changed to suit the
user.

1: Click on the Settings button on the

Labels panel. The Label Settings
dialog appears.

2: Click on the Colour button. The Select

Colour dialog appears.

3: Choose a colour from the swatches

by clicking on it or…

4: …click and drag the marker on the

Colour Wheel.

5: Click OK. The new colour appears in

the Colour button.

Continued.. 861 . 864

LAS User Manual 863

Measurements: Choose the Parameter to Display:

Two options are available that determine

the range of parameters to be measured:

All Parameters which calculates a

value for every parameter against
every selected object, and...

Predefined Parameters that

calculates only a commonly used
range of parameters.

The selection is made in Select Results:

Go there... 856

The Parameter options that will be

available on the Select Measurement
Parameters dialog will depend upon the
range selected.

1: Click on the browse button to the right

of the Parameter text box.

2: If the Predefined Parameters option

has been previously selected, the
Select Measurement Parameters
dialog will display only the most
commonly used parameters.

3: If All Parameters had been chosen,

all of the possible measurement
parameters will appear - a much
longer list.

4: Select the parameter to display by

clicking to enable the appropriate
check box.

5: Click OK.

6: If All Parameters are displayed, click

and drag the scroll bar to reveal more
of the list.

7: The dialog can be enlarged by

clicking and dragging the small arrow
on the botton right corner of the
dialog.

Continued… 865

864 © 2011 Leica Microsystems (Switzerland) Limited

 Measurements: Labelling Mode:

The Labelling Mode controls the features

on the image that are labelled.

1: Click on the arrows to the right of the

Mode Listbox and...

2: ... from the options click to select:

All Features: All the features

highlighted on the Binary Output
Image are labelled regardless of
whether they fall inside limits or not.

Accepted Features: Features are

labelled that fall within specified limits.
For example, if the Area parameters
are set to find image features no
smaller than 50px2 and no greater
than 150px2, then only those features
that fell within the parameters would
be labelled. All others would be
ignored.

Rejected Features: Use this option to

label features that fall outside the set
parameters. Using the example
above, features smaller than 50px2
and greater than 150px2 would be
labelled.

None: Turns off labelling.

Continued... 864

LAS User Manual 865

Measurements: Setting Parameters Limits with Filters:

Most Parameters can accept lower and

upper limits so that only features falling
within the scope of the limits are found
and labelled.
These range pair values can be saved
together with all of the other settings as a
Configuration to be restored and used at
any time.

To Create a New Filter:

The list of available parameters will
depend upon the All or Predefined option
in Select Results.

1: Click on the New Filter button and…

2: …on the Set Measurement

Parameters dialog expand the
Parameter Set and…

3: …click in the checkbox to the left of

the desired Parameter.

4: Click OK.

5: The chosen Parameter name

appears in the Filter List with the
default lower and upper values.
These values are based upon the
image in the Viewer.

Select All or Predefined Parameter in

Select Results: 866 Go there... 857

Continued… 867

866 © 2011 Leica Microsystems (Switzerland) Limited

 Measurements: Setting Parameters Limits with Filters: Continued:

1: Set the required lower and upper

limits for the Filter by clicking in the
limit text box and typing a new value.

2: A different Filter for all appropriate

Parameters can be added to the Filter
List by repeating the process.
All parameter filters in the list will be
used during measurement
processing.

Filter Tolerance:
The lower and upper limits of a Filter can
have an applied tolerance - in terms of
pixels for un-calibrated images and µm for
calibrated images - above and below the
limit value - by:

3: …clicking the Up/Down arrows to the

right of the Pixel Limit to reach the
required value.

Continued... 868

LAS User Manual 867

Measurements: Filter Tools:

1: Delete a Filter by clicking on it in the

list and then clicking the Delete Filter
button.

2: Restore Filter limits to their default

image-based settings. When this
button is clicked a warning (3)
appears; Click OK to continue.

4: Restore Filter limits to values based

upon the current image. The lower
and upper limits actually measured on
the image are used in the filter. For
example, if the Area Filter was
originally set to 500 lower and 2500
higher, and a measurement yielded
an feature low of 549 and an feature
high of 2317, these two values would
be used in the Filter. Again, a warning
appears for confirmation.

5: Clear all Filters. Requires

confirmation.

Save the Filters as a Configuration: Go

there… 902

Continued... 869

868 © 2011 Leica Microsystems (Switzerland) Limited

 Measurements: Displaying Results in the Grid:

1: Measurement results can be displayed on screen by 3: The Grid headings are determined by the range of
clicking to enable the Grid View. results chosen in Select Results – either a
Predefined range or All Measurements. These can
2: Click on an entry in the Grid to reveal the feature on be further filtered by selecting those which are to be
the image the label of which will be shown in a displayed on the Grid.
contrasting colour. In the same way, clicking on a
feature on the Binary Output Image will highlight the 4: Most headings when clicked will display a small
results for that feature in the Grid. If a sequence is arrow to the right. Clicking on the arrow will sort the
being measured, the appropriate image will be results low-to-high or high-to-low.
automatically displayed.
Pressing the keyboard Delete key will remove the Select Results to display on the Grid: Go there… 856
selected feature from the Grid and measurement
results but it will be coloured as not included.
Reinstate the feature in Binary Editing using the
Keep tool.

LAS User Manual 869

Histogram:

The Histogram function groups results

together and displays them graphically as
a vertical or horizontal bar chart or as a
pie chart.

1: Select Histogram by clicking on the

entry in the main menu.

The Histogram control panel is divided

into three parts:

2: The Tool Bar with the display

Features or Fields button (3) and the
Display Histogram button.

4: The Settings group and…

5: The Bins configuration. On entry, the

Bins – the groups into which the
results will be collected – are
configured and the upper and lower
Bin values (6) shown on the display.

7: Each Bin can be colour coded and to

show how the features are to be
allocated to Bins, they too can be
coloured accordingly by enabling the
Show Colour Coded Image check
box.

Continued... 871

870 © 2011 Leica Microsystems (Switzerland) Limited

 Histogram: Create a New Histogram:

1: Click on the New Histogram button on

the tool bar. The resulting file will
contain the current settings and both
the Field and Feature definitions.
These are used in the report.

2: On the Create New Histogram dialog,

click inside the Name text box and
type a new name for the Histogram.

3: Click OK.

4: The name of the new Histogram now

appears in the Histogram List which is
revealed by clicking on the arrows to
the right of the Current list box
window.

Continued… 872

LAS User Manual 871

Histogram: Create a New Histogram: Continued:

1: To select the parameter for the X-

Axis, click on the Browse button to
the right of the X window and…

2: …on the Select Measurement

Parameters dialog, …

3: …expand the required parameter

group by clicking on the arrow to the
left of the heading and…

4: …clicking to choose the parameter

that will be used as the X (horizontal)
scale.
Repeat the process for the Y-Axis
(vertical) scale.
The list of parameters is determined
by the selections made in Select
Results - All 873 or 873 Predetermined
Parameter 873 list: 873 Go there... 856

5: Click on the Show Histogram button

to display the Histogram (6).

Continued… 873

872 © 2011 Leica Microsystems (Switzerland) Limited

 Histogram: Bin Setup:

1: There are three options for the Bin

scales, selected by clicking on the
arrows to the right of the Type list
window…

2: …and clicking on the required option:

Linear uses the value range between

the lower and upper results and
divides by the number of bins
selected.

Log uses the same value range and

bin count but applies logarithmic
increments.

User Defined allows the value range

to be set manually by:

3: Clicking to disable the Auto function...

4: …clicking in the Lower Limit text box

and typing the lowest result to be
displayed, and…

5: …repeating the procedure for the

Upper Limit. These settings can be
used to exclude some values from
the Histogram.

6: Set the number of bins required by

clicking inside the Bin text box and
typing a value or using the Up/Down
arrows to the right of the box.

Continued… 874

LAS User Manual 873

Histogram: Bin Setup: Continued:

1: Enabling the Auto function by clicking

the check box, will allow the software
to determine the scale values and Bin
colours.

2: Disable the Auto function and click

User Defined to set the Bin colour to a
personal preference.

3: The Define Bins dialog presents two

options…

4: …the Auto Range will configure the

Bin colours to preset shades,
whereas…

5: …Set All Same Colour displays…

6: …the Select Colour dialog so that all

of the Bins use the same display
colour. Choose a colour from the
wheel, palette or slider and…

7: …click OK.

8: The Define Bins dialog changes to

show the selected colour.

9: Click OK to finish.

10: The default Bin labels are called

'Bin' plus a sequential number, but
that name can be changed by clicking
in the Bin Name text box and typing a
new name.

Continued… 875

874 © 2011 Leica Microsystems (Switzerland) Limited

 Histogram: Display and Label Options:

1: Click the Save Histogram button on

the tool bar to save the settings so
far. Any subsequent alterations to the
Histogram settings can be saved and
retrieved at a later date to replicate
the Histogram display.

There are two check boxes at the bottom

right of the Histogram display that will
expand the scope of the display:

2: Show Axis Option when checked

reveals the Y Axis configuration
panels (4) to determine the display
mode, style and scales.

3: Enabling Show Bin Details displays

the results alongside the Histogram
and also the configuration panels for
the results and the Histogram labels.

Continued… 876

LAS User Manual 875

 Histogram: Y-Axis Options: Log and %:

The user has extensive control on how the results are

displayed on the Histogram so that data can be 3: Click inside the Range text box and type a value to
presented in the most effective way. set an appropriate scale.

The Y-Axis scale has three options - %, Log and Auto - On Illustration (B) to display the results as a percentage
as well as a Range value to display the results of all the measurements:
appropriately.
4: Click to check the % check box and enter an
1: Click to enable the Show Axis Options. appropriate Range value.

On Illustration (A) to use a Logarithmic scale:

2: Click to check the Log check box. Continued... 877

876 © 2011 Leica Microsystems (Switzerland) Limited

 Histogram: Y-Axis Options: Auto Configuration:

The program can setup the scale and the

bin ranges by using the Auto function:

1: Click on the Auto check box to enable

it.

2: With the Limits button selected the

range of results allocated to each bin
is shown along the X-Axis.

3: Click the Names button to display the

Bin Name along the X-Axis.

Continued... 878

LAS User Manual 877

Histogram: Display Settings:

1: Enter a title for the Histogram by

clicking in the Title text box and
typing.

2: The Mode establishes how the results

are spread across the bins. Click on
the arrows to the right of the Mode
header and...

3: ...click to select the required option:

Differential displays the results in

each bin as a direct value against the
Y-Axis scale.

Cumulative+ adds the results

progressively across the Histogram to
display an ascending ramp.

Cumulative- subtracts the results

across the bins resulting in a declining
ramp.

4: Choose the display Style (illustrated

on the following page) by clicking on
the arrows to the right of the Style
header and from the menu...

5 ...clicking to select a style.

Continued... 879

878 © 2011 Leica Microsystems (Switzerland) Limited

 Histogram: Display Settings: Pie and Horizontal Bar:

Examples of (1) Pie Chart display style and (2) Horizontal Bar chart.

LAS User Manual 879

 Histogram: Show Bin Details and Statistics:

Both the details of the results in each bin and a list of The statistics are revealed by clicking on the Statistics
statistics are available by clicking to enable the Show tab (3).
Bin Details check box (1).

Click on the Bin Details tab (2) to display a Continued... 881

comprehensive breakdown of the results in each bin.

880 © 2011 Leica Microsystems (Switzerland) Limited

 Histogram: Field and Feature Options:

The illustrations so far have shown the

results for all of the measured features,
but the Field option will display results
based upon the Field(s) in terms of the Y-
Axis parameter.

Switch between displaying Field or

Features by:
1: Click on the small arrows to the right
of the Field/Feature header and...

2: Click to select the display option

required. There is no need to close
the Histogram before changing
display options.

3: The Field Histogram.

Continued... 882

LAS User Manual 881

Histogram: The Toolbar:

1: Create a New Histogram.

2: Edit Histogram Settings allows the

name of the Histogram currently
selected in the list to be changed.
Click in the Name text box on the
dialog and type a new unique name.
Click OK.

3: To delete all of the Histogram settings

click on the Trash Can. Confirm the
deletion on the dialog. This action
cannot be reversed.

4: Save the Current Histogram settings.

5: Open Histogram settings. Click on the

arrows to the right of the Current drop
down menu and click to select the
required Histogram from the list. Click
the Open button to load the settings.

6: Show the Histogram. Displays the

current Histogram - hide it by using
the usual controls on the upper right
hand of the Histogram display.

882 © 2011 Leica Microsystems (Switzerland) Limited

 Histogram: Reference Data:

User Reference Data is attached and

displayed on the Sequence Process
Report.
The entries are simply text strings – and
alpha/numeric characters are acceptable
– entered into the appropriate text boxes
on the Reference Data panel.

1: Click on the Reference Data entry in

the main menu to reveal the entry
panel (2). Seven default topic text
boxes are available – Experiment,
Preparation, Specimen, Keywords,
Result, Observation and Technologist
- although others to suit the user may
be added.

3: Click in a topic text box and type an

entry.

4: The Reference Data is saved by

creating a Configuration.

Create a Configuration: Go there... 902

Create a Report: Go there... 885

LAS User Manual 883

Histogram: Reference Data Edit and Hide, Reveal or Add a Topic:

To edit any of the Reference Data

topics
1: Click on the Edit button.

2: On the dialog (the heading shows the

topic selected) type the revised text.
Line returns are acceptable.

3: Click OK.

4: Edit another topic by navigating with

the Previous and Next buttons.

To Hide, Reveal or Add a New Topic:

5: Click the User Define button. The
dialog shows the existing topics and
their display status. Click on a Topic
check box to hide or display it.

6: Add a new Topic by clicking the New

button. On the dialog type the new
Topic Name. It will immediately
appear in the list.

7: Hide or reveal the selected Topic with

the Visibility button.

8: Click to reveal all topics.

9: Click to hide all topics.

10: Click the Up/Down buttons to move

the selected Topic up or down the list.

11: Click Delete (Trash Can) to remove

the selected Topic.

12: Click OK.

884 © 2011 Leica Microsystems (Switzerland) Limited

 Create Report: Select Report Contents:

Reporting in LAS Image Analysis is both

powerful and comprehensive. The reports
save all of the data generated by a
Processing Sequence in a Microsoft Excel
spreadsheet. A standard Excel-compatible
template is supplied with Image Analysis
and data can be stored using it providing
Excel is installed on the user's computer.

1: Click on the Create Report entry in

the main menu to open the control
panel.

2: The Report Contents are selected

from the list by enabling the check
boxes. Click on the arrow to the left of
the headings to reveal the contents
and…

3: …click to select the content item in

the check box.

4: Click and drag the scroll slider to

display further headings.

Template styling and layout can be

changed by users using the basic Excel
syntax, but before attempting changes
make a copy of the original and work on
that. See following page.

Continued… 886

LAS User Manual 885

Create Report: Select Images and Template:

1: Images can be displayed in the report

by clicking to enable the Image
Sequence or Groups header and
then…

2: …clicking to select the individual

images. No images are selected as a
default with a maximum of 5
selectable.
It is recommended that the number of
images selected be kept to a
minimum otherwise problems with
Excel memory usage could result.

Selecting the Template:

3: Click on the Browse button to the
right of the Template window and…

4: …on the Select Template Dialog click

to select the required template. A
comprehensive template (LAS
Analysis Template 1.xlt) is supplied
with LAS Image Analysis. It can be
found at:

My Computer>Shared Documents>
Leica Application Suite>
Analysis>Templates

...and can be copied to another file

name and used as the basis for end
user templates.

5: Click Open and the template name

appears in the Template window.

Continued… 887

886 © 2011 Leica Microsystems (Switzerland) Limited

 Create Report: Report File name:

1: To select a file in which to save the

Report, click on the Browse button to
the right of the Report window and…

2: …on the Select a Report File dialog

navigate to the required folder,…

3: Type a file name and…

4: ...click Open. The default file path is:

My Computer>Shared Documents>
Leica Application Suite>
Analysis>LAS Analysis Report.xls

...but can be changed to any location

using Windows navigation.

5: If the file name already exists, confirm

(or abort) the overwrite.

6: Click the Display Report check box to

display the report immediately.

7: Click Export to generate the report.

Continued… 888

LAS User Manual 887

Create Report: Report Example: Analysis Summary Front Page:

1: The Image Analysis Template comprises 6 individual Excel sheets indicated by tabs along the bottom edge.
Specific data in the sheets is linked to the front page to provide an analysis.

2: The Analysis Summary Front Page. Layout detail can be altered by the user.

888 © 2011 Leica Microsystems (Switzerland) Limited

 Create Report: Report Example: Results:

LAS User Manual 889

Create Report: Report Example: Results:

890 © 2011 Leica Microsystems (Switzerland) Limited

 Create Report: Report Example: Statistics:

LAS User Manual 891

Create Report: Report Example: Histogram:

892 © 2011 Leica Microsystems (Switzerland) Limited

 Create Report: Report Example: Images:

Excel display of a selected image with the

Binary Masks produced during a Process
Sequence.

LAS User Manual 893

 The Process Sequence: Introduction and Toolbar:

The Processing Sequence feature of LAS Image Optionally, a comprehensive report can be generated
Analysis takes groups or sequences of images and and displayed when processing is complete.
applies any or all of the other tools and their settings Individual images can also be grouped and processed
automatically to produce rapid results, very often without even if they have differing tool settings.
any further user intervention.
The sequence can be programmed to halt at any tool to
For sequences – colour or monochrome - initial analysis allow refinement – deleting or separating features for
is carried out on a single image with the program example – before resuming the sequence.
‘remembering’ each step. Then, with a single button click
all of the images will be processed using the step The Processing Sequence can be saved as a
settings. Configuration to be recalled and re-run at any time.

894 © 2011 Leica Microsystems (Switzerland) Limited

 The Process Sequence: Toolbar:

1: Create a New Processing Sequence.

Clicking this button clears all existing
sequence information so needs to be
confirmed on the dialog.

2: Run Processing Sequence. Starts an

existing sequence.

3: Clicking Next Step stops the

automatic run and allows the user to
move through the sequence step-by-
step. Click the Run button to resume
automatic processing.

4: The Pause button will stop the

sequence at any time. Click it again to
resume.

5: Stop the sequence. Requires

confirmation on the dialog.

6: The Update button refreshes the

processing sequence if any changes
are made to individual images.

Continued… 896

LAS User Manual 895

The Process Sequence: Selecting Image Sequences:

1: The individual images shown in the

Gallery illustration are part of a
sequence with…

2: …the composite labelled MultiStep.

There are a total of 25 images in the
sequence.

3: If necessary, click the arrow to the

right of the Sequence panel header to
expand it.

4: Load the first image to the Sequence

Image List by clicking the Append
button.

5: The image name appears in the list.

6: The program ‘recognises’ the image

as part of a sequence and the
Append Sequence button becomes
active. Click to load all of the images
to the list.

Continued… 897

896 © 2011 Leica Microsystems (Switzerland) Limited

 The Process Sequence: Selecting Image Sequences: Continued:

1: Remove individual images from the

list by clicking to select the image and
then clicking the Remove button.

2: To clear the list completely, click on

the Reset button.

3: Sort the images in numerical order by

enabling the Sort check box.

4: To output a sequential list containing

all of the image results, click the
Options button and...

5: ...from the Process Settings menu,

enable the Accumulate Results check
box.
If it is left unchecked results from
individual images are displayed on
the Grid during the Processing
Sequence and at the end by clicking
on an entry in the Sequence List.

6: The Export Report check box when

enabled will automatically create a
report when processing is finished.

7: To display a Histogram for each

image as it is processed click to
enable the Show Histogram check
box.

8: Enable the Refresh Display check

box to refresh the display between
each image analysis.

9: The Grid results display will be

updated for every image if the
Refresh Results check box is
enabled.

10: Click the 'X' to close the Process

Settings menu.

Setup Report settings 898 : Go there... 885

Continued... 898

LAS User Manual 897

The Process Sequence: Selecting Image Groups:

To load individual images into the Image

List:

1: Press and hold either the Ctrl or Shift

key on the keyboard and…

2: …click on each of the thumbnails in

the Gallery of the images to be
included. Those selected will have a
brown frame.

3: Click on the Append button and...

4: ... the image names will appear in the

list.

Continued… 899

898 © 2011 Leica Microsystems (Switzerland) Limited

 The Process Sequence: Create a New Processing Sequence:

1: Click on the New Sequence button.

Because this will delete all existing
Processing Sequence results…

2: …confirm a new set of

measurements by clicking Yes at the
dialog. Click No if the current settings
are going to be required later and
save them to a configuration.

3: Select a typical image from the list

and apply the processing tools in the
usual way. As settings are made, the
program keeps track of them
displaying a green tick mark to the left
of the name to indicate normal
processing with the current settings.

4: Some images may require specific

additional processing – for instance,
artefacts may need to be removed on
an image-by-image basis using the
Binary Edit tool. The Processing
Sequence can be configured to halt at
any tool or to skip a tool completely by
clicking to the left of the tool name
until…

5: …a red cross appears to indicate this

tool should be skipped or…

6: … two green vertical bars are shown

to denote that the Processing
Sequence will pause at this tool on
every image for further manual
changes to be made.

Saving settings as a Configuration: Go

there... 902

Continued… 900

LAS User Manual 899

The Process Sequence: Images with Different Settings:

Images in a sequence or group can have

different filters or setting applied to them;
The settings are automatically applied by
the Processing Sequence.

1 & 2: In the illustration two images from

the same sequence have different
Threshold settings. Those for the first
image would apply to all images in the
sequence or group because they
were made first.
Subsequently, the Threshold for
image (2) was changed but this would
not affect the remaining images, only
image (2).

3: When changes are made to an

individual image the Update button
must be clicked to apply the new
settings to the Process Sequence.

Continued… 901

900 © 2011 Leica Microsystems (Switzerland) Limited

 The Process Sequence: Working on Other Images:

Sometimes, during the setup of a

Processing Sequence it may be
necessary to work on an image not
included in the Sequence List. LAS Image
Analysis is flexible enough to allow
processing on a separate image whilst
retaining the Processing Sequence
information.

1: The image, although part of the

sequence, has not been included in
the Sequence Image List. However,
work can be carried out on the image
without affecting the Process
Sequence.

2: Having completed processing the

separate image, create a new
Configuration. This will include all of
the current settings including those of
the separate image, so its data can
be recalled later.

Starting the Processing Sequence:

3: With the settings complete, click on
the Run Sequence button. Each
image will be processed in turn and
the results displayed in the Grid if it is
enabled. The image currently being
processed is highlighted in the Gallery
.

4: At any time the process can be halted

and stepped through single process
at a time by clicking on the Next
button.

All of the Process Sequence data is saved

in a Report as a Microsoft Excel file: Go
there… 885

LAS User Manual 901

Configurations: Tools:

The current settings for most editing

panels can be saved with the archive as a
Configuration to be retrieved at a later
date. Retrieved Configuration settings are
applied to automatically to the tools

1: Each configuration has a unique

name and can be accessed by
clicking on the arrow to the right of
the Current Configuration window and
from the drop down list…

2: …clicking to select the required

configuration.

3: Click the Display Configuration

Settings button to list the
configuration details.

Save Configuration:
Saves all of the current settings and
process sequences in a unique file.

4: To save the current settings as a new

configuration, click on the New button
and…

5: …type a unique name for the new

configuration.

6: Click OK to save the setting.

7: Click on the Save Configuration

button The new configuration appears
in the drop down list.

Continued... 903

902 © 2011 Leica Microsystems (Switzerland) Limited

 Configurations: Edit, Remove and Print:

Edit Configuration:
A Configuration name can be changed by:

1: Select the configuration to be

changed from the drop down list.

2: Click on the Edit Configuration button.

3: On the Edit Configuration dialog,

change the name by clicking in the
text box and typing a new name
and…

4: …clicking OK.

5: Click the Save Configuration button.

Remove a Configuration:
A Configuration can be removed from the
list by:

6: Selecting the configuration to be

deleted from the drop down list.

7: Clicking the Delete (Trash Can)

button.

8: Confirm the deletion and the

Configuration will be removed
permanently. The operation cannot be
reversed.

Print a Configuration:
Check that the printer is on and connected
to the computer. The configuration is
captured to an Excel spreadsheet and
printed from there.

1: Select the Configuration to be printed

from the drop down list. When the
Configuration settings appears on an
Excel spreadsheet, use the print
function to print the settings list.

LAS User Manual 903

Appendix:

This Appendix contains useful reference

information. The word 'Feature' is used to
indicate parts of the image that are of
interest and need to be measured.

904 © 2011 Leica Microsystems (Switzerland) Limited

 Parameter Descriptions: Number and Size:

All measurements are quoted in calibrated

units.

Number:
1: Number: Carries out a count of all the
selected objects on the Binary Output
Image.

Size:
2: Area: Measures the Area of every
selected object.

3: Convex Area: Derived from the mean

Feret diameter approximating to the
enclosing polygon.

4: Equivalent Circle Diameter: The

diameter of a circle that has the same
area as the feature.

Continued... 906

LAS User Manual 905

Parameter Descriptions: Size: Continued:

1: Length: The greatest distance

between parallel lines drawn through
2 points on a feature’s boundary
regardless of orientation. Also called
Max Feret Diameter.

2: Breadth: The shortest distance

between parallel lines drawn through
2 points on a feature's boundary
regardless of orientation.

3: Orthogonal Feret: The Feret diameter

perpendicular to the Max Feret
diameter or length.

4: Feret 0: The greatest horizontal

distance (width) measured in the
horizontal direction.

5: Feret 90: The greatest vertical

distance (height) measured between
parallel lines in the vertical direction.
Feret 0 and Feret 90 are the
equivalent of the width and height of a
Bounding Box around the feature.

Continued…. 908

906 © 2011 Leica Microsystems (Switzerland) Limited

 Parameter Descriptions: Shape:

1: Roundness: The perfect circle has a

notional value of 1.0. Any variations
from the circular are reflected by an
increase in the notional value.

2: Aspect Ratio: Object length divided by

object breadth or Feret Length/Feret
Breadth.

3: Fullness Ratio: The square root of the

object area divided by the objects
Convex Area. v(A/CA).

Continued… 909

LAS User Manual 907

Parameter Descriptions: Size and Boundary:

1: Curve Length: The Curve Length can

measure the actual length of an
irregular feature – a piece of string for
example - the curves of which may
overlap.

2: Curve Width: Measures the width of

the same piece of string.

Boundary:
3: Perimeter: Count in µm of the
distance around the Perimeter of a
feature. It includes all inlets and
projections and compensates for the
edge orientation.

4: Convex Perimeter: Derived from the

Feret diameter and approximates to
the perimeter of the enclosing
polygon.

5: Vertical Projection: The pixel count

representing the shadow cast by a
feature if light were impinging upon it
from a vertical direction.

6: Horizontal Projection: The pixel count

representing the shadow cast by a
feature if light were impinging upon it
from a horizontal direction.

Continued… 907

908 © 2011 Leica Microsystems (Switzerland) Limited

 Parameter Descriptions: Position:

FCP = Feature Count Point and

represents the lowest and rightmost pixel
in the feature.

1: X-FCP is measured from the left-

hand edge of the image to the
Feature Count Point, and…

2: Y-FCP is measured from the top

edge of the image to the Feature
Count Point.

Centroid is the Centre of Mass of an

feature. Its position is measured from the
edges of the image by the co-ordinates…

3: X Centroid: The distance from the left

hand edge, and…

4: Y Centroid: The distance from the

feature’s top edge.

The feature's position co-ordinates on the

image are measured with:

5: X-Min and X-Max: The boundary pixel

closest to and furthest from, the left
hand edge of the image.

6: Y-Min and Y-Max: The boundary pixel

closest to and furthest from, the top
edge of the image.

Continued… 910

LAS User Manual 909

Parameter Descriptions: Orientation:

1: Orientation: The angle of the Feret

Length to the horizontal.

2: Orthogonal Orientation: The angle

perpendicular to the Orientation
(Feret Length) - essentially the
Orientation minus 900).

3: Derived Orientation: The angle

representing a line plotted through a
range of pixel co-ordinates.

Continued… 911

910 © 2011 Leica Microsystems (Switzerland) Limited

 Parameter Descriptions: Topology and Intensity:

Topology:
1: Forks: The number of downward
projecting fork-shaped limbs.

2: Joins: The number of points at which

a projection (limb) joins the feature.

3: Tops: The count of upward-extending

projections (limbs).

4: Ends: The number of downward-

extending projections (limbs).

Intensity:
5: Integrated Grey: Sum of the grey
value of all of the pixels within the
feature.

6: Grey Mean: Sum of the value of all of

the pixels in the feature (Integrated
Grey) divided by the number of pixels
in the feature (Equivalent to Area).

7: Grey Variance: Result of the value of

the darkest pixel in the feature
subtracted from the lightest pixel
value.

LAS User Manual 911

 Macro Runner

LAS Macros automate image processing, analysis and Macro programs are run within the LAS environment either
measurements for quantitative microscopy. The using the LAS Macro Runner or in combination with LAS
richness of image processing functions in LAS can be Image Analysis. LAS Macros are included in the analysis
adapted to a diverse range of imaging tasks. LAS Macro sequence at the step for Image Processing or Binary
allows repetitive tasks to be customised to the needs of Processing. This combination makes a complete application
particular applications, optimising imaging solutions in a solution that can be repeatedly used by operators with no
wide range of fields. This versatile software processes specialist knowledge of LAS Macros.
images obtained by Leica digital cameras and digital
microscopes by the software of the Leica Application By adding the versatility of LAS Macros to the automation of
Suite. the LAS Image Analysis sequence or to the simplicity of the
LAS Macro Runner, provides an efficient solution to
The LAS Macro Editor defines the instructions for image demanding and unconventional tasks in analytical
processing, binary processing and measurement. An microscopy.
LAS Macro routine with its image processing instructions
is very much like a conventional computer program. The For detail information about the optional LAS Macro Runner,
difference is that you do not create it by typing in its please refer to the LAS Macro Editor help file. Go there... 912
statements character by character; rather, you create it
interactively, using facilities from the panels of the LAS
Macro Editor. An instruction is created automatically and
inserted into the LAS Macro by pressing the Insert
button next to the instruction in the Pending Instruction
window. There is no need to write any software code!

912 © 2011 Leica Microsystems (Switzerland) Limited

 Phase Expert

Optional module Phase Expert has been designed to measure precisely up to ten
different phases – regions of the image that can be identified by their homogenous
colour or grey level.

For example, regions of differing reflectivity in oil-shale; Colours due to the polarised
light of different constituents in a rock section or stain variations in tissue or bone
sections. Phase Expert can determine the occurrence of these phases both in terms of
the overall image or with reference to just one of the selected phases.

Samples are typically embedded in a resin billet with the image face ground and
polished to good reflective flatness.

Single or multiple images can be processed automatically once any required grey
processing filters specified and thresholds set.

Results can be displayed on screen in a variety of formats both as graphics and as

tabular in the Grid. And they can be saved as a report to Microsoft Excel Spreadsheet
for easy distribution. Displayed results and reports provide wide scope for tailoring to the
user’s needs.

Phase Expert works in conjunction with Image Analysis which must be installed,
licensed and enabled.

LAS User Manual 913

Phase Expert: Step by Step:

These steps represent the sequence users should follow to

successfully carry out Phase measurements.

Because Phase Expert works in conjunction with Image Analysis, in

most steps links are provided to both modules:
Red, will take the reader to the appropriate place in the Image
Analysis (Automatic) help file for more detailed information, and
Blue, connects to help files that are specific to Phase Expert.
Sometimes, it will be beneficial to follow both links if they are
provided.

Set up the Phases:

Each of the Phases to be measured is colour-coded and that colour
is used on the processed image to distinguish it from other Phases.
Names and colours of the Phases can be selected by the user.
Up to 10 different Phases can be chosen for measurement.
Phase Expert help >>> 916

Select the Images to Process:

One or more images can be processed in sequence simply by
selecting the Thumbnail(s) in the Gallery and then clicking the
Append button on the Select Images to Measure panel.
Image Analysis help >>> 782 Phase Expert help >>> 920

Choose the Reference:

Phase Expert provides two paradigms for determining the
measurements:
Measure each Phase against the Field – the entire image, or:
Measure each Phase against a selected Phase called the
Reference.
If the entire Field is to be used check that none of the phases have
the (Ref) marker set against and go to the next step.
Phase Expert help >>> 919

Set the Measure Frame:

The Measure Frame determines the part of the Field that is
measured and analysed.
Phases lying outside the Measure Frame will be ignored.
For Phase Expert the usual option is Entire Image (Field).
Image Analysis help >>> 851 Phase Expert help >>> 922

Image Processing Pre-Filter:

A wide and powerful range of filters to improve Phase boundaries
and recognition making measurement and analysis faster are more
precise.
Image Analysis help >>> 797

Continued... 915

914 © 2011 Leica Microsystems (Switzerland) Limited

 Phase Expert: Step by Step: Continued:
Adjust Thresholds:
Adjust Thresholds allows the upper and lower pixel values for each Phase to
be adjusted so that they accurately represent the desired boundaries. The
Phase detection process uses the pixel range values set during Threshold
adjustment.
Clear, easy to use Histogram shows each Phase in its own colour with a
simple Bar display beneath.
Range adjustment can be made by dragging on the Histogram or by
entering values.
Two modes – Continuous and Overlapping – are available.
Image Analysis help >>> 814 Phase Expert help >>> 923

Binary Processing Pre-Filters:

The Binary Pre-Filters provide the tools for modifying the Binary Processed
Image in the same manner as Greyscale Pre-Filters work on the original
image but with finer precision – small, individual details can be targeted and
modified.
Discard and Combine details.
Fill Holes and Separate Touching details.
Image Analysis help >>> 830 Phase Expert help >>> 929

Binary Image Editing:

Binary Image Editing is a tool collection that provides the methods for working
directly on processed Binary Images to add, remove, select and de-select
details.
Facilities also include drawing and filling shapes as well as grouping
features.
Image Analysis help >>> 838 Phase Expert help >>> 930

Results and Histogram:

There is a wide range of options for displaying Phase Expert results both as
graphics – Bar and Pie charts – and tabular in Detail and Summary.
Create personalised layouts and save as re-usable Configurations.
Display tabular results for multiple fields as aggregates or individual fields
and phases.
Image Analysis help >>> 870 Phase Expert help >>> 931

Reference Data:
A comprehensive range of Data Items can be appended to the Phase Expert
results that will identify important details such as the Project Name, the
Specimen and how it was prepared. Enter the information in the Reference
Data dialog.
Administrators can add to the supplied list of data headings to comply with
corporate demands.
Image Analysis help >>> 883 Phase Expert help >>> 939

Create Report:
Phase Expert Reports are created and saved in Microsoft Excel spreadsheet
format. Excel must be installed on the computer to create the report but only
the Excel Viewer is needed to display it. Images can be included and each
report is structured to show the results comprehensively on individual sheets.
A flexible Template is supplied with Phase Expert.
The default Template can be modified to suit the user’s needs.
The report can be displayed as it is created.
Image Analysis help >>> 885 Phase Expert help >>> 940

LAS User Manual 915

 Phase Expert: Set up the Phases:

Phase Expert measurements can

function in either of two modes:

A: Field Mode: In which each of the

phases Green (1), Red (2) and Blue
(3) in the diagram are measured with
respect to the entire Visible Field (4).

B: Reference Mode: The phases Blue,

Red and Yellow are measured with
respect to another phase coloured
Green (4) in the illustration, that
generally encompasses the others.
The Field parameters are ignored in
the measurement comparisons but
are reported.

5: Reference Mode is selected by

clicking the Tick button on the tool
bar. Clicking again turns off
Reference Mode.

6: When Reference Mode is active the

tag ‘Ref’ appears next to the chosen
phase. In the illustration the word
Reference has been typed in by the
user.

Continued… 917

916 © 2011 Leica Microsystems (Switzerland) Limited

 Phase Expert: Set up the Phases: Continued:

Each phase to be measured is colour-coded

on the binary output image. Set up the
phases by:

1: Click on the arrow to the right of the

Phase header to reveal the panel. On
opening a single phase with the default
name Phase 1 is automatically
displayed.

For some jobs a single Phase will be

sufficient, but many images will contain
several Phases all of which need to be
identified. In these cases add further
Phases as described on the following
pages.

2: Change the Phase Name by clicking in

the name text box and typing a new
name.

3: To change the Phase Colour, double-

click on the colour and…

4: …the Colour dialog appears. Choose a

colour from the swatches, wheel or hue
slider or type the required values.

5: Click OK.

6: An example of a Phase Name and

Colour change.

Continued… 918

LAS User Manual 917

 Phase Expert: Set up the Phases: Continued:

1: Add further Phases by clicking on the

Create New Phase button. Change the
Phase Name and Colour using the
procedure described on the previous
page.

As additional Phases are added they are

given a sequential number shown in the
left-hand column. The lowest number
represents pixel values close to 0, the
black end of the greyscale. Each
additional Phase represents a higher
span of pixel values so the three Phases
in the illustration might represent the
greyscale pixel values:

Phase 1: Red: 0 (Black) to 72.

Phase 2: Blue: 73 to 154.
Phase 3: Green: 155 to 255 (White).

...for example.

The initial range values are established

by software based on a broad analysis of
the image. The user will adjust them
during a later step.

2: Delete a Phase by clicking to highlight it

and then clicking on the Trash Can
(Delete) button. If a Phase is deleted by
mistake, immediately click the Create
New Phase button and it will be restored.

3: Navigate through the Phases using the

Previous (Left) and Next (Right) buttons.

Continued... 919

918 © 2011 Leica Microsystems (Switzerland) Limited

 Phase Expert: Choose the Reference Phase:

In this example, flakes of two minerals - dark

grey (1) and light grey (2) on the image, exist
blended together. The task is to determine
what percentage of each is represented by
the blend. The area of both minerals
together (3) is represented by a combined
phase called the Reference.

The first step is to measure the area of

both minerals to determine the combined
Reference..

The next step is to measure Mineral 1

separately and finally...

...measure Mineral 2 separately.

4: The combined measurement - the

Reference - is colour-coded red. Mineral
1 is coloured Blue and Mineral 2, green.
The colours have been set up as phases
with appropriate names.

5: Set the Reference phase - the combined

measurement - by clicking on the phase
and then on the Set Reference button.
(Ref) appears against the phase to
indicate it is the Reference.

To see the outcome of this example: Go

there… 926

Continued… 920

LAS User Manual 919

Phase Expert: Select the Images to Measure:

1: On the Processing Sequence click on

Select Images to Measure.

2: Click on a Thumbnail in the Gallery. If a

single image is to be processed it does
not have to be added to the Sequence
Image List. Continue only if multiple
images are going to be processed.

3: Click to Append button on the Select

images to measure dialog.

Repeat the steps to add more images to

be measured as a batch.

All selected images must have the same

type - for example .bmp, .png, .jpg -
although it is recommended that images
of type tif or jpg are used because these
are not compressed. Sometimes
artefacts of the compression process are
seen in formats using compression.

4: Images can be selected in any order

from the Gallery. To sort them into
numerical sequence click to enable the
Sort check box.

5: Use the Remove button to remove a

single selected image from the list, or
click the Reset button to remove all
images.

920 © 2011 Leica Microsystems (Switzerland) Limited

 Phase Expert: Multiple Image Processing:

When multiple images have been added to

the Image List, the process so far has been
carried out on just one of them. But the
program 'remembers' the settings so by
clicking on the Run Sequence button (1) all of
the images are processed one after the other
and measured using the same settings. This
is sometimes called Batch Mode.

The image (2) shows all of the phases

displayed together. Use the Show All check
box on the Adjust Threshold panel to see this
result.

For further information about the Processing

Sequence see Image Analysis help: Go
there... 894

Continued... 926

LAS User Manual 921

Phase Expert: Set the Measure Frame:

1: In the Processing Sequence click to

select Measure Frame.

2: On the Measure Frame panel click on

the arrows to the right of the header.

3: Select the Entire Image option from the

drop down menu. Although other options
are available and can be used, Entire
Image is the usual selection for Phase
Expert.

922 © 2011 Leica Microsystems (Switzerland) Limited

 Phase Expert: Adjust Threshold:

Adjust Threshold allows the pixel range

values of each phase to be adjusted so that
they accurately represent the phases
boundaries. The phase detection process
uses the upper and lower pixel values set
during the Threshold adjustment.

1: Click to select Adjust Threshold on the

Processing Sequence menu.

Colour images use the colour wheel dialog

(2) that allows Hue and Saturation to be set
on the Colour Wheel (3) and Intensity
controlled with the Slider (4). Actual values
can be typed into the Colour Range text
boxes (5).

Monochrome or greyscale images use the

Histogram panel (6) that also has Colour
Bars that, in some instances show how pixel
values overlap each other. Values can be
typed into the text boxes (8). The value range
on the Histogram can be adjusted by clicking
and dragging on the vertical dashed bars
when the cursor is represented by a double-
ended arrow (9), or the entire range can be
shifted up or down when the four-ended
arrow (10) is displayed.

The following pages illustrate Threshold

adjustment for both colour and mono images.

Continued… 924

LAS User Manual 923

 Phase Expert: Adjust Threshold: Field Mode:

In this example three phases are going to be

detected and measured with respect to the
entire visible field, hence Field Mode.

On original image (1), the three areas of

interest are the yellow flecks, the orange
speckles and the pale blue blobs that
surround the yellow area. Each area will be
detected in turn starting with the yellow flecks
– Phase 1 detected in Red.

2: On the Phase panel, click to select the

Phase 1 (Red).

3: Enable Interactive and Show Binary and

click the Reset button to remove any
existing detection.

4: Disable Accumulate.

5: Click on part of the area of interest – in

this case the yellow flecks – and drag a
small rectangle. The pixel values
contained within the rectangle are
immediately displayed on the Colour
Wheel and in the text boxes and the
binary image (7) will be shaded with the
phase colour – in this case Red..

6: Click to enable Accumulate. This will add

any further selections to the existing.
Keep repeating step (5) to include any
(yellow flecked) area not yet detected.
Make fine adjustments on the Colour
Wheel, Intensity Slider and in the Colour
Range text boxes.

7: The final image with Phase 1 (Red)

detecting all of the yellow flecks.

Continued… 925

924 © 2011 Leica Microsystems (Switzerland) Limited

 Phase Expert: Adjust Threshold: Field Mode: A Completed Sequence:

The process for detecting the two remaining

phases – orange speckles and pale blue
blobs, Phase 2 and 3 – is the same as
threshold detection for Phase 1 but selecting
relevant areas on the image.

1: Click to select the phase to be detected.

2: Select and detect an appropriate area of

the image and adjust the threshold on
the Colour Wheel controls.

3: The result of Adjust Threshold for the

orange speckles – Phase 2 Blue, and…

4: …for Phase 3 Green, the pale blue

blobs.

Continued... 926

LAS User Manual 925

 Phase Expert: Adjust Threshold: Reference Mode: Detect the Reference Phase:

1: Click on the Reference phase on the

phase list. This indicates the phase
that is going to be selected. In this
example the Reference is going to
include both Mineral 1 and Mineral 2.

2: Click to enable Show Binary,

Interactive and also on Reset to clear
any automatic detection.

3: Click to disable (OFF) Accumulate.

4: Disable the Continuous function on

the Histogram.

5: Select an area that represents either

of the minerals, click and drag to
create a region. All of the pixels
values within the region will be
displayed on the image and on the
Histogram.

6: Click to enable (ON) Accumulate and

repeat procedure (5) on another part
of the minerals that is not yet
detected. The Accumulate feature
will add the selection to the previous
values. Continue like this until all
areas of the two minerals are
detected and combined.

7: Adjust the detected phases by

dragging the vertical dotted lines on
the Histogram until only the areas
covered by Mineral 1 and Mineral 2
are represented. Everything else is
ignored.

8: On the Histogram the different

phases are allowed to overlap
because Continuous is disabled.

9: The Reference phase binary image.

Continued… 927

926 © 2011 Leica Microsystems (Switzerland) Limited

 Phase Expert: Adjust Threshold: Reference Mode: Continued:

Detect Mineral 1 that is colour-coded Blue

on the phase list.

1: Click to select the Blue (Mineral 1)

phase.

2: Follow the steps on the previous

page to start the detection but this
time select a region that represents
only Mineral 1, repeating the
selection until all of the phase has
been detected.

3: Fine tune the detection with the

Histogram controls – notice how the
Mineral 1 phase (Blue) overlaps the
Reference (Red) phase.

4: The Mineral 1 phase binary image

after detection.

Continued… 928

LAS User Manual 927

 Phase Expert: Adjust Threshold: Reference Mode: Continued:

Detect Mineral 2, colour-coded Green on

the phase list.

1: Click to select the Green (Mineral 2)

phase.

2: Follow the steps on the previous

pages to start the detection but this
time select a region that represents
only Mineral 2, repeating the
selection until all of the phase has
been detected.

3: Fine tune the detection with the

Histogram controls – notice how the
Mineral 2 phase (Green) overlaps
the Reference (Red) phase but butts
against the highest value for the
Mineral 1 phase (Blue).

4: The Mineral 2 phase binary image

after detection.

That completes the Threshold phase

detection for this example.

Continued… 929

928 © 2011 Leica Microsystems (Switzerland) Limited

 Phase Expert: Binary Processing Pre-Filters:

The Binary Processing Pre-Filters

provides several filters and a range of
tools that can quickly and easily improve
a binary image - combining or separating
detail, filling holes, separating phases that
are touching and reconstructing
'fractured' phases.

1: Click on the Binary Processing Pre-

Filter option on the Processing
Sequence menu to open the Pre-
Filter panel.

For detailed information and

techniques about Binary Processing
Pre-Filters, see Image Analysis help:
Go there... 797

LAS User Manual 929

 Phase Expert: Binary Image Editing:

Binary Image Editing is a tool collection

that provides the methods for working
directly on Binary Images to add, remove,
select and de-select features. Facilities
also include drawing and filling shapes as
well as grouping features.

1: To use the Binary Editing tools, click

on the Binary Image Editing option on
the Processing Sequence menu.

For detailed information about Binary

Editing, see the Image Analysis help:
Go there... 838

930 © 2011 Leica Microsystems (Switzerland) Limited

 Phase Expert: Results and Histogram: Phase

1: Click to select Results & Histogram on

the Processing Sequence menu.

2: Click on the arrows to the right of the

Phase/Field header and…

3: …from the drop-down list click on the

Phase option.

4: Click on the Show Histogram button.

5: The Phase Histogram shows the phases

and their results plotted against the
parameter chosen for the Y axis.

With Phase Histogram selected, the X

axis always shows the Phases and
cannot be changed. The Y axis which
represents the measured parameter, can
be user selected. Go there… 933

6: For details of other Axis and Bin options,

see Image Analysis 873 help.

Continued… 932

LAS User Manual 931

 Phase Expert: Results and Histogram: Field Histogram:

1: Click on the arrows to the right of the

Phase/Field header.

2: From the drop-down menu click to select

the Field option.

3: Click the Show Histogram button.

4: The Field Histogram is displayed. This is

the result of the entire image – Field –
plotted against the parameters chosen
by the user for both the X and Y axes.

5: Selecting the parameters for the

measurement axes: Go there… 933

6: The Y axis and Display Settings dialog

revealed by enabling the Axis Options
check box, and...

7: ...the Bin Details panel when the Show

Bin Details check box is enabled.

Continued… 933

932 © 2011 Leica Microsystems (Switzerland) Limited

 Phase Expert: Results and Histogram: Select the X and Y Parameters:

With the Phase option selected for the

Histogram display, only the Y axis parameter
can be changed. For the Field option both X
and Y parameters can be selected.

1: Click on the Browse button against the

axis to be changed. If the button is
greyed out the option is not available.

2: The Select Measurement Parameters list

will depend upon the display option
chosen and the axis to be altered. Click
to select the parameter required.

3: Click OK.

Continued... 934

LAS User Manual 933

 Phase Expert: Tabular Results: The Field example:

If multiple fields are measured, results can be

aggregated or shown for individual fields by:

1: Clicking on the Options button on the

Select Images dialog and ...

2: ...clearing/enabling the Accumulate

Results check box on the Select Images
to Measure panel.

The check box affects the display only so

there is no need to re-process the
images.

3: Click on the Phase Details tab to reveal

the results as each Phase compared to
the Field or compared to the Reference
Phase is one has been set up.

4: Click on the Phase Summary tab to

show each Phase as a set of user-
defined statistics.

To change the oder of the results - low-to-

high or high-to-low, double-click a column
header.

Continued… 936

934 © 2011 Leica Microsystems (Switzerland) Limited

LAS User Manual 935
 Phase Expert: Tabular Results: The Field example: Continued:

If multiple fields are measured, results can be

aggregated or shown for individual fields by:

1: Clicking on the Options button on the

Select Images dialog and ...

2: ...clearing/enabling the Accumulate

Results check box on the Select Images
to Measure panel.

The check box affects the display only so

there is no need to re-process the
images.

3: Click on the Phase Details tab to reveal

the results as each Phase compared to
the Field or compared to the Reference
Phase is one has been set up.
Click on the Phase Summary tab to show
each phase as a set of user-defined
statistics.

To change the order of the results - low-to-

high or high-to-low, double-click a column
header.

Continued… 937

936 © 2011 Leica Microsystems (Switzerland) Limited

 Phase Expert: Tabular Results: The Reference example:

Shown below are the tabular results for the Reference 2 & 3: ...Minerals 1 (44.4%) and 2 (55.6%) that together
example. add up to 100%.

1: Click on the Details tab to show the overall 4: Click on the Summary tab to show the basic statistics.
relationship between the Reference phase
(Highlighted) - always shown as representing 100% of Continued… 938
the area - and...

LAS User Manual 937

 Phase Expert: Results and Histogram: Select the Tabular Display Parameters:

1: To change the Phase Details column

header parameters or the Phase
Summary statistic parameters, click on
the appropriate results tab.

2: Click on the Tool Tab and, depending

upon the results tab selected the Select
Phase Details or Select Phase Summary
dialog will appear.

3: Enable the required check boxes to

include the parameter required.

4: Click OK.

The Show All option when enabled will select

all of the options.

Click on the Hide All button to clear all of the

options before starting a new range of
selections.

938 © 2011 Leica Microsystems (Switzerland) Limited

 Phase Expert: Reference Data:

A comprehensive range of Data Items

can be appended to the Phase Expert
results that will identify important details
such as the Project Name, the Specimen
and how it was prepared. Enter the
information in the Reference Data dialog.

1: Click on the Reference Data option

on the Processing Sequence menu
to reveal the Reference Data dialog.

Administrators can add to the

supplied list of data headings to
comply with corporate needs (2).

For details refer to Image Analysis 883

help:

LAS User Manual 939

 Phase Expert: Create Report:

1: Click to select Create Report on the

Processing Sequence menu.

2: On the Create Report panel, click to

select/de-select the images to be
included in the report.

3: Nominate a Report Layout Template by

clicking on the browse button and…

4: …navigating to and clicking to select a

template. A default template is supplied
with Phase Expert and is located in the
Excel Templates\Phase\LAS Phase
Expert Templates folder.
The supplied template can be modified to
suit the user’s requirements and saved
under a separate file name.

5: Click Open.

6: Specify where the report is to be saved

by clicking the browse button to the right
of the Report text box and…

7: …navigating to the destination folder, ...

8: …typing a file name for the report and

clicking Open.

Continued… 941

940 © 2011 Leica Microsystems (Switzerland) Limited

 Phase Expert: Create Report: Continued:

1: To display the report automatically as it is created, 3: The front page of the report displayed in Microsoft
click to enable the Display Report button. Excel.

2: To create the report click the Export button.

Continued… 942

LAS User Manual 941

Phase Expert: Create Report: Continued:

1: Sample page from the Microsoft Excel report.

2: The Excel Spreadsheet tab structure.

942 © 2011 Leica Microsystems (Switzerland) Limited

 Phase Expert: Configurations:

The current settings for most editing

panels can be saved with the archive as
a Configuration to be retrieved at a later
date. Retrieved Configuration settings are
applied to automatically to the tools

1: Each configuration has a unique

name and can be accessed by
clicking on the arrow to the right of
the Current Configuration window
and from the drop down list…

2: …clicking to select the required

configuration.

3: Click the Display Configuration

Settings button to list the
configuration details.

Save Configuration:
Saves all of the current settings and
process sequences in a unique file.

4: To save the current settings as a

new configuration, click on the New
button and…

5: …type a unique name for the new

configuration.

6: Click OK to save the setting.

7: Click on the Save Configuration

button The new configuration
appears in the drop down list.

Continued... 944

LAS User Manual 943

 Phase Expert: Configurations: Continued:

Edit Configuration:
A Configuration name can be changed
by:

1: Select the configuration to be

changed from the drop down list.

2: Click on the Edit Configuration

button.

3: On the Edit Configuration dialog,

change the name by clicking in the
text box and typing a new name
and…

4: …clicking OK.

5: Click the Save Configuration button.

Remove a Configuration:
A Configuration can be removed from the
list by:

6: Selecting the configuration to be

deleted from the drop down list.

7: Clicking the Delete (Trash Can)

button.

8: Confirm the deletion and the

Configuration will be removed
permanently. The operation cannot
be reversed.

Print a Configuration:
Check that the printer is on and
connected to the computer. The
configuration is captured to an Excel
spreadsheet and printed from there.

1: Select the Configuration to be printed

from the drop down list. When the
Configuration settings appears on an
Excel spreadsheet, use the print
function to print the settings list.

944 © 2011 Leica Microsystems (Switzerland) Limited

 Grain Expert

The grain sizes in a metal, plastic, composite or material is an important factor in

determining its strength or machinability. LAS Grain Expert allows you to measure the
average grain size of visible grains on the surface of a specimen. Grain boundaries are
identified on the basis of contrast in the image. Grains can be measured using a number of
different standards and techniques using automatic methods to identify boundaries. For
example, on appropriate images:

Automatically identify grains and measure their size.

Count the intersections of grain boundaries with a test line

Mark the diameter of selected grains on an image

Edit the image to trace the shape of selected grains on an image

Compare a sample image with a selection of typical example images to select the most
appropriate boundary identification method.

LAS Grain Expert calculates:

Average grain size, expressed in terms of a grain size number.

Mean grain area.

Maximum and minimum grain size.

Confidence level.

Relative accuracy.

The results obtained depend on the standard and technique employed.

LAS User Manual 945

 Grain Expert: Step by Step:
Grain Expert must be installed, licensed and
enabled. Click on the Analysis Workflow (1)
and then on the Grain Expert tab (2).

If the tab is not present, Grain Expert has either

not been installed or is not enabled: Refer to
the LAS Installation Guide.

These steps represent the sequence users

should follow to successfully carry out Grain
measurements.

Because Grain Expert works in conjunction with Image Analysis, in most steps links are provided to both modules:
Red, will take the reader to the appropriate place in the Image Analysis (Automatic) help file for more detailed
information, and Blue, connects to help files that are specific to Grain Expert.
Sometimes, it will be beneficial to follow both links if they are provided.

Select the Images to Process:

One or more images can be processed in sequence simply by selecting the Thumbnail(s) in the Gallery and then
clicking the Append button on the Select Images to Measure panel.
Image Analysis help >>> Grain Expert help >>> 947

Identify the Grain Boundaries:

A range of Reconstruction Images are supplied with Grain Expert to help to simplify and speed the task of finding the
grain boundaries.
Additional close refinement can then be achieved with the Threshold and Sensitivity controls.
Grain Expert help >>> 949

Binary Image Editing:

Repair boundaries, remove features and generally fine-tune the Binary Image prior to analysis.
Image Analysis help >>> Grain Expert help >>> 957

Select Standard:
Grain Expert provides configurations for all of the internationally accepted standards for grain analysis together with a
wide range of statistical methods.
Grain Expert help >>> 958

Results and Histogram:

Display the results using graphical Histograms - bar and pie charts available - as well as Tabular data in the Grid
conveniently grouped under detail and statistic tabs for both Fields and Grains.
Image Analysis help >>> Grain Expert help >>> 961

Reference Data:
Add user and analysis references to make the job totally product and company specific. The references are
automatically included in the report.
Image Analysis help >>> Grain Expert help >>> 967

Create Report:
Reports are created using Microsoft Excel and a suitable template. A standard template is supplied with Grain Expert
and can be tailored to user and corporate needs.
Image Analysis help >>> Grain Expert help >>> 968

946 © 2011 Leica Microsystems (Switzerland) Limited

 Grain Expert: Select Images to Measure:

1: On the Processing Sequence click

on Select Images to Measure.

2: Click on a Thumbnail in the Gallery.

3: Click to Append button on the Select

Images dialog to add the selected
image to the Sequence Image List.

Repeat the steps to add more images

to be measured as a batch.

All selected images must have the

same type - for example .bmp, .png, .
jpg - without compression, and the
same resolution.

4: Images can be selected in any order

from the Gallery. To sort them into
numerical sequence click to enable
the Sort check box.

5: Use the Remove button to remove a

single selected image from the list,
or...

6: ...click the Reset button to remove all

images.

Continued...

LAS User Manual 947

Grain Expert: Select Images to Measure: Continued:

Additional controls on the Select Images

panel are check boxes – click to enable,
click again to disable - reached by:

1: Click on the Options button to

reveal...

2: ...the Process Settings pop-up menu.

3: Accumulate Results: Adds the results

of the current pass to those of the
previous. Two or more images have
to be selected.

4: Export Report: At the end of the

analysis run an Excel Report is
generated and displayed. Excel must
be installed on the computer.

5: Show Histogram: Displays the

Histogram at the end of the analysis
run.

6: Refresh Display: Updates the display

as processing proceeds
independently of the Interactive/Apply
setting on each sequence panel.

7: Refresh Results: Updates the Grid as

processing proceeds. Turn the Grid
on by clicking to enable the Grid
display button (8) on the Side Tool
Bar.

9: If multiple images are part of a

sequence, the software recognises
this and…

10: …the Append Sequence button

becomes active. Click the button and
the selected range is imported into
the Image List. This avoids having to
load images separately.
To select a range of images in the
Gallery, click on the first image and
then holding down the Shift key click
on the last. All of the images between To select a range of images separately, click on the first image and
will be selected. then holding down the Ctrl button click on each of the others.
If Sort is enabled, the sequence will be sorted numerically.

Continued…

948 © 2011 Leica Microsystems (Switzerland) Limited

 Grain Expert: Identify the Grain Boundaries:

To speed grain boundary detection,

several Reconstruction Images are
provided with Grain Expert. By matching
one of these to the selected specimen, a
more appropriate and faster software
algorithm can be used.

1: Click to select Identify Grain

Boundaries in the Processing
Sequence menu.

2: On the Identify Boundaries panel,

click to enable Show Binary to view
the processed image.

3: Click to disable Interactive to prevent

processing whilst matching a
Reconstruction Image to the
specimen.

4: Click the Select button.

5: The first Reconstruction Image

appears.

6: Move back and forth through the

images until one appears that more
closely matches the specimen than
any of the others. It does not have to
be identical.

7: Click OK.

8: Click on the Apply button and the

initial boundary detection will appear
in the binary window.

Continued…

LAS User Manual 949

Grain Expert: Identify the Boundaries: Continued:

1: In the example the Dual phase

brightfield Reconstruction Image has
been chosen as quite closely
matching that of…

2: …the specimen.

3: Click OK and…

4: …a scaled version of the

Reconstruction Image appears in the
Identify Boundaries window.

Continued…

950 © 2011 Leica Microsystems (Switzerland) Limited

 Grain Expert: Identify the Grain Boundaries: Polarised Illumination:

Grain Expert supports both greyscale and

colour images, but using conventional
bright- and darkfield illumination the grains
will appear only as various shades of grey.
Images captured using polarised light
techniques will display in colour and the
Etched Polarised Reconstruction Image
should be selected.

1: Click to select Identify Grain

Boundaries in the Processing
Sequence menu.

2: On the Identify Boundaries panel,

click to enable Show Binary to view
the processed image.

3: Click to disable Interactive to prevent

processing whilst finding the
Reconstruction Image.

4: Click the Select button.

5: The first Reconstruction Image

appears. Move through the
reconstruction samples to find
Etched, Polarised.

6: Click OK.

7: Click on the Apply button and the

initial boundary detection results will
be displayed in the binary window.

LAS User Manual 951

Grain Expert: Select the Boundary Colour:

The colour used to show the grain

boundaries can be chosen to suit both the
user and the image.

1: Click on the Colour box on the

Identify Boundaries panel. The Select
Color dialog appears.

2: Choose a new colour by dragging the

target mark on the Colour Wheel,

3: …clicking to select a Swatch or…

4: …typing Values.

5: Click OK. The new colour appears in

the Colour box.

6: Click to enable the Show Binary

check box and…

7: …the new colour appears on the

binary output image.

Continued…

952 © 2011 Leica Microsystems (Switzerland) Limited

 Grain Expert: Detect the Boundaries:

The Threshold and Sensitivity controls

together, serve to detect the boundaries
between individual grains. Time spent at
this and during the Binary Editing stage to
identify the boundaries as closely as
possible, will result in a more accurate
analysis.

Set the Sensitivity to Mid-range:

This is the starting setting to provide initial
boundary detection and will be adjusted
later:

1: If necessary, click to enable Show

Binary. This will display the binary
output image with the detected
boundaries shown in the selected
colour.

2: Click to enable Interactive update.

3: Click, hold and drag the Sensitivity

slider to about the mid-point on the
track.

4: Part of the original image and…

5: …after the Sensitivity has been

changed. At this stage many of the
boundaries remain undetected – the
original grey instead of red.

Continued…

LAS User Manual 953

Grain Expert: Detect the Boundaries: Continued:

Depending upon the Reconstruction

Image chosen, for greyscale images an
additional control will be available to the
user - the Threshold button that can fine-
tune the boundary detection using a
Histogram.

1: The binary image with Sensitivity set

from the previous step.

2: On the Identify Grain Boundaries

panel, click the Threshold button.

3: The Threshold Remove Phase

Histogram appears representing the
image with the total black pixels
(Value 0) to the left and the white
pixels (Value 255) to the right.

The vertical dotted bars and the

sliders beneath the Histogram
represent the span of pixel values
included in the binary image. Pixels
with a value greater than 151, the
setting on the illustration, are ignored.

4: Click on the Auto button. This will

automatically detect detail in the
image that could represent grain
boundaries.

5: The binary image after Auto detect -

far more boundaries have been
detected and the entire image is more
refined.

Continued…

954 © 2011 Leica Microsystems (Switzerland) Limited

 Grain Expert: Detect the Boundaries: Continued:

1: The binary image following the Auto

detect stage.

The Histogram Remove Phase dialog can

be used to refine the detection process by:

2 & 3: Dragging the Histogram sliders to

include or reject pixel values. On the
illustration, the black and dark grey
range has been excluded by dragging
the left slider to value 43. The mid-
and light grey range has been
increased by dragging the right slider
to value 183. In this way it is possible
to detect boundaries that the Auto
detect did not find.

4: Complete the detection by gradually

dragging the Sensitivity slider to the
left – approaching 0 – until there is no
perceptible improvement in the
boundary detection.

5: The binary output image so far.

Continued…

LAS User Manual 955

Grain Expert: Detect the Boundaries: Colour Images:

When colour images are selected, the

Grain Boundaries panel has an additional
control - the Noise Tolerance slider.

1: Click, hold and drag the Sensitivity

slider to achieve the best boundary
detection possible, and then refine the
detection by...

2: ...dragging the Noise Tolerance slider

to the left or right.

3: Part of the original image with Noise

Tolerance set very low and possibly
more boundary or 'edge' detection
than is really necessary.

4: Noise Tolerance set too high at 7 and

some boundaries have been missed.
For this image a setting somewhere
between the two would be appropriate
and adequate.

Continued…

956 © 2011 Leica Microsystems (Switzerland) Limited

 Grain Expert: Binary Image Editing:

Binary Image Editing provides the tools for

working directly on binary images to add,
remove, select and de-select features.
Facilities also include drawing and filling
shapes as well as grouping features. Use
Binary Image Editing to 'repair' incomplete
boundaries and mark phases that should
not be included in the analysis.

1: On the Processing Sequence menu,

click on the Binary Image Editing
entry.

2: The Binary Image Editing panel

appears. There are two groups of
buttons:

3: Mode and…

4: Tools. Some of the Mode buttons

affect the way that the Tools behave.

5: The Size window controls the line

thickness for some of the Tools used
for drawing.

6: The colour of the binary output image

is user selectable.

7: The fast-edit tool combination of

Erase and Draw Line, can be used to
remove detected boundaries - the
image is not modified, only the binary
overlay.

8: Click on the image and draw a short

line intersecting the boundary to be
removed.

9: The boundary is automatically

detected and removed.

For detailed information about the Binary

Editing tools and how to use them, read
Image Analysis help: Go there...

Continued...

LAS User Manual 957

Grain Expert: Select Standard and Method:

Three Standards are available within

Grain Expert:

ASTM E112,
JIS G 0551/2 and
ISO 643 2003.

Select the Standard:

1: Click on the Select Standard entry on
the main menu.

2: The Select Standard panel appears.

3: Click on the arrows to the right of the

Standard header and…

4: …from the drop down list, click to

select the required standard.

Choose the Method:

5: Click on the arrows to the right of the
Method header.

6: From the drop down list, click to

select the Method required. Examples
of the Method structures and the
display results are shown on the
following pages.

Continued…

958 © 2011 Leica Microsystems (Switzerland) Limited

 Grain Expert: Select Method Diagrams:

The following Method diagrams

and result displays are
applicable to the analysis of the
entire Field:

1: On the Results and

Histograms panel, click on
the arrows to the right of
the Field/Grain header and
select the Filed option.

2: Planimetric Method.

3: Vertical Lines Method.

4: Horizontal Lines Method.

Continued...

LAS User Manual 959

 Grain Expert: Method Diagrams: Continued:

Continued from the previous

page...

1: Select the Field option on

the Results and Histogram
panel.

2: 3 Circles Method and…

3: Intercept Method.

Grain Results:
4: To display the Grain
results binary output image
in which each grain size is
colour coded, on the
Results and Histogram
panel click on the arrows to
the right of the Field/Grain
header and select Grain
from the options.

5: The Grain results binary

output image.

Continued…

960 © 2011 Leica Microsystems (Switzerland) Limited

 Grain Expert: Results and Histogram: Grain:

1: Click to select Results & Histogram

on the Processing Sequence menu.

2: Click on the arrows to the right of the

Grain/Field header and…

3: …from the drop-down list click on the

Grain option.

4: Click on the Show Histogram button.

5: The Grain Histogram shows the

grains and their results plotted against
the parameters chosen for the X and
Y axes. Selecting the X and Y axes
parameters: Go there…

For details of other Axis and Bin options,

see Image Analysis help.

Continued…

LAS User Manual 961

Grain Expert: Results and Histogram: Field:

1: For this analysis 5 fields were

chosen.

2: On the Results and Histogram panel,

click on the arrows to the right of the
Grain/Field header and from the
options…

3: …click to select Field.

4: click on the Show Histogram button.

5: The Field Histogram shows the 5

fields along the X axis with their
measured area on the Y axis.

Continued…

962 © 2011 Leica Microsystems (Switzerland) Limited

 Grain Expert: Results and Histogram: Select the Histogram Parameters:

With the Field option selected for the

Histogram display, only the Y axis
parameter can be changed - the X axis
will always read Image Number. For the
Grain option both X and Y parameters
can be selected.

1: Click on the browse button against

the axis to be changed. If the button
is greyed out the option is not
available.

2: The Select Measurement

Parameters dialog appears. The
items in the list will depend upon the
display option chosen and the axis to
be altered. Click to select the
parameter required.

3: Click OK.

Continued...

LAS User Manual 963

 Grain Expert: Results and Histogram: Grain Results:

1: If necessary, click on the Show Grid button on the 2 & 3: Click on the appropriate tab to reveal the results.
Side Tool Bar to display the results in tabular form. The result parameters can be selected to suit the job
and user: Go there…
Four tabs are displayed across the top of the Grid. To the
left the Grain results – Details and Statistics – and to the
right the Field results, Details and Statistics also (Shown Continued…
on the following page).

964 © 2011 Leica Microsystems (Switzerland) Limited

 Grain Expert: Results and Histogram: Field Results:

1: If necessary, click on the Show Grid button on the 2 & 3: Click on the appropriate tab to reveal the results.
Side Tool Bar to display the results in tabular form. The result parameters can be selected to suit rhe job
and user: Go there…
Four tabs are displayed across the top of the Grid. To the
left the Grain results – Details and Statistics (Shown on the
previous page) – and to the right the Field results, Details Continued…
and Statistics also.

LAS User Manual 965

Grain Expert: Results and Histogram: Select the Tabular Display Parameters:

1: To change the Tabular (Grid) display

column header parameters for Grain
or Field, click on the appropriate
results tab.

2: Click on the Tool Tab and, depending

upon the results tab selected the
Select Details or Select Statistics
dialog will appear.

3: Enable the required check boxes to

include the parameter required in the
displayed results.

4: Click OK.

The Show All option when enabled

will select all of the options.

Click on the Hide All button to clear all

of the options before starting a new
range of selections.

Continued...

966 © 2011 Leica Microsystems (Switzerland) Limited

 Grain Expert: Reference Data:

A comprehensive range of Data Items can

be appended to the Grain Expert results
that will identify important details such as
the Project Name, the Specimen and how it
was prepared. Enter the information in the
Reference Data dialog.

Administrators can add to the supplied list

of data headings to comply with corporate
demands.

For details refer to Image Analysis help:

Continued…

LAS User Manual 967

Grain Expert: Create Report:

The report is created using Microsoft

Excel which must be installed on the
computer.

1: On the Processing Sequence menu,

click to select Create Report.

2: The Select Content dialog lists all of

the information and images that can
be included in the report.

3: The list is divided into sub-sections –

click on the small arrow to the left of a
sub-section header to reveal all of the
section options.
To include an item in the report, click
to enable the check box to the right of
the item.

4: The list is extensive so use the scroll

bar to reveal more items (5). Images
tend to be large files that can make a
report unwieldy, especially if it is to be
transmitted electronically – by e-mail
for example – so where possible keep
the included images to a necessary
minimum.

Continued…

968 © 2011 Leica Microsystems (Switzerland) Limited

 Grain Expert: Create Report: Select the Excel Template:

The reports are created using an Excel

template; A standard template is provided
with Grain Expert that will be suitable for
many applications. It can be modified by
the user to reflect the job or corporate
style. Alternatively, use any appropriate
existing Excel template.

Locate the Template:

1: Click on the browse button to the right
of the Template text box.

2: Navigate to the Template folder…

3: …and file and click to select it.

4: Click on the Open button.

A default template is supplied with Grain

Expert and is located in the
Users/Excel Templates\Grain\ folder
with the name:
LAS Grain Analysis Expert Template.xlt.

The precise path may vary with the

installed operating system.

Continued...

LAS User Manual 969

Grain Expert: Create Report: Select the Report Save Location

1: Click on the browse button to the right

of the Report text box.

2: Navigate to the folder into which the

report file will be saved.

3: Type a name for the file and…

4: …click on the Open button.

5: If the report is to be displayed as

soon as it is created, click to enable
the Display Report check box.

6: Click on the Export button.

Continued...

970 © 2011 Leica Microsystems (Switzerland) Limited

 Grain Expert: Report Front Cover:

This report was created with Microsoft Excel using the the
template supplied with Grain Expert. The report can be
displayed and read (but not created) using Excel Viewer.

Continued...

LAS User Manual 971

 Grain Expert: Report Sheets:

This report was created with Microsoft Excel using the the 1: Depending upon the information included in the
template supplied with Grain Expert. The report can be report, it is divided into sheets access by clicking the
displayed and read (but not created) using Excel Viewer. tabs along the lower edge.

2: The illustration shows the Images sheet.

972 © 2011 Leica Microsystems (Switzerland) Limited

 Reticule

Reticule is an optional module that allows a precise pre- LAS Reticule has a wide and varied range of precision
defined measuring grid – a Reticule - to be overlaid on a uses:
live image.
Item counting and distribution.
Software generated: Completely independent of
eyepiece reticules. Comparison and location.

A fast and reliable method of selecting the best grid Sizing.

to suit the application.
Volume estimation.
A reticule can be captured and merged with the
image. Assessing image scale.

Fixed and Scalable versions available from a

comprehensive Library.

Reticule patterns are stored as standard svg format

allowing users to design and create to their own
special requirements.

LAS User Manual 973

Reticule: Fast Track:

Reticule Fast Track is a checklist of the

steps to take to get LAS Reticule
operational quickly:

1: Select and load the required

Reticules from the Library: Go there...
975

2: Calibrate the microscope: Go there...

268

3: Select the best live image format to

suit the application and hardware.
Good resolution but with an
acceptable refresh rate is the
essential aim: Go there... 240

If using a Stereo microscope the zoom is

adjusted, the Reticule drawings adjust in
size but due to image shift are no longer in
the correct position. Use the AX-Carrier
option to correct this,

Most suitable for use with a microscope

having automatic magnification readout.

974 © 2011 Leica Microsystems (Switzerland) Limited

 Reticule: Library:

The first task is to select the styles

required from the Reticule Library and
attach them to the Reticule software.

1: Click to select the Setup Workflow

and…

2: Select the Reticule tab. If it is not

present the module is not installed or
is not licensed.

3: On first use the Available Reticules

window will be empty, so to select the
styles…

4: …click on the Add button.

Continued... 976

LAS User Manual 975

Reticule: Available Reticules:

1: The Widows navigator dialog opens

with the Reticule Files folder
automatically selected.

2: A single style can be selected simply

by clicking on its file name, but for
multiple selections, hold down the Ctrl
key on the keyboard whilst clicking
individual files. The illustration shows
11 files selected and highlighted.

3: Click on the Open button.

4: The selected reticule files appear in

the Available Reticules window.

5: Select an individual style by clicking

its entry. It is highlighted and…

6: …displayed in the viewer.

7: To delete a reticule style, click to

select it in the list (5) and then click
the Remove button. If a style is
inadvertently deleted it can be
recovered from the library. See
previous page… 975

976 © 2011 Leica Microsystems (Switzerland) Limited

 Reticule: Types: Fixed:

Fixed reticules are non-movable and not

scalable. They are always centered in and
displayed to fit within the image window -
'fit to window' where the window is the live
image size.

For example, a crosshair displayed across

the centre of the image window remains
fixed and unchanged when the image is
moved (by moving the camera or
panning), or if the magnification is
changed.

Examples here show:

1: Weibel Reticule.

2: Squares Reticule one of several

variations and styles.

3: A Point Grid Reticule.

4: Mertz Reticule.

Fixed Reticules are defined in pixels.

Mertz, Point and Weibel are examples of

stereological grids.

Continued... 978

LAS User Manual 977

Reticule: Types: Scalable:

Scalable Reticules have an absolute size

associated with them and are defined in
physical units for example millimetres.
They can be positioned (moved) and
rotated by the user; but cannot be re-sized
manually.

If the image is moved or zoomed, the

centre of a Scalable Reticule will remain
over the feature in the image where the
user positions it and it will re-size to be
correctly scaled to the image. The scaling
information is obtained by reading it
automatically from the connected
microscope. So, for example, a
rectangular reticule placed over a feature
will remain centred over the feature and
scaled to the image when the image
zoomed in or out.

Scalable Reticule examples are:

1: Horizontal Bar 500µm

2: Circles 500µm

3: Cross 200µm.

978 © 2011 Leica Microsystems (Switzerland) Limited

 Reticule: Reticule Settings:

There are three colour options for the

reticule which are selected on the Reticule
Settings panel:

Default: Displays the reticule in the

colour defined by the Reticule
definition file.

Complementary: Automatically
determines the display colour based
upon the average hue of the image to
maintain a good contrast.

User Defined: Allows the user to

select the display colour as follows:

1: Click to enable the User Defined

Colour radio button.

2: Click in the Selected Colour window

and…

3: …the Windows Select Color dialog

appears. Choose a colour from the
wheel by clicking and holding the
‘target’ and dragging it to the desired
colour, or clicking to select a Basic
Color in the matrix. Precise colour
selection can be made by typing the
red, green and blue (RGB) values in
the text boxes, and a colour can be
saved by clicking the Add to custom
button.

4: Click OK and the selected colour

appears on the Reticule Settings
panel (5).

LAS User Manual 979

Reticule: Apply Reticule:

The reticule style to be used is selected

on the Acquire Workflow:

1: Click to select the Acquire Workflow

and the Camera tab.

2: The Active Reticule control panel

selects the desired reticule and
determines if it is to be displayed or
not. Like most other control panels, it
can be moved to any part of the
screen by clicking and holding on the
header bar and dragging it to the
required position. Return it to the
normal location by clicking the ‘X’ on
the header.

3: Click on the small arrow to the right of

the Active Reticule list box and all of
the reticule styles attached to the
module appear.

4: Click to select the style required.

5: Click to enable the Show Reticule

checkbox and the reticule will appear
on the live image.

980 © 2011 Leica Microsystems (Switzerland) Limited

 Reticule: Using Reticules:

1: The selected reticule in the chosen

colour on the live image.

2: Move the reticule by clicking on it (8

‘handles’ appear) and dragging it to
the desired location.

3: A Scalable Reticule can be rotated to

any angle by clicking, holding and
dragging on the Rotate ‘handle’.

Images captured from the Acquire

Workflow by either clicking on the Acquire
Image button or pressing function key F3
on the keyboard, will merge the reticule
with the image. To capture without the
reticule, either switch it off with the Show
Reticule checkbox or make the capture
from the Browse Workflow.

LAS User Manual 981

Reticule: Creating Reticules:

There are several vector graphic drawing packages Download and install Inkscape.
available that are capable of creating svg Reticule files.
An open source application - download and use the Run the program.
program free of charge but do not use it for commercial
gain - called Inkscape (Google for the latest releases), From the File option, select Document Properties
will produce Reticules to import into LAS. and set the default units to pixels (px), the Canvas
Size to Custom 640 x 640 (adjust as necessary to
This and the following page is a general guide; Open suit the LAS Viewer size and camera)
source software is constantly changing and improving so
some experimentation will be necessary. Click Fit Page to Selection.

Continued... 983

982 © 2011 Leica Microsystems (Switzerland) Limited

 Reticule: Creating Reticules: Continued:

Select a shape from the toolbar and

draw it on the canvas area. The
shape must not extend beyond the
canvas.

Use the various tools and dialogs to

change line weights and colours.

Save the drawing using the

navigation dialogs and the Plain svg
format.

Open the file in a simple text editor

- Notepad or Wordpad. Find the
lines that specify:

width="680px" and height="680px"

...and make sure the 'px' suffix is

present. LAS needs these units.
Save the file which will be a fixed
reticule.

To create a Scalable Reticule with

the file open in the text editor and
replace the width and height lines
with:

width="0.2mm" height="0.2mm"
viewBox="0 0 680 680"

...where 0.2mm is the scale required.

Do not change the pixel size. Save
the file.

Launch LAS and import the

'personalised' Reticule in the
normal way: Go there... 975

To determine just how Reticule files are

constructed, open a Reticule Library file in
Notepad and check that newly created
files have the same structure.

LAS User Manual 983

 Web Sharing

Leica Application Suite Web Sharing module streams live

images across a local network so that they can be seen in No special hardware required - only a standard DFC
real time by other users. camera on the Master microscope.

In the illustration, the Master PC (1) running Leica Other users – Clients – need only Internet Explorer
Application Suite with Web Sharing installed and enabled, with Microsoft Silverlight plugin to view the images.
streams the microscope image to the local network. Other
users – Clients – on the network view the image in real With the appropriate projection hardware, a Client can
time using only free software, Internet Explorer with the be used for large screen viewing in lecture halls and
Silverlight plugin. On the illustration the Clients are seminars.
numbered 2 through to 5 but almost any number of clients
can view the image simultaneously. Three image size options – 640 x 480, 800 x 600 and
1024 x 768 pixels.
Web Sharing is an optional module, free to evaluate for 60
days; After that a chargeable license is required. Clients can see Scale Bar and Master Cursor
movements, especially useful for pointing to areas of
Suitable for Local Area or Virtual Private networks, interest.
wired or wireless.
Image Capture feature for Clients so that they can
Only the Master PC needs Leica Application Suite and have a permanent, printable record on their own
the Web Sharing software. computer.

984 © 2011 Leica Microsystems (Switzerland) Limited

 Web Sharing: The Launch Panel:

The Web Sharing launch panel is on the

Camera tab of the Acquire Workflow.

1: Click on the Acquire Workflow.

2: If necessary, click on the Camera tab.

3: Click on the small arrow to the right of

the Web Sharing header to expand
the panel.

Continued… 986

LAS User Manual 985

Web Sharing: Cursor and Scale Bar:

Both the Cursor (and Scale Bar if it is

enabled) on the Master monitor are
available to the Clients.

If the Master Cursor is moved - to point

out a particular feature for example - the
movement will be reflected on the Client
monitors. If the Scale Bar is moved to a
new position, this too will change position
on the Client display.

If it is intended that a Scale Bar is visible:

1: Click the small arrow to the right of
the Calibration Settings header and…

2: …also on the Scale Bar header to

access the settings and in particular
to enable the Scale Bar by clicking
the Show button (3) to reveal the
Scale Bar (4).

When Web Sharing is active, most of the

other LAS functions are disabled to
ensure rapid refresh at the Client
monitors, so the Scale Bar setup can only
be carried out with Web Sharing disabled.

Scale Bar setup details: Go there…

Calibration: Process > Calibration: Go

there... 268

986 © 2011 Leica Microsystems (Switzerland) Limited

 Web Sharing: Camera Settings:

The Camera Settings for both Live and

Captured Formats, affect the speed at
which Web Sharing images are
downloaded to the Client monitors.
The Live Format affects the refresh rate
and the Captured Format the download
speed if the Client decides to save the
image. Images saved at a Client machine
will have the same resolution as the
Master Camera Captured Format.

Before enabling Web Sharing check the

Camera Formats; Start with a fairly low
resolution – 696 x 514: 3 x 3 Color Binning
– for example, and increase the
resolution if the refresh rate allows or
more detail is required at the Client
monitors.

1: Make sure that Web Sharing is

disabled.

2: Click on the Camera tab on the

Acquire Workflow.

3: Click on the small arrows to the right

of the Captured and Live Format
headers and…

4: …from the drop down menus choose

one of the lower settings.

More detail about Camera Formats: Go

there... 239

Continued… 991

LAS User Manual 987

Sharing Stored Images:

Usually, Web Sharing streams the current

live image, but it is possible to temporarily
disable the camera and instead stream
images that have previously been
captured.

The compression type of the stored image

does not matter, but the resolution should
be at least 1024 x 768.

The first step is to disable the camera and

replace it with the Demo Camera loaded
automatically with LAS Version 3.0
onwards. If Leica Application Suite is
running, close it down.

1: On the Desktop Task Bar click on the

Start button. The illustrations show
Windows XP layout but Vista is very
similar.

2: Click on the All Programs arrow.

3: Move the cursor to highlight Leica

Application Suite in the list of
programs.

4: On the sub-menu move the cursor to

License LAS and click the option.

5: The LAS Framework will load but

instead of launching the User
Interface will stop when the Main
Menu bar appears. Click on Options.

6: From the drop down menu click to

select Hardware Setup.

7: When the Hardware Setup dialog

appears, click on the small arrows to
the right of the Image Source header.

8: From the list of option click to select

Demo Camera and then click Save.

9: Launch the program by clicking on

the Leica Application Suite header.

Continued… 989

988 © 2011 Leica Microsystems (Switzerland) Limited

 Sharing Stored Images: Continued:

With LAS running:

1: …click on the Acquire Workflow.

2: Click on the Camera tab to reveal the

camera controls.

3: Click on the arrow to right of the

Image Controls header to display the
Camera Twain icon. Click it to open…

4: …the Demo Camera dialog.

5: Select the image to be shared by

right-clicking the Select image
caption.

Continued… 990

LAS User Manual 989

Sharing Stored Images: Continued:

1: On the Select Image File dialog

navigate to the folder that contains
the image to be shared and…

2: …click the arrow to the right of the

Files of Type window and chose the
file compression type from the list by
clicking it.

3: Click to select the image.

4: Click Open.

5: The Demo Camera dialog re-appears

with the path of the chosen image
and…

6: …the image itself displayed.

7: The image resolution is display under

the Captured Image details - it should
be at least 1024 x 768.

Close the Demo Camera dialog (small X

top right of the window) and follow the
normal Web Sharing procedure from
here. Go there... 991

990 © 2011 Leica Microsystems (Switzerland) Limited

 Web Sharing: Select Image Size and Enable:

1: If necessary, expand the Web

Sharing panel by clicking on the small
arrow to the right of the header.

2: Web Sharing has three size options –

that is the size that the clients will see
on their monitors. Larger sizes are
slower to download so the refresh
rate at the client will also be slower.
The actual refresh rate depends upon
the network hardware and the
number of active users.

Check that Web Sharing is disabled –

the Enable checkbox is blank – and
click the arrows to the right of the
header and from the drop down list
click to select the required size.

3: Click to enable Web Sharing.

4: When the software successfully loads

the Master Address appears on the
panel. The Address is unique to the
Master but can change every time
LAS is started. Right click on the
Address to reveal options to:

5: Copy the Address (URL) to the

clipboard from where it can be saved
or attached to an e-mail for example,
or…

6: …Open Address (URL) to test the

network connection by sending the
image back to the Master for display.

Continued… 992

LAS User Manual 991

Web Sharing: Client Check Network Connection:

A client can check if he is connected to

the Master by sending the Master Address
and waiting to see if there is a response.
The process called ‘pinging’ is carried out
at the Command level.

1: Hold down the keyboard Windows

key and at the same time press the R
key or…

2: …click on the Start button and from

the Start Menu…

3:… click on the Run option.

4: On the Run dialog type the word

'command’ in the text box and…

5: …click OK.

6: In the Command Window against the

> prompt type ‘ping’ and then the
numeric part of the master Address –
in the example 192.168.2.4. Do not
include the http or the /Las/ parts of
the string.

7: If connection is made successfully the

details will be displayed in the
window. An unsuccessful connection
will report ‘Address not found’.

Continued… 993

992 © 2011 Leica Microsystems (Switzerland) Limited

 Web Sharing: Client Software:

Clients do not need LAS or Web Sharing

software to receive images; All that is
required is Microsoft Internet Explorer
Version 6, 7 or 8 with the Microsoft
Silverlight plugin. Currently (January 2009)
no other browser supports Silverlight.

1: Launch Internet Explorer either from

a desktop shortcut or from Program
Files.

2: The illustration shows Internet

Explorer 6 with the entire address
string supplied by the Master Web
Sharing module (3) typed (or copied)
into the address text box. Ensure that
the last forward slash (/) is included.

4: If Silverlight is not installed the link to

Microsoft Silverlight appears. Click on
the arrow to…

5: …download the plugin (4.6Mbytes)

and follow the onscreen instructions
to install it.

Silverlight is automatically installed with

Internet Explorer versions 7 and 8.

Continued… 994

LAS User Manual 993

Web Sharing: Client Connection Status:

1: The illustration shows Internet

Explorer Version 7 with the live image
from the Master in the Web Sharing
window.

2: The Connection Status – either

buffering or connected - is shown on
the side panel with…

3: …and indicator in the top right hand

corner – yellow whilst buffering and
green when connected.

4: The Master Cursor will appear if it is

within the LAS Viewer area and also
the Scale Bar if it is enabled. If the
Master user moves the Cursor or the
Scale Bar the new positions are
displayed at the Client display.

Continued… 995

994 © 2011 Leica Microsystems (Switzerland) Limited

 Web Sharing: Client Restart and Acquire Image:

If connection is temporarily lost or the Master goes ‘off 3: …when a new Captured Image window appears…
line' to change images:
4: …right click on the image and from the menu click to
1: …click Restart to refresh the connection and current select Save Picture As…
image.

2: To copy the image click the Acquire Image button Continued… 996
and…

LAS User Manual 995

Web Sharing: Client Acquire Image: Continued:

1: On the Save Picture dialog, navigate

to the folder in which to save the
image.

2: Type a name for the image and…

3: Click on the Save button.

Images are saved only in the Windows

bmp format.

996 © 2011 Leica Microsystems (Switzerland) Limited

 Multi-User Package

This is an optional module for DM Motorised

microscopes. It has to be installed and
licensed before it can be used.

Using Multi-User Package, each user of the

microscope can create unique profiles and
store them for retrieval later. A profile
comprises the microscope hardware set up
together with individual settings such as the
light source and intensity for a specific
combination of objective and contrast
method.

Both Administrators and Standard Users can

create profiles but Administrators have an
added facility in that they can make their
profiles 'public' and available to all other
microscope users. Standard Users can
access only their own profiles.

Both Administrators and Standard Users can

nominate a Default Profile from their Profile
List that is automatically loaded when the
microscope is switched on. If a Default has
not been selected then the last set up used is
loaded.

The Multi-User Package is available by:

1: Clicking on the Setup Workflow and...

2: ...on the MUP tab.

Continued... 998

LAS User Manual 997

 Multi-User Package: Users and Controls:

The Multi-User software recognises users by

their Windows log-in so an Administrator or
Standard User can be identified, the
appropriate control panel displayed and
access allowed to the proper Profile List.

Standard Users:
The first time a Standard User opens the
Multi-User Package an empty Profile List is
created. After a task has been carried out on
the microscope, the settings can be saved as
a profile by clicking the Store profile button.

1: The Store new profile dialog appears.

2: Click in the Profile name text box and type

a unique name for the profile.

3: Click in the Short description text box and

enter a relevant note about the settings.

4: Click OK.

5: The Store Profile message appears

showing progress.

6: The profile is stored and appears in the

Profile List. The Type entry, User Defined
indicates that this profile is still private and
owned by the user.

The user can then make changes to the

microscope and store those as a separate
profile which will also be added to the List. So,
a 'library' of microscope settings are built up
that can be re-loaded at will, saving the users
time and guaranteeing consistency.

Continued... 999

998 © 2011 Leica Microsystems (Switzerland) Limited

 Multi-User Package: Reload and Delete a Profile:
Reload a profile:
The microscope settings stored in a profile can
be quickly re-loaded by:

1: Click on the required profile in the Profile

List.

2: Click the Reload profile button.

3: The Reload progress indicator appears -

there are usually sounds from the
microscope as well if objectives, filters and
focus are being changed.

Delete a profile:
To remove a profile from the List:

1: Click on the profile to be deleted in the List.

4: Click the Delete profile button.

5: Confirm or cancel the deletion on the

dialog.

6: Click OK. The deletion cannot be reversed.

Continued... 1000

LAS User Manual 999

 Multi-User Package: Default Profile:
To select a profile that will be used as the
Default switch on settings:

1: Click in the Profile List to select the profile

to be used as Default.

2: Click the Set default button.

3: An 'X' appears in the Default column on

the Profile List.

4: To remove the Default selector, click the

Restore default button.

If a profile is not selected as the Default, the

settings last used by the user will be loaded to
the microscope at switch on.

Continued... 1001

1000 © 2011 Leica Microsystems (Switzerland) Limited

 Multi-User Package: Profile Preview:

Profile settings can be displayed by clicking the Show Profile data is stored in HTML format in a folder created by
profile button - the Profile Preview (1) appears. Leica Application Suite. Copy the HTML to a destination of
choice by clicking the Save HTML button (3) and then
navigating to the required folder.

Continued... 1002
2: Print the settings by clicking the Print Profile button.

LAS User Manual 1001

 Multi-User Package: Copying a Profile:

Profile data is stored in HTML format in a folder created by 2: ... and then navigating to the required folder.
Leica Application Suite.
3: Give the copy file a name and...
The file can be copied to a folder of the users choice and
from there can be distributed to other users, for example 4: ...click Save.
by e-mail. Copy the HTML to a destination of choice by:

1: Clicking the Save HTML button... Continued... 1003

1002 © 2011 Leica Microsystems (Switzerland) Limited

 Multi-User Package: Administrator Publishing:
Administrators have the same tools as Standard
Users plus the facility to publish any profile from
their own list.

Publishing makes a profile available for sharing

among all the other computer users.

1: From the Administrator's Profile List the

profile to be published is clicked to select it.

2: Click the Publish profile button.

3: The profile becomes available to all users

and is marked as such with the 'Published'
label.

LAS User Manual 1003

Leica Microsystems (Switzerland) Ltd.
Stereo and Macroscope Systems
CH 9435 Heerbrugg
Switzerland

Telephone: +44 1223 411411

FAX +44 1223 210692
Hotline: +44 1223 401824
[email protected]
www.microscopy-imaging.com