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Análise de Sholl

This plugin automates Sholl analysis of neurons in ImageJ. It creates concentric circles around the selected soma point and counts intersections with the neuron. This allows faster, more thorough analysis than manual methods. The plugin files must be installed in ImageJ's plugins folder. Source images must be grayscale and have the neuron in a single color separated from the background. Scale must be set correctly for accurate results reporting. The plugin dialog allows setting analysis parameters like radius and step size. Results can be saved as an image, text, or copied to the clipboard.

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69 views4 pages

Análise de Sholl

This plugin automates Sholl analysis of neurons in ImageJ. It creates concentric circles around the selected soma point and counts intersections with the neuron. This allows faster, more thorough analysis than manual methods. The plugin files must be installed in ImageJ's plugins folder. Source images must be grayscale and have the neuron in a single color separated from the background. Scale must be set correctly for accurate results reporting. The plugin dialog allows setting analysis parameters like radius and step size. Results can be saved as an image, text, or copied to the clipboard.

Uploaded by

ds2
Copyright
© Attribution Non-Commercial (BY-NC)
We take content rights seriously. If you suspect this is your content, claim it here.
Available Formats
Download as PDF, TXT or read online on Scribd
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Sholl Analysis Plugin

This is a plugin for ImageJ that automates the task of doing Sholl analysis on a neuron.
Its internal algorithm is based upon how Sholl analysis is done by hand – it creates a
series of concentric circles around the soma of the neuron, and counts how many times
the neuron intersects with the circumference of these circles. However, since it can do
this process repeatedly, and in a relatively short amount of time, it allows for new
possibilities with the analysis that would be too tedious to be done manually.

Installing the Plugin


For ImageJ to be able to use the plugin, it must first be installed in the plugins folder
within the ImageJ application folder. On Windows machines, the ImageJ folder is
usually located in the Program Files folder, which is located in the root of the hard drive.
To access it, select the “Run” item from the Windows Start Menu, then when it appears
type “C:\Program Files” into the “Open” box, and click OK. On Macintosh machines,
the ImageJ folder is usually located within the Applications folder. To access it, double-
click on the Hard Drive icon on the Desktop, and then open the Applications folder in the
resulting window. Once the plugin is placed in the plugins folder, which is inside the
ImageJ application folder, the next time ImageJ starts up, an option to run the Sholl
Analysis plugin will exist in the Plugins menu.

There are two files that constitute the Sholl analysis plugin: ShollAnalysis_.class and
ShollAnalysis_.java. The .class file contains the actual plugin code that ImageJ runs, and
as such, it is required for the plugin to work. The .java file contains the human-readable
source code for the plugin, and is therefore optional. Those with a curiosity for seeing
how the code works or for making changes to the plugin may be interested in obtaining
and installing the .java file along with the .class file, but all that is necessary for the
plugin to work is the .class file. These files belong in the aforementioned plugins folder
within the ImageJ application folder.

Source Image Requirements


Since ImageJ can understand virtually any kind of image format, this plugin can be used
to analyze the image content of virtually any kind of file. However, there are a few
restrictions that the content of the image must adhere to, in order for the plugin to work
properly. The first restriction is that the image must be an 8-bit grayscale image. If it is
not already, ImageJ can convert the image to an 8-bit grayscale image by going to the
“Image” menu, opening the “Type” submenu, and selecting the “8-bit” option. The
second requirement is twofold: 1) That all of the pixels representing part of the neuron
are of the same gray value, and 2) That all of the pixels not representing part of the
neuron are of a different, or multiply different gray values. In other words, the neuron
must all be one solid color, while the rest of the picture (noise, background objects, etc.)
can be anything but that color. This can be, of course, a very restrictive condition on an
actual captured image of a neuron. For some images, thresholding can be used to satisfy
this condition, but for others, especially those with lots of background noise, it may be
necessary to trace the neuron first.
Preparing the Image
Before using the plugin, it is a good idea to make sure that the scale information
embedded in the image is correct, as the plugin uses this information to calculate and
report the results of its analysis. Specifically, the scale information is the conversion
factor between pixels and physical units of length (i.e. microns). This can be set by going
to the “Analyze” menu and selecting the “Set Scale…” item while the image window is
selected. If there is no known scale information for the image, the analysis can still be
carried out using pixels as the unit of length, but the usefulness of the results, especially
when comparing them results from different images, may be limited.

When the plugin displays its options dialog box, it shows the current scale information at
the top, to allow the user to ensure it is correct before continuing with the analysis. If it is
not, then the user must cancel the dialog box and fix the scale before continuing.

Using the Plugin

Step 1: Use the point selection tool to select a point on the image. This point will be
the center of all of the analysis circles that will be sampled. Ideally, it should
be placed in the center of the soma for the best results.

Step 2: Run the plugin from the “Plugins” menu.

Step 3: Set the analysis parameters in the resulting dialog box and click “OK”. Note
that all values are in the units of length displayed at the top (i.e. microns).
Details about the meaning of each analysis parameter are described below:

Starting Radius: The radius of the smallest analysis circle, along which the #
of intersections will be measured.
Ending Radius: The maximum value for the radii of the analysis circles.
This does not necessarily mean there will be an analysis
circle with this radius value. All it means is that no
analysis circles will have radii greater than this value.
Radius Step Size: The interval between radii of consecutive analysis circles.
This value may be set to zero for continuous intersections
measurements.
Radius Span: The margin around each radius value in which continuous
intersection measurements are made. These measurements
are then combined to calculate a single value for the # of
intersections at that radius. This is a break from traditional
Sholl analysis, since the intersections are being counted in
an area around the circumference of each analysis circle,
instead of only along the circumference. This value may be
set to zero to disable the span feature, in which case only
one intersection measurement is made for each analysis
circle along its circumference.
Span Type: The statistical function used to combine intersection
measurements within a span to produce a single value.
Options are mean or median.

Starting Radius

Step Size Radius Span


Ending Radius

Step 4: Save the results, which can be done in many ways. The user can chose to save
the resulting graph as a picture file by going to the “File” menu and selecting
the “Save” option while the graph window is selected. Or the raw data from
the analysis can be saved to a text file by clicking the “Save…” button on the
graph window. The raw data can also be copied to the system clipboard by
clicking the “Copy…” button on the graph window. Common spreadsheet
programs can import the raw data directly from the saved text file or from the
clipboard, to be used in further analysis.
Release Information

v1.0: Initial release of the plugin.

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