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Microscope Lab

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10 views

Microscope Lab

Uploaded by

yaseminoral06
Copyright
© © All Rights Reserved
We take content rights seriously. If you suspect this is your content, claim it here.
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Download as PDF, TXT or read online on Scribd
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B.

Çağrı Tosun
Nanobiotechnology Lab
Light Microscope
The “Compound” Part

• Simple microscopes have single magnifying lens (like a magnifying glass).


• Compound microscopes have two sets of lenses for magnification.

• Lens closer to the eye  ocular lens (magnifying power of 10x).


• Lenses closer to the object being viewed  objective lens.

The “Light” Part

• Bright-field light microscopes produce a dark image against brighter, backlit


background.
• Provide a 2-D image.
Stereo Microscope Electron Microscope
 A stereo microscope has a magnification  Approximate size of cell is 2 µm.
of about 10 X.  Objects smaller than 0.2 µm can be seen
 With a stereo microscope you can look at with an electron microscope. (Viruses,
the details of larger objects. cellular structures, organelles)
 Use electron illumination
 For example, wings of butterfly or an
entire leaf of flower
Scanning Electron Microscope (SEM) Transmission Electron Microscope (TEM)
 SEM use electron illumination.  TEM is electron illuminated.
 Specimen is coated with gold and the  The electron beams pass through thin
electrons bounce off . slices of specimen
 Give exterior view  gives a 2-D view.
 The image is seen in 3-D  It has high magnification and high
resolution.
 High magnification and high resolution.
Confocal Microscopy
• Confocal microscopy is used to study cell structures and their
functions. Confocal provides unprecedented microscopic details
of cells and tissues’ complex morphology and dynamics with
extremely high-quality superresolution images.
• Confocal imaging relies on the sequential collection of light.
1. Imaging System

Body tube
Eyepieces & Ocular Lenses
• Ocular lens magnifies the specimen 10x.

• You will always be looking through the ocular and


objective lens.
Magnification & The Objective Lenses
• Rotary nosepiece of your microscope has four
objective lenses attached.

• Scanning Power Objective Lens (red band)


• Magnifies objects 4x

• Low Power Objective Lens (yellow band)


• Magnifies objects 10x.

• High Dry Objective Lens (blue band)


• Magnifies objects 40x

• Oil Objective Lens (black&white)-forbidden!!!


• Magnifies objects 100x

OIL OBJECTIVE LENSES (100X) SHOULD NEVER BE USED WITHOUT IMMERSION OIL!!!
DO NOT USE OIL IMMERSION OBJECTIVE (100X) UNLESS OTHERWISE REQUIRED!!!
Total magnification

• Total Magnification = ocular lens x objective lens


(10x)

 if the 40x objective is in place, the total magnification would be ?


Field of View:
Area that one can see through the ocular and objective
Magnification Field of View details
2. Viewing System
How to Focus Image?
Working Distance:
specified distance of objective lens from the studied sample.
Magnification Working distance
Coarse and fine adjustment knobs are required for adjusting this distance
coarse adjustment: obtain an approximate focus
fine adjustment: provides exact and clear image
Quality of Microscope
1. Resolution
Ability of microscope to distinguish two points as seperate points. In other words,
resolution is the level of detail.
Determines how well specimen detail is preserved.
Limit of resolution with naked eyes 0,1 mm / microscope 0,2 µm.
Eg. The resolving power of a microscope is 1µm that means two points 1µm apart can
be seen as separate points. If they are closer together than that, they will blend
together into one point.
Resolving Power of the Human Eye, the Light Microscope,
and the Electron Microscope
2. Contrast
• the ability to see particular detail against the background
• based on the differential absorption of light by parts of the specimen
• the transparent specimen is stained with some special dyes.
3. Magnification
• Degree of enlargement
• Magnification power detailed images
1. Diameter of Field of View
• Observe the image of small pieces of graph paper under scanning (4X), low power (10X)
and high power (40X) objectives

Compare field of view for all objectives by counting the number of squares in the field
of view
Comparing Powers of Magnification and details of graph paper.
2. Letter ‘e’
The letter ‘e’ inverted when viewed under the microscope.
• Optical magnification inverts the virtual image of the specimen.
• Convex lenses (objective lenses)

Movement appears to be in opposite direction than actual movement

move slide

slide appears to move


3. Depth of Field
• The depth of field is the specimen thickness that is in focus at one focal setting (the
thickness of the specimen that is acceptably sharp at a given focus level)
• Decreases as magnification increases

It refers to 3-D effect that is viewable by changing the fine adjustment under
high power.
Using the microscope:
The microscope should stay in a permanent position
Never touch the lenses of the microscope with your hands
It should be carried in upright position with both hands
At the end of the experiment ;

1. bring the objective to 4X


2. lower the stage to the bottom
3. take out the slide from the microscope!!!

The lenses should be cleaned only with lens cleaning solution.

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