Lab 1

Introduction to the Compound Microscope

Microscopes are instruments designed to produce magnified visual images of small objects. You will use two types of microscopes in this laboratory, a compound microscope, and a dissecting microscope.

Parts of Microscope -Find each of these parts on your compound microscope.

• Eyepieces- 2 of these, they fit into microscope head. One eyepiece can be focused; the other cannot. Determine which is which. Also: one eyepiece has a pointer; one eyepiece has a small ruler inside, called an eyepiece reticule.
  
• Nosepiece (Objectives are mounted into the nosepiece)
• Objectives
  Dry objectives: 4X, 10X, 40X Oil immersion: 100X
  Total magnification depends on (objective magnification) x (eyepiece magnification)
• Stage- Where slides are placed; is raised and lowered with focusing knobs.
  Course and fine adjustments (calibrated for 2 µm)
• Condenser (sub-stage condenser)
• Condenser iris diaphragm
• Field diaphragm
• Light source
• Dust cover

Getting Started [NB: Always start with the 4X objective rotated into the viewing position].

1. Put slide on stage in slide holder; center the specimen under the objective looking directly at your slide if necessary (i.e., not looking through the microscope yet).

2. Turn on light (not too bright). Move specimen into view if it is not already there.

3. Open field diaphragm and condenser iris diaphragm all the way so that light fills your field of view.

4. Look at specimen using either the 4X or the 10X objective


• Adjust focus with focusing knob so that it looks clear to you
• Adjust distance between eyepieces so that you can see the specimen with both eyes If this adjustment is properly made, you should not be aware of two images-- you should see a single, fused image instead.
• Identify which eyepiece is the focusing eyepiece; which is the non-focusing eyepiece
• Refocus specimen using only the eye that looks through the non-focusing eyepiece Close eyelid to other eye, or mask it with your hand when you do this. Be sure that your viewing eye is fully relaxed (get the "stares;" focus on infinity).
• Switch viewing eyes. Adjust focus of focusing eyepiece so that this image is in sharp focus.
• View specimen with both eyes.

5. Repeat this procedure when you switch to the 40X and 100X objectives; the depth of focus of a lens decreases as magnification increases. Your eyes may focus optimally at different distances.

Remember: Do not try to view whole mount specimens with these high power objectives.

Koehler Illumination

All too frequently, sophisticated and well-equipped microscopes fail to yield excellent images because of incorrect use of the light source. Excellent illumination should be bright, glare-free and evenly dispersed in the field of view. Research quality microscopes, such as the ones available for your use in the lab, can achieve high quality images by use of Koehler illumination. This is particularly important when you are using the 40X and 100X objectives.

Adjust your microscope for Koehler illumination

1. While watching specimen, close field diaphragm most of the way

2. Using knob on condenser, raise condenser until edges of the partly close field diaphragm appear superimposed on the already-focused specimen. Edges of field iris diaphragm should appear sharply focused. Substage condenser is usually close to highest position.

3. If field diaphragm is not centered in field of view, use substage condenser centering screws to center the field diaphragm. Slowly open field diaphragm until it just disappears from view. N.B. This must be repeated for each objective. This adjustment produces a fully-lit and evenly-lit field of view.

4. Lift eyepiece out of body tube and look down tube at the back of the fully-lighted objective. While looking down the microscope tube, slowly open and close the substage condenser iris diaphragm. Closing this diaphragm "cut into" the periphery of the back of the objective. For best contrast, approximately 1/4 to 1/3 of the back of the lens should be occluded. Replace the eyepiece in the tube. N.B. This step too, should be repeated for each objective.

5. Adjust intensity of light by adjusting brightness of the bulb. Do not adjust the light by raising or lowering the condenser, or by closing the aperture iris diaphragm.

FIGURE: Diagram of light paths through the lenses during Koehler illumination.

Care of your Microscope

1. Microscopes are precision instruments that should be treated with care. Avoid banging or dropping parts. Carry it by the base, not by the neck that connects the stage and the nosepiece. But in case of an accident, please report trouble to your instructor so that proper repairs can be made.

2. Do not move your microscope once you have set it up. Jarring causes damage to the lamp. Place the microscope in a convenient position and leave it there while working.

3. Never force knobs or levers. All adjustments should be made smoothly. If you cannot understand why you are having difficulty, do not continue. Stop and get help.

4. Never remove parts and leave them lying about unidentified.

Cleaning up

1. Always rotate lowest power objective into position before putting microscope away.

2. Lower stage.

3. Reduce light intensity of bulb. This will prevent an unwanted surge of power through the bulb filament when the light is turn on again.

4. Clean accessible surfaces with lens tissue. Do not try to clean inside lenses or oculars, Inform your instructor about difficult cases, and the parts will be cleaned by an expert. N.B. Do not use solvents for cleaning. Dry lens paper should suffice. Use of improper solvents may dissolve material used to mount the lens.

5. Put on dust cover. Return microscope to storage cabinet.

6. Lock your microscope cabinet. Don't forget your key.

This page was posted 25 January 2003

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