Exercise No.

THE COMPOUND MICROSCOPE

PART III. VIEWING YOUR SPECIMEN

1. Prepare a wet mount using a very small letter “e” cut from a magazine page.

A. Describe in detail the process of preparing the wet mount. Provide an illustration
(picture or drawing with correct labels).

Carefully cut out a small piece of paper that contains the letter e with scissors.
With forceps, pick up the letter e-containing piece and put it in the center of a
clean microscope slide.
 Place a drop of water on the specimen.

 Place a cover slip on the slide. Put one edge of the cover slip into the
drop of water, and then slowly lower the cover slip over the specimen.

 Remove any air bubbles from under the cover slip by gently tapping the
cover slip.

 Dry any excess water before placing the slide on the microscope stage
for viewing.
  Prepare a wet mount in order to examine the letter under the microscope:
Place the specimen in the center of a clean slide.

 Observe the letter e under low power and high power. Scan the slide so
that you have observed the letter e from side to side and top to bottom of
the field of view.

B. How will the presence of air bubbles be prevented? What should be done in case
air bubbles are present? 

Place one edge of the coverslip against the slide at a 45° angle so that it meets
the edge of the water drop. To stop fingerprints, keep the coverslip by the corners. The
liquid should cover the entire surface of the coverslip. Lower the coverslip slowly over
the specimen. The goal is to keep air bubbles under the cover slip to a minimum. One
toothpick may be used to keep the lower edge in place while another is used to carefully
lower the slip into place. If air bubbles are trapped under the coverslip, gently tap it with
the eraser end of a pencil to release them.

C. What is the proper way of dealing with the following situations: there is too much
water and the cover slip is floating around and if there is too little water and some
of the space under the cover slip is still dry?

Simply place a drop of water on the top of the letter with a dropper. Because if
too much water is added, the cover slip will "float," resulting in an excessively thick water
layer. If there is insufficient water, the specimen may be crushed or dry out too quickly.
To avoid this situation, simply place the desired amount of water, which is only a drop,
and use a dropper.

D. How are liquid substances added to a wet mount?

Steps to how liquid substance added to wet mount

 Prepare a wet mount slide.
 Collect a drop of liquid substance with an eye dropper or pipette.
 Place a drop of liquid substance on one edge of the coverslip, and
the flat edge of a piece of paper towel
 on the other edge of the coverslip. The paper towel will draw the water ou
t from under the coverslip, and the
cohesion of the water will draw the stain under the coverslip.
 As soon as the stain has covered the area containing the specimen you a
re finished. The stain does not need to be under the entire coverslip. If the
stain does not cover the area needed, get a new piece of paper
towel and add more stain until it does.
 The slide is now ready for viewing.

2. Set up the microscope.

A. Enumerate the steps in setting up the microscope.  

PREPARING THE HEAD

• Before puning a specimen on the sta8e, turn on the illumination and set it to a comfortable
intensity level.
• Rotate the 1OX objective into position.

• Rotate eyepiece diopter ring to ’0’ (white line on CFWN eyepieces). See diagram above.
• Adjust interpupillary distance so both right and left images merge into one.

2.
 FOCUSING FOR YOUR EYES

• Place your specimen on the stage.

• Using the coarse knob, focus the 10X objective. Adjust with fine focus on smallest detail
visible.
• Position the 40X objective and fine focus.
• Note: If reticle is used, focus reticle. Bring specimen image into focus on reticle. Adjust other
eyepiece to match. Skip 4X procedure.
• Switch to 4X objective. Adjust diopters of eyepieces to accommodate.
• Recheck focus at 40X.

3.
SETTING THE CONDENSER FOCUS

• Using the field diaphragm control ring, close the field diaphragm to its smallest size.
Bring the image of the field diaphragm into focus with the condenser focus knob.

• Using the condenser centering screws, bring the field diaphragm image to the center of the
field of view.
Diaphragm must be centered before clearing from the field of view.

• Centering must be checked for each objective.

4.
CENTERING THE ILLUMINATION
• To focus and center the illumination remove the diffuser.
• Close the aperture diaphragm on the condenser.
• Use a filter (the ND or blue filter) as a mirror to observe the filament image on the underside
of the condenser. (For reflected light systems remove an eyepiece and look at image at
back of)

• Focus the filament image until it becomes sharp.

• After illumination is focused and centered, replace the diffuser.
Slide to focus Centering
screurs

Clamp screw Focusing

knob

SOW Lamphouse HMX-2 iarnpbozae

5.
CONTROLLING CONTRAST

AND DEPTH OF FIELD

This last step controls contrast and depth of field with the condenser aperture diaphragm.

• Remove one eyepiece. Look down the tube at the back of the objective.
• Adjust the aperture diaphragm so that it is just inside the opening (about 25% less than full
aperture).

Exit pupil of objective

Aperture diaphragm
Size of the condenser aperture diaphragm
 B. What is the importance of adjusting the interpupillary distance of the lenses in
a microscope with two oculars?  How will you know that the correct width is
achieved? Are we going to use both eyes in viewing?

The distance between the centers of your two pupils is measured as the
interpupillary distance. The distance between the binocular microscope's two
eyepieces must correspond to your interpupillary distance. Each person has a
unique interpupillary distance, and the microscope must be adjusted to
accommodate this. This applies to any binocular microscope. You must determine
the correct interpupillary distance and set the microscope for that distance during
your first session with a new microscope. During each subsequent session, you must
ensure that the interpupillary distance of the microscope is correct for you and adjust
it if it is not. This will yield better image quality and reduce eye strain. If you see two
images when looking through the eyepieces, the interpupillary distance is incorrect.
To fix it, move the eyepieces closer or farther apart until the two fields of light merge
to form a single circle of light. Your interpupillary distance is now correct.

D. While peering into the eyepiece, move the knob of the iris diaphragm towards and
away from you. Is there a difference in the brightness of the field? What causes the
difference?
Moving the lever to the left increases the light; moving the lever to the right
decreases the light. Forward/backward stage control: moves the mechanical stage
holding the slide forward and backward. At low power, the amount of light transmitted
to your eye is greatest. Light (and thus resolving power or the ability to discern two
nearby objects as separate) is reduced when you turn to a higher power. Make up for
it with light access (sometimes called the iris diaphragm).
On the lowest power objective, the field of view is the widest. When you increase
the magnification, the field of view narrows. On low power objective, you’ll see more
of an object.
On the lowest power objective, the depth of concentration is greatest. The depth of
concentration is reduced each time you turn to a higher power objective. As a result,
at higher magnification, a smaller portion of the specimen is in view.

3. View the specimen with the scanner.

F. Adjust the light for maximum contrast using the iris diaphragm. Can you see the entire
letter? What is the position of the letter when seen through the eyepiece?
 The magnification that will be used to display the letter will be determined by the
magnification that will be used. You can see the letter better with the low power objective
than with the high power objective, which just shows a portion of the letter. The letter will
appear upside down and backwards. Because of two sets of mirrors in the microscope,
the letter appears upside down and backwards. This means you'll have to pass the slide
in the opposite direction as the picture moves.

G. Using the stage control knobs, move the slide until you find the letter “e” in your field of
view. If you have markedly different eyesight between your eyes, how will you make
use of the diopter ring to gain sharp focus?

The diopter ring is used to make the eye have a clear focus on the specimen if
the two eyes have different vision. By changing each side based on the two eyes' vision
until they are both sharply focused.

4. View the specimen with the low-power objective.  

This is a 40x magnified picture of the letter e. The letter was placed in the proper orientation on this
slide. When viewed under a microscope, it appears to be upside down. Because of two sets of mirrors in
the microscope, the letter appears upside down and backwards. This means you'll have to pass the slide
in the opposite direction as the picture moves.

The slide moves to the left when you move it to the right on the stage, and vice versa. The slide
moves toward you if you move it away from you on the stage. Knowing how the controls will move the
slide will make it much easier and faster to locate objects on your slide.

5. View the specimen with the high-power objective.  

This is a 100x magnified picture of the letter "e." You'll notice that as the lens's power increases, your
field of view narrows.

When using a microscope at high magnification, the field of view shrinks. The magnification of
the objective lens has an opposite relationship with the field of view. With a higher magnification, the
specimen appears bigger since a smaller area of the sample is stretched out to fill the field of view of
your eye.

6. If the microscope has an oil immersion objective, then view the specimen using this lens.  

A. Turn the nosepiece so that no objective lens is directly over the slide.  Then place a
small drop of immersion oil on top of the cover slip. Provide an illustration (picture or
drawing with correct labels).
7. Turn off the microscope.  
a. Lower the stage and then remove the slide. Clean and dry the slide and cover slip.  
b. Clean and dry the stage and lenses USING THE RIGHT KIND OF PAPER!
c. Place the 4X objective over the stage, facing the front.
d. Center the mechanical stage and make sure it is fully lowered.
e. Unplug the microscope and wrap the cord correctly.
f. Cover the microscope with the dust jacket.
g. Return the microscope to the correct cabinet placing the oculars toward the BACK of the
cabinet.