The Microscope

We will be using a compound light microscope in this lab to view various cells and
tissues. It is very important that you learn to use the microscope correctly, and can
efficiently get images into the proper focus for study.

Know the following parts of the microscope and the function of each part.

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 Know the following functions of the microscope parts.

PART FUNCTION
Arm Used to carry the microscope; located between the
body tube and the base

Base Supports the microscope; used to carry the microscope

Body tube (Head) Supports the objective lens system and the ocular
lens; directs light toward the ocular lens

Coarse focus (adjustment) Brings the specimen into focus by raising or lowering the
stage; never use the coarse focus on high power!

Condenser Substage lens that concentrates light on the

specimen; the condenser control knob will raise and
lower the condenser to vary the light; the best position
for the condenser is just below the opening in the
stage

Fine focus (adjustment) Used for final focusing (fine-tuning the image)

Iris diaphragm Regulates the amount of light that passes through the
stage; located at the base of the condenser

Objective lenses Magnify the specimen for viewing; scanning lens (red
ring) magnifies 4X, low power lens (yellow ring)
magnifies 10X, high power lens (blue ring) magnifies
40X, oil immersion lens (white ring) magnifies 100X

Ocular lens (eyepiece) The lens (or lenses) you look through; magnifies the
specimen 10X; monocular microscopes have one
ocular lens; binocular microscopes have two ocular
lens
Revolving nosepiece Contains the objective lenses; use nosepiece to rotate
the correct objective lens into place over the opening in
the stage

Stage Flat platform where the slide is placed for viewing

Stage clips Hold the slide in place; two knobs below the stage
control the precise movement of the specimen on a
mechanical stage

Substage light Light source; a light dimmer dial on the base varies
the intensity of the light source

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Use and Care of a Compound Microscope
1. Always use both hands when transporting the microscope. Place one hand on the
arm and the other hand under the base. Make sure the cord is not hanging down
so you don’t trip over it.
2. Carefully place the microscope on your work table.
3. Carefully remove the dust cover and set it aside.
4. Place the cord behind the microscope then plug it into the outlet. Be very careful
not to trip over the cord or pull on it.
5. Once you have the microscope plugged in, sit down and find the light switch. The
light switch on your microscope is located on the side of the base (see
microscope picture above). Turn on the light switch and adjust the intensity of
the light with the light dimmer dial (see microscope picture above).
6. Turn and “lock” the scanning lens (4X objective) in place above the opening in the
stage. If the lens is in the correct place, you should see a bright circle of light when you
look through the eyepiece. This circle of light is your field of view.
7. Without looking through the eyepiece, lower the stage completely and carefully
place the microscope slide on the microscope stage with the specimen on the slide
directly over the opening in the stage. If your microscope has stage clips, secure
the slide under the stage clips. If your microscope has a mechanical stage, turn
the control knobs (below one side of the stage) to position the specimen directly
over the opening in the stage.
8. While looking through the eyepiece, carefully turn the coarse adjustment knob to
raise the stage until the specimen comes into view.
9. Use the fine adjustment knob for final focusing of the image.
10. Since the scanning lens allows you to see a larger portion of your slide in less
detail, you may need to increase your magnification. To switch to the next
magnification, carefully turn the revolving nosepiece until the low power lens (10X
objective) is locked in place.
11. Your compound light microscope is parfocal. If your image was in proper focus
before you changed the objective lens, you should only have to do minimal
focusing to get the image in focus with the new objective. You should only have
to use the fine focus knob on low power to get the image in proper focus.
12. If you need to increase your magnification to the high-power lens (40X objective),
simply turn the revolving nosepiece until the 40X objective is locked in place. Make
sure your image is in proper focus on low power before switching to the high-power
objective.
13. Use ONLY the fine adjustment knob to focus on high power. NEVER USE THE
COARSE FOCUS KNOB ON HIGH POWER! The high-power lens should be very
close to your slide when in proper focus. If you turn the coarse adjustment knob
while on high power, the objective could easily break your slide.
NOTE: If you lose the image on high power, go back to low power and
find/focus it before going up to high power again.
14. When you are finished with the microscope, return the scanning lens (4X) to
place. Lower the stage completely, remove the slide, and carefully place into
the correct box.
15. Carefully clean all the lenses with lens paper. Never use paper towels or any
other kind of paper/cloth on your microscope lenses as it may scratch the glass.

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 16. Carefully unplug the microscope and wrap the cord snugly around the base.
17. Place the dust cover over the microscope and carefully carry it with both hands
back to where you got it.

Viewing Objects Through the Microscope

Procedure:
1. Place a paramecium slide on the microscope.

2. Bring the slide into focus using the scanning (4X) lens.

3. Observe how much of the paramecium you can see. Make a drawing of your paramecium below.

Scanning Lens Observation:

4. Now, switch to low power (10X) and use the fine adjustment knob to focus the image.
Note: Before switching to the low power lens, make sure your paramecium is in the
center of your field of view.

5. Observe how much of the paramecium you can see. Make a drawing below.

Low Power Observation:

6. Lastly, switch to high power (40X). Make sure your paramecium is in the center of your low
power field of view before you switch to the high-power lens. If you lose the image, go back to
the low power lens, focus it, put it in the center, switch to the high-power lens, and use fine
focus only.

7. Observe how much of the paramecium you can see now. Make a drawing below.

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 High Power Observation:

Answer the parts of Question #8 with either “Increased” or “Decreased”

8. As you increased the magnification, what happened to the:
(A) working distance (distance between the lens and specimen)?
(B) field size (how much of the slide in your field of view)?
(C) image brightness and resolution (clearness of the image)?
9. Why is the letter “e “inverted when viewed under the microscope?
10. Identify the different types of microscopes and give their use.
11. Give the advantages and disadvantages in using the electron microscope.

Estimating the Size of the Field

1. Switch to the scanning lens on your microscope, lower the stage, and remove the slide.

2. Obtain a metric ruler and place it under the stage clips with the millimeters (smallest
marks) over the opening in the stage.

3. Switch to the low power lens and raise the stage until the millimeters are in focus.

4. Count the number of millimeters that will fit in your low power field of view. The
millimeters are counted as the spaces between the dark lines. Record the number
below.

Diameter of Low Power Field of View = mm

5. To calculate the diameter of your low power field of view in micrometers, multiply
your answer in #4 by 1,000.

Diameter of Low Power Field of View =

mm x 1,000 = µm

6. Lower the stage completely and remove the ruler.

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 7. Place the paramecium slide on the microscope and bring into focus using the low
power lens.

8. Choose one paramecium cell and estimate its length and width in µm (based on the
diameter of the entire field of view).

Example: if the low power field of view was 1000µm and the paramecium cell
occupied one-half of the field of view, you could estimate the length to be
500µm. If it occupied one-fourth of the field of view, you could estimate the
length to be 250µm.

Length = µm Width = µm
The Lens Effect

1. Place the “Letter e” slide in the stage clips (slide label right side up).

2. Get the letter “e” in focus using the low power lens.

3. Observe the position of the letter. Make a drawing below.

Letter “e” Observation:

4. How does the position of the letter viewed through the microscope differ from the
actual position of the letter on the slide?

5. Looking through the microscope, move your slide to the right.

In which direction does the image move?

6. Looking through the microscope, move your slide forward (away from you).

In which direction does the image move?

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 The mirrors in the microscope flip and reverse the image before it reaches your eyes.
This is called the lens effect. This may be a little confusing at first, but with practice
you will learn to move the slide in the desired direction with no problem

Perceiving Depth and the Depth of Field

1. Obtain a slide with three colored threads crossed and get the image into focus on low power.

2. Adjust the light with the diaphragm, if necessary, to increase the contrast of the threads.

3. Move the slide so that the area where all three threads overlap is in the center of your
field of view and adjust the image so that it is in clear focus.

4. Switch to the high-power lens and use the fine adjustment knob to fine-tune the image.
Move the slide so you can see all three colors at once. Note: you will only be able to
see a very small portion of each color.
5. Looking through the microscope, turn the fine adjustment knob to lower the stage just
until the threads are out of focus. Now, slowly roll the stage back up noting which
color thread comes into clear focus first, then second, then last. The first thread to
come into focus when you raise the stage is the one that is on top. The second thread
to come into focus is the one in the middle.

Record your observations, relative to which color of thread is uppermost, middle,

and lowest. Top thread color

Middle thread color

Bottom thread color

There is always a certain amount of distance between the cover glass and the slide. This
distance becomes more apparent when you increase the magnification of the specimen.

Put out your hand in front of you so you can see your hand and the wall behind it. Can
you get your hand and the distant wall into clear focus at the same time? No. This is
similar to what you are experiencing when trying to view a specimen on high power.
You cannot get the entire specimen into focus at the same time. The portion of the
slide that is in clear focus at any given time is the depth of field.

When you view the various tissue slides in upcoming lab exercises, you will likely be
viewing more than one layer of cells. When viewing these slides on high power, you
will need to move the fine adjustment back and forth slowly to view the entire slide.

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Determining Total Magnification
Total magnification is the total number of times the specimen you are viewing is
magnified. To determine the total magnification, you multiply the ocular lens
magnification times the objective lens magnification. On your microscopes, the ocular
lens always magnifies 10X.
Example: 10X (ocular) x 4X (objective) = 40X (total magnification)
Scanning Low power High power Oil immersion
Magnification of
Objective Lens

Magnification of
Ocular Lens

Total
Magnification

Preparation of a Wet Mount

1.
2. Obtain a clean microscope slide, a cover slip, and a toothpick.

3. Place a small drop of methylene blue stain on the clean microscope slide.

4. Take a clean toothpick and gently scrape the inside of your cheek.

5. Roll the toothpick with the cheek scrapings into the drop of stain on your slide and stir the
contents gently.

6. Put the toothpick into the coffee can provided on your lab bench.

7. Carefully lower a cover slip over the stain/cheek scrapings mixture.

Note: Slowly lowering the cover slip at about a 450-degree angle helps avoid air bubbles
getting trapped beneath the cover slip.

This is called a temporary, wet mount, since a cover slip was placed over a
specimen within a liquid.
8. Observe your cheek scrapings under the scanning lens (4X), the low power lens (10X),
and the high-power lens (40X). Remember to put the cells in the center of your field of
view before increasing magnification.

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 9. Find a few isolated cheek cells. You may see clumps of cells and/or crumpled cells. Try to
find a few whole, flat cells out by themselves and focus on them. You should see roundish
cells, with a stained nucleus in the center.

10.Make drawings of your cheek cells below.

11. When you are finished, return the scanning lens to position, lower the stage, remove the
slide and place the slide in the coffee can provided on your lab bench.

Scanning Observation:

Low Power Observation:

High Power Observation:

SELF-ASSESSMENT QUESTION 1:

1. Why is the letter “e “inverted when viewed under the microscope?

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2. Identify the different types of microscopes and give their uses.

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 _______________________________________________________________________

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3. Give the advantage and disadvantages in using the electron microscope.

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