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COMPOUND MONOCULAR
MICROSCOPE
The microscope is one of the principal tools of the biologist. It was invented through the efforts of Dutch scientist
Anton Van Leeuwenhoek. In the laboratory, the microscope serves as a very useful tool to help you discover the
fascinating secrets of the living world which the unaided eye cannot see. In this exercise, you will study the proper use
and care of this delicate instrument.
OBJECTIVES
At the end of the period, students should be able to:
1. identify the parts of a compound microscope and learn the function of each part;
2. manipulate the different parts correctly.
3. compute for the magnification of drawing.
4. differentiate the low power objective from the high power objective in relation to the size of the field of
vision, magnification, and resolving power.
MATERIALS
compound microscope human hair (gray, black, and colored)
glass slide medicine dropper
cover slip transparent or plastic ruler
colored threads small letter "e" cut out from an old newspaper
PROCEDURE:
Before studying the different parts of your microscope, bear in mind the following points:
1. Other students in different classes also use your microscope. Always inspect it before using and report to the
instructor any damage you can see.
2. The microscope is a delicate and expensive instrument. Exercise utmost care when using it so that other
students will be able to use it too.
3. Manipulate the microscope only when you are thoroughly familiar with it.
Focal length (mm), an optical constant of the lens system, is the distance from the center of the lens to the
point where parallel rays entering the lens are brought to a focus
Working distance (mm.) is the free space between the specimen surface and the objective.
Numerical aperture (N.A.) is a measure of the resolving power of an objective. An objective with 0.25 N.A.
allows the viewer to distinguish as separate 25000 lines per inch.
B. Microscope Manipulation
1. After familiarizing yourself with the parts of the microscope and the function(s) of each, you may now
proceed to manipulate it. Before you proceed, here are some reminders each time you prepare for
microscopic work:
a. Place the microscope in an upright position with the arm directed towards you. Make sure that it is at
least two inches from the edge of the table.
b. Make certain that the low power objective is in position (a slight "click" will indicate that the LPO is
already in place).
c. Adjust the diaphragm to its largest opening.
d. Look through the eyepiece with one eye while keeping the other eye open and adjust the angle of the
mirror to obtain good illumination.
Note: Keeping the eyes both open while using the microscope prevents eye strain.
2. Cut out a small letter "e" from an old newspaper. Prepare a wet mount slide of the letter "e" following the
procedure outlined in Figure 3.
3. Place the wet mount slide on the stage of the microscope with the letter "e" in its normal reading position.
See to it that the letter "e" is right in the center of the stage aperture. Secure the slide into place with the
stage clips.
4. Slowly turn the coarse adjustment knob downward to lower the body tube until the tip of the LPO almost
touches the slide. Do this while looking from the side to prevent the unlikely event of breaking the slide.
While looking into the ocular, slowly turn the coarse adjustment knob upward to raise the LPO until the
letter comes into focus. Turn the fine adjustment knob slightly to obtain a sharp image of the letter.
a. Draw the letter "e" in its normal size and position, and as seen under the LPO.
Fig.B.1 Letter “e” on the slide Fig.B.2. Letter “e” under LPO (mag. ____x)
b. Compare the size and position of the letter "e" as viewed under the LPO with the actual position of the
letter "e" on the slide.
C. Orientation
1. While looking into the ocular, move the slide slightly to your right. Next move it to your left. Then upward and
downward.
a. In which direction does the image of the letter “e” appear to move in each case?
2. Move the slide towards you and then away from you.
b. In what direction does the image appear to move in each case?
3. Rotate the nosepiece to put the high power objective into position. Make sure that as the objective moves into
position, it does not hit the stage or the slide. Listen for a "click" sound which means that the objective is in place.
Most microscopes today are parfocal. This means that once an image is in sharp focus under LPO, it remains
in focus under HPO without further adjustment. If the image is not in sharp focus, bring it into sharp focus with a
slight turn of the fine adjustment knob. If at first try you fail to see the image under HPO, refocus under LPO then
repeat the procedure for focusing with the HPO until you see the image.
Note: Do not use the coarse adjustment knob when the HPO is in place. The reason for this may be obvious if you
look at the distance between the HPO and the top of the slide. The distance is so close that a downward movement of
the objective has the danger of hitting and breaking the slide. With the fine adjustment knob, the upward and
downward movements of the objective are very slight and do not normally hit the slide or the stage.
E. Field of Vision
The field of vision is that circular lighted field where you see the image of the object or specimen.
1. Return the letter “e” to the center of the field of view. Without moving the coarse adjustment knob, shift the objective
from LPO to HPO by turning the nosepiece. Sharpen the focus using the fine adjustment knob. If necessary readjust the opening of
the diaphragm (or aperture disc) and the position of the mirror to improve light and contrast.
2. Draw the resulting image of the letter “e” under HPO as Figure E.1.
The resolving power of the microscope is its ability to distinguish two adjacent points as distinct and separate.
The depth of focus is a property of the microscope that indicates how deep the space the microscope can bring
into clear view at any one time.
The materials used for microscopic examination have depth, as well as length and width so that you need to focus
at different levels of the specimen.
1. Mount a gray and black hair across each other on a microscope slide. Add a drop of water and cover with a
cover slip. Focus under low power objective. Turn the fine adjustment knob back and forth slowly. Draw the
resulting image as Figure F.1.
a. Can you determine which hair is on top when looking through the microscope?
2. Move the slide to position the hair strands at the center of the field of vision. Shift to HPO and bring the hair
strands into sharp focus with the fine adjustment knob. Draw the resulting image as figure F.2.
b. Is the resolving power in the HPO as great as in the LPO?
3. Obtain a prepared slide with colored threads. Place the slide on the stage and focus under LPO the point where
all three threads intersect. Draw the intersection of the colored threads under LPO as Figure F.3. Rack the body tube
to the lowest position without hitting the slide. The threads may be out of focus. Slowly rack the body tube upward.
The first thread that comes into focus is the bottom one. Focus the other threads.
c. When one thread is in focus, are the other threads in focus too?
d. What is the color sequence of the threads from bottom to top?
4. Shift to HPO. Slowly rotate counter clockwise the fine adjustment knob until all the threads are out of focus.
Next, slowly rotate it clockwise. The first thread that comes into focus is the top one. Focus the other threads. Draw
any of the colored threads as Figure F.4.
e. When one thread is in focus, are the other threads in focus too?
f. In which objective is it easier to determine the sequence of the colored threads? Why?
Fig. F.1. Intersection of hair strands under LPO. Fig. F.2. Hair strands under HPO.
(mag. ____x) (mag. ___x)
Questions to Answer:
1. How can each objective be identified if the identification marks are removed?
2. How does a microscope magnify an object?
3. Explain why an inverted image is seen under a compound microscope?
4. What is the purpose of the oil in the oil immersion objective?
5. Why should the ocular micrometer be calibrated for each objective?
References:
Duka. I. A. and M. G. Diaz. 2007. Biology 1 Laboratory Manual: An Investigative Approach. 8 th ed. UPLB. pp. 6 – 13.
Fernandez, W.L. et al. 1986. General Microbiology Laboratory Manual. UPLB. pp. 1 – 15. UST General Microbiology
Manual
Format of Post Lab Report
Exercise No. 1
Compound Monocular Microscope
Name:
Date Performed
Date Submitted:
Instructor:
I. Objectives:
II. Materials:
III. Methodology: (Summarized form and should be in a passive and past tense form)