ACTIVITY NO #: 1

ACTIVITY TITLE: Microscopy

Objectives
 Identify the different parts of the compound microscope.
 Describe the function of each part.
 Demonstrate the correct use and care of a compound microscope.
 Be able to troubleshoot microscope.

STUDY

Introduction:
Light, or optical, microscopes are
essential for histological studies because
they allow us to visualize cells and
morphological features of tissues. Light
microscopes rely on glass lenses and visible light
to magnify tissue samples. It was invented in
XVII century, and has been improved over
the years, resulting in the powerful modern
light microscopes. The most remarkable
improvement has been getting better glass
lenses to obtain sharper and non-distorted
images, as well as adding devices to explore
new ways of visualizing tissular features.
Light microscopes contain two main
lenses: objectives and eyepieces (ocular).
Objective gathers the light that went through the tissue, whereas the eyepieces project the tissue image on the eye.
The total magnification is the result of multiplying the objective magnification by the eyepiece magnification. For
example, if we have a 40x objective (magnifies 40 times) and a 1x eyepiece (magnifies 10 times), the total
magnification is 400 times. More advanced microscopes can get 1000 to 1500 magnifications (100x objective
and 10x or 15x eyepiece). Some light microscopes may contain additional internal lenses between the objective
and the eyepiece that can change the total magnification. Magnification and resolution power must not be
confused, because not matter how we can magnify an image, including digital methods, the resolution cannot be
increased.
 Let’s find out how much you already know about this module. Briefly answer the following
questions.

Prelab Questions:
1. What are the three components of a microscope?

2. How electron microscope is different from light microscope?

3. Why is that microscopy is essential to histopathology?

RESEARCH

Materials:
Compound Microscope
Prepared tissue sample (Colon or Gall Bladder)

Procedure
1. Place the microscope on the bench squarely in front of you.
2. Obtain a prepare slide and place it in the clips on the mechanical stage.
3. Adjust the eyepieces on a binocular microscope to your own personal measurements.
a. Look through the eyepieces and using the thumb wheel, adjust the distance between the eyepieces
until one circle of light appears.
b. With the low-power (10x) objective in place, cover the left eyepiece with a small card and focus the
microscope on the slide. When the right eyepiece has been focused, remove your hand from the
focusing knobs and cover the right eyepiece. Looking through the microscope with your left eye,
focus the left eyepiece by turning the eyepiece adjustment. Make a note of the number at which you
focused the left eyepiece so you can adjust any binocular microscope for your eyes.
4. Raise the condenser up to the stage. On some microscopes, you can focus the condenser by the following
procedure:
a. Focus with the 10X objective.
b. Close the iris diaphragm so only a minimum of light enters the objective lens.
 c. Lower the condenser until you see the light as a circle in the center of the field. On some
microscope, you can center the circle of light (Figure 4) by using the centering screws found on the
condenser.
d. Raise the condenser up to the slide, lower it, and stop when the color on the periphery changes from pink
to blue (usually 1 or 2 mm below the stage).
5. Open the iris diaphragm until the light just fills the field.
6. Adjust the contrast by changing the diaphragm opening.
7. When you brought an image into focus with high power, rotate the turret to the next lens, and the subject will
remain almost in focus. All of the objectives (with the possible exception of the 4X) are parfocal; that is, when
a subject is in focus with one lens, it will be in focus with all the lenses. When you have completed your
observations under low power, swing the high-dry objective into position and focus. Use the fine adjustment.
Only a slight adjustment should be required.
8. Move the high-dry lens out of position, and place a drop of immersion oil on the area of the slide you are
observing. Carefully click the oil immersion lens into position. It should now be immersed in the oil (Figure
5). Careful use of fine adjustment knob should bring the object into focus.
9. Record your observations and note the magnifications.

(a) (b) (c)

FIGURE 4 Focusing the condenser. (a) Using low power, lower the condenser until a distinct
circle of light is visible. (b) Center the circle of light using the centering screws. (c) Open the iris
diaphragm until the light just fills the field. Precision Graphics, Johnson, et al (2014)
 ANALYZE

Data and Results

1. Label the parts of the compound microscope.

2. From the labelled parts of microscope in number 1, identify which parts are under mechanical,
magnifying and illuminating parts and give its general functions.
Mechanical Parts Magnifying Parts Illuminating Parts
 Functions Functions Functions

3. Examining the slide stained in H&E.

a. Draw your observation at the center as it appears in the low and high-power field.

ACTION
Critical Thinking and Application

1. Lisa was assigned to view the tissue sample of the small intestine under the compound
microscope. She started to operate and observe the specimen; however, she said everything was dark,
and she could not see anything. What could be the problem?

2. Your teacher asks you to help Antonio because he has a problem operating the microscope, especially
high-power objectives. He said that everything disappears every time he views the specimen under
HPO. How will you resolve Antonio's issue?
 3. Mirabel complained that the prepared slide given to her had no specimen to observe; she added that it
was all blurry. However, when the prepared slide was examined under a different microscope, there was a
specimen, and it was crystal clear. What could be the problem with Mirabel's microscope?

REFERENCES:
Lecture notes in medical technology. http://mt-lectures.blogspot.com/2017/08/lecture-13-morphological-
examination-of.html
Atlas of plant and animal histology. https://mmegias.webs.uvigo.es/02-english/6-tecnicas/6-
optico.php#:~:text=Light%2C%20or%20optical%2C%20microscopes,light%20to%20magnify%20tissue
%20samples.
ThomasTKtungnung. (February 13, 2020). Blood smear preparation and staining practical lab.
https://www.youtube.com/watch?v=KSs0SMfERuA
Spitalnik, P.F. Histology laboratory manual. Vagelos College of Physicians & Columbia University.