Name : ___________________________________ Course/Year: __________ Date Performed: __________

Section : ___________ Group No. _______ Date Submitted : __________

Instructor : _________________________ Rating: _________________

Activity No. 1
Proper Use and Handling of Microscope

I. Introduction:
The applications of microscopy in the forensic sciences are almost limitless. This is due in large
measure to the ability of microscopes to detect, resolve and image the smallest items of evidence, often without
alteration or destruction. As a result, microscopes have become nearly indispensable in all forensic disciplines
involving the natural sciences. Thus, a firearms examiner comparing a bullet, a trace evidence specialist identifying
and comparing fibers, hairs, soils or dust, a document examiner studying ink line crossings or paper fibers, and a
serologist scrutinizing a bloodstain, all rely on microscopes, in spite of the fact that each may use them in different
ways and for different purposes.

II. Objectives:

After the activity, you should be able to:

1. Identify the parts of the microscope and their functions.
2. Become familiar with the 3 variations of light microscopy.
3. Determine total magnification of the specimen, using various objective lenses.
4. Be able to switch objective lenses, while focused on the specimen, without moving the stage.
5. Handle the microscope safely and clean it.
6. Explain principles and terms used in microscopy and focusing.
III. Materials:
Microscope glass slide glass cover

IV. Procedure:
A. Safe Handling and Operation
1. When moving your microscope, always carry it with both hands (Figure 1). Grasp the arm with one hand
and place the other hand under the base for support.
2. Keep one hand on the arm, and one on the base. Do not carry anything else with a microscope.
3. Clean ALL objective lenses and the ocular with lens paper BEFORE you even place a slide on the stage, and it
is a good idea to wipe the condenser lens also. The last person using the microscope may have left it dirty:
it is imperative to begin with cleanliness.
4. Turn the revolving nosepiece so that the lowest power objective lens is "clicked" into position.
5. Place the microscope slide on the stage and fasten it with the stage clips. You can push down on the back
end of the stage clip to open it.
6. Using the coarse adjustment, lower the objective lens down as far as it will go without touching the slide!
Note: Look at the slide and lens from the side when doing this (see Figure 2).
7. Look through the eyepiece and adjust the illuminator (or mirror) and diaphragm (Figure 3) for the greatest
amount of light.

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8. Slowly turn the coarse adjustment so that the objective lens goes up (away from the slide). Continue until the
image comes into focus. Use the fine adjustment, if available, for fine focusing.

7. Move the microscope slide around so that the image is in the center of the field of view and readjust the mirror,
illuminator or diaphragm for the clearest image.

8. You should be able to change to the next objective lenses with only slight focusing adjustment. Use the fine
adjustment, if available. If you cannot focus on your specimen, repeat steps 4 through 7 with the higher power
objective lens in place. DO NOT ALLOW THE LENS TO TOUCH THE SLIDE!

9. The proper way to use a monocular microscope is to look through the eyepiece with one eye and keep the other
eye open (this helps avoid eye strain). If you have to close one eye when looking into the microscope, it's ok.
Remember, everything is upside down and backwards. When you move the slide to the right, the image goes
NOTE: Most microscopes are parfocal; that is, the slide should be in focus (or nearly so) when you switch from
low-power to high-power. If you have properly focused your microscope at low-power and your slide is properly
prepared (i.e. the object is flattened by a coverslip), you should be able to switch automatically from low-power to
high without fear of having the high-power objective lens scraping or touching the slide.An important factor in
using the microscope is the distance between the object viewed and the objective lens of the microscope. This is
called the working distance. As you increase magnification the working distance becomes less.to the left!

10. Do not touch the glass part of the lenses with your fingers. Use only special lens paper to clean the lenses.
(read the page on keeping your microscope clean).

11. When finished, raise the tube, click the low power lens into position and remove the slide.

Remember, microscopes are expensive scientific instruments. Handle them properly and carefully and
they will last for many years!

B. Parts of A Compound Microscope:

1. OCULAR LENS or EYEPIECE On a binocular scope there are two ocular lenses, one for each eye.
These lenses magnify the image at 10Xpower. The power of the ocular lens multiplied by the
objective lens gives the total magnification of the microscope.
2. ARM A support for the upper portion of the scope. It also serves as a convenient carrying handle.
3. MECHANICAL STAGE CONTROLS A geared device to move the slide (placed in the slide clamp)
precisely.

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4. COARSE ADJUSTMENT KNOB A rapid control which allows for quick focusing by moving the
objective lens or stage up and down. It is used for initial focusing.
5. FINE ADJUSTMENT KNOB A slow but precise control used to fine focus the image when viewing
at the higher magnifications.
6. BASE The part of your microscope that sits on a level, stable support.
7. NOSEPIECE A circular plate with 4 objective lenses that can be rotated into position for different
magnifications.
8. OBJECTIVE LENS Four separate lenses that magnify the image (4X, 10X, 40X and 100X)
depending on the objective in use. The lens is positioned just above the object being viewed.
OBJECTIVE POWER OBJECTIVE NAME
4X SCANNING
10X LOWPOWER
40X HIGH POWER
100X OIL IMMERSION
9. SLIDE CLIP A clip to hold the slide on the stage.
10. STAGE A platform for placement of the microscope slide.
11. CONDENSER A lens that concentrates or directs the light onto the slide.
12. IRIS DIAPHRAGM CONTROL A lever (or rotating disk) that adjusts the amount of light
illuminating the slide. Use just enough light to illuminate the object on the slide and give good contrast.
13. LAMP The light source.
14. LAMP SWITCH Turns the lamp on and off.

Draw and Label the Different Parts of A MICROSCOPE

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Guide Questions:
1. Why move the 10X objective lens into place when putting the microscope back into the cabinet?
2 How is the image of an object seen through the high- power objective different from the image seen through
the low-power objective?
3. What would be the total magnification of a specimen using the 40X objective lens?
4. What is parfocal?
5. Why reduce your light when using the 10X objective lens?
6. What happens to the light intensity as you increase magnification?
7. What term is used to describe the feature of the microscope that makes it possible to move among the
objective lenses with just MINOR focusing?
8. Which objective lens is also called the oil-immersion lens?
9. Why do we have to clean all objective lenses before using the microscope?
10.Why do we have to look at the side and not at the center when when lowering the objective lense using the
coarse adjustment knob?

Conclusion:

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 The simple microscope, or magnifying glass, comprising a single converging lens, was known in ancient times, but the first
compound microscope is thought to have been invented by the Dutch spectacle-maker Zacharias Janssen around 1590. However,
because of the aberration unavoidable in early lens systems, the simple, single-lens microscope held its own for many years,
Anton van Leeuwenhoek (16321723) constructed many fine examples using tiny, near-spherical lenses. His single-lens
microscopes were capable of magnifying up to 300 times. With them he discovered microorganisms, thereby founding the
science of microbiology and providing the basis for the germ theory of disease. Probably the greatest of the early microscopists,
however, was the Italian Marcello Malpighi (16281694), who is generally regarded as the founder of histology. Compound
microscopes incorporating achromatic lenses became available from the 1840s.

Light microscopes continued to be refined, with the development of the phase-contrast microscope, for example. However, the
next major advances were instruments that used electrons instead of light the transmission electron microscope (TEM),
invented in the early 1930s, and the scanning electron microscope (SEM), invented in the mid-1960s.

MAGNIFICATION AND RESOLUTION

In using the microscope it is important to know how much you are magnifying an object. To compute the total magnification of
any specimen being viewed multiply the power of the eye piece (ocular lens) by the power of the objective lens being used. For
example, if the eye piece magnifies 10X and the objective lens magnifies 40X, then 10 x 40 gives a total magnification of 400X.
The compound microscope has certain limitations. Although the level of magnification is almost limitless, the resolution (or
resolving power) is not. Resolution is the ability to discriminate two objects close together as being separate. The human eye
can resolve objects about 100 m apart. The compound microscope has a resolution of 0.2 m under ideal conditions. Objects
closer than 0.2 m are seen as a single fused image.
Resolving power is determined by the amount and physical properties of the visible light that enters the microscope. In general,
the greater the amount of light delivered to the objective lens, the greater the resolution. The size of the objective lens aperture
(opening) decreases with increasing magnification, allowing less light to enter the objective lens. Thus, you will probably find it
necessary to increase the light intensity at the higher magnifications.

DEPTH PERCEPTION AND THE MICROSCOPE

Any microscopic object viewed has depth as well as length and width. While the lens of your eye fully adjusts to focus on an
object being viewed and provides you with a three dimensional interpretation, the lenses of a microscope are focused
mechanically and can only see in two dimensions (length and width). For example, if the specimen you are examining has
three layers of cells, you will only be able to focus on one cell layer at a time. In order to perceive the relative depth of your
specimen use the fine adjustment to focus through the different planes (the three cell layers) individually to build a three
dimensional picture or interpretation of your specimen.
THE FIELD OF VIEW: A CIRCLE
When you view an object under the microscope you will observe that it lies inside a circular field of view. Each different
magnification has a different sized field of view. If you determine the diameter of the field of view you can estimate the size of
an object seen in that field. As you increase the magnification, the field of view
(and diameter) gets proportionately smaller. As a consequence, a critter that appears small under scanning power may appear
large under high power. The actual size of the critter did not change, only the space in which you placed it for viewing.

Low-power field of view

diameter is 1.8 mm

Ruler (mm)

2 mm

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CARE OF THE MICROSCOPE
Your microscope is an expensive instrument that must be given proper care.
Always follow these general instructions when using a microscope.
1. Always carry the microscope with both hands, one hand under the base, and the other on the arm. When getting ready to put the
microscope away, always return it to the low-power or scanning-power setting.
2. When setting the microscope on a table, always keep it away from the edge.
It is generally best to clear your lab table of items that are not being used.
3. The lenses of the microscope cost almost as much as all of the other parts together. Never clean them with anything other than
lens paper.
4. Please report any microscope damage or irregularity in its operation at the beginning of the period so that the repair costs will
not be charged to you.
You are responsible for the microscope while you are using it.

SETTING UP & VIEWING OBJECTS THROUGH THE MICROSCOPE

1. Place the low-power objective in position if it is not so fixed. In changing from one objective to another, you will hear a click when the
objective is set in proper position.
Make certain that the lenses are clean.

2. Check preliminary lighting

a. Turn on the substage light.
b. Position the condenser as high as it will go by turning the substage adjustment.
This provides for a maximum of light.
c. Open the iris diaphragm by means of the lever beneath the condenser, which is below the stage of the microscope.

3. Place the slide for viewing

a. Make certain that the lower-power objective is in place.
b. Lower the stage (away from the objective) with the coarse adjustment.
c. Place a properly prepared slide (see below) on the stage and secure with the stage clips or mechanical
stage depending upon which is present on your microscope.
d. Place the slide with the object directly above the brightly illuminated substage condenser.
4. Focus: Proper focusing technique
a. Viewing the stage from the side, use the coarse adjustment knob to raise the stage until
the stop is reached that will prevent further movement of the stage.
b. Looking through the eye piece, or ocular, lower the stage slowly by turning the coarse adjustment knob counter-clockwise until the
object is in focus.
c. Use the fine adjustment to bring the object into sharp focus.
d. If necessary readjust the amount and intensity of light for comfortable viewing.

5. Increasing Magnification: Switching from Low to High Power

a. First, be sure your slide is focused under low-power.
b. Leave the slide where it is and switch to high-power. Watch from the side to make sure that
the objective lens does not touch the slide. The object should be in focus.

NOTE: Most microscopes are parfocal; that is, the slide should be in focus (or nearly so) when you switch from low-power to high-power. If
you have properly focused your microscope at low-power and your slide is properly prepared (i.e. the object is flattened
by a coverslip), you should be able to switch automatically from low-power to high without fear of having the high-power objective lens
scraping or touching the slide.
An important factor in using the microscope is the distance between the object viewed and the objective lens of the microscope. This is called
the working distance. As you increase magnification the working distance becomes less.
6. Re-Focus under High Power
a. ONLY USE THE FINE ADJUSTMENT.
When the high-power objective is being used, never use the coarse adjustment.

7. Remove the slide

a. Switch the objective to either scanning or low-power.
b. Lower the stage using the coarse adjustment.
c. Remove the slide from the stage.
Note: Never remove a slide while under high-power.

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Answer to guide questions.
1. The objective lens should be returned back to its original position in order to avoid scratches when the objective lens and the
aperture hit
each other.
2. The image seen on an HPO is more clear since the object is magnified several times..
3. The magnification depends on the eyepiece lens times the objective lens magnification which is 40X.
4. "Parfocal" refers to objectives that can be changed with minimal or no refocusing.
5. You would reduce your light when using the 10x objective because it will reduce eye discomfort and will increase contrast.
6. To magnify (enlarge) the image, light rays have to be spread further apart - therefore, the image becomes dimmer,
or light intensity decreases. Lower magnification will have a higher light intensity.
7. parfocal
8. 100X objective lens
9. In order to maintain the quality of the image being viewed in the microscope coz dust will deteriorate its quality.
10. In order to avoid the objective lens touching the glass slide that could destroy your specimen and might scrath the aperture
of the microscope thus lowering the quality image of the specimen.

Answer to guide questions.

1. The objective lens should be returned back to its original position in order to avoid scratches when the objective lens and the
aperture hit
each other.
2. The image seen on an HPO is more clear since the object is magnified several times..
3. The magnification depends on the eyepiece lens times the objective lens magnification which is 40X.
4. "Parfocal" refers to objectives that can be changed with minimal or no refocusing.
5. You would reduce your light when using the 10x objective because it will reduce eye discomfort and will increase contrast.
6. To magnify (enlarge) the image, light rays have to be spread further apart - therefore, the image becomes dimmer,
or light intensity decreases. Lower magnification will have a higher light intensity.
7. parfocal
8. 100X objective lens
9. In order to maintain the quality of the image being viewed in the microscope coz dust will deteriorate its quality.
10. In order to avoid the objective lens touching the glass slide that could destroy your specimen and might scrath the aperture
of the microscope thus lowering the quality image of the specimen.

Answer to guide questions.

1. The objective lens should be returned back to its original position in order to avoid scratches when the objective lens and the
aperture hit
each other.
2. The image seen on an HPO is more clear since the object is magnified several times..
3. The magnification depends on the eyepiece lens times the objective lens magnification which is 40X.
4. "Parfocal" refers to objectives that can be changed with minimal or no refocusing.
5. You would reduce your light when using the 10x objective because it will reduce eye discomfort and will increase contrast.
6. To magnify (enlarge) the image, light rays have to be spread further apart - therefore, the image becomes dimmer,
or light intensity decreases. Lower magnification will have a higher light intensity.
7. parfocal
8. 100X objective lens
9. In order to maintain the quality of the image being viewed in the microscope coz dust will deteriorate its quality.
10. In order to avoid the objective lens touching the glass slide that could destroy your specimen and might scrath the aperture
of the microscope thus lowering the quality image of the specimen.