PRACTICAL 1- MICROSCOPY PROTOCOLS

1. THE COMPOUND MICROSCOPE

PURPOSE
To learn the structure and basic use of the compound microscope
MATERIALS
 Compound microscope
 Slides and cover slips
 Newsprint
 Scissors
NOTE: The compound microscope is a delicate instrument and can easily be damaged.
Always carry it carefully with both hands, one under the base and the other holding the
arm.
OVERVIEW
The microscope is a biologist's basic tool. It has been developed to help explore the world
of living things too small to be seen with the naked eye. Early microscopes had only one
lens and were difficult to use. The biggest problem was magnification. The more power-
ful the lens needed for greater magnification, meant the closer the viewer's eye had to be
to the lens. At very high magnification, the lens almost touched the eye. The early micro -
scope user had to be very steady. A major advance in microscopes came with the inven-
tion of the compound microscope. It has two sets of lenses, which magnify objects much
greater than a single lens.
PART A: STRUCTURE OF THE MICROSCOPE
The compound microscope has four basic parts: the lens system, the focusing system, the
stage, and the lighting system. The Lens System One of the two sets of lenses is the ob -
jective lenses. They work similarly to the lens of the early, simple microscope. The ob-
jective lenses make the initial or primary magnification. They are located in the nosepiece
of the microscope. Inscribed on each objective is the magnification or power of that lens.
This tells the number of times the lens magnifies the image. For example, if you are look -
ing at a strand of hair with a 4X (four-power) lens, the hair will appear four times its ac-
tual size. Your microscope probably has at least two objective lenses. Some microscopes
 have as many as four objectives. Rotate the lenses in the nosepiece until they click into
position. The objective lens in use is always the one directly under the body tube.
Usual powers for objective lenses are:
I. 4X; The scanning lens
II. 10X; The low power lens
III. 40X; The high power lens
IV. 100X; The oil immersion lens. NOTE: This lens should not be used without special in-
structions from your teacher.
The second kind of lens in the microscope is the ocular – sometimes called the eyepiece.
This lens is located at the top of the body tube. The ocular serves as a small telescope,
magnifying the image made by the objective lens. This enlargement is called the second-
ary magnification. The magnification of the ocular may be 5X, 10X, 15X, or 20X. The
most common power used in microscopes is the 10X ocular. Examine the ocular of your
microscope. Do not remove it from the body tube. If the power is not stamped on the top
portion of the ocular, you should assume that it is 10X. The total magnification of the mi-
croscope is determined by multiplying the primary magnification (from the objective) by
the secondary magnification (from the ocular). For example, if the objective lens is 10X
and the ocular is 5X, the total magnification is: 10X x 5X = 50X.

PART B: USING THE MICROSCOPE

Put the low power objective in place. Look through the ocular and adjust the light so that
you see a uniformly bright field of view. The field of view, also called the field, is the
area you see through the lens.
If you see specks of dirt in the field, clean your lenses with lens tissue. Now prepare a
slide to view under the microscope. Cut a lowercase "e" from a newspaper and place it in
the centre of a clean slide.
Put a drop of water on top of the letter. Next, place the edge of a cover-slip against the
water, and with a pencil gently lower the cover-slip over the "e."
Placing the cover slip in this manner prevents bubbles from forming. Be sure that the bot-
tom of the slide is dry. This type of slide is called a wet mount. Place the slide under the
stage clips, so that the "e" is right side up. You are now ready to focus on the "e."
Caution: If the lens is dirty or you get water on it, gently wipe it with lens tissue. Never
use facial tissues. Lenses are made of soft glass and scratch easily.
Caution: Be sure that the objective does not touch the slide-both the lens and slide can be
damaged. Do not look through the eyepiece while lowering the objective toward the
stage. It is difficult to judge through the eyepiece how far the objective is moving.
Caution: Never use the coarse adjustment knob in high power. The objective is very close
to the slide in high power, and coarse adjustment could cause the objective to hit the
slide.
Focusing always begins with the lower power (10X) objective. First, click the low power
objective into position in the nosepiece. Then, looking at the side of the microscope, turn
the coarse adjustment knob until the objective is as close as possible to the slide without
touching it.
Now look through the ocular and turn the coarse adjustment knob in the direction that
will move the objective away from the stage. The "e" will come into approximate focus.
To sharpen the focus, turn the fine adjustment knob back and forth. Are you surprised
that the borders of the letter "e" are far from perfect? Note the position of the letter
through the microscope. The letter on the slide is right side up.
Now, look at the "e" under high power. First, under low power, centre the "e" in the field
of view. Switch to high power by turning the nosepiece until the high power objective
clicks into place. Sharpen the focus by turning the fine adjustment knob. If you cannot
find the "e" under high power, try this. Look through the ocular and move the slide
slightly. If this does not bring the "e" into view, move the slide in other directions.
When you are finished using the microscope, remove the slide from the stage. Rinse the
slide and cover slip with water. Dry the slide with a paper towel and not lens tissue.
Glass cover slips should be air dried to prevent breakage. Return both to their proper
places. Finally, be sure the microscope is on low power and put it away. Remember to
carry the microscope carefully, with one hand under the base and the other holding the
arm.