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Name: ____________________
Using a Compound Light Microscope
Background Information: The microscope is an important scientific tool. It enables a person to observe
things too small to be seen with the unaided eye. Anything that you want to look at with a microscope can be
called your specimen, and the nature of your specimen may dictate what sort of microscope you need to see it
with. If you are looking at the outside of your specimen, you want a scope that scans the surface - a scanning
microscope images light (or other radiation) reflected off the surface of a specimen. To look inside, you need to
get light (or other radiation) to go through your specimen - it needs to be thin, semi-transparent, or both. A
microscope that sees images as light or electrons passes through a specimen is a transmission microscope.
Light microscopes, which you'll be using, use light to produce their images. Scanning light microscopes are
called dissecting microscopes, and there are many kinds of light-transmission 'scopes, named for the way the
light is delivered. Our lab has mostly bright field compound microscopes - specimens are seen against a bright
background, and several (compound) magnifications can be chosen by rotating the objective lenses. A
compound microscope has two lenses and a light source that illuminates the object being observed. In this type
of microscope, light passes through the specimen, or object being viewed. One lens, the objective lens, causes
the light rays coming from the specimen to spread apart, forming an enlarged image of the object. The second
lens, the ocular, focuses and further enlarges the image from the objective lens. .The magnification of the
objectives - commonly 4X, 10X, 40X, and sometimes 100X(we don’t have this one) - are magnified again by
the eyepiece or ocular lens - usually 10X - so the total magnification for the different objectives are 40X
(4x10), 100X (10x10), 400X (40x10), and 1000X (100x10).
When working with a compound microscope, you may use specimens that have been prepared in one of two
ways. A prepared slide is made to be permanent and can be purchased from a supply warehouse. A wet mount
slide is made for temporary use and can be made and used during a lab period. To improve contrast, the
microscopist uses stains or dyes that bind to cellular structures and absorb light.
Safety: Put on a laboratory apron. Always handle the microscope with extreme care. You are responsible for
its proper care and use. Use caution when handling microscope slides, as they can break easily and cut you.
Never use direct sunlight as a light source for a compound light microscope with a mirror. The sunlight
reflecting through the microscope could damage your eye. We will use a microscope with an illuminating bulb.
Observe proper laboratory procedures when using electrical equipment.
Materials (per group):
Compound light microscope
Cover slips
Erlenmeyer flask (water)
Toothpicks
blank microscope slides
lens paper
Prepared slides
Cotton Swabs
Elodea (aquatic plant)
Pipette
Methylene blue stain
Procedure:
Part A: Care of the compound light microscope
1. There is a proper way to carry the microscope. Always carry the microscope with both hands. Grasp
the arm of the microscope with one hand and place your other hand under the base. Always hold the
microscope in an upright position so that the ocular cannot fall out. Place a microscope on your
workbench at least 10 cm from the edge. Position the microscope with the arm facing you.
2. Rules to remember:
*The lowest power objective should be in position both at the beginning and the end of
microscope use. In other words, always store the microscope with the scanning lens in place.
*Use only lens paper to clean dirty or smudged lenses. Do not use your shirt or a paper towel.
*Keep the stage clean and dry to prevent rust and corrosion.
*Do not remove parts of the microscope or force parts past the point the move easily.
*Keep the microscope dust-free by covering it after use.
*Report any malfunctions to the teacher.
Part B: Identification of Parts and their uses
Identify the following parts on your microscope. Use the diagram to help you with this.
1. Eyepiece (ocular lens): Top most series of lenses through which an object is viewed. What is the
magnifying power of the ocular lens on your microscope? (Look for the number on the side followed by
the “x”.
2. Body tube: holds nosepiece at one end and eyepiece at the other end; conducts light rays.
3. Arm: Supports upper parts and provides carrying handle.
4. Revolving nosepiece: Turns to change from scanning, to low, to higher power objective lenses.
5. Objectives (objective lenses):
1. Scanning power objective (shortest lens): Holds scanning lens used to view the whole slide.
What is its magnifying power?
2. Low-power objective (medium in length): Holds low power lens used to view the object in
greater detail. What is the magnifying power of this lens on your microscope?
3. High-power objective (longest in length): Holds high power lens used to view the object in even
greater detail. What is the magnifying power of this lens on your microscope?
4. Oil immersion objective: Holds oil immersion lens and is used in conjunction with immersion oil
to view objects with the greatest magnification. **Our microscopes do not have these lenses. It
is rarely used in a Biology I class.
6. Coarse-adjustment knob: Knob used to bring object into approximate focus; used only with lowpower objective.
7. Fine-adjustment knob: Knob used to bring object into final focus.
8. Diaphragm or Iris diaphragm control lever: Controls amount of illumination used to view the object.
9. Light source: An attached lamp that directs a beam of light up through the object.
10. Base: The flat surface of the microscope that rests on the table.
11. Stage: Holds and supports microscope slides.
12. Mechanical stage: A moveable stage that aids in the accurate positioning of the slide. Does your
microscope have a mechanical stage?
13. Mechanical stage clip control knobs: Two knobs that are usually located on the stage. One knob
controls forward/reverse movement, and the other controls right/left movement.
Part C: Use of the Microscope
Total magnification: Total magnification is calculated by multiplying the magnification of the ocular lens
(eyepiece) by the magnification of the objective lens that is in place.
*Calculate the total magnification if the microscope is on the scanning power objective.
*Calculate the total magnification if the microscope is on the lower power objective.
*Calculate the total magnification if the microscope is on the higher power objective.
Focusing the microscope:
You should now begin to learn an important skill that will significantly increase your comfort. You must learn
to view through the eyepiece with both eyes open! Whether you have a monocular microscope (one eyepiece,)
or a binocular microscope (two eyepieces,) start from the beginning to use both eyes while viewing. Try this
experiment. Close one eye and squint (as you usually would while looking through the microscope) as long as
you can. I can assure you it will not be very long. It is tiring and uncomfortable, to say the least. And how long
do you think you will enjoy looking through your microscope with this posture? You get the picture. Now, try
this. Open both eyes, but place your hand over one eye. Isn’t that more comfortable? You could probably do
this indefinitely. If you were spending hours each day looking through a microscope, which would you prefer?
Every professional microscopist has mastered this skill. Just train yourself to always keep both eyes open. It
may seem difficult, but your eye will automatically shut out the image from the eye not used for viewing
through the monocular microscope, and with the binocular microscope, both eyes will focus on the image. Like
riding a bicycle, once you learn it, you never forget.
Always start and end with the Scanning Power Objective lens and the stage all the way down! Place a prepared
slide on the microscope stage, with the specimen directly over the center of the glass circle on the stage (directly
over the light). You will see a dark line that goes part way across the field of view, try turning the eyepiece.
That dark line is a pointer that will be very valuable when you want to point out something to your lab partner,
or your teacher! Adjust the Diaphragm as you look through the Eyepiece, and you will see that MORE detail
is visible when you allow in LESS light! Too much light will give the specimen a washed-out appearance.
TRY IT OUT!!
Slowly move the coarse adjustment knob. This will move the stage further away from the lens. Adjust it back
and forth until you get the clearest image possible. If it is still a little fuzzy, use the fine adjustment knob to
clear it up.
If you would like to view the specimen with a higher power magnification:
Make sure the image is in the center of your field of view. While watching from the side, turn the nosepiece
and change the scanning lens to the lower power lens. Make sure that the lens will not hit the stage, as this can
damage the lens. The microscope is “parfocal” which means you should be able to switch powers and be in
focus with very little adjustment. DO NOT TOUCH THE COARSE ADJUSTMENT KNOB ONCE YOU
HAVE CHANGED FROM SCANNING TO A HIGHER POWER LENS. Turning the coarse adjustment
knob could cause the stage to hit the objective lens. This can scratch the lens or crack your slide. You can focus
by turning the Fine adjustment knob. Repeat these steps to switch to the high power objective.
Tips on Making Good Drawings:
1. Don’t even think of starting your drawing unless you have a PENCIL! Drawings in PEN are
UNACCEPTABLE! This is for two reasons:
(a) You can erase pencil!
(b) You can shade in areas more easily in pencil.
2. Each drawing must include clear, proper labels! In the upper left hand corner of each circle include the
specimen name as written on the slide label. In the upper right hand corner, include the magnification (100x or
400x).
Elodea
100x
3. Labels should start on the outside of the circle. The circle indicates the field of view as seen through the
eyepiece. All arrows should end with the point touching the object to be labeled!
4. Animal cells should always include at least the following labels: Cell membrane, Nucleus, Cytoplasm.
5. Plant Cells should always include at least the following labels: Cell membrane (if distinguishable), Cell wall,
Nucleus, Cytoplasm, and Chloroplast.
6. I don’t want you to Look at the cells; I want you to SEE them! In order to do that, you MUST: Apply your
knowledge of cell structure to your drawings! An unlabeled drawing is nothing more than
scratches on a piece of paper!
(notice the angle of the coverslip in the diagram)
I. Animal Cells: Prepare a temporary wet mount of your cheek cells. To do this you will obtain a clean, blank
glass slide and cover slip from the materials table. You will use a toothpick or cotton swab to gently rub the
inside of your cheek to get some cells. Smear the cells onto the middle of the glass slide. Place a drop of
methylene blue stain. Lower the cover slip at an angle (45 degree) to avoid trapping air and causing air bubbles
in your slide. If you have too much stain, call me over and I will show you a trick for pulling some out from the
cover slip. Focus to get the best view. Sketch a few cheek cells on high power. Label the cell membrane,
nucleus, and cytoplasm.
II. Plant Cells: Obtain a clean, blank slide and cover slip form the materials table. Use the forceps to remove a
leaf from the growing tip of an Elodea pond plant and prepare a wet mount slide. Add a drop of water to the
slide; place the leaf in the water droplet. Cover the specimen with the small cover slip. Lower the cover slip at
an angle (45 degree) to avoid trapping air and causing air bubbles in your slide. Focus to get the best image.
Sketch a few plant cells. Label the cell membrane/cell wall, the nucleus, the cytoplasm, and the chloroplasts.
III. Protozoa and other cool stuff: You may be able to see small micro-organisms swimming around in the
pond water. Make a wet mount slide of some of the water from the plant bag. Search for living organisms that
you might find swimming around. Use a depression slide for this sample. Please note that you do not need to
add a drop of water since the specimen is liquid. Sketch/describe what you see.
IV. Other: You may now make and view temporary mount slides or view more prepared slides of your
choosing. Please make notations of what you see. I encourage you to take pictures of your specimens with
your cell phones. If you get some good ones, send them to me.
Name: __________________________________
Date: ____________________
Questions:
1. Describe the image of an object seen through the high-power objective as compared to that seen through the
low-power and scanning lenses.
2. Why should you never use the coarse adjustment when focusing the high-power objective lens?
3. Everything you see through the microscope is “inverted” (upside down and backwards). If you viewed an
organism moving toward the bottom of the slide and then it moved to the right what was it actually movement?
4. How did the size and shape of the cheek (animal) cells compare with the Elodea (plant) cells?
5. State 2 procedures which should be used to properly handle and care for a light microscope.
6. Explain why the specimen must be centered in the field of view on scanning power before moving on to the
next power.
7. If a microscope has an ocular that is 20X and a high-power lens that is 43X what would be the total
magnification on high power?
8. Briefly describe how to make a wet mount slide of a specimen.
9. What happens to the amount of light when you change from a lower to a higher power?
10. How could you adjust for this?
Name that part:
*The part of the microscope that changes from one objective lens to another. _____________________
*Controls the amount of light passed through the system. ____________________________________
*Also known as the eyepiece because it is where you view the specimen. ________________________
*The knob that clears up an image on any power. ___________________________________________
*Carries the image from the objective to the ocular lens. _______________________________________
Drawings and notations about specimens viewed in lab activity today. Don’t forget the proper way to label.