Dangerous Ideas and Forbidden Knowledge, Spring 2008
Lab 1
Microscopy, Cells,
and Micropipettes
OBJECTIVES:
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Learn how to use and care for the compound microscope.
Identify and explain the function of the parts of the compound microscopes.
Prepare wet mount slides.
Understand of the following terms: prokaryote, eukaryote, cell, cell membrane, cell wall,
cytoplasm, membrane-bounded organelle, nucleus, chloroplast
Compare and contrast prokaryotic and eukaryotic cells
Compare and contrast plant and animal cells
Observe some interesting organisms!
Learn how to use micropipettes
We humans are highly visual creatures, obtaining much of our information about the
world around us by using our eyes. To understand many things we need to visualize them: we
must see them. Not surprisingly, then, some of the most useful tools in biology, and indeed in
science in general, are those that allow us to visualize objects, processes, and phenomena. This is
one of the main advantages of the powerful computers of today, because their power allows us to
visualize things, and thus explore them, as never before possible. But even that technology
cannot replace one of the most fundamental tools of biology, the microscope. With the
microscope we can see things that are otherwise too small for us to see, and thus study them.
Understanding the nature of cell structure and function is important to an understanding
of organisms. All organisms are composed of cells, whether they exist as single cells, colonies of
cells, or in multicellular form. Cells are usually very small, and for this reason, a thorough
understanding of subcellular structure and function has been possible only through advances in
electron microscopy and molecular biology.
In today’s lab you will explore with your lab partners the different basic kinds of cells
that exist and learn how to correctly use a light microscope. You will be comparing specimens of
cells in terms of size, color, and other characteristics. We will then discuss these differences.
GENERAL GUIDELINES
1.
Work in pairs. However, each student in the pair should have his/her own microscope and
do the procedures.
• If you do not have experience with a compound microscope, pair up with someone
who does.
• If you have experience with a compound microscope, pair up with someone who
does not.
• Please pay careful attention to how you use and care for these instruments. Failure to
properly handle the microscopes may lead to a deduction of points from your lab
score. Microscopes are delicate and expensive optical instruments and must be
handled properly and carefully. Prepared slides are also expensive and must be used
thoughtfully as well.
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Dangerous Ideas and Forbidden Knowledge, Spring 2008
2.
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8.
Do not keep your coats, bags, and other personal belongings (items not used in the lab) on
the lab table. You will need room on the lab bench for your lab materials, lab manual,
notebook, and microscope.
Obtain microscopes from the microscope cabinets. Always carry the microscope in an
upright position; the lenses you look through slide into the body tube and can easily fall out
(slides left on the microscope by inconsiderate students can also fall to the floor if the
microscope is tilted). With one hand, grasp the arm of the microscope and place the other
underneath to support the base. Please be patient while you are waiting for students ahead
of you to get their microscopes and please do not get in the way of individuals carrying
microscopes.
Unwrap the cord and plug the microscope into an outlet. Save the band used to hold the
cord! Arrange the cord so that it does not hang over the edge of the bench or can get
snagged by someone, sending it crashing to the floor.
Calculating magnification:
• Multiply the power (magnification) of the eyepiece by the power of the objective lens
to obtain the total (total) magnification.
• The power of the objective lenses is written on them. It usually is followed by an
“x”, and is an integer, NOT a decimal number.
• The power of the eye piece is 10x.
• Always include in any sketches the total power or magnification under which you
observed the specimen. Always include the “x” as part of the magnification (e.g.,
10x, 400x).
Use lens paper ONLY to clean ocular and objective lenses!
When finished, be sure that
• the scope is turned off.
• all slides are removed and returned to their proper container in the correct orientation.
• the stage is wiped clean.
• the scope is set with the lowest power (4X) objective in place.
• the stage is lowered as much as possible.
• the cord is properly secured.
• the scope is returned to its proper place in the microscope cabinet.
If your microscope does not work, tell your instructor, or the lab technician, or put a note
on it indicating what you think is wrong with it. If you find a slide on your microscope
stage please give it to your instructor or place it in the lost slide box.
PROCEDURES- PART I: LETTER E SLIDES
1. Obtain a “letter e” slide. Place this prepared slide on your stage, using the specimen
holder to keep hold it in place. Turn your microscope on, adjusting the light intensity if
necessary. Check to make sure that your 4X objective lens (the shortest one) is locked in
place above your slide. Before you look through the ocular lens, check to see that your
“e” is in the light path. Use the stage controls (X and Y axis knobs) to adjust the position
of your slide if necessary.
2. Now look through your ocular lens and adjust the coarse focus knob until your image is
nearly in focus. Note that you may want to also use the fine focus knob to really sharpen
up the image. You are now looking at your “e” at a total of 40 X magnification. (You
ocular lens has magnified the image 10 times; the objective lens has magnified it 4
times.)
3. Once you have the image in focus, center it in the middle of your field of view, and turn
the revolving nosepiece until your next shortest objective, the 10X lens, is locked in
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Dangerous Ideas and Forbidden Knowledge, Spring 2008
place. You may need to again adjust the fine focus knob to sharpen the image. Note that
you should NOT manipulate the coarse focus once you have left the 4X objective lens.
4. Repeat step 3, moving your nosepiece until the 40X objective is locked in place, again
adjusting only the fine focus knob. Our microscopes have an additional lens, a 100X
objective that we will not be using today. If you’d like to try this lens, please ask an
instructor or TA for assistance.
PROCEDURES- PART 2: ANIMAL CELLS
Now that you’re comfortable on the microscopes, it’s time to move to something more
interesting: our own cells! We’ll be examining our cheek cells. These are the same cells we’ll
be using as our DNA source next week.
NOTE: Because you will be isolating cells from your cheek, and the cheek cells are considered
biohazardous material, please follow the clean-up instructions carefully!
Step One: Prepare a “wet mount” slide with of your cheek cells.
1.
Obtain a clean glass slide
2.
Place one small drop each of water &
methylene blue in the center of a clean glass
slide.
3.
Gently rub the inside of your cheek with the
blunt end of a toothpick.
4.
Stir the cheek material (you may not be able
to see any) into the methylene blue in the
center of the slide.
5.
Apply cover slip. (Touch one edge of the
cover slip to the edge of the drop of liquid.
Hold the cover slip while the liquid runs
along the entire edge. Then gently lower the
cover slip over the specimen.)
6.
Dispose of toothpick in the beaker provided.
Step Two: Examine your cheek cells.
These cells are flat with irregular shapes or contours.
Some of the thin, flat edges of the cells may be folded in your preparation. Many of the cells will
probably be clumped together. It is difficult to observe the cells clumped in large masses or
groups. Bacteria may be present on some cells; these cells will appear to be covered by small,
dot-like or stick-like structures.
1. Place the cheek cell slide on the microscope and examine it under the 4X
objective, as you did with the “letter e” slide. Focus the cells and adjust the light
intensity as necessary.
2. Observe these cells using the low power objective. Move the slide so that an
individual cell is in the center of the field of view. Then move the 10X objective
into place and bring the cell into focus using the fine-focus knob. You may wish
to also examine these cells using the 40X objective. Note that your worksheet
asks you to record your observations of these cells.
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Dangerous Ideas and Forbidden Knowledge, Spring 2008
PROCEDURES- PART 3: PLANT CELLS
Our plant model today will be a common aquatic plant, Elodea. To observe these plant
cells you will need to prepare a slide:
1. Obtain a clean slide and place a drop of water on it. At the front of the room, you’ll find a
small dish containing Elodea. Carefully tear a small piece of a leaf off, and place this
torn leaf on in the drop of water on your clean slide. Gently place a coverslip on the slide
as you did with the cheek cells.
2. Follow the procedures you used with the cheek cell and letter e slide to observe these plant
cells, starting with your 4X objective and moving up one lens at a time. Note that these
leaves are several cells thick; it will be easiest to see individual cells at the torn edge of
the leaves. Your worksheet again asks you to sketch and observe these cells.
PROCEDURES- PART 4: BACTERIA CELLS
Your instructor will set-up slides of bacteria cells on a microscope at the front of the
room. These bacteria are dead and have been stained with a colored dye to make them more
visible. To really see these cells, we need to examine them with the maximum magnification
possible on a light microscope (1000X). To do this, a special technique, oil immersion, is
necessary. Observe these cells (but please do NOT adjust the objective lenses) and answer the
questions on your worksheet.
PROCEDURES- PART 5: POND WATER
If time permits, prepare an additional slide by placing a drop of pond water on a slide and
applying a coverslip as above. Take some time to observe the variety of organisms present in this
microscopic world. Select one organism to draw and describe as indicated on your worksheet.
PROCEDURES- PART 6: USING THE MICROPIPETTES (TIME PERMITTING)
Some of our work this quarter will involve accurately measuring very small quantities of
fluids. To handle microliter volumes, we will use special instruments called micropipettes.
Recall that 1 milliliter is 1/1000 of a liter. One microliter is 1/1000 of a milliliter. Your
instructor will show you how to use the micropipettes, and give you an opportunity to practice
using some colored dyes. Keep the following in mind whenever you use the micropipettes.
1. Place a new, disposable pipette tip on the end of your micropipette each time you use it.
2. Always check that you have the correct micropipette for the job. We use 3 types of
pipettes. The pipette labeled “2-20” can accurately pipette between 2 and 20 microliters.
The pipette labeled “20-200” can pipette between 20 and 200 microliters. And your
largest pipette, labeled “200-1000” can accurately pipette between 200 and 1000
microliters.
3. Check to be sure that your pipette is set for the correct volume. These pipettes are
adjustable; you can adjust the desired volume by turning the black dial on the upper part
of your micropipette.
4. When your micropipette is correctly adjusted, use your thumb to depress the plunger to the
first stopping point. Holding your thumb down, place the tip below the surface of the
fluid you wish to pipette. Slowly release the pressure from your thumb, allowing the
pipette to draw fluid up into the pipette tip.
5. To transfer this fluid, place your pipette tip in a tube and slowly depress the plunger to
expel the fluid.
With your group members, practice pipetting various volumes (2 microliters, 10 microliters, and
20 microliters) of dye onto the plastic dishes provided.
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Dangerous Ideas and Forbidden Knowledge, Spring 2008
Lab 1 Worksheet
Name: ___________________
Microscopy, Cells, and Micropipettes
1. If you are observing a sample using your 10X objective lens, the total magnification of the
image you observe is ________X.
2. Draw a single, isolated cheek cell (at high power) in the
space at right. As you’ll do with any biological drawing,
include the name of the organism, magnification, color, a
description of the shape, and a description of any
structrues that you can see inside the cells.
Specimen: ____________________
Total Magnification: ____________
Description: ___________________
______________________________
3. Draw a portion of the Elodea leaf you observed (at high
power) in the space at right. As you did above, be sure to
identify your specimen, note your total magnification, and
describe and label any structures you can identify within
the cells.
Specimen: ____________________
Total Magnification: ____________
Description: ___________________
______________________________
4. Compare and contrast the structure of the animal and plant cells. What do you notice that they
have in common? In what ways are they different?
5. How large are the bacteria cells compared to the other cells you have observed today? Keep in
mind that you have observed these cells at 1000X, and the others at 100X or 400X. Do they
appear to have anything in common with plant and animal cells? How are they different?
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Dangerous Ideas and Forbidden Knowledge, Spring 2008
6. In the space at right, draw one organism you found in the
pond water sample, including labels and descriptions, as
always.
Specimen: ____________________
Total Magnification: ____________
Description: ___________________
______________________________
Do you think the organism you selected is most likely a plant, animal, bacteria, or
something else? Briefly explain your reasoning.
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