Procedure
Equipment and Supplies
(Do this in groups of 3 or 4. This list is for one group.)
1. A well populated culture of Paramecium. You need 40
ml per group.
2. A series of small test tubes. Label them 0, 1, 5, 10,
15, 30, 45, and 60.
3. Formalin. Put 1 ml in each small test tube.
4. A tube of artist's water color. Get this at an art
supply store. It comes in a container that resembles a small tooth
paste tube. Get lampblack or a red pigment that is not based
on cesium or chromium or other heavy metal.
5. A compound microscope and a supply of microscope slides and cover
slips.
6. Glass or plastic pipettes with bulbs. Be sure that the
pipettes used on the live cells never come in contact with formalin.
Even a small droplet of formalin will kill the cells and thus stop
feeding.
7. A clock or stopwatch for timing.
Procedure
1. Mix a little water color with spring water or autoclaved tap
water to produce a few ml of a distinctly red suspension. You will
add the water color to the 40 ml culture in just a moment. But
first, remove 5 ml and put it in the tube labelled "0" that
contains formalin. Mix it up well. Now immediately
add
0.5 ml of that suspension to your Paramecium culture.
This is time 0. You just took the Time 0 sample.
2. At the designated times remove another 5 ml from your
culture of Paramecium containing water color and pipet it into the
appropriately labeled tube (1, 5, 10, 15, 30, 45, and 60 minutes).
Mix each well. The formalin will kill the cells, thus stopping food
vacuole formation. It will also preserve the Paramecia for a little
while so you don't have to do the counts (steps 3 & 4 below) on the
same day as you do the transferring.
3. Let the tubes stand undisturbed for at least 30 minutes after you
added the cell suspension to the formalin. During this time
the cells, now dead, will settle to the bottom of the tube. Anytime
from 30 minutes to 24 hours after Step 2, remove the dead cells form a
tube. They will be piled up at the bottom of the tube.
Put a Pasteur pipette with a bulb so that the tip is right at the
bottom. (Squeeze the bulb BEFORE you stick the pipet in the
tube.) Now release the bulb and the paramecia will be sucked into
the tube.
4. Put the material you sucked up on a microscope slide and add a
cover slip. Now look under low power at the dead paramecia on the
slide. The red water color will be in the medium surrounding the
cells, but look more closely and you will see that some of the cells
contain food vacuoles in which there is red water color. This is a
"labelled vacuole." Count the number of labeled vacuoles
in at least 10 Paramecia. Arrange your data this way for each time
you sample.
Cells – Time 0
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Number labeled vacuoles
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1
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2
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3
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4
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5
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6
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7
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8
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9
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10
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Background
Paramecium eat bacteria which they ingest by
phagocytosis. A group of bacteria is swept into a membrane-bound
pocket at the end of the cytostome, and then the pocket pinches off, forming
a food vacuole.
Then the food vacuoles circulate through the cytoplasm where lysosomes empty
digestive enzymes into the food vacuole. When all the digestible
material has been transported out of the vacuole and into the cytoplasm, the
remaining material is dumped out of the cell by exocytosis at a special
place on the cell membrane called the cytoproct or cytostome.
Study of the process of food vacuole formation and loss can be carried out
very easily using artists' water colors to label the vacuoles.
This can be done with students at many levels of biological and mathematical
ability from simply asking what the maximum number of food vacuoles is, or
graphing the number of food vacuoles as a function of time, to more
sophisticated kinetic studies including the development of equations to
describe the process. Refer to the paper by Berger and Pollock to learn how to do the mathematically more sophisticated experiments.
Once the basic experiment has been done, lots of inquiry is possible of the
sort that asks what the effect of some treatment is on food vacuole
formation or loss. For example, what is the effect of mild osmotic
shock on the shape of the graph?
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