Pierce College
Putman/Biol 241
Name: ________________________________
Lab 03 SGOs: The Cell: Transport Mechanisms (10 points)
Pierce College Student Outcome:
Lab Outcome 2: Perform a variety of cell physiology experiments; predict results of tonicity tests that utilize
erythrocytes and varying concentrations of saline solutions; describe the differences between simple diffusion,
facilitated diffusion, osmosis and filtration.
By completing and understanding this lab, you should be able to do the following:
1. Observe the diffusion of a liquid through a colloid and explain what is happening. Give the
relationship between molecular weight and the rate of diffusion.
2. Observe the diffusion of dye through water and explain the comparative rate of diffusion of
dye through agar and through water.
3. Explain the results of an osmometer demonstration.
4. Observe and explain the results of placing a red blood cell in an isotonic, hypotonic and
hypertonic solution.
5. State whether newsprint can be read through test tubes containing 0%, 0.9% and 5% saline
solution plus blood, and why, based on the results of SGO 4 above!
Exercise 3.1: Diffusion of a Liquid Through a Colloid.
Introduction and Methods: In this experiment, you will determine which has a higher rate of
diffusion through the solid gel, agar: KMnO4 (potassium permanganate) or methylene blue. For
reference, KMnO4 has a mass of 158.04 g/mole and methylene blue has a mass of 319.86 g/mole.
Which do you think? Record your hypothesis in the lab report (end of lab). Make the hypothesis
simple and do not include a reason for your answer. A hypothesis such as, “Methylene blue will
have a higher rate of diffusion than KMnO4,” would be a good hypothesis. “I think that (or) I
believe that methylene blue will have a higher rate of diffusion than KMnO4 because it’s a
protein stain and sticks to agar,” would be a bad hypothesis; the “I think that” and “I believe
that” are irrelevant, and the statement after “because” would require an additional experiment.
Enter your hypothesis before moving on with the experiment! You will not be penalized for an
incorrect hypothesis!
A little on the scientific method. In science, we construct hypotheses to be the best
possible explanations for natural phenomena. We then construct experiments that always must
have, as one possible outcome, the falsification of the hypothesis. We run the experiment. If the
results do not support the hypothesis, then we say the hypothesis has been falsified or disproved
and we either modify it or throw it out completely. If the results of an experiment support the
experiment, we say the results “support” the hypothesis, or the results “suggest” the hypothesis is
the most likely explanation; we don’t use the word “prove” because the next experiment must
have, as one outcome, the possible falsification of the hypothesis, even if the falsification of the
hypothesis is highly unlikely. To write that a hypothesis has been “proven” is contrary to the
scientific method as this implies that further experiments cannot falsify the hypothesis; you must
always have, as one possible outcome, the experimental falsification of the hypothesis.
Putman/Pierce College Biol 241 Lab 03/20150728/Page 1 of 9
So, to test your hypothesis, work in
groups of two. From the supply table obtain
M
K
a petri dish containing agar and a soda
straw. Make two wells in the agar. To do
this, about 1/3 the way across the Petri dish,
plunge the straw straight down into the agar,
twist, and bring it straight out. Repeat
another third the way across so that you
have two wells as in the figure to the right.
From the supply table, obtain a grease pencil or permanent marker and mark on one the
plastic on one side of the dish an “M” for methylene blue and on the other side “K” for KMnO4,
so you can remember which is which. You will also need a 15 cm ruler.
From the supply table, obtain a dropper bottle of methylene blue and one of KMnO4.
Careful: Don’t get KMnO4 in your mouth or eyes as it is toxic! First, note how the drops come
out of the dropper bottles; you are going to fill the “M” well with methylene blue and the “K”
well with KMnO4 and you don’t want any spillage. Carefully fill each well with the
corresponding reagent; note the time from a clock—this is your time zero. Enter the time in the
lab report.
Determine what time it will be when 60 minutes will have elapsed and enter this time in
your lab report. While you are waiting for the reagents to diffuse through the agar, move on the
the other exercises in this lab.
At the end of 60 minutes, measure the distance each reagent has diffused from the edge
of the well, in millimeters, and record these values in your lab report. Then calculate the rate as
distance, in mm, divided by time, in minutes, and enter this value for each in the table in the lab
report.
Answer the questions in the lab report.
Exercise 3.2: Diffusion of Dye Through Water.
Introduction and Methods: Kinetic energy is heat, the energy of motion. Diffusion is the random
movement of particles from high concentration to low concentration, from an ordered state to a
disordered state, as powered by kinetic energy. Diffusion moves materials from high to low
concentrations through the aqueous media of the body, inside cells, in extracellular fluids, in the
lymph, cerebrospinal fluid, plasma, etc.
At the demo table will be a large Petri plate containing water. It will be positioned on a
couple of plastic rulers so that the 0 mm mark will be in the center of the plate. Be careful not to
bump or breathe on the setup!
When the class is ready, the instructor will ask for a time keeper. A few crystals of
KMnO4 will be placed at time 0 on the 0 mm mark. At 5 minutes, the time keeper will say “time”
and another student will read the distance the reagent diffused through the water. Record this
value in your lab report, calculate the rate of diffusion of KMnO4 through the water by dividing
the distance traveled by 5 minutes, and answer the questions.
Putman/Pierce College Biol 241 Lab 03/20150728/Page 2 of 9
Exercise 3.3: Diffusion Through a Semipermeable Membrane
Introduction and Methods: Osmosis is the diffusion of water
through a semipermeable membrane. An osmometer is a
device used to measure pressure resulting from osmosis.
At the demo table, there will be an osmometer already
setup. The osmometer will consist of a thistle funnel attached
to dialysis tubing (dt) containing saturated sucrose (ss),
immersed in distilled water (dw). Dialysis tubing is a
semipermeable membrane that allows water through but not
sucrose. Your instructor will have recorded time zero at the
zero mark on the osmometer. During lab, observe the
osmometer, note the height of the column, in mm, and the
time, in minutes it took the column of fluid to reach that
height; record these values in your lab report and calculate the
rate at which the column of fluid is moving. Then answer the
questions.
dw
ss
dt
Exercise 3.4: Microscopic Observation of Cells in Solutions of Different Tonicity.
Introduction and Methods: For this lab, we will be using blood. Take appropriate precautions:
Wear gloves. When finished, any materials that touched blood should go in the biohazard
containers. Your instructor will give you further instructions regarding this.
Obtain a microscope slide, three toothpicks, three coverslips and a permanent marker or
grease pencil. Mark a dot in the upper right-hand corner of the microscope slide.
At the supply table will be aqueous (water) solutions of 0%, 5% and 0.9% saline. Follow
these directions very carefully:
Place a single drop of 0% saline on
the microscope slide as indicated to the
right, followed by a droplet of blood from a
toothpick dipped 2 to 3 mm in a blood tube.
Gently mix the blood into the aqueous
0%
5%
0.9%
solution, then dispose of the toothpick in
biohazard. Add a coverslip on the specimen.
Place a single drop of 5% saline on the microscope slide as above, mix, dispose of the
toothpick in the biohazard and add a coverslip.
Last, place a single drop of 0.9% saline on the microscope slide as above, mix, dispose of
the toothpick in the biohazard and add a coverslip. Note: 0.9% saline is normal saline and is
isoosmotic to blood plasma.
Once you have made your slide, first observe the erythrocytes at 0.9% treatment. Focus
on lowest power first, then at 100x, then finally at 400x. Describe the shape and condition of the
cells in the lab report. Using your mechanical stage, move your slide over and examine the
erythrocytes in the 5% treatment. Describe the shape and condition of the cells in the lab report.
Finally, without refocusing, using your mechanical stage, move the slide over to examine the
erythrocytes in the 0% treatment. Describe the shape and condition of the cells in the lab report.
Putman/Pierce College Biol 241 Lab 03/20150728/Page 3 of 9
Again, if the erythrocytes were focused at the 5% treatment, you will not need to refocus at the
0% treatment.
Answer the questions provided.
Exercise 3.5: Macroscopic Observation of Cells in Solutions of Different tonicity.
Introduction and Methods: At the demo table, you will find three test tubes: Test tube 0% will
contain two drops of blood mixed in 2 mL of 0% saline solution (distilled water), test tube 0.9%
will contain two drops of blood mixed in 2 mL of 0.9% saline solution and test tube 5% will
contain two drops of blood mixed in 2 mL of 5% saline. Also at the table will be a scrap of
newsprint. You will hold the newsprint behind each of the test tubes and observe if you can read
the newsprint through the blood solution.
Before you go to the demo table, construct a hypothesis stating which solution you will
be able to read newsprint through and which you will not be able to read newsprint through. A
good hypothesis might be: "I will be able to read newsprint through the 5% saline-blood solution
but not through the 0.9% and 0% saline-blood solutions." Important: Do not include any possible
reasons explaining your projected results! A bad hypothesis would be: "I will be able to read
through the 5% saline-blood solution because the blood cells settle to the bottom, but I will not
be able to read through the 0.9% and 0% saline-blood solutions because the blood cells coagulate
and remain in solution." You leave explanations for the discussion portion of your lab!
Please make sure the lab is in order before you leave!
Putman/Pierce College Biol 241 Lab 03/20150728/Page 4 of 9
Biol 241
Lab 3 Report: Microscopy
(10 points)
Name: ___________________________________
Date: ________________ Lab Section: ________
Exercise 3.1: Diffusion of a Liquid Through a Colloid.
Hypothesis: ___________________________________________________________________
Results:
Reagent:
Time
Time Elapsed (min)
Distance Traveled (mm)
0 min
0 mm
Rate: Distance
(mm)/Time (min)
60 min
Reagent:
Time
Time Elapsed (min)
Distance Traveled (mm)
0 min
0 mm
Rate: Distance
(mm)/Time (min)
60 min
Discussion:
1. Which had a higher rate of diffusion through the agar, methylene blue or KMnO4? Why?
2. Did your results disprove your hypothesis or support your hypothesis?
Putman/Pierce College Biol 241 Lab 03/20150728/Page 5 of 9
3. Even if your results supported your hypothesis, why can’t we write that the results of an
experiment “proved” the hypothesis?
Exercise 3.2: Diffusion of Dye Through Water.
Results:
Distance KMnO4 traveled
through water (mm) in 5 min
Rate of diffusion of KMnO4
through water: mm/min
Discussion:
1. What was the rate of diffusion of KMnO4 through water?
2. What would increase this rate?
3. Give an example of how or where diffusion might move a substance in a biological
system?
Putman/Pierce College Biol 241 Lab 03/20150728/Page 6 of 9
Exercise 3.3: Diffusion Through a Semipermeable Membrane.
Results:
Time (min)
Height of Column
(mm)
Osmotic rate,
mm/min
Discussion:
1. Why did the column of water rise? Explain fully!
2. What was the energy source powering the rise in the column of water?
Exercise 3.4: Microscopic Observation of Cells in Solutions of Different Tonicity.
Results:
Treatment:
0.9%
5%
0%
Shape/condition of
erythrocytes:
Discussion:
1. In which treatment (if any) did water move out of the cells? What was the evidence for
this?
Putman/Pierce College Biol 241 Lab 03/20150728/Page 7 of 9
2. In the treatment you mentioned in #1, why did water move out of the cells?
3. In which treatment (if any) did water move into the cells? What was the evidence for
this?
4. In the treatment you mentioned in #3, why did water move into the cells?
Exercise 3.5: Macroscopic Observation of Cells in Solutions of Different Tonicity.
Results
Write your hypothesis here before beginning this part of the lab: __________________________
______________________________________________________________________________
______________________________________________________________________________
Results: Follow the methods above and record your results here:
2 drops blood + % saline
Can read newsprint through
solution: Yes or No
0%
0.9%
5%
Putman/Pierce College Biol 241 Lab 03/20150728/Page 8 of 9
Discussion: Based on your previous microscopic observations of what happened to red blood
cells when they were placed in 0%, 0.9% and 5% saline solutions, answer the following
questions:
1. Through which percent saline solution(s) could you read newsprint? Why?
2. Through which percent saline solution(s) could you not read newsprint? Why? How did
osmosis affect these blood cells?
3. Did your results support or disprove your hypothesis?
4. If your results supported your hypothesis, why can't we write that your results "proved" your
hypothesis?
Putman/Pierce College Biol 241 Lab 03/20150728/Page 9 of 9
Putman/Pierce College Biol 241 Lab 03/20150728/Page 10 of 9