TRAINING LAB-MAKING A SERIAL DILUTION/USING A
COLORIMETER
Name________________________________________
Background- Working in the field of science you will learn to use many special techniques and scientific tools. In
this lab you will be exposed to a few of the common techniques and tools used in laboratories throughout the world.
1. You will be trained to complete a SERIAL DILUTION of a liquid
2. You will be trained to measure liquids using a PIPETTE.
3. You will be trained to use a scientific instrument called a COLORIMETER.
Procedures:
Part 1- How to Measure Liquids with a Pipette and a Graduated Cylinder.
1. A GRADUATED CYLINDER and a PIPETTE
Pipette
are both used for measuring liquids,
1. You want to put 4 mL of liquid A in a
however, they measure in slightly opposite
test tube.
ways. You should be familiar with how to
2. Suck up liquid A into the Pipette until
use both of these important science tools.
you reach the 0 mL mark near the top. BE
Graduated Cylinder
EXACT! (Your supervisor will show you
1. You want to put 4 mL of liquid A in a
how to get the liquid into the pipette.)
test tube.
3. Place the tip of the filled Pipette over
2. Pour liquid A into the Graduated
your test tube.
Cylinder until you reach the 4 mL mark.
4. Slowly release liquid A out of the
BE EXACT!
Pipette into your test tube and STOP when
3. Pour the 4 mL of liquid A from the
liquid A reaches the 4 mL mark on the
Graduated Cylinder into your test tube.
Pipette. BE EXACT!
5. DONE – 4 mL in your test tube.
4. DONE – 4 mL in your test tube.
5. There should be no liquid left in the
5. There will still be some liquid
Graduated Cylinder.
remaining in your Pipette (just empty it
back into the liquid A container.)
Part 2- How to Preparing a Serial Dilution
1. You will need a beaker of water, a set of six test tubes, a test tube rack, 5mL pipette, a pipette bulb, and
small labels for your test tubes.
2. Your goal is to take the blue liquid (a 100% food color solution) and DILUTE it with water to make it less
concentrated, BUT, you will need to know the blue liquid’s exact concentration when you are finishing
adding water - a difficult task!. A SERIAL DILUTION, however, is a quick and easy lab technique that
allows you to know the exact concentration of the liquid as you dilute it with water. After making a serial
dilution you end up with several solutions each one diluted with more and more water and each one exactly
ONE-HALF the concentration of the solution before it (if the original solution is 100%, the next solution you
make will be 50% concentration, the next 25%)
3. Get 6 test tubes. Label them 1 through 6 using very small pieces of tape near the top of the test tubes.
4. Place 4mL of the 100% blue food coloring solution into tube 1.This is your 100% solution. You are finished
with tube 1.
5. Next, place 4mL of the 100% blue food coloring into tube 2.
6. Add 4mL of water to tube 2 (tube 2 should now contain 8mL of liquid). Place your thumb over the top of the
tube 2 and turn it upside down a couple of times to mix.
7. Measure out exactly 4mL of the solution from tube 2 and put it into tube 3. Tube 2 now has 4mL of a lighter
blue solution left in it- ½ of 100% or a 50% solution (50% food color, 50% water).
8. Add 4 mL of water to tube 3 and mix as before.
9. Measure out 4 mL of the solution from tube 3 and put it in to tube 4. Tube 3 now has 4 mL of yet a lighter
blue solution left in it – ½ of 50 % or a 25% (25% food color, 75% water)
10. You should just keep going in the same way, making more and more dilute solutions. When you are
finished making # 6 it will still have 8 mL of solution. Simply measure out 4 mL of the solution and pour it
down the sink.
11. Look closely at the colors of your six tubes. It’s easy to see how a serial dilution works- each tube contains
exactly ONE-HALF the concentration of the tube before it. You have made a 100% solution, 50% solution,
25% solution, 12.5% solution, 6.25% solution, and 3.12% solution- A SERIAL DILUTION!! EASY!!!
12. Rinse out your pipette before putting it away.
Part 3 – Using a Colorimeter
1. Read the rest of these instructions while waiting your turn to use the colorimeter so you will know what else
to do.
2. When it’s your turn to use the colorimeter take your serial dilution tubes with you and be prepared to record a
series of numbers (Absorbance) in table 1 below.
3. A COLORIMETER does nothing but measure the amount of light that is absorbed by a sample (the blue
liquids in this experiment). A light on one side shines through the cuvette (small plastic box) and your
sample, and a sensor on the opposite side detect how much light passed through. The amount of light that
comes through is called % TRANSMISSION. The amount of light that does not come through is called
ABSORBANCE. It makes sense that a very dark liquid will absorb most of the light, so it should have a very
low % transmission and a very high absorbance. A light liquid will absorb very little of the light, allowing
most of it to pass through, so it should have a high % transmission and a low absorbance.
4. So, why are we measuring how much light will be absorbed by our solutions? This information can help you
determine the concentration of unknown solutions that you did not make- but more about this later!
5. Why is a colorimeter so expensive? Because it is so accurate! The colorimeter can even detect that clear glass
absorbs some light and doesn’t let it all pass through.
6. FOLLOW THESE STEPS TO USE THE COLORIMETER.
a. Turn on the calculator, then press the “APPS” button, then scroll down until DataMate is
highlighted, then press “Enter”.
b. Use arrows on the colorimeter to have green light on under 635 nm
c. Find the BLANK in the beaker near the colorimeter. The BLANK is simply a cuvette filled with
plain, clear water. We want to measure how much light the blue food coloring in your samples will
absorb- NOT the plastic in the cuvette or the water we mixed in. you will use the blank and “tell”
the colorimeter to ignore the plastic and water in its measurements. Wipe off the lower part of the
blank’s cuvette with a paper towel (fingerprints can block the light). Raise the lid on the
colorimeter; slowly push the blank all the way in the chamber, then close the lid. The absorbance
(shown in the top right hand corner on the calculator screen) should read “0” with the blank in
place. You need to “zero” the colorimeter by pressing “1”, then pressing “3”, then pressing “1”,
then pressing “Enter”. These steps may need to be repeated 3-4 times You have now “told” the
machine to ignore the glass and water! Remove the blank cuvette and prepare to test your samples.
d. Fill the empty cuvette with your liquid from the tube with the lowest concentration made with the
graduated cylinder, tube G6. Wipe off the cuvette to remove fingerprints. Put the cuvette in the
colorimeter, close the lid and record the absorbance in the data table. Notice that the number of the
tubes in the data table is the opposite of the order you are measuring them! Remove the cuvette
and pour the liquid back into the test tube.
e. Fill the empty cuvette with your liquid from the tube with the next lowest concentration made with
the graduated cylinder, tube G5. Wipe off the cuvette to remove fingerprints. Put the cuvette in the
colorimeter, close the lid and record the absorbance in the data table. Remove the cuvette and pour
the liquid back into the test tube.
f. Repeat for your remaining samples made with the graduated cylinder.
g. Repeat for your samples made with the pipette.
h. When you have finished press “6”, then press “Enter”, the turn the calculator off.
9. When you have finished, rinse out ALL of your tubes and remove the labels. THANKS!
1 (100%)
Tube Number and % Concentration
2 (50%)
3 (25%)
4 (12.5%)
5 (6.25%)
6 (3.12%)
AbsorbanceDilutions made with
pipette
Part 4- Graphing Your Results
1. Watch the video available on Blackboard or Labshares to help you graph the results using graphing
software.
2. % concentration will be on the x-axis and absorbance will be on the y-axis.
3. If you measured your serial dilution perfectly, your points should line-up to make a straight line that
angle upward to the right. However, your points will be more scattered if any of your serial dilutions
measurements were off -- even by just a tiny bit.
4. Use the computer program to draw a “best fit,” straight line through the data points (the line pass
through the middle of all the points so there are an equal number of points above and below the line).
A “best fit” line is sort of an average to show where your points probably should be if there were no
measurement errors.
5. Display the equation for this line.
6. You will use this line and equation to determine the % concentration of the unknown solution.
7. Put this in your teacher’s Labshares Hand-In folder to be printed.
Part 5- How to Determine the % Concentration of the Unknown Solution
*IMPORTANT-READ ME* Your graph is also very important because it can be used as a science tool.
Suppose I gave you a test tube with blue food color in it. I then ask you to tell me EXACT %
concentration of this blue liquid. Is this possible?? YES!
Here is what you would do:
1. Place the test tube with the unknown blue liquid in the colorimeter and determine its absorbance
(remember - absorbance is related to concentration!).
1. 2. Use your graph and locate the absorbance of the unknown blue liquid on the Y-axis. Use a ruler to
draw a light, dotted line from this point over to your best-fit line on the graph.
2. Next, move the ruler and draw a light, dotted line from this point on your best-fit line down to the Xaxis.
3. The place that your light, dotted line meets the X-axis is the % concentration of the unknown blue
liquid, however, you must accurately estimate this number. For example – if the line ends up between
6.25% and 12.5% on the X-axis you must decide if the solution is 7%, 8%, 9%, 10%, or 11%
concentration.
Questions-Serial Dilution\Colorimeter
name__________________________
1. What would the % concentration of blue food coloring be if you had continued your serial dilution one
more time and made a tube # 7?
____________________________________
2. You place 30mL of a 60% sugar solution in beaker #1. You next place 30mL of the 60% sugar solution
in beaker #2. You are now ready to begin making a serial dilution. What will be the % concentration of
the sugar solution in the beaker #2 when you are finished?
_____________________________________
3. In the Training Lab you just completed, you added 4mL of water to 4mL of blue solution each time you
completed a Serial Dilution. However, if you had been using 10mL of blue solution in the test tubes, you
would have needed to add 10mL of water each time for the Serial Dilution to work properly. Likewise, if
you have 100mL of a solution and you wanted to complete a Serial Dilution you would need to add
100mL of water to this solution –always diluting it by one-half. Now for the question – in question #2
how much water should you add to beaker #2 to begin you Serial Dilution?
____________________________________________
4. You have an 80% starch solution in a beaker and you complete a serial dilution of this solution. When
you are finished, you have six beakers (numbered 1-6). Beaker # 1 has some of the original 80% starch
solution in it. What will be the % concentration of starch in beaker #6?
_____________________________________________
5.
You decide to test beaker #6(from question #4 above) to make sure it still contains starch. From a
previous lab, what test will you complete to see if starch is present?
6. Container #1 contains 25mL of liquid. Swimming in this container is 10,000 Giardia. You complete a
series of serial dilutions until you finally end up with a container that holds fewer than 1000 Giardia.
What container # will this be?
________________________________
7. Why is it important to wipe off each cuvette before placing it in the colorimeter?
8. You place a test tube filled with black paint into the colorimeter. What do you think the absorbance
would be? Explain your answer.
9. You forgot to used the blank and adjust the colorimeter before testing the absorbance of your samples.
How will this most likely affect the absorbance readings of your samples?(mark one of the following
answers)
o my sample’s absorbance readings will probably be higher than they should be
o
my sample’s absorbance readings will probably be lower than they should be
o
my sample’s absorbance readings will not be affected in any way
MAKE SURE YOU HAVE READ PART 5 OF THE TRAINING LAB FOR THESE QUESTIONS
10. I give you a test tube filled with an unknown concentration of blue food color solution and ask you to tell
me its concentration. You know exactly what to do! You walk over to the colorimeter and insert the test
tube. The absorbance on the colorimeter reads 0.80. What is the % concentration of this blue food color
solution? HINT- you need to use your graph!
__________________________________________
11. You test another blue solution of unknown concentration and find it has an absorbance of 0.25. What is
the % concentration of this unknown blue food color solution?
__________________________________________
12. One more to make sure you’ve got it! A blue solution’s absorbance is 0.47. What is the % concentration
of this unknown blue food color solution?
__________________________________________