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Lab 1 - Laboratory Techniques-1

Lab 1: Laboratory Techniques
Biology is a hands‐on science. While the type of activity you might perform as a biologist will vary
depending on the field you enter, there are some basic techniques that all beginning lab students should
understand. By the end of this lab, you should be able to:
Keep yourself safe in the lab.
Identify the parts of the microscope and describe their functions.
Calibrate a micrometer and make measurements with a microscope.
Estimate cell concentration using a haemocytometer.
Choose the right pipette for the job at hand and be proficient in its use.
Properly balance and operate a centrifuge.
There are four exercises in this handout, each with a pre‐lab or in‐lab portion (or both). At the end of the
handout are worksheets on which you should write your responses to the questions.
Pre-lab exercises: Before attending the lab, go through the entire handout and complete all the pre‐
lab exercises and questions (look for the arrows ). Fill out corresponding pre‐lab questions on the
Worksheets below and bring your answers to the lab.
In-lab exercises: These exercises will be set up as a series of stations where you will perform a specific
task to familiarize yourself with the more common instruments or techniques used in a biology lab. Each
station is a stand‐alone task, so you can start at any in‐lab exercise and complete them in any order. All of
the in‐lab exercises and questions (Worksheets, pages 14 to 16) will be completed during your lab session.
Exercise 1: Lab Safety
Research biologists often spend a lot of time working in the laboratory. As well as the usual workplace
hazards, common biology lab hazards include being exposed to hazardous chemicals or biohazardous
materials, working around an open flame, being attacked by an iguana (or other organism of study), etc.
The undergraduate laboratory has been designed to keep these hazards to a minimum, but you still have to
take the following precautions:
NO eating or drinking in the lab
Lab coat and goggles (or safety glasses) must be worn when using noxious chemicals.
NO open‐toe shoes are allowed in the lab.
Broken glass must be disposed of in a broken glass trash receptacle (not the normal trash).
Know the locations of first aid kit and fire blanket (under cabinet nearest to Ms. Mouradian’s office)
and fire extinguisher (on large post in centre of the laboratory). An eyewash station is by the sink.
Be aware of what other students are doing around your workspace to help prevent collisions and
© Dora Cavallo-Medved, Department of Biological Sciences
keep your work area clean and leave it clean for the next group. This includes desktops, benchtops, test
tubes and other glassware. Leave microscopes clean, with the lowest objective in place.
in the event of fire or other major safety threat, there are fire alarms located at the doors to each
stairwell. Proceed calmly to the stairwell located to the left of the lab as you exit the door. Make your
way down the stairs and step well away from the building when you exit, leaving the walkway/driveway
clear for emergency personnel.
 Pre-Lab Exercise 1: Lab Safety
As mentioned on the first day of class, before you can perform any lab exercises for this course you MUST
complete the “Online UWindsor Undergraduate Laboratory Safety Awareness Training”. You can access
this training course online at http://cleo.uwindsor.ca/ccc/labsafety . To login you will need to enter your
UWindsor username (UWinID) and password (the one you use for your University of Windsor email). Then
follow the instructions to complete the training. Once you have completed the training you will be required
to take an online quiz and you must score 90% or greater to pass (you can take the quiz multiple times).
Upon passing you will receive a Completion Code indicating that you are now certified to perform labs in
this course. Please provide your completion code to your TA/GA when you come to your first lab. Failure to
do so will mean that you cannot complete any labs for this course. If you have any questions regarding the
online training please refer to the Laboratory Safety Bulletin that was provided to you on the first day of
Exercise 2a: Parts of a Compound Light Microscope
The microscope has probably contributed more FIGURE 1: Parts of a compound light microscope.
to the development of biology as a science than
any other instrument. Microscopes of one kind
a. light source
or another are used by scientists in every field of
biology from genetics to ecology. Start to master b. condenser lens with
iris diaphragm
these skills today so you can build and expand
c. horizontal stage
on them throughout your biological studies. In
this course, you will use these skills in the d. objective lens
upcoming “Fractionation” lab and in the “Mitosis
f. ocular lens
and Meiosis” lab.
body tube
The “Light” Part of the Microscope
h. arm
Light travels from the light source in the i. coarse focusing knob
microscope’s base through an iris diaphragm
fine focusing knob
into a condenser just below the stage. The
k. base
condenser focuses the rays of light to provide
better image resolution. The intensity of the light
is adjustable using the light source’s rheostat and the amount of light reaching the specimen is adjustable
using the iris diaphragm. The more light that is allowed to enter, the less contrast (or detail) you will be able
to observe; when the iris is fully open, the image is flooded with light, and definition is lost due to “burn
out.” Typically, you’ll want to close the iris diaphragm as far as possible in order to give maximum contrast
while still allowing sufficient light through to see the object.
Lab 1: Lab Techniques
The “Compound” Part of the Microscope
A “compound” microscope is one with two or more lenses. The ocular lens is the one closest to the eye,
and usually magnifies objects ten times (10X) their actual size. The objective lenses are a collection of
lenses located on the rotary nosepiece. There are usually three or four objective lenses, each allowing for
different degrees of magnification (e.g. 4X, 10X, 40X, etc.). The total magnification is a product of both
lenses’ powers. So, for the 4X lens, the total magnification would be:
Total magnification = ocular magnification x objective magnification
Total magnification = 10X x 4X
Total magnification = 40
 Pre-Lab Exercise 2a: Parts of a Compound Light Microscope
Proceed to the worksheets found at the end of this document and answer the Pre‐Lab Exercise 2a
Exercise 2b: Use of a Compound Light Microscope
The following guidelines will help you safely and effectively operate your microscopes in exercises 2c to 2d
so please read this section carefully prior to attending labs.
Basic Precautions: Do You Have $3000? No? Read this.
A microscope appears sturdy, but is as fragile (and expensive) as a nice camera. Use care in handling your
microscope at all times:
When carrying your microscope,
put one hand on the base and one on the arm. Keep it upright and
close to your body.
When placing the microscope on the bench,
position it away from the edge of the table, and take
care to ensure that someone can’t accidentally get entangled in the electric cord.
When cleaning the lenses, only use lens tissue. Never use paper towels, facial tissues, Kimwipes® or
any other type of paper or cloth to clean the lenses – they’ll scratch the glass. Fold the lens tissue at
least twice to prevent skin oils from getting on the lens.
Don’t damage the high powered objective (long lens) by bumping it against a slide or a specimen.
Always have your low power objective (short lens) in place when:
 placing and removing slides
 taking the first look at your specimen
 using the coarse adjustment
Basic Use of a Compound Light Microscope
You will perform these steps for Exercises 2c and 2d.
Get Set Up
1. Clean all the microscope’s lenses (for guidelines, see “Basic Precautions,” above).
2. Plug in the microscope and turn on the light source.
Lab 1: Lab Techniques
3. If it isn’t already in position, rotate the nosepiece until the low or scanning power (i.e. 4X) objective lens
is in line with the body tube. You’ll feel a “click” when the lens is properly positioned.
The first time you use a microscope in this lab
4. If you look through the eyepieces and see two images, the interpupillary distance is not right for you.
Slide the eyepieces closer together or farther apart until the two fields merge to form a single circle of
light. Record the value from the window above the right eyepiece to use the next time you use a
Interpupillary distance (mm): ______________________
5. Focus the eyepieces:
Rotate the diopter adjustment ring on the left eyepiece until the number on the side matches your
interpupillary distance.
Cover your right eye and bring the object into clear, sharp focus using the coarse and fine
adjustment, as needed (see “Focus on the Specimen,” below).
Cover your left eye and bring the same object into focus using the diopter ring of the right eyepiece.
Record the diopter setting to use the next time you use a microscope.
Right diopter value: __________________
Note: Keep these two values ‐‐ interpupillary distance and diopter value – in a safe place
because you will use them again in future labs.
Focus on the Specimen
6. Looking from the side of the microscope, make sure the slide is centred in the light from beneath the
7. Always start with the low power (4X) objective lens. Looking through the eyepieces, make sure the
target object is centred in the field of view (the circle of light you see through the microscope).
8. Use the coarse focusing knob (the big one) to bring the image into focus. Only use the coarse focusing
knob with the low power (4X) objective lens.
9. Fine tune the clarity with the fine focusing knob (the small one).
Tip: Keep both eyes open when examining a specimen – it won’t tire you out as much as if you
were squinting.
Adjust the Lighting
10. Remove the eyepiece and close the iris diaphragm until you see a dark ring that takes up about 1/5th of
the original field of view, and then reinsert the eyepiece.
Focus on the specimen with the iris wide open. Slowly close the iris until the field of view just begins to
Note: You must readjust the lighting each time you switch lenses.
Lab 1: Lab Techniques
Tip: Learn to use the illuminating system on your microscope correctly; most of the problems
you will have focusing will be due to incorrect adjustment of light.
Switch to a Higher Power Objective Lens
11. Looking from the side of the microscope, slowly rotate the 10X power objective into place. Because this
is a longer lens, you need to be sure that the objective does not touch the slide; this could damage the
slide (relatively cheap) or the lens (relatively expensive).
12. Adjust the iris diaphragm (see “Adjust the Lighting,” directly above).
13. Adjust the fine focus. Most microscopes are parfocal and parcentered, meaning that the image should
remain focused and centred when you switch between objective lenses, but you may need to make
very slight adjustments to the fine focus.
Tip: When in doubt, back it out. If the object seems to disappear after switching to a higher
powered lens, switch back to a lower powered objective and try again.
 Pre-Lab Exercise 2b: Use of a Compound Light Microscope
Take a virtual tour of a compound microscope at the following website:
http://www.udel.edu/biology/ketcham/microscope/scope.html. After completing the tour, take a
screenshot with both the “At view checklist” and “Through view checklist” completed and bring the print
out to your lab along with your worksheets. (See http://eduscapes.com/tap/topic7.htmif you don’t know
how to take a screenshot.)
Exercise 2c: Measuring a Microscopic Object
An important part of describing what you see through a microscope is giving details about the size of the
specimen. In order to do this reliably, you will need to be able to calibrate a micrometer and measure
microscopic objects – skills that will be used in the upcoming “Fractionation” lab.
To determine the size of the object you are viewing you use two pieces of equipment:
ocular micrometer: a small glass disk in the eyepiece that has uniformly spaced lines etched into it.
stage micrometer: a glass slide with a small ruler etched into it. In your case, the ruler is divided into
fine increments of 0.01 mm at one end and 0.1 mm units on the remainder (Figure 2).
FIGURE 2: Stage micrometer.
0.01 mm
Lab 1: Lab Techniques
0.1 mm
Example of How to Calibrate an Ocular Micrometer
1. Prior to starting, please set up
FIGURE 3: Calibrating the micrometers for the 4X objective lens.
the microscope following steps 1
2.5 stage micrometer divisions
to 13 of “Exercise 2b: Use of a
Compound Light Microscope,”
and then put the stage
micrometer slide onto the stage.
2. With the 4X objective lens in
place, look through the eyepiece
and align the lines at the left side
of the stage micrometer with the
lines at the left edge of the
ocular micrometer (Figure 3).
Stage micrometer
Ocular micrometer
1 ocular micrometer division
3. Determine how many divisions of the stage micrometer fit into a single division of the ocular
micrometer (4X objective lens). In Figure 3:
1 ocular unit = 2.5 stage units
Since every stage unit is equal to 0.1 mm (Figure 2) then for the 4X objective lens:
1 ocular unit = 2.5 stage units x
0.1 mm
1 stage unit
1 ocular unit = 0.25 mm
Note: You have to recalibrate the ocular micrometer for each objective.
4. You can then use that calibration to determine the size of
any object, as in Figure 4. The cell highlighted by the circle
drawn around it is about 0.75 ocular units, so:
0.75 ocular units x
0.25 mm = 0.1875 mm
1 ocular unit
FIGURE 4: Measuring a cell. Puccinia garminis,
Black rust uredospores, www.inra.fr.
In-Lab Exercise 2c: Measuring a Microscopic Object
1. Prior to starting, please set up the microscope following steps 1 to 13 of “Exercise 2b: Use of a
Compound Light Microscope,” and then open the iris diaphragm to its brightest setting.
2. Calibrate the ocular micrometer for the 4X objective (see “Example of How to Calibrate an Ocular
Micrometer,” above). Repeat for the 10X and 40X objectives. Fill out Table 1 on your worksheet.
3. Switch back once more to the 4X objective, as you’ll be inserting a slide (see “Basic Precautions” in your
Pre‐Lab). Put the prepared slide with the letter “e” on to the stage and adjust the focus and lighting.
4. Measure the letter “e” in ocular units. Repeat for the 10X and 40X objectives. Fill out Table 2 on your
Lab 1: Lab Techniques
Exercise 2d: Estimating Cell Concentration Using a
Knowing the number of cells in a solution is so critical to biology that numerous methods of estimating cell
concentration have been developed. The most widely used type of microscopic counting chamber is a
haemocytometer, so‐called because it was originally designed for performing blood cell counts. A
haemocytometer is simply a thick microscope slide with a chamber where the solution is held.
The counting area has a laser‐etched grid of lines dividing it into nine large squares (double line, Figure 5),
which are easily seen with a 4x objective. If you switch to a 10X objective you can see that the middle large
square is square is subdivided into 25 medium sized squares (single line, Figure 5) that are further
subdivided into 16 even smaller squares. The large square measures 1 mm x 1 mm in area and the chamber
is a constant 0.1 mm in depth, which means that each of the large squares is:
1 mm x 1 mm x 0.1 mm = 0.1 mm3 = 0.0001 ml in volume (v).
FIGURE 5: Haemocytometer grid.
To use a haemocytometer, you simply count the total number of cells in a
large square (n) and then divide that number by the volume of the large
square (v) to determine the average concentration (c) of the mixture.
c = n/v
c = cell concentration in cells/mL
n = average number of cells counted
v = volume of square (mL)
Therefore, if you counted a total of 75 cells from one large square, your cell concentration would be:
c = 75 cells/0.0001 mL
c = 750,000 cells/mL
Note: To be more accurate, one would normally take the average number of cells from at least four large
In-Lab Exercise 2d: Estimating Cell Concentration Using a Haemocytometer
1. Prior to starting, please set up the microscope following steps 1 to 13 of “Exercise 2b: Use of a
Compound Light Microscope.” Using 70% ethanol, clean the haemocytometer and coverslip and then
blot dry with a Kimwipe®.
2. Place the coverslip on the haemocytometer, letting it rest on the raised supports.
3. Gently mix the solution to ensure the yeast cells are suspended evenly in the solution.
4. Draw up a small amount of solution with a Pasteur pipette (like an eyedropper) and place the pipette
tip at the notch at the edge of the slide. Do not squeeze the bulb; just allow the chamber to fill through
capillary action.
Lab 1: Lab Techniques
5. Ensuring the 4X objective lens is in place, put the prepared haemocytometer onto the stage and adjust
the focus and lighting.
6. Increase the strength of the objective lens to 10X or 40 X. Because the slide is unusually thick, take extra
care that the lens doesn’t touch the cover slip.
FIGURE 6: Counting cells on the
7. Count the cells within the boundaries of one large square (double line,
Figure 5). What about the cells on the boundaries? In order to avoid
double‐counting, normal practice is to include cells overlapping the top
and left lines (Figure 6, grey), but not those overlapping the bottom or
right lines (Figure 6, white).
boundary of the haemocytometer grid.
8. Repeat once more and fill out Table 3 on your worksheet.
Exercise 3: Using a Pipette
Pipettes are glass or plastic tubes used to transfer liquids of volumes between FIGURE 7: Common types of pipettes.
1 and 100 mL from one container to another. Improper pipetting technique is
a major source of laboratory error. You’ll be using pipettes to make up
solutions in the upcoming “Organic Molecules” and “Fractionation” labs, so
this is a good chance to perfect your technique.
There are many different types of pipettes, each with their own, specialized
use. Three of the more common types (Figure 7) are:
serological pipette: A full 10 mL liquid is expelled when all the liquid is
 Mohr pipette: Gradation marks stop at a baseline before the tip of the
pipette. To transfer 10 mL, the pipette is only drained to this baseline. If
you expel the whole volume you would deliver “10 mL and a bit.”
 Pasteur pipette: Like an eyedropper, it transfers uncalibrated volumes up
to 2.5 mL. Also called a “transfer pipette.”
Almost everything you need to know in order to use the pipette correctly is
printed on the neck of the pipette (see Figure 8):
FIGURE 8: Pipette specifications.
maximum volume of liquid that can be
transferred (also coded by colour in the
Size of
coloured rings).
size of the divisions on the pipette.
10 ml in 1/10
blowout rings are often included to indicate
TD 20°C 37034
whether all the liquid in the pipette should be
expelled (e.g. serological pipette).
The specifications for the pipette in Figure 8 indicate that all liquid in this pipette must be expelled, and that
the pipette is calibrated in 1/10 mL divisions and will deliver up to 10 mL.
Lab 1: Lab Techniques
As well as different kinds of pipettes, there are different kinds of pipetters, which are used to “suck up” and
discharge liquids from a pipette. You will be using a Pipette Pump. This type of pipette has colour coded
barrels indicating the pipette sizes it is designed to handle:
Blue for volumes 2 mL and smaller (≤ 2 mL)
Green for volumes larger than 2 mL (>2 mL)
Note: Be sure to use the appropriate pipette and the appropriate Pipette Pump to deliver the
volume desired.
In-Lab Exercise 3: Using a Pipette
1. Use a pipette to transfer 3.5 mL of the orange/red solution to a test tube. Make sure you choose the
correct pipette and Pipette Pump to handle the 3.5 mL volume.
2. Assemble the pipette and Pipette Pump together. Place the tapered end of the pipette into the liquid.
Ensure that the tip stays in the liquid the entire time you are filling the pipette.
Tip: If you are using a serological pipette, ensure that you use the wheel to raise the white top of
the pump approximately 1 cm before you put the tip in the liquid to ensure you will be able to
expel all the liquid from the tip of the pipette.
Hold the Pipette Pump in one hand and place your thumb on the wheel of the Pipette Pump. Use
your thumb to slowly rotate the wheel downward, causing the liquid to rise into the pipette.
Note: Check how your
pipette is numbered. Is FIGURE 9: Pipette gradations. Zero at the top (top figure) or the bottom (bottom figure).
zero at the top or bottom?
If zero is at the top, then to
get 3 mL of liquid, for
3 mL
instance, the meniscus
should be at the 7 mL mark
(Figure 9, top); if zero is at
the bottom, draw the
liquid up to the 3 mL mark
(Figure 9, bottom).
Take the tip of the pipette out of the liquid and move the pipette to the test tube.
Use your thumb to rotate the wheel upward, expelling the liquid.
Tip: What kind of pipette are you using? If you are using a serological pipette, make sure you
expel all the liquid. If you are using a Mohr pipette expel the liquid only until the bottom of the
meniscus touches the baseline.
Remove the pipette from the Pipette Pump.
3. Use another pipette to transfer 0.3 mL of the blue solution to the same test tube and mix gently.
4. In a second fresh test tube, combine 2.5 mL of the yellow solution and 0.2 mL of the red solution and
mix gently.
Lab 1: Lab Techniques
5. Bring both test tubes to your lab instructor for checking.
6. Proceed to worksheets found at the end of this document and answer the in In‐Lab Exercise 3
Exercise 4: Using a Centrifuge
Centrifugation is a process used to separate materials that are mixed together in a solution. A centrifuge is
widely used in biochemistry, cellular and molecular biology and medical labs to precipitate cells and viruses,
separate subcellular organelles, and isolate macromolecules such as DNA, RNA, proteins, or lipids. Since the
heaviest materials in any mixture will fall to the bottom and the lightest rise to the top, scientists use a
centrifuge to increase the effective gravitational force on the material and speed up the process. You use a
similar application when you do your laundry and use the spin cycle to force water out of your clothes so
they will dry faster. You will be using a centrifuge in the upcoming “Fractionation” lab to separate organic
In-Lab Exercise 4: Using a Centrifuge
1. Swirl the flask of yeast first to make a homogenous solution. Then fill a test tube with 3 mls of yeast
suspension. Describe the mixture before centrifugation in Table 5 on your worksheet.
2. Make a counterbalance for your sample, matching the masses (not volumes) as closely as possible:
Weigh the test tube you prepared with your sample.
Put a new test tube on the scale and use a Pasteur pipette (like an eyedropper) to fill the test tube
with water until your blank weighs the same as your sample did.
Note: If the centrifuge spins with unbalanced tubes, it may permanently damage the
3. Put the balanced tubes into the centrifuge directly opposite from each other.
4. Close the lid, lock the lid and set the time to spin your samples to 2 minutes. The machine will
automatically start
5. Keep the lid closed until the centrifuge has completely stopped spinning.
6. Remove your tubes, being careful not to jar them so the suspensions don’t remix. Describe the mixture
after centrifugation in Table 5 on your worksheet.
7. It is important to be able to separate the two components without causing the solid (pellet on the
bottom/side of the tube) to go back into solution (supernatant). You will need to master this skill for
future labs, so use this as an opportunity to practice. Using a Pasteur pipette (like an eyedropper), try to
carefully remove all of the supernatant from the pellet without disturbing the pellet. Deposit the
supernatant in a waste container.
8. Add 3 mls of water to your pellet and mix to get the pellet back into solution. Now you can discard the
solution down the sink.
Lab 1: Lab Techniques
NAME: _________________________________
STUDENT #: __________________________
DATE: _______________
MARK: ______________
TA/GA Name: ______________
Pre-Lab Exercise 1: Lab Safety
Complete the Online UWindsor Undergraduate Laboratory Safety Awareness Training” at
http://cleo.uwindsor.ca/ccc/labsafety and bring your “Completion Code” to lab so that it can be
recorded by your TA/GA.
Pre-Lab Exercise 2a: Parts of a Compound Light Microscope
Using the theory above and the online tutorial in section 2b, match the definitions on the left with the
words on the right (not all the words on the right have a matching definition).
a. The platform that holds the slide up beneath the objective
b. Controls the contrast in order to achieve a high image
c. The lens in a microscope closest to the specimen.
d. The part of the microscope that you look through. Usually has
a 10X magnification level.
e. Moves the microscope stage up and down to bring the slide
sharply into view. Use only with the low power objective lens
in place.
f. Focuses the rays from the microscope's built‐in illumination to
provide better image resolution.
1. parfocal
2. condenser
3. objective lens
4. fine focusing knob
5. stage
6. ocular lens
7. iris diaphragm
8. quadruple nosepiece
9. coarse focusing knob
Pre-Lab Exercise 2b: Use of a Compound Light Microscope
Print a screenshot of the http://www.udel.edu/biology/ketcham/microscope/scope.html website, with
both the “At view” and “Through view” checklists completed. Bring a printout of this screenshot to your
lab. (See http://eduscapes.com/tap/topic7.htm if you don’t know how to take a screenshot.)
Lab 1: Lab Techniques
In-Lab Exercise 2c: Measuring a Microscopic Object
TABLE 1: Calibration
TABLE 2: Measurements of the letter "e"
1. While viewing the letter “e”, how is it oriented as seen through the eyepiece of the microscope?
2. How does the image move when the slide is moved to the right or left?
3. What happens to the brightness of the view when you go from low to high power?
4. Why is it difficult to locate an object starting with a high power objective?
5. Why is it necessary to calibrate the ocular micrometer for each objective?
6. How does the area of the field of view change with increasing power of the objectives?
7. If a cell has a diameter of 5 μm at 50X, what is its diameter at 400X? __________________________
Lab 1: Lab Techniques
In-Lab Exercise 2d: Estimating Cell Concentration Using a
TABLE 3: Estimation of cell concentration.
In-Lab Exercise 3: Using a Pipette
1. List the type of pipette and Pipette Pump you would use to transfer these solutions?
PIPETTE (1, 2, 5 OR 10 ML)
0.35 ML NAOH
5.6 ML KOH
8.2 ML KCL
3.5 ML HCL
2. At the pipette station, there are several stands with pipettes clamped to them. For each of these, write
down the volume contained in the pipette.
a. ________________
b. _______________
c. __________________
In-Lab Exercise 4: Using a Centrifuge
1. Enter your description of the mixture before and after centrifugation in Table 5 and hypothesize as to
the composition of the different layers.
TABLE 5: Description of samples before and after centrifugation.
2. What can happen if the tubes in a centrifuge are of different masses or are not placed directly opposite
each other?
Lab 1: Lab Techniques
Abramoff, P. and R.G. Thomson. 1991. Laboratory Outlines in Biology‐V. WH Freeman and Company, New
York, NY, USA.
Campbell, N. and J. Reece. 2005. Biology, 7th edition. Pearson Publishing, San Francisco, CA, USA.
Helms, D.R., C.W. Helms, R.J. Kosinski, and J.R. Cummings. 1998. Biology in the Laboratory, 3rd edition. WH
Freeman and Company, New York, NY, USA.
Mooney, T.A., P.E. Nachtigall and S. Vlachos. 2009. Sonar‐induced temporary hearing loss in dolphins,
Biology Letters in press. doi: 10.1098/rsbl.2009.0099
Morgan, J.G. and M.E. Brown Carter. 2008. Investigating Biology: Laboratory manual, 6th edition. Pearson
Education Inc., San Francisco, CA, USA.
Vodopich, D. and R. Moore. 2005. Biology Laboratory Manual, 7th edition. McGraw Hill Higher Education,
Boston, MA, USA.
Lab 1: Lab Techniques