1997 WWLPT Biology Institute:
Life Cycles: Reproduction & Embryological Development
The Effects of "Recreational"
Drugs on the Development of Chick Embryos as a Model for Human Embryogenesis
& John Colvin
(The PDX Starbuck-a-roos)
Target Age or Ability Group Audience
Materials & Equipment
Background [Prior Knowledge
or vocabulary necessary to complete activity]
The Student Lab
Method of Evaluation/Assessment
References Including Web Addresses
This activity has two different approaches to observing chick embryogenesis.
The first part is a modification of Dunn's
embryological research in which a chick is grown "shell-less" or outside
of its egg shell. A growth chamber is constructed using a 4" PVC plastic
pipe, HandiWrap, and a sterile petri dish as a cover for student observation
of chick embryogenesis. Part two is an extended
activity in which students study the effects of various "recreational"
and/or "over-the-counter drugs," e.g., caffeine, aspirin, ethanol, nicotine,
etc., on embryonic development This is done by injecting small concentrations
of these drugs into 72 hour chick eggs.
Many of the students we teachers come in contact with come from diverse
backgrounds and experiences. Students are beginning to develop their own
points of view that may differ from their parents, teachers, and other
adults. This is a time of experimentation in their lives, e.g., alcohol,
smoking, and "recreational drugs." Many believe that they are "indestructible"
and that they have a full life ahead of them. All to often tragedy strikes
and they face the reality that we all are truly mortal. Also, a number
of students are becoming sexually active and are unaware of what consequences
their actions have upon future generations. The goal of these of activities
are to help form, modify, and possibly change student behavior.
In the first part of the activity, we are using the chick embryo as
a model of human development. In the first 24 hours of chick development
several critical systems "come on line." These critical systems include
the nervous, circulatory, gastrointestinal, and urogential. In humans the
same systems develop in the first six to eight weeks. It is these systems,
especially brain, spinal cord, and heart development, that we want students
to focus upon.
In the second part of the activity, we are using the chick embryo to
demonstrate that even "over-the-counter" and/ or "recreational" drugs have
an effect on the development of critical systems. The objective is to reinforce
the concept of taking "care of your baby before it is born."
Some students may strongly object to the use of vertebrate animals for
experimentation. See the end of this activity for further information on
Target Audience or Age Group
General to Advanced Biology students
Grade Level: 10+
Instructions/Special Precautions: to
Required of students
Teachers Guide for the Preparation of Materials
Readings and research on chick and human embryology. Familiarity with early
development of nervous, circulatory, gastrointestinal, and urogential systems.
Formulae for Avian Physiological Salt Solution:
1 1/2 to 2 hours are needed to construct the "shell-less" chambers. Get
materials from a local hardware store. Construct enough chambers for small
groups of two (or four) students.
Make arrangements with a local hatchery for at least one fertilized egg
per group of students; plus a couple of extra per class (accidents happen
and sometimes eggs are infertile). If collecting eggs from several
sources, e.g., student's home, etc., have them store the eggs in refrigerator,
about 18 degrees C, immediately upon collecting from the hens
until you have sufficient numbers. The eggs can remain in the refrigerator
for up to one to two weeks. On the day of the experiment let all the eggs
warm to room temperature at the same time.
Fertile chick eggs can be ordered from Carolina
Equilibrate the incubator(s) to 37.5 - 38.5 degrees C for at least one
day. Make sure a large bowl of water is in the incubator. The
water will prevent the eggs from dehydration which is one of the causes
Rotate the eggs 180 degrees twice a day during incubation. This rotates
the embryo within the shell and prevents adhesion of embryonic membranes
to the inside of the shell. Furthermore, rotating the eggs stimulates
extra-embryonic membranes and promotes fluid redistribution among various
parts of the egg during incubation.
Class time needed
Initial set-up will take two 45 minute or one 90 minute period.
Follow-up sessions of 5 to 10 minutes per day for recording observations
for the duration of the activity.
Hazards/Precautions for Shell-less environment
Chick mortality is high - 50% by 18.5 days, few hatch in 21 days
Egg needs to be rotated - try rocking it at 30 degree angles 2 or 3 times
1% CO2 environment - use Alkaseltzer tablet(s) in water w/in
80 % humidity - put apparatus w/in enclosure w/ water
Ca++ replacement, esp. by day 14 - 18 -- can use shell fragments
w/ intact shell membrane
Infections: use 70% Ethanol to "sterilize" HandiWrap and egg shell, before
cracking open. Sterilize chamber and petri plate in pressure canner or
Chick embryos before 72 hours have a low survival rate (trade-off for early
Suggested Classes of 1% Stock Solutions for Testing Common Drugs:
Caffeine, Nicotine, Alcohol, Aspirin (order from Carolina
[Prior Knowledge or Vocabulary Necessary to Complete Activity]
This activity helps students answer:
The Student Lab to
Observations of Early Embryogenesis in Chicks in a
"Shell-less" Environment (modified from "Chicken Little" - Kristal
Watts, Sam Barlow H.S. Gresham, OR, USA and based upon research by Bruce
Construction of PVC "Shell-less" Chick Embryo
||Sterile petri dish TOP
|Shell-less chamber w/ clamp
||70% Alcohol swab
|6" square of plastic HandiWrap
|Nile Blue Sulfate
||Stain Neutral Red Stain
Setting up the chamber:
1. Place the HandiWrap square into the shell-less chamber. Make a depression
with the center so that the bottom is just off of the table surface.
Placing chick into chamber:
2. Secure the HandiWrap with clamp below the top rim by ½" to
leave room for the petri dish TOP.
3. Swab plastic wrap with 70% alcohol and let air dry completely.
1. Working in small groups get one fertilized chicken egg. Crack the
shell with a sharp blow on the table’s edge. Put the contents of the egg
into the depression by separating the two halves of the shell much as one
would place an egg into a frying pan for "sunny-side-up" eggs. Under ordinary
circumstances, the yolk will orient by gravity, with the embryonic disc
uppermost. The embryo is now ready to be observed.
Note: since cooling will adversely affect development, keep the
embryo out of the incubator for no longer than necessary. If observations
exceeds 2 minutes position a lamp above the chamber. The heat will keep
the embryo warm until you return it to the incubator. Keep the chamber
covered with the petri dish to avoid contamination.
2. Add a drop or two of 0.01% Neutral Red or Nile Blue Sulfate directly
over the blastoderm. Gradually during a period of five to ten minutes,
the embryonic cells will concentrate the dye. In so doing the thicker
parts of the embryo will become darker than the thinner cellular areas.
Hensen's node, the primitive streak, neural folds, brain vesicles, etc.,
will gradually become visible.
3. Observe daily for five to seven days. Write and illustrate a journal
of your observations. Identify and label the following structures: blastoderm,
yolk sac, allantois, chalaza, somites, albumen, Hensen’s node, primitive
streak, neural folds and brain vesicles.
The Effects of "Recreational Drugs" on the Development
of Chick Embryogenesis
This experiment involves the injection of living eggs with various "recreational
drugs" including caffeine, alcohol, nicotine, and aspirin to see their
possible effect on embryonic development. In fact, this is one of
the standard industrial tests for the biological activity and permissible
dosage of pharmaceuticals before they are released for human use.
1. Select four 72-hour eggs from the incubator. Candle each egg before
Serial Dilutions for Injections in Chick Eggs
How to Assemble the
2. Swab each egg with 70% alcohol to limit bacterial contamination.
Allow the egg to remain in a horizontal position for a minute or two to
dry and allow the embryo to rotate to the top.
3. Sterilize the scissors blades by dipping them in 70 % alcohol
and pass it through an open flame.
4. Hold the egg with one hand and hold one point of the scissors
near its tip in the other. With a twisting movement, drill a small
hole through the blunt end of the egg. The hole should be large enough
to admit a hypodermic needle.
5. Place the egg in a egg carton with the hole on top and (put
a drop of 70% alcohol on the hole to keep it sterile while) preparing the
remaining eggs and solutions for injection.
6. Using a felt tip marking pen, label each egg A, B, C, or D. This
indicates what concentration of your assigned drug is in each egg. Egg
"D" is the control.
7. Your group will do a serial dilution of your
assigned drug. Using your syringe, make a dilution of 1 unit of your drug
to 9 units of Ringer's solution into a sterile container—this is 0.1%.
Repeat the dilution by taking the 0.1% solution and make another 1:9 dilution—this
is 0.01%. Repeat the series for solutions "B" and "C."
Source: M.V. Tyrode
||Final Dilution in Eggs
||0.5 ml of .1%
||0.5 ml of .01%
||0.5 ml of .001%
8. Using a sterile syringe draw out 0.5 ml of a solution and inject
into the egg. The needle should just pierce the vitelline membrane.
9. Seal the hole in the shell with a piece of adhesive tape.
10. Incubate the eggs for 5 to 7 days at 37.5 to 38 degrees C--- and
then open to examine the embryo. Evaluate the experimental results
by recording them on the white board for class results.
11. Record the chemical name of the compound tested and evaluate
the effects at all dilutions on a 5-point scale as follows:
++ Development strongly enhanced as compared with control egg
+ Development slightly enhanced as compared with control
0 No detectable effect
- Development slightly impaired as compared with control
-- Development stopped or embryo killed at time of injection
of Evaluation/Assessment to
Experiment #1: Observe daily for five to seven days. Write and
illustrate a journal of your observations. Identify and label the following
structures: blastoderm, yolk sac, allantois, chalaza, somites, albumen,
Hensen’s node, primitive streak, neural folds and brain vesicles.
Experiment #2: Students will compare and contrast normal embryological
development to eggs that have been exposed to different concentrations
of "recreational drugs." Comparisons will be observed, described, sketched,
and measured in their journals.
Ideas to top
Experiment #2 could be done in the "shell-less" environment and the effects
of the "recreational" drugs could be observed at all stages of early development.
The risk is chick higher mortality.
Consider using Zebrafish embryos http://zfish.uoregon.edu/
References Including Web Addresses
2. Deeming, C. and Ferguson, M. 1991, "Egg incubation:
its effects on embryonic
development in birds and reptiles", Cambridge
University Press, New York.
3. Dunn, Bruce E., 1991 "Egg Incubation: Its effects on embryonic
development in birds and
reptiles," Demming and Ferguson. Cambridge
University Press, Cambridge.
4. Hanks J.H., and Wallace R.E., (1949) Proc. Soc.
Exp. Biol. Med., 71:196-200.
5. Marthy H.J. 1990, "Experimental Embryology in Aquatic Plants
and Animals", Plenum
Press, New York.
6. Rugh, R., 1962, "Experimental Embryology - Techniques
and Procedures" Burgess
Publishing Company, Minneapolis.
7. Tyrode M.V., (1910) Arch. Int. Phar., 20:205-223.
8. Zagris, N., Duprat, A.M., and Durston, A., 1995 "Organization
of the Early Vertebrate
Embryo," Plenum Press, New York.
Some people have moral and ethical objections to the use of vertebrate
models for biological research. This is an issue that you as the teacher
have to be proactive and comfortable addressing. You will also need the
support of your administrator(s) and parents. You will also need to not
only provide justification for this lab; but also alternatives in which
students can receive similar information.
For further information on bioethics go to the 1992
Woodrow Wilson Institute Earth Ethics home page or Animal
Use Case Studies.
For further information on the use of poultry as biomedical models go to
Models in Biomedical Research
Construction of PVC "Shell-less" Chick Embryo
Chamber: to top
Cut 3" off of a 3" diameter PVC plastic pipe for your stand and cut another
1" off for the clamp. Split the 1" piece open into a "C" shape.
Assembling the Egg Candling
Box: to top
A candling box can be prepared most simply by cutting an oval hole two-thirds
the diameter of an egg in the top of a paper box of sufficient size to
hold an ordinary desk lamp inside. The lamp should have the shade
removed or turned so light can shine up. Since the effectiveness
of the candling box is dependent upon having a maximum amount of light
pass through the egg against a black background, the hole in the box should
be lined with soft black felt against which the egg can be pressed snugly.
The box should be placed in a shaded corner or dark room for maximum effective
use. Look for the shadow of the embryo, the vitelline veins and the sinus
terminalis. Use only candled eggs with a proven embryo in them.
Each dilution is to be 1/10th or 1.0 x 10-1 less
concentrated than the previous. For students unfamiliar with this process
it is best to let them practice with well plates or small test tubes with
food coloring. The following is an activity that demonstrates this:
1. Using an eye dropper place one drop of concentrated food coloring
into the first well. Add nine drops of water. This gives you a 1 to 10
or 10-1 dilution from the original.
2. Take one drop of the new dilution and place it into the next well
and add nine drops of water. This gives you a 1 to 100 or 10-2
dilution from the original.
3. Repeat step 2 until you get the desired final dilution.
Ordering from Carolina Biological
Supply: to top
Pricing was based on 1997 Science/ Math Catalogue No. 66. Phone: 1 (800)
note: may use substitutes from local pharmacies or other local sources,
e.g., Caffeine = No-Doze, Nicotine = soaked cigarettes, Salicylic Acid
||Fertile chick eggs
||order two weeks before use; highest fertility -- Feb.-June p.74
||$20.95/ 100 gm
||$63.25/ 100 ml
||$6.55/ 500 ml
||$15.96/ 500 gm
||1 % Neutral Red
||$8.65/ 120 ml
||Nile Blue Sulfate
||$13.65/ 25 ml
||$7.75/ 25 pcs
||27 G, 1/2" needles
||$13.25/ 50 pcs
Jim Hashimoto & John S. Colvin
Jim Hashimoto has been teaching for 6 years at Franklin High School.
He received his B.S. in Biology from Western Oregon State College in 1991
and his currently working on his M.S.T at Portland State University.
His areas of scientific interest include animal physiology, aquatic invertebrates
and natural resource conservation. He is a member of NSTA,
OSTA, NABT, and The Nature Conservancy. He has been involved in several
research and professional development activities: ISMTE Project,
1991-1993; DOE TRAC program, Richland, WA in 1993; Green City Data Project,
1994; Access Excellence Program, San Francisco, CA in 1995 and is currently
participating in the Woodrow Wilson National Fellowship Program in Princeton,
N.J. For further information contact Jim at email@example.com
John S. Colvin teaches high school biology and integrated biology to
sophomores at Sam Barlow High School in Gresham, OR. Lately he has been
involved in the Partners in Science Program, Research Corp. Tucson, AZ
doing medical research on sperm motility at the Oregon Regional Primate
Research Center. For further information contact John at firstname.lastname@example.org