HATCHING AND DISPLAYING AVIAN EMBRYOS
R. A. Voitle, H. R. Wilson, L. W. Kalch and C. R. Douglas
Construction of a Still Air Incubator .......................... 1
Incubating and Hatching Eggs ................
Fertile Eggs ...........................
Hum id ity ...............................
Position of Eggs .......................
Candling Incubated Eggs ................
Incubation Time Required for Hatching ..
Displaying Living Chicken Embryos .............................. 9
The Preparation and Preservation of Embryos .................... 11
Landmarks of Embryonic Development ............................. 14
Preparation of Colored Chicks in the Shell ..................... 16
Preparation and Demonstration of Various Anatomical Systems
of the Fowl ..... ........................................ ... 18
Skeleton ................................................. 18
Soft Organs ............................................... 19
Mimeograph Series No. PY 69.4
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Every year there is an increasing demand for information on the
construction of still-air incubators, hatching, preserving and displaying
avian embryos. The stimulation behind these demands is the student's
ever increasing thrust for knowledge about nature coupled with the fact
that the chicken supplies a neatly packaged embryo which requires only
the addition of heat, moisture and air to stimulate development and
CONSTRUCTION OF A STILL AIR INCUBATOR
An attractive, inexpensive incubator can be constructed by using
cardboard boxes as the basis for construction. This design uses a
lightbulb as a source of heat and a commercial thermostat to regulate
the temperature. As a source of moisture,a cake pan filled with water
Materials and Supplies:
All the dimensions mentioned below must be adjusted to the size of
the boxes used.
1 box 18" x 14" x 13" in height (corrugated)
1 box 16" x 12" x 12" in height (corrugated)
(The larger box should be 2" greater in both length
and width and 1" higher)
1 glass 18" x 14" (size of larger box)
" Hardware cloth 20" x 16" (4" wider and longer than smaller box)
Light socket (porcelain)
Electric cord 7' 20'
Male electric plug
Thermostat (wafer type)
Water Pan (Cake pan should cover at least half of the bottom of
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Light bulb (60-75 watt)
1" Masking tape (freezer tape)
Insulating material (newspapers)
Tin Snips (wire cutters)
Knife and/or scissors
3/4" block of wood 4" x 4"
Round rolled oats box or fruit juice can (this acts as chimney
Newspapers for bottom should be several layers
Aluminum foil (to line inside of box)
Cut flaps from smaller box and attach the thermostat to one of
the longer sides making sure that the wafer clears the upper edge
by at least -". A portion of one of the flaps can be glued to the
bottom of this box to fill in the space where the inner flaps do
not come together.
Apply glue on inside of the bottom around the center portion of
the larger box. Now center the smaller box inside the larger box and
place weights in the bottom to ensure bonding. There should be at least
2" between the sides of the inner and outer box (1" would be better).
Make a hole through both boxes for the rod that holds the wafer of the
thermostat (be sure that the alignment is correct). Make another hole
through both boxes 1" from the bottom and centered so that the electric
cord can pass through.
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Lightly stuff the area between the boxes with insulating material
(newspapers are fine whole or shredded in strips). Do not bulge sides.
Draw a line all the way around on the outside of the larger box
at the level of the upper edge of the smaller box.
With a knife or single edged razor blade, cut along this line.
Make sure the cut only goes through the outer cardboard layer and nicks
the corrugated portion. You must not cut through the inner layer of
Now you must make diagonal cuts in the corners of the larger box
so the flaps will fold neatly (diagram).
Fold the flaps inward along the cut portion.
Mark these flaps where the" come in contact with the inner edge of
the smaller box and cut along these lines.
Tape the flaps with 1" masking tape to the edge of the smaller
Cut a piece of hardware cloth (-" mesh) so that it is 4" wider
and 4" longer than the inside diameter of the smaller box.
Using the tin snips cut a 2" square from each corner of the
hardware cloth. Then bend the projecting pieces of cloth so they form
legs to support the screen. It should fit snugly in the bottom of the
Wire the light socket so that the flow of electricity goes through
the thermostat (see Figure 1). The light socket should be attached
to a wooden base (4" x 4" block).
Figure 1. Wiring Diagram for Still Air Incubator
Socket == Plug
Line the box with aluminum foil, place several layers of newspaper
in the bottom, place cake pan under screen, assemble thermostat on
back wall, insert light bulb and place rolled oats box around light.
Cut 4 ventilation holes (1" i. diameter) about level with the
top of the eggs. Cut covers 1-" 2" out of extra cardboard to
regulate air-flow and attach to the inner box with brads.
Plug in ! ! !
Regulate temperature so that it ranges between 101 and 1030 .
Check unit periodically for several hours to make sure it is functioning
properly before setting eggs.
Place eggs on hardware cloth around the light and lay the glass
on top of the box as a cover.
For a more permanent type of incubator, refer to the University of
Florida Agricultural Extension Service Circular # 335, "A Small Display
Incubator" by L. W. Kalch.
INCUBATING AND HATCHING EGGS
A supply of fertile eggs (one to two dozen) may be obtained from
hatcheries or from any poultry flock where roosters are being kept with
the hens. Eggs purchased from food stores are not fertile and therefore
will not hatch.
Hatching eggs should be incubated as soon as possible after they
are laid. They can, however, be stored up to seven days with good
results when held at a temperature of 500 to 600 F and a relative
humidity of 70 to 75% (45 550 wet bulb reading).
A still air incubator should be operated at a temperature of
from 101 to 1030 F, with the bulb of the thermometer at the top of
the eggs. When the temperature is higher than this the incubation
period will be decreased and if lower then it will be increased and in
either case hatchability will be reduced. If the temperature of the
embryo exceeds 1040 F, death may occur.
It is necessary that the temperature be kept as close to 1010 1030
F as possible thus the need for an accurate thermometer and a rcliz!cle
heating and control unit.
Each incubator is different. Through experience the operator will
be able to determine the best operating conditions for his particular
incubator. For example, if the first hatch of chicks comes off at 20
days instead of the normal 21 days, the incubator was too warm and the
operating temperature should be lowered about 10 F for the next hatch.
If the chicks required more than 21 days to hatch, the incubator was too
cool and the temperature should be raised about 10 F. (Normally chicks
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pip the shell on the 19th day, emerge from the shell on the 20th
day, and are dry and fluffed out and ready to be removed from the
incubator on the 21st day.)
Do not place the incubator hn drafts or direct sunlight or allow
it to be subjected to other conditions that might cause fluctuations
To prevent the eggs from drying out too rapidly, moisture must
be added to the air. This can be done by filling the small pan with
water to a depth of one inch and placing it under the egg tray.
Figure 2 shows the comparative size of the air cell in an egg
on the seventh, fourteenth and eighteenth days of incubation. If
the air cell is too slow in enlarging, too much moisture is present.
On the other hand, if the air cell is too large, more moisture is needed.
Size of the air cell can be determined by candling (placing the egg
before a strong light see page 8).
It is advisable to sprinkle the eggs lightly with warm water at
the time of hatching to prevent the chicks from sticking to the shell.
This is done when the first egg pips. Best results are achieved when
the relative humidity is 60% (860 F wet bulb) for the first 18 days
and then increased to 65% (900 F wet bulb) for the remainder of the
incubation period. 7
Size of air cell on the seventh,
fourteenth and eighteenth days
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Position of Eggs:
Eggs will hatch poorly unless they are turned several times each
day from the first through the seventeenth day of incubation. This
prevents the developing embryo from sticking to the shell. In this
small incubator there are two ways that this may be done. The eggs
may be marked on one side and laid in the incubator with the marked
side up. Turn the eggs an odd number of times (3 or 5) each day.
This will prevent having the egg laying in the same position each
night, which is likely to be the longer period. All eggs should have
the marked side up at the same time and all eggs should have the marked
side down at the same time. This system will insure getting all eggs
The other less desirable method of turning eggs would be to place
the eggs small end down in egg cartons which have had the top removed.
To turn the eggs the carton may be tilted, first one direction and then
After fertilization the egg contains all the materials necessary
for embryonic development, except sufficient air. The embryo obtain
the necessary oxygen for growth from the air that passes through the
tiny pores i the egg shell.
In operating the incubator you should be sure that the machine is
properly ventilated at all times. There should be 4 vents (openings
1" in diameter) for ventilation. It is desirable to adjust these
openings; halfway the first 18 days of incubation and fully open the
final three days( see page 4). Be sure that the incubator is placed
in a well ventilated room.
Candling Incubated Eggs:
Incubated eggs are candled to determine whether or. not they are
fertile and to check on the growth and development of the embryo.
Fertility is more easily determined in white shelled eggs, which may
be candled on the 3rd or 4th day of incubation. Brown shelled eggs
should be checked on the 5th or 6th day of incubation.
When candling fertile eggs on the 4th or 5th day of incubation the
embryo appears as a small reddish area with blood vessels extending
out into the egg. At this stage the embryo gives the appearance of a
large red spider.
Infertile eggs will appear clear with no embryo development. In
some eggs the embryo will die after a few days of development. In
such cases the blood will accumulate at the outer ends of the blood
vessels and will form a blood ring or a streak. After a few days this
will turn black and the egg starts to decompose.
When checking embryo development the large end of the egg should
be placed to the opening of the chandler since the embryo will be
found near the air cell. The candling of eggs should be done in a
Egg candlers may be purchased from poultry equipment dealers or
they may be made. A satisfactory chandler may be made by placing a
60 watt light bulb in a small box with a hole drilled in the side. This
opening should be 1" to 1" in diameter with the light bulb centered
directly on the hole. A rolled oat box lined with aluminum foil and
with a "door" cut i the top (to let heat escape) may be used.
INCUBATION TIME REQUIRED FOR HATCHING
Ducks (except Muscovy) ......................
Bobwhite Quail ..............................
Coturnix Quail ..............................
DISPLAYING LIVING CHICKEN EMBRYOS
Live embryos make an interesting exhibit for fairs or science
exhibitions if properly prepared. In preparation for the display, the
student must arrange his setting schedule so that all the embryos he
wishes to display will be the proper age on the day of the show. It
is suggested that 3,5,7 and 10 days would be the appropriate ages to
display the embryos. Approximately 6 eggs should be set for every age
embryo to be displayed. This allows for infertility, poor technique,
embryonic mortality and leaves extra embryos available in case some
should dry out too much during display. A good method of determining
setting time is to count back from the show date, the days in age of the
embryo to be displayed. Two to three hours should be allowed for warmup
of eggs in determining setting time.
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Materials and Supplies:
Incubated (fertile) eggs
Egg carton (to hold eggs) or an elevated pedestal for each age
of embryo, near the glass top of your
incubator. (3/4" x 2" x 2" blocks
with a l!" hole bored in the block)
Small soft artist's brush
Distilled or soft water
All instruments that will penetrate the egg scissors, tweezers,
etc. should be placed in boiling water for 30 minutes and then dipped
To prepare for the removal of the shell, candle the egg and mark
the outline of the air cell. Using a knife or scissors, carefully punch
a small hole in the center of the air cell outline. If a chandler is
not available and you do not wish to construct one, a small hole may be
punched in the center of the large end of the egg. Proceeding carefully,
enlarge the hole, taking care not to rupture the inner shell membrane.
The shell and outer shell membrane should be renoved to the very edge
of the junction of the shell, air cell, and inner shell membrane. Again,
care must be taken not to rupture the inner shell membrane. At this
point you may proceed in either of two directions. Visibility is greatly
reduced by the inner shell membrane. This membrane can be removed,
but some skill is necessary to do so without rupturing the blood vessels.
One procedure is to place a drop of distilled or soft water on
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the membrane near the edge of the shell. Grasp the membrane with the
tweezers gently at its junction with the shell, being careful not to
grasp any blood vessels or albumen. Now very gently and very slowly
pull the membrane away from the shell, blood vessels, and albumen
(the use of the second pair of tweezers may be valuable here). The
membrane should be completely removed to the size of the shell opening.
The disadvantage to this method is that eggs prepared this way
tend to dry out more rapidly which results in the death of the embryo;
however, the embryos will remain in good condition (in the incubator)
for several hours, which is generally long enough for the period of
display. It should be pointed out here that the removal of the inner
shell membrane should be delayed as long as possible. The ideal situation
is to remove the membrane immediately before the show.
If the first method proves to be undesirable, a second method
is to brush the inner membrane with !ptroleum jelly. This method
helps to retard evaporation. At the incubation temperature (101-1030 F)
petroleum jelly melts and makes the membrane relatively transparent.
Visibility is not as good as with the peeling method but until some
skill is attained, the petroleum jelly method is more likely to be
It is possible that your specimens could dry out during prolonged
display; this should cause no problems for you have set 6 eggs for
every embryo age an' a': that is necessary is to open a new embryo
of the proper age.
THE PREPARATION AND PRESERVATION OF EMBRYOS
Chick embryo displays at fairs, etc., always seem to be popular
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and educational. The great changes in size and form that take
place from day to day are quite startling. It is possible to preserve
embryos so that they maintain their original size and shape for several
years. The natural color of the embryo is bleached away by the prepara-
tion, but this does not seem to detract too much from the display because
the size and shape of the embryo are the most striking aspects of the
exhibit. The preservant used is made by mixing 1 part (by volume) of
f ormalin (40% formaldehyde) with 3 parts (by volume) of water. Embryos
of various ages are freed of yolk, albumen, and extra-embryonic membranes
and placed in the formaldehyde solution for fixing and preserving. The
solution should be changed twice, or as required for it to remain clear.
Materials and Supplies:
Incubated (fertile) eggs
Egg chandler (optional)
Formalin (40% formaldehyde) CAUTION, AVOID EXCESS INHALATION
Water (soft or distilled)
2 or more bowls or petri dishes
Jars (baby food)
The student should allow for infertile eggs, embryonic mortality
and accident by setting at least 3 eggs for each embryo wanted for
preservation. The setting schedule can be arranged so that the embryos
of the desired age come off on a convenient day. Count back the number
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of days of age for each embryo from the date the embryo is to be prepared
and this is the setting date. Embryos can be prepared conveniently
from 5 to 19 days of age to give practically a complete observable
record of the chick development from fertilization to hatching at 21
days. It would be an advantage to start with the older embryos and
proceed backward. In this way, technique and confidence would improve
as the embryos became more fragile.
To prepare the embryo from the incubated egg, crack the shell
as if to fry an egg. This should be done over a sink. Let the contents
of the egg drop gently into your hand. You can now allow the great mass
of albumen and much of the yolk to escape through your fingers, taking
care not to allow the embryo to slip through too. The embryo and the
remaining yolk, albumen and extra-embryonic membranes are then transferred
to one of the bowls which contains some of the saline (9 grams non-
iodized table salt in 1000 cc of water) solution. Here the embryo is
g ently swished around in the solution to free it of undesirable material.
The structure (umbilical) attached to the abdomen must be clipped to
remove the embryonic membranes. Next, transfer the embryo to a second
bowl which contains the formaldehyde solution. Again, the embryo is
gently washed in an attempt to rid it of all extraneous material. The
embryo is then transferred to a third (or back to the first if it has
been washed) bowl containing the saline solution, and gently washed a
third time. These processes are repeated until the embryo is clean.
Now fill a clear jar (baby food) with the diluted formalin solution
(3 parts water to 1 part formalin) and drop in the embryo. Do not be
alarmed if the embryo floats as this is normal until its tissues absorb
some of the solution. Check the preserved specimens at intervals and
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transfer them to a fresh diluted formalin solution if the solution is
cloudy or contains floating tissue, etc. Continue these transfers until
the solution remains clear and free of excess floating tissue. When
this point is reached, fill the jars to the top and close them tightly.
Never leave the embryo exposed to air for any extended period.
Label the embryo indicating specie, age, and other pertinent data.
It is suggested that the labels be placed on the top of the cap to leave
an unobstructed view of the specimen.
Following are listed some of the changes that take place during
embryonic development. It might add to your exhibit if this were
reproduced and the descriptions that apply to your embryonic ages were
LANDMARKS OF EMBRYONIC DEVELOPMENT
Between oviposition and
Division and growth of living cells
Segregation of cells into groups
of special function
Several hundred cells
No growth; stage of inactive
embryonic life (if temperature is
kept below physiological zero =80 F)
Appearance of primitive streak
Appearance of primitive gut;primordial
germ cells appear in germinal crescent
Appearance of vertebral column
Appearance of neural groove
Appearance of somites
Appearance of blood islands;vitelline
Appearance of eye (6 somites)
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Appearance of heart
Appearance of amnion (protection)
Appearance of one (14 somites)
Cranial flexure and torsion evident
Heart beats;beginning of yolk sac
Appearance of amnion;chick has
turned on left side
Appearance of nasal pits
Appearance of posterior limb buds
Appearance of anterior limb buds
Aprr anre of aW!antnis;
(a) cm':-yonic respiratory system
(b) hold's kidney excretions
(c) aids in caborption of albumen
(d) connected to future cloaca
Appearance of tongue and esophagus
Appearance of proventriculus,
gizzard, reproductive organs and
differentiation of sex
Appearance of beak and egg-tooth
voluntary movement;duodenal loop
Allantois and chorion serosaa)
anastomnse and lay against shell;
ceca and digits appear
Feather tracts appear
Embryo bird-like in appearance;
growth of allantois nearly complete
Beak begins to harden
Abdominal walls established, loops
of intestine are seen hanging in
Appearance of scales, claws, down
and muscle;cartilaginous skeleton
is complete and ossification is
Embryo turns head toward blunt end
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Small intestines taken into abdomen
Beak, nails and scales relatively
cornified;albumen practically gone
and yolk increasingly important
Amniotic fluid decreases;head
under right wing with beak toward
Transfer of embryo to hatching
trays turning no longer required
Yolk sac begins to enter body
cavity and amniotic fluid disappears
(embryo swallows it) ; beak pierces
air cell and lungs function slightly
Yolk sac drawn completely into
body;pulmonary respiration starts;
allantoic circulation and respiration
cease and allantois dries up
(a) head under right wing
(b) legs toward head
(d) shell quite brittle as some
calcium of the shell is used
to build skeleton
THE PREPARATION OF COLORED CHICKS IN THE SHELL
E. A. Schano, Cornell University, has pointed out that the main
purpose in coloring embryos is to provide a practical method of
identifying the young of individual hatches during early development
and to enable the observation of their movements after they leave the
nest. In wildlife management studies where identification and obser-
vation of ducks is extremely difficult because of natural protective
coloration, identification and observation are made easier by
coloring the embryos with bright dyes.
These coloring techniques lend themselves very nicely to
exhibition in science fairs.
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Materials and Supplies:
1 large beaker (1,000 cc) or pan
1 small beaker (250 cc) or water glass
1 hypodermic syringe (see science teacher) and 1 No. 20
1 bottle merthiolate or alcohol
1 food coloring kit
Incubated (fertile) eggs
1 No. 3 dental drill or knife with sharp point.
1 chandler (see page 8)
Place water in the large beaker or pan and heat to boiling. Loosen
the tops on the dye bottles and place them upright in a small beaker
or glass. The tops must be loosened so the bottles will not explode
when heated. Put syringe, needle drill or knife blade in the container
with the bottles of dye. Add about a half inch of distilled water to
the container, then place the small container in the larger container
and boil for 30 minutes. Remove from the heat, pour off water and
allow contents to air cool.
Select fertile eggs which have been incubated to the 18th day.
Candle the eggs to determine the location of blood vessels. Find a point
about 1" from the small end of the egg with no blood vessels, daub an
area about the size of a half dollar with merthiolate or alcohol to
disinfect it. Dip the tip of the No. 3 dental drill or the knife blade
into the merthiolate or alcohol, place against the egg in the area with
no blood vessels which was disinfected and rotate until a hole is drilled
just through the shell. Assemble the syringe and needle. Be careful
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not to contaminate these (particularly the needle) by contact with the
fingers or foreign objects. Dip the needle in the merthiolate and then
into the dye and withdraw about -cc of dye into the syringe. Insc'1
only the tip of the needle through the membranes, (be sure you are
through the membranes). To prevent overflow, very slowly inject the
dye into the egg. Remove the needle and seal the hole with cellophane
tape, fast drying cement, wax or bandage tape and return the egg to the
Use the dyes as they come from the bottle or combine them to
obtain unusual colors. You might find it desirable, on the day of the
exhibition, to have some colored chicks that were hatched the previous
day 'n combination with hatching and unhatched eggs, for the purpose of
demonstrating that the chicks were colored in the shell and not dipped
after hatching. Chicks from breeds that have white or yellow down are
most satisfactory for dyeing. The skin will also be colored and colored
droppings may be noticed for a few days. The dye has no detrimental
effects on the chick and the care required is normal.
PREPARATION AND DEMONSTRATION OF VARIOUS ANATOMICAL
SYSTEMS OF THE FOWL
Preparation of a skeleton for exhibition:
A mature chicken should be used for this preparation since fully
ossified bones give the best results.
The bird should be humanely sacrificed and as much tissue and muscle
as possible removed from the bones. Care should be taken not to damage
the bones. The remaining tissue can be loosened from the bone by
boiling the skeleton in water for about three hours. The bones
should be scrubbed clean and with a small brush. Soaking the bones
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in chlorine bleach for about one hour will whiten and further clean them.
After drying, the bones may be assembled with wire and glue and
mounted on a small rod. The skeleton may be coated with a thin coat
of varnish for protection.
Staining of embryo skeletons of intact specimens:
Remove feathers or down by dropping embryo in a sodium sulfide
solution for a few minutes and then wash the specimen thoroughly.
Fix specimen by transferring to a jar of 95% alcohol for at least
3 days then dehydrate with 100% alcohol for 5-8 hours.
Place the embryo in acetone to extract the fat. Several changes
will be necessary before all the fat is removed. Continue changes
until the fluid remains clear.
Clear the specimen in 2% potassium hydroxide (KOH). Change the
sdution until it remains clear for at least two days.
Stain the specimen by placing it in alizerin red solution made
up in potassium hydroxide solution. Prepare a stock solution of 1 gm.
alizerin red in 100 parts of water. Use 1 ml. of this stock solution
to 100 ml. of 2% KOH. Staining will be complete within 12-24 hours.
Over-staining may be somewhat corrected by placing specimen in sunlight.
Final clearing is accomplished by infiltrating the specimen with.
glycerin. Start with a solution of 1 part glycerin to 4 parts KOH
for 2-3 days. Gradually increase glycerin until the specimen is
mounted in pure glycerin.
Preparation of soft organs for exhibition:
Some of the hollow organs (e.g., the respiratory system and cir-
culatory system) may be demonstrated by casting them (molding true-to-
life models) with different materials. One of the most successful
materials is methyl methacrylate. A procedure for casting organs with
this material has been described by Taylor, Boone and Barnett (The use
of methyl methacrylate for casting hollow organs of the chicken, by
R. 0. Taylor, M. A. Boone and B. D. Barnett. Poultry Research Series
No. 15, 1962. Poultry Science Department, S. C. Agr. Exp. Sta.,
Clemson Agricultural College, Clemson, South Carolina).
A Method of Color Marking Young Waterfowl, by C. D. Evans. Journal of
Wildlife Management 15:101-103, 1951.
A Small Display Incubator, by L. W. Kalch. University of Florida
Extension Circular, 1961.
Charts, Models and Slides on the Chick Embryo. Turtox Service Leaflet.
General Biological Supply House, 761-763 East 69th Place, Chicago 37,
Early Embryology of the Chick, 4th edition, by B. M. Patten. Blakiston
Company, New York, New York.
Embedding Specimens in Transparent Plastic. Turtox Service Leaflet No. 33.
General Biological Supply House, 761-763 E. 69th Place, Chicago 37,
Embryological Studies for the High School Biologists. The American
Biology Teacher 16(1):16-17.
Foundations of Embryology, 2nd edition, by B. M. Patten. McGraw-Hill,
Inc., New York, 1964.
Incubation and Brooding of Chickens. USDA Bulletin No. 1538.
Incubation, Fixation and Mounting of Chick Embryos. Turtox Service
Leaflet No. 17. General Biological Supply House, 761-763 East
69th Place, Chicago 37, Illinois.
Lillie's Development of the Chick, (revised), by H. L. Hamilton. Henry
Holt and Company, New York, New York.
The Avian Embryo, by A. L. Romanoff. The MacMillan Company, New York,