EXPERIMENTAL CHICK EMBRYOLOGY ^^25 



moval of the vitelline memtrane ) in and out of a wide-mouthed pipette, thereby removing 

 most of the adherent yolk. Still others play a gentle stream of medium on the surface of 

 the inverted blastoderm, blowing the yoli: granules away. 



The early stages are particularly fragile and the less handling the better. In most 

 instances the peripheral yolk may be trimmed away with scalpel, scissors, or even with 

 needles, and the yolk adherent to the underside of the area pellucida is negligible. It 

 la particularly difficult to handle the blastoderms of less than l8 hours without damage. 



Later Embryos : Chick embryos of 56 or more hours of incubation are rather simple to 

 remove from their eggs. The entire egg mass is broken Into a finger bowl of Ballne solu- 

 tion cracking the underside of the shell after allowing the egg to remain motionless for 

 several minutes. The embryo, unless it Is so far advanced that it is heavy, will float to 

 the upper surface. Excise the embryo as In the manner described above or. If the embryo Is 

 well formed, grasp the yolk-sac umbilicus with forceps, cut It distally to the forceps, 

 rupture other membranes, and draw the embryo away from its yolk. If the embryo Is young 

 it may be pulled into a submerged watchglass and transferred to a fresh finger bowl of 

 solution. If It is advanced. It may be transferred In an ordinary teaspoon whose bowl has 

 been perforated with many small holes. 



EXAMINATION OF EXCISED CHICK BLASTODERMS 



The early blastoderms may be transferred to a watchglass In several drops of Locke's 

 or saline medium (wanned to 58°C.), oriented with needles into the proper position and 

 then the medium sucked off by means of a fine bore pipette, while encircling the blasto- 

 derm. In this way the blastoderm will be flattened onto -the dry bottom of the watchglass. 

 Immediately add fresh medlxim (at 58°C.) so that it flows beneath the blastoderm, lifting 

 it up on the surface tension of the fluid. If 0.001^ phenol red (pH indicator) is added 

 to the medium, the slightly purple tinge of the alkaline medium will provide an excellent 

 background for greater clarity of the chick embryo structures, wltho^it adding any toxic 

 factor. Such a watchglass may be placed on a wanning stage or on an electrically control- 

 led heating stage and the living embryo examined for a considerable period of time. 



If one is interested In general morphology of these early stages it is advisable to 

 mount embryos of the same age In the normal position, and also upside down, so that, in 

 the latter Instance, one has a view directly into the intestinal portals. 



Such a mount may be made on a glass slide providing a out-out in filter paper Is made 

 of just such size as to frame the area pellucida and mask out the area opaca. If such a 

 blastoderm Is first Inverted, pulled up onto the slide (while the slide is submerged In 

 solution), most of the medium drained off, and the filter paper frame added, the embryo 

 will remain flat. The yolky margins of the blastoderm will adhere firmly to the filter 

 paper. A cover glass may be added providing Its comers are elevated slightly by bits of 

 Permoplast, to prevent crushing. The embryo may now be hydrated with the appropriate 

 medium. It must be remembered, however, that the chick embryo cannot acquire sufficient 

 oxygen from any aqueous medium directly, and that it can be "drowned", particularly if It 

 has already developed its own circulatory system. The ideal environment is a closed 

 space, completely humidified, with the blastoderm floating on the surface of a nutrient 

 medium. 



The later stages may be examined in the manner of any vertebrate form. After about 

 5 days the embryo will take on a definite avian appearance, and It will shortly become 

 possible to make a dissection to study the internal organs. This is so because the car- 

 tilage and bone will not have developed. There is very little cartilage, even at 8 days. 



PERMANENT PREPARATIONS OF CHICK EMBRYOS 



The primitive streak stages can be best fixed while still on the egg, by dropping 

 the fixative onto the blastoderm gently from above. Since fixation renders the blastodenn 

 rather brittle, it is best to cut it out within a few minutes after fixation, and then 

 transfer it to a Syracuse dish with fresh fixative for the requisite time. (Use instru- 

 ments other than those for operating purposes.) 



