Ul;? 



EXPERIMENTAL CHICK EMBRYOLOGY 



CHORIO-ALIAKIOIC MEHBR»ME 

 AHH I ON 



HP OF SHELL 



LBUNEM 



LAYERS OF THE SHELL 

 SHELL HEHDRAKE 

 CHORIO-ALLAHTOIS 



PIPETTE USED TO 



TRANSFER THE 



GRAFT THROUGH THE 



SHELL OPENING 



ALLANTOIC BLOOD VESSEL 



Diagram illustrat liijd, tlie metliod of chorio-allantoic grafting. Host embryo 

 is generally incuoated for 8 to J days, the groft may be any anlage (e.g., 

 the limb bud of 72 hour stage). Tlie graft is generally recovered at about 

 tlie I'Jth day, just before the time of hatching of Xhe host when the mem- 

 oranes begin to dry up. 



and the further possibility that the graft may develop a parasitic relationship to the 

 host, thereby utilizing some of the nutritional requisities of the host and Indirectly 

 alter the normality of the host environment. 



Nevertheless, the chorio-allantois is proving of value in the study of mammalian iso- 

 lates (Nicholas and Eudnick, 1955) and in the culturlng of bacteria and viruses (Good- 

 pasture, 1958). Experience has Indicated that the h8 hour eye and the 72 hour limb are 

 the most satisfactory anlage to transplant. 



THE PBOCEDUBE : 



1. Preparation of the hosts : Begin incubation of fertile host eggs, properly 

 marked, about 8 days before the operation. Turn the eggs twice daily. 



2. Preparation of the donors : Begin Incubation of donors on the 5th or 6th day of 

 host incubation, so that the donors will be either 1+8 or 72 hours along at the 

 time the hosts are 8 days incubated. Mark the eggs, and rotate twice daily. 



5. Preparation of the host for the graft : (Work fast under the most sterile con- 

 ditions) 



a. Candle the host eggs, select and mark the regions most suitable for a 

 graft. The forked Junction of large blood vessels at some distance from 

 the embryo should be indicated by a pencil drawing on the shell. 



b. Prepare a window in the shell, under aseptic conditions (see directions 

 for this above). Bemove the shell in one piece. Moisten the shell mem- 

 brane with several drops of sterile Locke's or chick Einger'a solution. 

 This Is necessary in order to avoid Injury to the underlying and aome- 

 tlmes adherent chorio-allantoic membrane, and consequent mpturing of the 

 blood vessels. Cover the aperture with a sterile cover glass, or replace 

 the shell, and return the (host) egg to the incubator while preparing the 

 graft. 



k. Preparation of the transplant : (l^e - 1+8 hours, limb - 72 or more hours) 



a. Candle the donor to determine the approximate age. Crack the underside 

 of the shell on the side of a finger bowl containing about 100 cc. of 

 sterile chick Ringer's solution. When the blastoderm has moved around 

 the dorsal position, quickly excise it, separate it from the vitelline 

 membrane, and transfer it in a wide-mouthed pipette to a sterile watch- 

 glass under a dissection microscope. Confirm the age and stage of 

 development. 



