272 EMBRYOLOGICAL METHODS. 



position in the incubator. The progress of the development may be 

 followed up to the fifth day through the window. 



A description of further developments of this method, with figures of 

 special apparatus, will be found in Anat. Anz., ii, 1887, pp. 583, 609. 



See also PATON, Journ. Exper. Zool., xi, 1911, p. 469 (cultivation of 

 the embryo in vitro). 



594. Preparation. During the first twenty-four hours of incuba- 

 tion, it is extremely difficult to separate the blastoderm from the 

 yolk, and they should be fixed and hardened together.* In later 

 stages, when the embryo is conspicuous, the blastoderm can easily 

 be separated from the yolk, which is very advantageous. To 

 open the egg, lay it on its side and break the shell at the broad end 

 by means of a sharp rap ; then carefully remove the shell bit by 

 bit by breaking it away with forceps, working away from the broad 

 end until the blastoderm is exposed. The egg should be opened in 

 salt solution, then lifted up a little, so as to have the blastoderm 

 above the surface of the liquid ; the blastoderm is then treated 

 with some fixing solution dropped on it from a pipette (1 per cent, 

 solution of osmic acid, or Ranvier and Vignal's osmic acid and 

 alcohol mixture, iodised serum, solution of Kleinenberg, 10 per 

 cent, nitric acid, etc.). By keeping the upper end of the pipette 

 closed, and the lower end in contact with the liquid on the blasto- 

 derm, the blastoderm may be kept well immersed for a few minutes, 

 and should then be found to be sufficiently fixed to be excised. 

 (Of course, if you prefer it, you can open the egg in a bath of any 

 fixing liquid [10 per cent, nitric acid being convenient for this 

 purpose] of such a depth as to cover the yolk ; and having exposed 

 the blastoderm, leave it till fixed [fifteen to twenty minutes] ; but 

 I think the procedure above described will generally be found more 

 convenient.) 



The egg is put back into the salt solution, and a circular incision 

 made round the embryonic area. The blastoderm may then be 

 floated out and got into a watch-glass, in which it may be examined, 

 or may be brought into a hardening liquid. 



Before putting it into the hardening fluid, the portion of vitelline 

 membrane that covers the blastoderm should be removed with 

 forceps and shaking. 



* ANDREWS (Zeit. wiss. Mik., xxi, 1904, p. 177) separates the blasto- 

 derm at this stage by injecting picro -sulphuric acid (not any rapidly 

 acting fixative) firstly, between the blastoderm arid the vitelline mem- 

 brane, so as to separate the two above, and then between the blastoderm 

 and the yolk, so as to free the blastoderm below and float it up. This 

 done, the membrane may be incised and the blastoderm removed. The, 

 injection is best done with a pipette having a fine point bent upward^ 



