376 EMBRYOLOGY 



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 we 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. 



Fixation in 10 per cent, nitric acid has the advantage of greatly 

 facilitating the separation of the blastoderm. The acid snould 

 be allowed to act for ten minutes, after which it is well to bring 

 the preparation into 2 per cent, solution of alum [cf. Hofmann, 

 Zeit. wiss. Mile, x, 1893, p. 485). Mitrophanow {Anat. Hefte, 

 xii, 1899, p. 200) fixes with nitric acid of 3 per cent. ; Suschkin 

 {Nouv. Mem. Soc. Nat. Moscow, xvi, 1899, p. 34) with sublimate ; 

 FiscHEL {Morph. Jahrb., xxiv, 1896, p. 371) with Rabl's platino- 

 sublimate, § 81 (embryos of the duck) ; Patterson {Biol. Bull. 

 Wood's Hole, xiii, 1907, p. 252) with picro-sulphuric acid con- 

 taining 8 per cent, of acetic acid, for an hour (ova of Columba) ; 

 HosKTNS {Kansas Univ. Sci. Bull., iv, 1907, p. 176), after removing 

 shell, for five to fifteen minutes in a mixture of 3 parts of 10 per 

 cent, formol with 1 of 10 per cent, nitric acid, and then excises the 

 embryo. 



In order to counteract the turning up of the edges of the blasto- 

 derm that generally happens during the process of hardening, it 

 is well to get the blastoderm spread out on the convex surface of a 

 watch-glass, and leave it so during the hardening. 



For hardening Henneguy prefers the osmic acid and alcohol 

 mixture of Ranvier and Vignal, or Flemming's mixture followed 

 by successive alcohols. 



Stain and imbed by the usual methods. 



Up to about the fiftieth hour embryos may be mounted entire 

 in glycerin or balsam. 



818. M. Duval's Orientation Method {Ann. Sc. Nat, 1884, 

 p. 3). In the early stages of the development of the ova of Aves, 

 before the appearance of the primitive streak, it is difficult to 

 obtain a correct orientation of the hardened cicatricula, so as to 

 be able to make sections in any desired direction. Duval, 

 starting from the fact that during incubation the embryo is almost 

 always found to be lying on the yolk in such a jiosition that the 

 big end of the egg is to the left and the little end to the right of 

 it, marks the position of the blastoderm in the following way. 



