CHAPTER XXV. 273 



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

 facilitating the separation of the blastoderm. The acid should 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. Mik., 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.j xxiv, 1896, p. 371) with Rabl's platino-sublimate, 

 76 (embryos of the duck) ; PATTERSON (Biol. Bull. Wood's Hole, 

 xiii, 1907, p. 252) with picro-sulphuric acid containing 8 per cent, 

 of acetic acid, for an hour (ova of Columba) ; HOSKINS (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. 



595. 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 position 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. 



With a strip of paper 5 millimetres wide and 50 millimetres long 

 you construct a sort of triangular bottomless box. You lay this 

 on the yolk, enclosing the cicatricula in such a position that the 

 base of the triangle corresponds to what will be the anterior region 

 of the embryo, and its apex to the posterior region ; that is to say, 

 if the big end of the egg is to your left, the apex of the triangle will 

 point towards you. You now, by means of a pipette, fill the paper 

 triangle with 0-3 per cent, solution of osmic acid. As soon as the 



preparation begins to darken you put the whole egg into weak 

 M. 18 



