Chapter XVI: Some Embryological Methods 117 



cult to separate the blastoderm from the egg during the first 24 hours of 

 incubation and it is advisable therefore, to fix and harden both together 

 and to remove the blastoderm later.* The blackened area affords a 

 convenient means of orienting the preparation for sectioning. 



4. For the Embryology of Teleosts the following are the most useful 

 mounted stages : 



I. Whole mounts. Two-, 4-, 8-, 16-, 32-, and 64-cell stages (only the 

 blastodisc segments); early periblast; late periblast; early germ -ring; 

 embryonic shield; various stages of early embryos, such as embryos of 

 45, 50 and 60 hours. 



II. Sections (paraffin). Four, 16, and 32 cells (vertical sections parallel 

 to the first plane of cleavage); late- cleavage (vertical sections); early, 

 mid, and late periblast (vertical sections); transverse and sagittal sec- 

 tions of early germ-ring, embryonic shield, early embryo, late germ-ring, 

 and closing of blastopore, respectively. 



All stages may be fixed in picro-acetic (reagent 23, Appendix B) for 

 30 to 40 minutes. Later stages may also be fixed in Hermann's fluid 

 (reagent 26) 2 to 4 minutes. Eggs fixed in Hermann's fluid should be 

 washed for an hour in frequent changes of water, and the membrane of 

 each egg should then be pricked with a needle before passing it through 

 the series of alcohol. The eggs are finally preserved in 83 per cent, 

 alcohol. (Child finds that fixation for about a minute in 10 per cent, 

 acetic acid saturated with corrosive sublimate, followed by 10 per cent, 

 formalin, gives good results without the yolk becoming hard.) 



Before the preserved material can be mounted in toto or sectioned, 

 the essential part (the blastoderm) must be dissected off under a dissect- 

 ing lens by means of sharp needles. If the blastoderms are to be 

 mounted entire they may be passed down through the alcohols (see Wal- 

 ton's device, memorandum 4, chap, iii), stained in Conklin's hematoxylin 

 (reagent 48, Appendix B), then dehydrated and mounted in the usual 

 way. To avoid crushing the objects, the cover-glass should be supported 

 by means of bits of broken cover. Material which is to be sectioned 

 may be stained in toto or the sections may be stained on the slide. In 

 the latter event, to facilitate orientation, it is necessary to tinge the 

 blastoderms slightly with Bordeaux red or some other cytoplasmic stain 

 unless the fixing reagent has already done so. For the same reason it is best 

 to imbed the material in a watch glass, arranging it near the bottom of the 

 paraffin mass so that one can see with a microscope how to shape the 



* Andrews (Zeittchrift filr wissenschaftliche Mikroskopie, Vol. XXI, 1904, p. 177) injects 

 picro-sulphuric acid (1) between the vitelline membrane and the blastoderm and (2) 

 between the blastoderm and the yolk, by means of a pipette which has a fine upcurved 

 point. The blastoderm may then be readily freed from the yolk. This operation should 

 be performed before the egg has been subjected to the action of any reagents. 



