380 EMBRYOLOGY 



then pierce the inner chorion with fine scissors or needles, and 

 gently press out the ovum. Harden in alcohol. 



FiCK {Zeit. wiss. ZooL, Ivi, 1893, p. 529) uses a mixture of 

 250 parts of 1 per cent, chromic acid, 1 of acetic acid, and 750 of 

 water. 



827. Triton (Scott and Osborn, Quart. Journ. Mic. Soc, 

 1879, p. 449). The albumen is here present in the form of several 

 concentric coats, which are very delicate. Incise each of them 

 separately with fine scissors, turn out the ovum, and fix it in 

 solution of Kleinenberg. 



Hertwig {Jen. Zeit. Naturw., 1881-2, p. 291) puts the eggs 

 into a mixture of equal parts of 2 per cent, acetic acid and 0*5 

 per cent, chromic acid. After ten hours he incises the membranes, 

 opening one end of the inner chorion, and turns out the embryos 

 and brings them into successive alcohols. 



MiCHAELis {Arch. mik. Anat., xlviii, 1896, p. 528) fixes ova, 

 with their envelopes, in a mixture of concentrated sublimate 

 solution and concentrated picric acid, 20 parts each, glacial 

 acetic acid 1, and water 40, but removes the envelopes before 

 bringing into alcohol. 



828. Salamandra (Rabl, Morphol. Jahrb., xii, 2, 1886, p. 252). 

 For his more recent methods see § 834. 



Gronross {Anat. Anz., xiv, 1898, p. 461) fixes the ova with 

 a mixture of 50 parts each of saturated sublimate and 0-5 per 

 cent, chromic acid with 1 part of acetic acid. 



829. Rana. Gatenby has found that the following mixture 

 often gives very good results for the eggs (not embryos) of Rana 

 temporaria ; it dissolves away the albumen coat, preserves yolk 

 and mitochondria, and leaves the eggs soft enough to cut in 

 paraffin with a rotary microtome : — ■ 



Bichromate of potash of 2 per cent. . 100 c.c. 



Chromic acid of 1 per cent. . . . 100 ,, 



Nitric acid . . . . . . 6 ,, 



Use at least 40 c.c. to twenty or thirty eggs for fifteen to twenty-four 

 hours. Shghtly shake, and the albumen coats fall off if not already 

 dissolved. Wash out for about one hour in running water and then 

 upgrade from 70 per cent, alcohol (one half-hour), 90 per cent, (one 

 hour), to absolute alcohol, two changes of one hour each. Clear in 

 benzol for fifteen minutes. Add chips of wax and place in thermostat 

 for half an hour. Transfer to pure wax for one-half (to three-quarters) 

 of an hour. Avoid unnecessary heat. 



The eggs thus treated can often be cut 6 (jl on a rotary microtome 

 provided with a sharp knife. The method is indicated where large 

 numbers of stages of the early development of the frog are required for 

 junior class purposes. The main objection to the method is that the 

 chromic acid attacks pigment. Fertilisation and segmentation stages 

 and general cytology are often extremely good ; stain in any way. 



O. Hertwig {Jen. Zeit. Naturw., xvi, 1883, p. 249). — The ova are 

 thrown into nearly boiling water (90° to 96° C.) for five or ten minutes. 



