METHODS OF HARDENING AND PRESERVING. 363 



'very finest grade of oil-stone. No oil should be used, but instead a mixture of 

 equal parts of glycerin and water. Before the knife is honed it must be made as 

 clean as possible. The oil-stone itself also must be cleaned with equal care, and 

 the mixture of glycerin and water should, if necessary, be filtered before using 

 to keep it free from dirt. A single particle of dirt may be the cause of making 

 many microscopic notches in the edge of a knife. A knife is well sharpened 

 when its edge appears smooth and straight under a magnifying power of twenty- 

 five diameters. 



The microtome knife should be as unlike a razor as possible. It must have 

 a very thick back and be as heavy and rigid as practicable, so that the actual 

 cutting-edge may be as steady and inflexible as it can be made. Knives of suit- 

 ably heavy construction are now furnished with all the best microtomes. 



Methods of Hardening and Preserving. 



The two most generally useful methods for preserving embryos are with 

 Zenker's and Tellyesnicky's fluids. Good results may be had with the other 

 reagents. Specimens preserved with picro-sulphuric acid have the advantage of 

 staining readily. To study the medullary sheaths of nerve-fibers, as is necessary 

 to follow the development of the fiber tracts in later stages, the specimens must 

 be preserved in Miiller's fluid. Flemming's and Hermann's fluid are valuable, 

 especially for cytological study, but are applicable only to small pieces. 



I. Zenker's Fi^uid. 



Formula : Corrosive sublimate, 5 g™- 



Potassium bichromate, i gm. 



Sodium sulphate, I gm. 



Water, loo c.c. 



Add 5 c.c. of glacial acetic acid to the fluid immediately before using. 



The fluid does not have great penetrating power, but may be used for embryos 

 of 25 mm. The amount of fluid should be from ten to twenty times the volume 

 of the specimen, and better results are obtained if the fluid is changed after a few 

 hours. Chicks of the first and second days are hardened in two to four hours ; 

 embryos of 6 to 8 mm. in eight to ten hours; embryos of 12 mm. in twenty-four 

 hours ; larger embryos in thirty to forty hours. After the proper interval in 

 Zenker's fluid the specimens must be removed and washed in running water for 

 twelve to twenty-four hours. Transfer to 50 per cent, alcohol for one to three 

 hours, then to 60 per cent., 70 per cent., and 80 per cent. It is indispensable to 

 remove now the excess of corrosive sublimate by adding sufficient tincture of 

 iodine to give the alcohol the color of port wine; if the iodine disappears,. it must 

 be renewed. After from one to three days, according to the size of the specimen. 



