APPENDIX. 



633 



with good penetrating power. Fixation is complete in from 24 to 48 hours Wash in run- 

 ning water a few hours, then transfer to graded alcohols. The tissues take a good differen- 

 tial stain (haematoxylin and eosin). 



8. Perenyi's fluid. 



Nitric acid, 10 per cent 4 parts. 



Alcohol 3 parts- 



Chromic acid, 0.5 per cent , parts. 



Small embryos are fixed in from 6 to 12 hours. Wash in several changes of 70 per cent, 

 alcohol (24 hours) and preserve in 80 per cent, alcohol. This fixative is claimed to be espe- 

 cially good for segmenting ova. 



9. Zenker's fluid. 



Potassium bichromate 2.5 grams. 



Sodium sulphate i gram. 



Mercuric chloride 5 grams. 



Glacial acetic acid 5 c.c. 



Water 100 c .c. 



This should be freshly made or the acetic acid added to a stock solution of the other ingre- 

 dients just before using. This is a good fixative for both nuclei and cytoplasm, but has a 

 tendency to cause shrinkage. Fixation of small embryos (up to 20 mm.) is complete in from 

 12 to 24 hours. Wash in running water for 12 hours and transfer to 70 per cent, or 80 per 

 cent, alcohol. The mercuric chloride always forms precipitates in the tissues, but these 

 can usually be removed by adding a little tincture of iodine to the alcohol from time to time 

 until the alcohol retains the iodine color. Then change to fresh alcohol. 



III. HARDENING. 



.Although most fixatives tend to harden the tissues, it is almost always necessary to 

 harden them further. For this purpose alcohol is used. With delicate tissues it is advisa- 

 ble to use grades of 30, 50, 70, and 80 per cent., leaving the specimen in each grade for 

 several hours. 



IV. PRESERVATION. 



Alcohol (70 to 80 per cent.) is much used as a preserving fluid. After several months, how- 

 ever, tissues are likely to lose their staining qualities to some extent. A better preserving 

 fluid, in which specimens may remain almost indefinitely, is made of equal parts alcohol (95 

 per cent.) glycerin and water. Objects fixed in formalin may be preserved in the formalin 

 and then passed through the graded alcohols in preparation for embedding. 



V. EMBEDDING. 



Paraffin should be used for small objects or when serial sections are to be cut. Celloidin 

 is very convenient for larger specimens, especially when a large number of sections is to be 

 cut for class purposes. 



Celloidin embedding. Place the hardened specimen in 95 per cent, alcohol for 24 hours, 

 and then in equal parts alcohol (95 per cent.) and ether for the same length of time. Put 

 into thin celloidin (5 per cent, solution of celloidin in equal parts alcohol and ether) for sev- 

 eral days. Then put into thick celloidin (10 per cent, celloidin in equal parts alcohol and 

 ether) for 24 hours or longer. The time objects should remain in the different solutions 



