THE FERTILIZATION PROCESS IN THE SNAIL. 



ston's rubber-asphalt-paraffin, solutions of xylol and paraffin, 

 chloroform and paraffin, etc., followed by pure paraffin, and pure 

 paraffin alone were tried with varying degrees of success. It later 

 became evident that most of the standard methods of infiltrating 

 and imbedding tissues will give good results, if the tissue has been 

 properly prepared to receive the medium. 



The greatest difficulty encountered in imbedding the ovotestis 

 may be properly charged to the cavernous liver and its surround- 

 ing membrane. Once these structures are freed of the clearing 

 agent, infiltration is certain and complete if the tissue is left in 

 the medium sufficiently long. These difficulties may be overcome 

 by cutting the ovotestis into small pieces. 



It is very difficult to obtain desirable serial sections of eggs 

 which have been oviposited unless the albumen is properly treated 

 before dehydration and infiltration are attempted. After the 

 vitellus has been successfully fixed, enough of the albumen must 

 be removed to permit the best infiltration of both the vitellus and 

 the remaining albumen. Good serial sections of fertilization 

 figures were obtained by treating previously fixed whole eggs with 

 the following solutions : 2 per cent, potassium bichromate 100 cc., 

 0.5 per cent, chromic acid 100 cc., and nitric acid (C.P.) 6 cc. 

 After this treatment the egg membrane and much of the albumen 

 were cut away with a very sharp instrument made from a piece 

 of a safety razor blade. Later, the vitelli were simply blocked out 

 in this manner and the blocks imbedded without previous treat- 

 ment in the above solution. Neither of these methods is favorable 

 for the study of polar bodies and the former is the less favorable 

 for the study of chromatin. 



c. Staining. The difficulties in staining and studying the slides 

 are the result of the affinity of the polymorphic yolk granules for 

 nuclear stains. After using a number of stains, Heidenhain's 

 iron haematoxylin was adopted, alone or with Gage's acid fuchsin, 

 lichtgriin, eosin or some other counterstain. Benda's crystal 

 violet, gentian violet, Delafield's haematoxylin and safrranin were 

 used with indifferent success. 



II. COPULATION. 



Pond snails, especially L. s. apprcssa, copulate freely through- 

 out the year in laboratory cultures. On three different occasions 



