The Development of Phascolosoma. 149 



When egg--laying- begins, great care should be taken to avoid 

 an excess of sperm. When a shower of eggs is thrown out by a 

 female, it is well to catch them in a small dish before they have 

 time to settle, and to transfer them into a large quantity of fresh 

 sea- water. Otherwise, if the water is white with sperm, superfetation 

 is likely to occur, or the embryos later become covered with a 

 growth of bacteria. 



Artificial Fertilization. Large individuals should be 

 opened, and the contents of the coelom of each allowed to fall into 

 a small vessel of sea-water. When a female with an abundance of 

 eggs is found, the maturer oocj^tes should be allowed to settle to 

 the bottom, whereas the smaller oocytes and coelomic corpuscles 

 should be decanted after a few seconds, and before they have had 

 time to sink. After several washings, the eggs will be ready for 

 fertilization. 



The vessels into which the coelomic fluid of males have fallen 

 should stand for several minutes, until the coelomic corpuscles have 

 settled. A small quantity of water is then to be poured from the 

 top of each vessel that proves on examination to contain active 

 spermatozoa. This mixture of active sperm from several individuals 

 should be added sparingly to the dishes which contain ova. In a 

 successful fertilization a very small proportion of the eggs will be 

 found to develop. 



By this method, one is able to study in the daytime the clea- 

 vage of the beautifully transparent egg of Ph. vulgare. Owing to 

 the difficulty in distinguishing with certainty the poles of the egg 

 after the four-cell period, I was obliged to begin each series of 

 observations with this stage. This makes it necessary to keep the 

 egg under continuous observation for five or six hours, in order to 

 carry the cleavage to the 48-cell stage. I did this by encircling 

 the eggs with a soft thread which supported the cover glass, while 

 a free end served as a wick to supply fresh seawater to replace 

 that which was lost by evaporation. Eggs thus treated live only 

 four or five hours, owing to the increasing salinity of the sea-water 

 in the preparation. Ziegler's compressorium, or some similar appa- 

 ratus, is therefore desirable for observing the later stages. 



For cytological work in which it is desirable to have a large 

 proportion of the eggs fertilized, it is necessary to await a deposit 

 of mature eggs by the female, or to secure an individual in wliich 

 the nephridia are distended with eggs. I have frequently found, 



