FERTILIZATION REACTION IN ARBACIA PUNCTULATA. 259 



to stream forth. In about five minutes, the suspension was 

 strained through cheese cloth to remove the pieces of the ovaries. 

 The eggs were then allowed to settle. When they had settled, 

 the supernatant liquid was poured off and the dish again filled 

 with fresh sea water. This process was repeated three times so 

 that all traces of the coelomic fluid and the tissue juices were 

 practically eliminated. Eggs were not used unless the normally 

 fertilized control showed above 95 per cent, membrane formation 

 and a good viability through the first cleavage. 



Males were cut in the same way and placed in a Syracuse 

 glass with the genital pores down. The sperm flows from the 

 pores in a creamy mass, which, when mixed with sea water in 

 the proportion of one drop of dry sperm to 25 c.c. of sea water 

 gives a grayish, opalescent suspension which is described in this 

 paper as a 1 : 25 sperm suspension. One drop of this mixture 

 when added to one drop of eggs in 7.5 c.c. of sea water gives 

 100 per cent, membrane formation with no polyspermy and such 

 an insemination is spoken of as a I : 25 : 7.5. insemination. 



Concentrated stock solutions were made up as percentage of 

 metallic salt in distilled water by means of the volumetric flask. 

 The solutions used in the experiments were made up by the 

 addition of the stock solution to sea water. Care was taken 

 with those metals which required high concentrations, that the 

 proportion of sea water and stock solution in the mixture used 

 should not alter the osmotic pressure to any appreciable extent. 

 Solutions were not used for more than five days save in the case 

 of AuCl 3 and PtCl 4 where the rarity of the metal demanded the 

 use of the one solution. 



When an insemination is spoken of as immediate, i.e., made 

 in the solution itself, 7.5 c.c. of the solution was put into a 

 watch crystal and one drop of eggs and one drop of 1 : 25 sperm 

 suspension added at opposite sides of the dish. The dish was 

 immediately rotated to insure thorough mixing of the gametes. 

 In the viability tests, the eggs were normally inseminated in 

 sea water and one drop of these was then added to 7.5 c.c. of 

 the solution. The dish was then rotated as stated for the 

 immediate inseminations. 



A number of comparative tests were made in which, owing to 



