560 FRANK R. LILLIE 



large quantities and the eggs then appear to have disposed of their 

 entire store, for 'washed fertihzed eggs no longer produce it. The 

 conditions are important in their bearing upon the question of 

 permeability of the egg-membrane with reference to fertilization. 

 The spermatozoa are very efficient indicators of substances leav- 

 ing the egg. In the case of Nereis it can be shown that there is a 

 sudden increase of permeability at the moment of fertilization, 

 but in Arbacia such evidence is lacking. 



The conditions in Arbacia may be shown by the following experi- 

 ment : Eggs were washed free from all fragments of ovary and placed in 

 about 20 times their own bulk of sea-water and divided in three lots, 

 a, h and c. The sea-water over them agglutinates Arbacia sperm in- 

 stantaneously. This test was made at 9.30 a.m. The supernatant fluid 

 was then removed and the eggs washed in 20 times their bulk of sea- 

 water. At 9.53 the supernMant water was tested and found agglutina- 

 tive. 9.58, a and b were again washed. 10.04, supernatant fluid again 

 agglutinative. 10.05, lot a was fertilized with 2 drops of sperm. 10.10, 

 a and b tested again; both agglutinate. 10.16, a and b washed again. 

 10.23, supernatant fluid of both agglutinates. 10.30, a and b washed 

 again. 10.36, both agglutinate sperm; b is rather more effective. 10.40, 

 washed a and b again. 10.45, both agglutinate; b more effective. 10.51, 

 washed a and b again. 11.00, both agglutinative; a more than b. 11.08 

 to 1 1.20, other tests of a and b show a somewhat more effective. 



The experiment shows both fertilized and unfertilized eggs of 

 Arbacia to be constantly secreting a substance into the sea-water 

 which agglutinates the spermatozoa. The substance must be 

 effective in very minute quantities for the amount of water used 

 in each of the eight washings was at least ten times the bulk of the 

 eggs, yet as soon as the eggs had settled to the bottom of the vials 

 used the supernatant fluid contained the agglutinin in appre- 

 ciable amounts. How long this process keeps up in Arbacia I 

 cannot say; and too much reliance cannot be placed on the result 

 for the fertilized eggs because a large proportion of the eggs failed 

 to fertilize or at least to segment. 



The conditions in Nereis are quite different; the experiments 

 showed that shortly after the eggs are taken they charge at least 

 ten times their bulk of sea-water with an easily detectable amount 

 of sperm agglutinin. But if the eggs are washed once usually no 

 more agglutinin canlDe demonstrated. If now the eggs are stirred 



