554 FRANK R. LILLIE 



In the case of Nereis it was not possible to reach such conclu- 

 sive results, because the sexually mature female is practically a 

 bag of eggs, and one cannot obtain other organs for testing. I 

 cut up five females that had shed their eggs in 10 cc. of sea- water. 

 In spite of efforts to get rid of all eggs, a considerable number 

 were in the water. For control I used the eggs of two females in 

 100 cc. sea-water. On test in half-an-hour the fluid from above 

 the eggs was found to be about ten times as agglutinative as the 

 fluid from the bodies of the spent females. So that it is certain 

 that other tissues do not produce much sperm agglutinin and it is 

 probable that they do not produce any. The small amount 

 present could be accounted for by the few eggs included, and per- ' 

 haps by egg secretions absorbed by the tissues. 



4. FIXATION OF THE AGGLUTININ BY SPERMATOZOA 



The next question was whether the agglutination reaction as 

 described has the usual characters of a chemical reaction? The 

 general result is (1) that an agglutinated sperm suspension in 

 which reversal has occurred is not capable of re-agglutination by 

 addition of more of the agglutinating substance and (2) that 

 the agglutinating substance disappears from an agglutinated 

 suspension if not present originally in excess. 



As regards the first point, the earlier experiments were con- 

 cerned entirely with the form and conditions of the reaction, and 

 the agglutinating substance was always used, as later results 

 showed, in excess. It was not possible to get a repetition of the 

 agglutination reaction under these circumstances. But one can 

 get a repetition of the reaction in a sperm suspension by addition 

 of successive small amounts of the agglutinating substance, until 

 the reaction is complete, as the following experiment shows: 



September 4, 1912. Arbacia. 2 cc. of a creamy active sperm suspension 

 was agglutinated with 5 drops of an egg-extract prepared as follows : The 

 ovaries of three females were cut up in about three times their volume of 

 distilled water and allowed to stand about thirty minutes. Then the 

 water was filtered off and made isotonic with sea-water by the addition 

 of concentrated sea-water (proportions of 58 to 42 parts) . This made a 

 very strong agglutinating extract. After reversal of the agglutination 

 described above the agglutination was repeated by addition of a drop of 



