STUDIES OF FERTILIZATION. 135 



polyspermy. It is not necessarily the first spermatozoon that 

 agglutinates that effects the initial discharge, but this will depend 

 on physiological condition of the spermatozoon, and also perhaps 

 on local differences of physiological condition on the surface of 

 the egg. 



5. The Effect of Copper on Agglutination of Spermatozoa. 



The objection might be raised that copper inhibits by pre- 

 venting agglutination of the spermatozoon to the egg, and not 

 in the later activation of the fertilizin. The effect of copper on 

 the agglutination reaction of the spermatozoa was therefore 

 studied: Egg-water (of 1,600 agglutinating units) diluted 100 

 times or more with 1/500,000 copper chloride in sea-water has 

 the same agglutinating effect on a sperm suspension in the same 

 copper solution as when no copper is present either in the egg- 

 water or sperm suspension. Agglutination will occur even in 

 the presence of one part of copper chloride to 25,000 parts of 

 sea-water, though more slowly. Copper chloride in the concen- 

 trations employed in the fertilization experiments has no notice- 

 able effect on rate or duration of the agglutinating reaction. 



The inhibiting action of copper must then occur after the 

 agglutination of the spermatozoon to the egg, as was assumed 

 in the preceding discussion. 



6. Protective Action of Egg-Water Against Copper. 



If copper chloride inhibits by combining with the fertilizin 

 of the egg, then egg-water which contains fertilizin should protect 

 against the inhibition of fertilization by deviating the copper 

 from the fertilizin in the egg to that in the egg-water. Now it 

 is known that egg-water contains a sperm-activating, a sperm- 

 aggregating and a sperm-agglutinating substance (Lillie, '14, '19), 

 and I have identified the sperm-agglutinating substance with 

 fertilizin. These considerations led to experiments on the pro- 

 tective action of egg- water on the copper inhibition, in which 

 definite positive results were obtained. 



The egg-water is obtained by placing eggs in sea-water which 

 receives their secretions. The strength of the egg-water depends 

 on egg-concentration primarily, and also on time to a certain 

 extent; it may be measured, as far as the sperm-agglutinating 



