SPERM AGGLUTINATION AND FERTILIZATION. 27 



It is highly improbable that the agglutinating substance had been 

 destroyed in five of the six, and retained in the only one (No. 5) 

 of the extracts which was made from ovaries containing some 

 ripe ova. So far as these observations go, the jelly of immature 

 ovocytes is free of agglutinating substance. 



Again on June 15 I made extracts from ovaries of three females 

 in two of which ripe ova were practically absent, the third had 

 a few. Tested the same day the two former extracts had no 

 sperm agglutinating properties; the third gave slight agglu- 

 tination. 



The females appeared to mature slightly earlier than the males, 

 so that for these experiments I was forced to use rather thin 

 sperm suspensions (mixed more or less with immature spermato- 

 zoa) , which were probably not as delicate indicators as one could 

 wish. However the difference between the ovaries containing 

 ripe ova and those without was perfectly distinct. Later when 

 fully ripe males could be had all ovaries contained ripe ova. 



The following observation also tends in the same direction: 

 June 27, 1914 — Three females were selected, of which number i 

 was the ripest attainable, the eggs flowing freely out of detached 

 ovaries, and very few ovocytes occur; numbers 2 and 3 were the 

 least mature attainable; number 2 had very few detachable ova, 

 mostly late ovocytes with a sprinkling of ripe eggs; number 3 

 had quite a few detachable ova with a large proportion of ripe 

 eggs. The ovaries of all three were cut up equally, and sea-water 

 added to each to make 10 c.c. When the ova and ovaries had 

 settled they stood at 1.5 c.c. in i, at 1.3 c.c. in 2, and 1.5 c.c. in 3. 

 After five hours, tests of the agglutinating strength of the super- 

 natant fluids were made with clear fresh sperm. 



No. I gave a lo-second reaction at 1/800 dilution. 



No. 2 gave a 6-second reaction at i/io dilution. 



No. 3 gave a 7-second reaction at 1/40 dilution. 

 Thus No. I is 80 times the strength of 2 and 20 times the strength 

 of 3. In general the fertilizin production is proportional to the 

 ripeness of the ovaries. 



There is not the slightest doubt in my mind about the demon- 

 strative character of these observations. The appearance of 

 agglutinating substance in the jelly of Arbacia eggs is secondary, 



