528 



SCIENCE 



[N. S. Vol. XXXVI. No. 929 



ways: (1) In a test tube or vial containing 2 

 or 3 c.c. of a milky sperm suspension, as above 

 described. (2) Drops of the sperm suspension 

 and of the egg-extract may be placed side by 

 side on a slide and connected under the micro- 

 scope, so as to observe the inception and prog- 

 ress of the phenomenon. (3) The best method 

 of studying the detail of the phenomenon is 

 to mount some of the sperm suspension be- 

 neath a long cover glass supported by glass 

 rods 0.5 to 1 mm. in diameter, and to inject a 

 drop of the egg-extract with a capillary pipette 

 into the suspension. If this be observed 

 under a low power of the microscope, one sees 

 that the drop fills with sperm exhibiting in- 

 crease of activity, which immediately agglu- 

 tinate into small masses, which then unite 

 with great rapidity to form larger masses 

 spaced throughout the drop. At the margin 

 of the drop and the sperm suspension a ring 

 of agglutinated sperms forms which ruptures 

 in numerous places, each segment contracting 

 to form a mass, so that the continuous ring 

 becomes a chain of beads visible to the naked 

 eye. The ring forms in a second or two and 

 breaks into masses in two or three seconds. 

 The agglutination is totally reversible, so far 

 as the visible effect is concerned, and the freed 

 spermatozoa, or some of them, appear to re- 

 gain full activity; moreover, the spermatozoa 

 between the masses are in active movement 

 throughout. 



The reversal of the reaction consists in the 

 resolution of the agglutinated masses into 

 their individual cells, but this does not mean 

 a return to their original physiological condi- 

 tion; for if the agglutinating solution (egg- 

 extract) be sufficiently strong, after a period 

 of intense stimulation followed by agglutina- 

 tion and reversal, the movements of the sper- 

 matozoa gradually cease entirely and in ten 

 minutes they are no longer capable of fertil- 

 izing ova dropped in with them. 



The agglutination is a factor of (1) the 

 activity and relative density of the spermato- 

 zoa in the suspension, and (2) of the strength 

 of the agglutinating solution. The more 

 active the sperms and the greater the density 

 of the suspension (within certain limits) the 



more rapid is the onset of the agglutination 

 and the larger the agglutinated masses. The 

 agglutinating medium may of course be made 

 to vary in strength, and hence in agglutina- 

 ting power by various means, e. g., by crushing 

 ovaries and eggs in about their own bulk of 

 sea-water, or extracting in distilled water, very 

 powerful solutions may be made. 



The agglutinating agent is not readily de- 

 stroyed by heat. After boiling and maintain- 

 ing at a temperature of 95° to 98° C. for about 

 70 minutes, the agglutinating action of a 

 strong extract was very much reduced but 

 was not entirely lost. 



No other tissue of the sea-urchin, so far as 

 observed, produces an agglutinating agent for 

 the sperm of the species : (1) The serum from 

 the body-cavity, whether of males or females, 

 is entirely neutral and the spermatozoa swim 

 in it normally. But the serum forms a power- 

 ful agent for extracting the agglutinin from 

 the eggs after the latter are removed from 

 the ovary, though in the intact animal in 

 which the eggs are separated from the serum 

 by the ovarian membrane no extraction of 

 agglutinin takes place. (2) Large pieces of 

 the intestine were cut up in sea-water or in 

 distilled water, the latter especially extracting 

 colored matters in large quantity; but no 

 sperm agglutinin could be detected in the 

 filtrate. 



The agglutination reaction fixes the agglu- 

 tinin, as is readily proved by the fact that it 

 disappears from an agglutinated sperm sus- 

 pension, if not present in excess. Whereas an 

 equal dilution with sea-water of the original 

 egg-extract is still highly agglutinative. The 

 agglutinative substance, therefore, presumably 

 enters into combination with some sperm sub- 

 stance acting in this respect similarly to its 

 antigen. 



II. The results are essentially the same for 

 Nereis, except that the visible agglutination 

 is not reversible to the same extent, a differ- 

 ence which is probably of purely secondary 

 significance. Moreover, the agglutinin is pro- 

 duced by the eggs in sea-water only in small 

 quantities before fertilization, but in very 

 large quantities at the moment of fertiliza- 



