270 STUDIES IN IMMUNITY. 



Tube c. Rabbit alexin 2 c.c; normal rabbit serum, 55 degrees, 2 c.c. (Same 

 as "b" without sensitizer.) 



To each tube is added one drop of hen blood. Hemolysis occurs rapidly in " a " 

 and "b." It is only partial in "c" after 3 hours. It is evident, then, that the 

 anti-alexin can have no effect on the sensitizer, which, in the experiment, is only 

 one-twentieth of the volume of the antiserum. It is evident, too, that the sensitizer 

 acts energetically although present in small amount and diluted in 4 c.c. of fluid. 

 In the rest of the article, for the sake of simplicity, these control experiments need 

 not be insisted on in each instance. 



We then determine the potency of the anti-alexin. For this 

 purpose varying amounts of alexin (say from 0.05 to 1.2 c.c.) are 

 added to a constant volume (0.3 of a cubic centimeter) of anti- 

 alexic or of normal serum (55 degrees). The following tubes are 

 prepared : 



A. Tubes containing 0.3 of a cubic centimeter of anti-alexin: 

 a. Alexin, 0.05 c.c.; 6. Alexin, 0.1 c.c.; c. 0.2 c.c.; d. 0.3 c.c.; 



e. 0.4 c.c.; /. 0.5 c.c.; g. 0.6 c.c.: h. 0.7 c.c.; i. 0.8 c.c.; j. 0.9 c.c.; 

 k. 1.2 c.c. 



B. Same as preceding tubes, with heated normal rabbit serum 

 (non-anti-alexic) replacing anti-alexin. 



To each tube is then added 0.2 of a cubic centimeter of sensitizer 

 and an hour later one drop of a suspension of hen corpuscles in 

 NaCl. The mixtures are kept at room temperature (18 C.). 



In the tubes that do not contain anti-alexin, hemolysis takes 

 place rapidly in, say, 15 minutes in those tubes containing most 

 alexin, and in 30 minutes in the one with the smallest amount. 

 Consequently 0.05 of a cubic centimeter of alexin represents the 

 minimal dose for hemolysis in a half hour, which may be taken as 

 a unit. We may add that the dilution of this small amount of 

 alexin in heated serum, one volume of alexin in ten of heated serum, 

 for the sake of mensuration has no effect on its activity. 



We may now consider the tubes containing anti-alexin. Let 

 us consider first the final reading, say on the following day, in order 

 to allow for the maximal action of the smallest amounts 'of free 

 alexin. On the following day hemolysis is complete in the tubes 

 containing 0.5 of a cubic centimeter or more of alexin; it is partial 

 in the 0.4 c.c. tube and slight in the 0.3 c.c. tube. Two-tenths of 

 a cubic centimeter or less shows no hemolysis. 



It may be noted that such results argue somewhat against the 

 hypothesis of a combination in fixed proportion. If there is slight 



