THE NATURE OF HEMOLYSINS 101 



bring about an optimum surface for the adsorption of the anti- 

 gen. The variations in the quantity of antigen necessary to 

 produce hemolysin in the various broths used bear out the impor- 

 tance of the colloidal properties of the menstruum to which the 

 artificial antigen was added, and the fact that broth that has 

 been passed through a Berkefeld filter requires the addition of 

 more antigen than the unfiltered broth to make it hemolytic 

 seems also to emphasize the importance of surface in the pro- 

 duction of artificial hemolysin. 



Another point that seems at first sight to afford a distinction 

 between the natural and artificial hemolysin is the clouding 

 that occurs with certain doses of the antigen. This variation is 

 however only apparent and can be avoided by emulsification of 

 the antigen before its addition to the broth, or by the presence 

 in the broth of just the proper surface. at the time of the addi- 

 tion of the antigen. Clouding depends in part on the rate at 

 which the antigen is added and upon the manner of adding it — 

 a considerably larger amount can be introduced without forma- 

 tion of a cloud if the emulsification be made drop by drop slowly 

 and with constant gentle motion. 



The points in which the natural and artificial antigens resemble 

 each other are as follows (a) both are comparably hemolytic; 

 (b) both are inactivated by heat at approximately the same 

 temperatures. The natural lysin of B. megatherium is usually 

 inactivated by heating to 56°C. for thirty minutes, though in 

 some specimens it was found to require 60°C. for the same 

 length of time, while the artificial hemolysin when containing 

 40 to '60 mgm. of antigen per liter is inactivated at from 60° to 

 65°C. for one-half hour. These temperatures are sufficiently 

 close together for the discrepancy to be accounted for by the 

 crudeness of the artificial methods. Streptolysin was found to 

 inactivate at 70°C. for two hours by Besredka and by Ruediger 

 when serum medium was used and then diluted with salt solu- 

 tion before being passed through a Berkefeld filter. In our 

 serum-broth medium the streptolysin was inactivated at 65°C. 

 for one-half hour. We are forced to believe that the colloidal 

 state of the medium has considerable effect on the temperature 



