The Influence o f Salt- Concentration on Haemolysis. 399 



In considering the results obtained during this part of the investigation, it 

 is necessary to bear in mind the action of hypotonic or salt-free media on 

 complementary sera, especially the phenomenon known as complement- 

 splitting. 



An observation made by Sachs and Terruuchi in 1907, whilst studying the 

 inactivation of complement in a salt-free medium, has an important bearing 

 on the problem in hand. These observers found that guinea-pig serum alone 

 was capable of causing a more or less marked degree of lysis of ox corpuscles 

 in a salt-free medium, made isotonic by the addition of 7*8 per cent. ' 

 saccharose, while the same serum had no lytic action in normal saline 

 solution. They were not, however, prepared to admit that this phenomenon 

 was really comparable to the haemolysis which takes place in a mixture of 

 red cells, haemolytic antibody and complement. They found that, while the 

 complement alone produced haemolysis in saccharose solutions, but not in 

 normal saline, complement acting together with a specific haemolysin always 

 produced more lysis in saline than in saccharose solution ; and, in some cases, 

 lysis was entirely absent when a complete haeinolytic system was allowed to 

 act in a saccharose medium, but well marked when the same system inter- 

 acted in normal saline solution. They noted, however, that inactivation for 

 30 minutes at 55° C. destroyed the power of the complement to produce this 

 abnormal haemolysis, and suggested that it might be due to " a peculiarity of 

 the normal haemolysin." 



A large number of experiments were undertaken in order to further 

 investigate this phenomenon. It is only necessary to present here the main 

 results obtained; but -it should be noted that different specimens of serum 

 obtained from presumably normal guinea-pigs gave very varying results, 

 while a single given specimen often showed marked changes within 2-4 to 

 48 hours though stored on ice. 



The specimens of serum vary in two ways, in their power of inducing lysis 

 in salt-free media and in their resistance to the anti-complementary action of 

 such media ; and these two variations do not run parallel. Thus, a given 

 specimen of serum may be actively lytic in a salt-free medium, but "be easily 

 inactivated by this same medium in the absence of red cells, while another 

 specimen may be feebly lytic but suffer only a slight degree of inactivation, a 

 third being actively lytic and markedly resistant to the inactivating action of 

 the salt-free medium. 



The specimens of guinea-pig serum used in these experiments were absorbed 

 at 0° C. with sheep's red cells for a period varying in different experiments 

 from one to two hours, in order to remove any trace of normal haemolysin 

 that might be present. It was found that the great majority of sera so treated 



