406 



Mr. W. W. C. Topley. 



saccharose solution, are incubated for one hour at 37° C, they undergo no 

 haemolysis. 



(2) ]STo antibody capable of combining with red cells at 0° C. is involved, 

 since the supernatant fluid after one hour's absorption on ice is capable of 

 haemolysing fresh red cells. 



It is important to note that in this experiment the complement is not 

 appreciably affected by the dilution with salt-free saccharose solution at 0° C, 

 since when a tube containing this mixture was centrifugalised the supernatant 

 fluid was capable of haemolysing added red cells on further incubation at 

 37° C. 



A somewhat different result was, however, often obtained. While the 

 deposit from a tube containing red cells and complement in saccharose solution, 

 which had been kept for one hour at 0° C, never showed any trace of 

 haemolysis when suspended in fresh saccharose solution and incubated for a 

 second hour at 37° C, it frecpaently happened that the supernatant fluid 

 failed to produce more than a trace of haemolysis when incubated at 37° C. 

 with fresh corpuscles. In such cases it was always found that, if the tube 

 containing complement alone in saccharose solution was similarly kept for 

 one hour at 0° C, then centrifugalised and the supernatant fluid added to 

 fresh red cells, only a trace of haemolysis occurred upon further incubation at 

 37° C. It is therefore clear that the failure of lysis is due to the action 

 of the salt-free medium on the complementary serum, and not to any fixation 

 of the complement by the red cells in these cases. At the same time, it was 

 found that if the supernatant fluid from one of these tubes containing 

 complement only was added to the deposit from a tube which, during the 

 preliminary treatment at 0° C, had contained both red cells and complement, 

 then a degree of lysis resulted during the second hour's incubation at 37° C, 

 which corresponded with the haemolysis which occurred when a tube 

 containing red cells and complement in saccharose was incubated at 37° C, 

 or kept on ice for one hour and then shaken and incubated for another hour 

 at 37° C. 



This clearly indicates that some part of the serum has been precipitated 

 and is present with the red cells in the deposit. In other words a certain 

 degree of complement-splitting has taken place. 



From other experiments performed during this part of the investigation 

 this does not seem to be the whole explanation. It was found in many 

 cases that, when a tube containing complement alone in saccharose solution 

 was kept on ice for one hour and then well shaken, the contents failed to 

 hamiolyse fresh red cells, though still producing lysis of the deposit from 

 an iced saccharose tube containing red cells and complement. It seems clear, 



