EXPERIMENTS TO SHOW THE NATURE OF HAMOLYSINS 541 



cells which had been separated by the centrifuge were mixed with a 

 little normal salt solution after freeing them as much as possible from 

 fluid. Then a little alexin in the form of normal serum was added. 

 After remaining thus for two hours at 37 C. these cells had all 

 dissolved. 



In this experiment, therefore, the red cells had combined with all 

 the substance sensibilatrice, entirely freeing the serum of the same. 

 That the action was a chemical one, and not a mere absorption, was 

 shown by the fact that red blood cells of other animals, rabbits or goats 

 for example, exerted no combining power at all when used instead of 

 the sheep cells in the above experiment. The union of these cells, more- 

 over, is such a firm one that repeated washing of the cells with normal 

 salt solution does not break it up. 



The second important question solved by these authors was this: 

 What relation does the alexin bear to the red cells? They studied this 

 by means of a series of experiments similar to the preceding. 



EXPERIMENT 2. Sheep blood was mixed with normal i. e., not 

 hsemolytic goat serum. After a time the mixture was centrifuged and 

 the two portions tested with the substance sensibilatrice to determine the 

 presence of alexin. It was found that in this case the red cells acted 

 quite differently. In direct contrast to their behavior toward the sub- 

 stance sensibilatrice in the first experiment, they now did not combine 

 with even the smallest portion of alexin, and remained absolutely 

 unchanged. 



EXPERIMENT 3. The third series of experiments was undertaken 

 to show what relations existed between the blood cells on the one hand 

 and the substance sensibilatrice and the alexin on the other, when both 

 were present at the same time, and not, as in the other experiments, when 

 they were present separately. This investigation was complicated by the 

 fact that the specific immune serum very rapidly dissolves the red cells 

 for which it is specific, and that any prolonged contact between the 

 cells and the serum, in order to effect binding of the substance sensi- 

 bilatrice, is out of the question. Ehrlich and Morgenroth found that 

 at C. no solution of the red cells by the hremolytic serum takes place. 

 They therefore mixed some of their specific hsemolytic serum with sheep 

 blood cells, and kept this mixture at to 3 C. for several hours. No 

 solution took place. They now centrifuged and tested both the sedi- 

 mented red cells and the clear supernatant serum. It was found that 

 at the temperature to 3 C. the red cells had combined with all of the 

 substance sensibilatrice, but had left the alexin practically untouched. 



It still remained to show the relation of these two substances to the 

 red cells at higher temperatures. At 37 to 40 C., as already mentioned, 

 haemolysis ocpurs rapidly, beginning usually within fifteen minutes. 

 It was possible, therefore, to leave the cells and serum in contact for 

 not over ten minutes. Then the mixture was centrifuged as before. 

 The sedimented blood cells mixed with normal salt solution showed 

 haemolysis of a moderate degree. The solution became complete when 

 a little normal serum \va> added. The supernatant clear fluid separated 



