13 



The fact that the filtrates have a decided effect on leucocytes has 

 also been shown in a second experiment. 



Experiment. Two cubic centimeters of a rich leucocyitc exudate 

 were placed in each of five small test tubes and centrifugated. The clear, 

 supernatant' fluid was now decanted from each tube and replaced by 2 

 c. c. of bouillon, heated ascites-serum or filtrate from a 24-hour ascites- 

 serum streptococcus culture. The leucocytes in each tube were evenly 

 suspended in the fluid, the tubes placed in a water bath at 37 C. and kept 

 there for I to i l /2 hours. Now .3 c. c. or normal rabbit serum (guinea- 

 pig serum when guinea-pig's leucocytes were used) was added to each 

 tube, the tubes inoculated with a loopful of a non-virulent streptococcus 

 broth culture and plates made at intervals, the tubes being kept in the 

 incubator. 



The following table shows the results of an experiment: 



Immed. 4-5 hrs. 20 hrs. 



Leucocytes -f 2 c. c. filtrate B104 + .3 c. c. normal serum. .2000 3200 Many 



Leucocytes -f 2 c. e. filtrate 10 A + .3 c. c. normal serum . .1200 2000 Many 



Leucocytes + 2 c. c. filtrate 300 + .3 c. c. normal serum ..1100 24 Many 



Leucocytes + 2 c. c. ascites serum -j- .3 c. c. normal serum . .1000 140 Many 



Leucocytes -f- 2 c. c. bouillon -f- .3 c. c. normal serum 1800 GO Many 



The numbers B10-1, 10A and 300 refer to different strains of streptococci. 

 B104 has a high virulence and 10A a medium virulence; both these strains pro- 

 duce liemolysin in heated serum, although 10A has never been passed through 

 animals. It was recently isolated from suppurating axillary glands. 300 is 

 non-virulent, and does not produce hemolysin. 



The suspensions of leucocytes in filtrates from serum cultures of 

 virulent streptococci have no effect on streptococci while the suspension 

 in filtrates from non-virulent organisms, in heated ascites-serum, and in 

 bouillon destroy large numbers of cocci. 



These experiments do not always give positive results, but there is 

 a small percentage of tubes in which the filtrate has apparently no effect 

 on the leucocytes. This is not surprising, however, when we consider 

 the fact that in about .TOO filtrates which I have tested for hemolysis, 

 there were nearly 10 per cent, that had no hemolytic properties, whereas 

 some of them were so active that .005 c. c. completely laked i c. c. of a 

 2]/2 per cent, suspension of rabbit's washed corpuscles in two hours. 



TEST TUBE EXPERIMENTS WITH HUMAN LEUCOCYTES. 



The foregoing experiments show that the leucocytes and opsonin are 

 the most important, if not the only, factors concerned in the destruction of 

 streptococci in the body of infected rabbits and guinea-pigs. In strepto- 



