412 BIOLOGY OF PNEUMOCOCCUS 



with which the specific protective antibodies are associated, and 

 furnished the foundation for the future development of methods 

 for the isolation and concentration of the protective substance. 

 From the serum of horses immunized with pneumococci of Types I 

 and II over a period of one to two years, Avery was able, by add- 

 ing ammonium sulfate to 38 to 42 per cent saturation, to precipi- 

 tate completely the immune bodies. The protein fraction holding 

 the protective antibodies did not correspond to the euglobulin of 

 the serum since the fraction was not rendered insoluble by one- 

 third saturation with ammonium sulfate or by complete saturation 

 with sodium chloride. In 1924, Felton, 896 by the use of water acidu- 

 lated with tartaric acid, separated the bulk of protective sub- 

 stance from antipneumococcic serum of Types I, II, and III. The 

 precipitate appeared to be composed largely of globulin, which in 

 the state of purity obtained at the time by Felton had an isoelec- 

 tric zone between 6.6 and 7.5. 



Banzhaf, 70 by Felton's method of dialysis, separated approxi- 

 mately 90 per cent of the protective bodies from the fraction of 

 globulins precipitable between the limits of 30 and 50 per cent satu- 

 ration with ammonium sulfate. In further studies, Felton 397 " 400 ' 403 

 found that the water-insoluble globulin retained its protective 

 power after repeated purification, and that the globulin could be 

 largely freed from accompanying serum proteins through precipi- 

 tation with appropriate concentrations of sodium sulfate and by 

 ethyl alcohol. 408 The possible protein nature of the immune sub- 

 stance was suggested by the fact that its protective value was di- 

 minished by the digestive action of pepsin, trypsin, pancreatin, 

 and papain (Felton and Kauffmann, 1927). 426 



Goodner, 528 by the addition of appropriate amounts of water, 

 was able to precipitate the protective antibody as well as agglu- 

 tinins from antipneumococcic serum. The dilution of serum, after 

 preliminary tests to determine the proper amount of water to be 

 added, was carried out at a temperature of approximately 4°. 

 After carrying the precipitate through a refining process, the re- 



