8 BULLETIN 340, IT. S. DEPARTMENT OP AGRICULTURE. 



serum has an extraordinary potency. It should be noted that it 

 does not follow that those rabbits which receive the smallest serum 

 doses should die, since not infrequently they may remain- alive when 

 the rabbits receiving larger closes succumb. 



This method of standardization has not been proved as accurate 

 and reliable as the test recommended by Ascoli, and which has been 

 employed in the experimental work with serum prepared in con- 

 nection with our experiments. In this test a 24-hour-old attenuated 

 bouillon culture is used, which is of such virulence that when intro- 

 duced subcutaneously in a 0.25 c. c. dose into 350-gram guinea pigs 

 it will kill them in from two to three days. These test cultures must 

 be previously standardized in such a way that they will kill guinea 

 pigs which 24 hours previously have been injected intraperitoneally 

 with 2 c. c. of normal serum. Guinea pigs treated in the same man- 

 ner and with the same dose of titrated standardized immune blood 

 serum must remain alive. 



The testing of the serum is carried out on six guinea pigs, eacji 

 receiving intraperitoneally 2 c. c. of the serum to be tested, and 24 

 hours later the established dose of the test culture is injected sub- 

 cutaneously in the axillary region. The serum is considered satis- 

 factory for immunization purposes if at least four of the guinea pigs 

 remain alive over six days while the control animals die within 

 three or four days. For protective and curative purposes in man, 

 only such serum should be selected which, by carrying out the same 

 conditions of the test, protect the guinea pig in 0.5 to 1 c. c. doses. 



EXPERIMENTAL DATA. 



HYPERIMMUNIZATION OF HORSES. 



On September 8, 1914, two horses, Nos. 48 and 96, were vaccinated 

 against anthrax according to Pasteur's method. On September 29 

 these two horses were given approximately 0.01 of a loopful of 

 virulent anthrax bacilli subcutaneously. Horse No. 48 showed no 

 apparent reaction following the injection. Horse No. 96, however, 

 developed local anthrax at the point of inoculation. The swelling 

 became enlarged and there was a considerable area of edema below 

 the same. This condition persisted for approximately a week, and 

 finally disappeared. The animal, however, showed no appreciable 

 rise in temperature during this period. 



The following table gives in detail the process of hyperimmuni- 

 zation : 



