432 



PATHOGENIC MICROORGANISMS 



quoted above. 102 .In order to do protection experiments properly, 

 it is necessary, in the first place, to produce powerful serum; in 

 the second place, it is important to raise the virulence of the test 

 strain to a high point. We may state from our own experience 

 that not all streptococci can be raised to the same degree of potency 

 for mice. Dochez and Avery raised the virulence of a number of 

 strains, by continuous mouse passage, to a point at which 0.000001 

 to 0.00000001 c.c. of a broth culture would kill the animals in twenty- 

 four to forty-eight hours. Workers in our own laboratory have 

 occasionally been able to do this with individual strains, but most 

 strains could not be raised to a virulence approaching this. The 

 organisms are best grown in ascitic broth. 0.5 c.c. of serum is 

 injected intraperitoneally, according to the technique of Neufeld, 

 twenty-four hours before injection of the organisms. On the follow- 

 ing day, a series of virulence controls is inoculated intraperitoneally 

 with appropriate quantities of organisms, and the serum animals 

 are injected, intraperitoneally, with doses of twenty-four-hour cul- 

 tures ranging from 0.001 to 0.00000001 c.c. (though these quantities 

 must be adapted to the known virulence of the strain). Animals 

 that survive for a period of five days are considered by Dochez and 

 Avery as sufficiently protected. The following protocol, taken from 

 the paper by Dochez, Avery and Lancefield, illustrates the method. 



PROTOCOL I 



In this protocol is shown the titration of the serum of a sheep immunized against 

 Strain 23. The culture employed for infection was an 18-hour broth culture of 

 No. S 23, which had received eighteen passages through white rats and mice. Each 

 mouse had received 0.5 c.c. of immune serum 24 hours previous to infection. 



VIRULENCE CONTROL 



PROTECTIVE POWER OF SERUM S 23 



In the tables D. indicates died, S. survived. 



10 - Dochez, Avery and Lancefield, Jour. Exper. Med., 30, 1919, 179. 



