GENERAL PHENOMENA OF IMMUNITY 23 



bitten by a small number of infected ticks or by injecting intraven- 

 ously a small amount of blood from an infected animal. 



Somewhat similar to the above examples is infection with attenu- 

 ated virus. Such attenuation may be obtained by prolonged cultiva- 

 tion on artificial media, by heat, by passage through animals, by 

 desiccation, by the use of chemical agents, and by pressure. If heat 

 be employed for attenuation, rather than for killing the organisms, 

 it must be properly adjusted. Toussaint employed this method in 

 his early experiments with anthrax in which he heated infected 

 blood to 55 C. for ten minutes. This method, however, is not re- 

 liable, probably because of variations in the resistance of individual 

 members of a culture of any given organism. Heat may be applied 

 also during the cultivation of organisms upon artificial media, a 

 method practised by Pasteur in producing anthrax vaccine. The 

 heat must be of such a degree as to permit growth of the organisms, 

 but at the same time reduce the virulence. As has been pointed 

 out before, the cultivation of organisms upon artificial media through 

 many generations leads to a reduction of virulence. This latter 

 method was employed by Pasteur in the development of the vaccine 

 for chicken-cholera. The attenuation of smallpox virus by passage 

 through the calf so reduces virulence that the virus may safely be 

 inoculated into man. Pasteur found that the virus of swine ery- 

 sipelas could be attenuated by passage through rabbits, and it is 

 well known that the passage of rabies virus through dogs and 

 through monkeys reduces its virulence. An excellent example of 

 attenuation by desiccation is found in the preparation of anti-rabic 

 vaccine. For this purpose the virus is raised by passage through 

 rabbits to a standard degree of virulence, the " virus fixe." The 

 spinal cord of a rabbit so infected is desiccated at 25 C. over KOH. 

 This method of attenuation is so delicate that there are distinct 

 variations in virulence between fragments dried for five, six, and 

 seven days, as well as virus dried for thirty-five, thirty-six, and 

 thirty-seven days or intervening periods. The longer the desicca- 

 tion the greater the reduction of virulence and the greater the safety 

 of the inoculation. Attenuation by the use of chemicals, such as 

 phenol, potassium bichromate, and sulphuric acid, has been prac- 

 tised. Chemical attenuation may also be applied to toxins, as in the 

 use of iodine terchloride and potassium iodide. A pressure of eight 

 atmospheres at a temperature of 28 to 39 C. has been employed 

 for the attenuation of anthrax cultures, but is probably not widely 

 applicable, is difficult, and possesses no superior advantages. 



Immunization with Dead Bacteria. In the study of immune 

 processes it was finally found that killed bacteria could be used for the 

 production of immunity. The organisms may be killed by heat or 

 by chemicals. In either case, it is necessary so to apply these agents 

 as to kill the organisms without destroying their proteins. The 

 use of heat sufficiently high to destroy spores leads to destruction 

 also of the proteins, and therefore the method does not apply to 



