PRACTICAL APPLICATIONS 40! 



1884, ver y soon after the discovery of the V. cholera by Koch. 

 His results were of doubtful value, his vaccines being made of 

 cultures of feeble virulence, and perhaps impure. The method 

 was placed on a scientific basis by Haffkine, who showed the 

 necessity for the use of cultures of great virulence. These are 

 prepared by passage through guinea-pigs. A more than lethal 

 dose of a laboratory culture is injected into the peritoneum of a 

 guinea-pig, and the peritoneal fluid (rich in vibrios) is collected 

 after death. This fluid is incubated in a thin layer, so as to allow 

 of thorough aeration, for fifteen hours, and is then administered 

 intraperitoneally into a second animal. After about twenty or 

 thirty passages the culture will have attained its maximum 

 virulence, the lethal dose being some -^ of the original. Its 

 potency falls off in some ten days, and a few further passages are 

 required to restore it. 



The treatment is commenced by a dose of attenuated virus. 

 This is prepared by cultivating an ordinary laboratory stock in 

 broth at 39 C. in conditions of complete aeration. An inocula- 

 tion on agar is made every day, until (after a few days) the fluid 

 is found to be sterile. The process is now recommenced, using 

 the last agar culture that grew, and after several generations a 

 culture of very feeble virulence is obtained. It causes oedema, 

 but no necrosis, when injected under the skin. The vaccines are 

 prepared by cultivating the organisms on agar slants of definite 

 size (10 centimetres long) for twenty- four hours, and emulsify- 

 ing with 8 c.c. of broth, or 6 c.c. of 0*5 per cent, solution of 

 carbolic acid. The dose is i c.c. One or two injections of the 

 attenuated virus, followed by one of the exalted, all at intervals 

 of three to five days, may be given, or the exalted virus only may 

 be used. The injections cause moderate fever, headache, and 

 general malaise, and local tenderness, swelling and enlargement of 

 the corresponding lymph glands, all of which pass off in a few days. 



This method (with various slight modifications with regard to 

 dosage) has now been used on a very large scale in India, with 

 strikingly good results. The immunity lasts for at least a year, 

 and probably decidedly longer if large doses of strong vaccines 

 are used, and, what is somewhat unusual, it manifests itself more 

 in a reduction of the incidence of the disease than in the 

 case- mortality. The value of the method is best seen from 

 statistics from -isolated regions in which some persons were 

 vaccinated and others not, all living under the same conditions. 



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