334 A MANUAL OF BACTERIOLOGY 



to twelve days at 37 C. Various details have to be 

 attended to in order to obtain toxin of maximum toxicity. 

 The use of meat some days old has been advocated, or 

 of acid beef-broth in which B. coli has been grown for 

 twenty -four hours, in order to eliminate the glucose (p. 32). 

 L. Martin makes use of " peptone " prepared by the auto- 

 digestion of a pig's stomach with dilute hydrochloric acid. 

 To the cultures 10 per cent, of a 5 per cent, solution of 

 carbolic acid are added, the bacilli are allowed to deposit 

 by standing for forty-eight hours, and the culture is filtered 

 through paper ; in this way filtration through a filter-candle 

 is dispensed with. Less than 0-01 c.c. of the toxin should 

 kill a 250-grm. guinea-pig in three to four days. Selected 

 horses which have been tested with mallein and tuber- 

 culin, and kept under observation for some time to ensure 

 that they are healthy, are then inoculated with this 

 filtrate, commencing with a dose of 0-01 to 0-1 c.c. accord- 

 ing to the toxicity of the toxin, or 20 c.c. of the toxin 

 together with 10,000 units of antitoxin may be given for 

 the first three doses. Individual horses vary in their 

 susceptibility to the toxin, so that care has to be exer- 

 cised with the first injections. The injections are given 

 subcutaneously over the shoulder, and produce local 

 swelling with some rise of temperature and general dis- 

 turbance, lasting two or three days. When this has 

 passed away the inoculation is repeated, a larger dose 

 being administered provided the reaction due to the 

 former one was not too severe. After the first two or 

 three doses, the dose of toxin given may usually be 

 double the previous one, so that after six or eight 

 weeks treatment it may attain 500 c.c. or more. 

 Individual horses vary much in their capacity to yield 

 antitoxin : on the whole those that are moderately 

 sensitive to the toxin seem to produce most antitoxin ; a 

 horse to be of value should after three months' treatment 



