156 Immunity 



that such purely antitoxic serums are inferior to those con- 

 taining other immunity products, such as bacteriolysins, 

 and recommends that the whole culture instead of the 

 filtered culture be used in the immunization of the animal. 

 If this is done, the bacteriolytic effect is added to the anti- 

 toxic effects of the serum. 



The serum may be used to prevent or to cure diphtheria. 



The antitoxin is commercially manufactured at present 

 by immunizing horses against increasing quantities of diph- 

 theria toxin until the proper degree of immunity has been 

 attained, then withdrawing the antitoxic blood. The details 

 are as follows: 



I. The Preparation of the Toxin. The toxic metabolic products 

 of the Bacillus diphtheriae are for the most part freely soluble, and 

 are therefore best prepared in cultures grown in fluid media. The 

 medium best adapted to the purpose is that recommended by Theobald 

 Smith.* 



To make it, the usual meat infusion receives the addition of a 

 culture of Bacillus coli, and is stood in a warm place overnight. The 

 colon bacilli ferment and remove the muscle and other sugars. The 

 infusion is then made into bouillon, titrated so that the reaction 

 equals + 1.1 when tested with phenolphthalein. It then receives an 

 addition of 0.2 per cent, of dextrose, and is sterilized in the autoclave. 

 To secure the b.est toxic product, the bacilli at hand must be carefully 

 studied and that naturally possessing the strongest toxicogenic power 

 employed for the cultures. The greatest toxicity seems to develop 

 between the fifth and seventh days. If the culture is permitted to 

 remain in the incubating oven beyond this period, the toxin gradually 

 is transformed to toxoid and its activity declines. The fatal dose for 

 a 250-300 gram guinea-pig should be about 0.001 c.c. given hypo- 

 dermically. 



II. The Immunization of the Animals. All commercial manu- 

 facturers of diphtheria antitoxic serums now use horses, as recom- 

 mended by Roux, instead of the sheep, dogs, and goats with which 

 the earlier investigators worked. The horse is readily immunized, 

 gives an abundant supply of blood which clots readily and yields a 

 beautiful clear amber serum. 



The horse selected should be in perfect health, and should be tested 

 with mallein and tuberculin to avoid obscure glanders and tuberculosis. 



A small dose of the toxic bouillon say 0.1 c.c. should be given 

 in the beginning, as one occasionally finds exceptionally susceptible 

 animals that will succumb to larger doses. If a marked local and 

 general reaction follows, it may be better to try another animal. If 

 no reaction is brought about, the immunization is carried on as rapidly 

 as possible. The toxin is injected hypodermatically into the tissues 

 of the neck, the skin being thoroughly cleaned and disinfected before 

 each injection. The doses are cautiously increased and may often be 

 doubled each day. If any unfavorable symptoms arise, treatment 

 must be interrupted for a day or two. The animal yields good anti- 

 toxic serum when it can endure several doses of 500 c.c. of the strong 

 toxin mentioned above. 



HI. Bleeding. When the withdrawal of a small quantity of 

 blood by a hypodermic needle introduced into the jugular vein shows 



* " Journal of Experimental Medicine," May and July, 1899, p. 373. 



