274 INFECTION AND IMMUNITY 



TETANUS ANTITOXIN 



Production of Tetanus Antitoxin. The production of tetanus 

 antitoxin is, in every way, analogous to that of diphtheria antitoxin. 

 It is necessary in the first place to produce a powerful tetanus toxin. 

 The methods of procuring this will be discussed in the section 

 upon tetanus toxin. Suffice it to say here that the most satis- 

 factory method of obtaining toxins consists in cultivating the bacilli 

 upon veal broth containing five-tenths per cent to two per cent 

 sodium chlorid and one per cent pepton. It has been advised, also, 

 that the broth should be neutralized by means of magnesium car- 

 bonate rather than with sodium hydrate. The bacilli are cultivated 

 for eight to ten days at incubator temperature and the broth filtered 

 rapidly through Berkefeld filters. The toxin may be preserved in 

 the liquid form with the addition of five-tenths per cent carbolic 

 acid, or may be preserved in the dry state after precipitation with 

 ammonium sulphate. 



It is necessary to determine the strength of the poison. This is 

 done according to v. Behring 14 by determining the smallest amount of 

 toxin which will kill a white mouse of twenty grams weight within 

 four days. This is most easily done by making dilutions of the toxin 

 ranging from 1 :100 to 1 :1,000, and then injecting quantities of 

 0.1 c.c. of each of these dilutions subcutaneously into white mice. 

 In this way, the minimal lethal dose is ascertained. 



For the actual production of antitoxin, horses have been generally 

 found to be the most favorable animals. The horses should be 

 healthy and from five to seven years old. The first injection of 

 toxin administered to these animals should be attenuated in some 

 way. Various methods for accomplishing this have been in use. In 

 America, the first injection of about ten to twenty thousand minimal 

 lethal doses 15 (for mice of twenty grams weight) is usually made 

 subcutaneously together with sufficient antitoxin to neutralize this 

 quantity. In Germany, v. Behring uses, for his first injection, a 

 much larger dose of toxin to which about 0.25 per cent of terchlorid 

 of iodin has been added. Immediately after an injection, the animals 

 will usually show a reaction expressed by a rise of temperature, 



14 v. Behring, Zeit. f . Hyg., xii, 1892 ; Deut. med. Woch., 1900. 



15 According to Park the "horses receive 5 c.c. as the initial dose of a toxin 

 of which 1 c.c. kills 250,000 grams of guinea-pig, and along with this a sufficient 

 amount of antitoxin to neutralize it. ' ' 



