222 INFECTION AND IMMUNITY 



Standardization. — The universal prophylactic use of tetanus antitoxin 

 has, as in the case of diphtheria antitoxin, necessitated its standardiza- 

 tion. A variety of methods are in use in different parts of the world. 

 In the following description the American method only wiU be consid- 

 ered as laid down under the law of July, 1908, and based upon the work 

 of Rosenau and Anderson ^ at the United States Hygienic Laboratories 

 at Washington. 



In conjunction with a committee of the Society of American Bac- 

 teriologists, these authors have defined the unit of tetanus antitoxin as 

 follows: 



The unit shall be ten times the least amount of serum necessary to 

 save the life of a 350 gram guinea-pig for ninety-six hours against the 

 official test dose of standard toxin. The test dose consists of 100 minimal 

 lethal doses of a precipitated toxin preserved under special conditions 

 at the hygienic laboratory of the Public Health and Marine Hospital 

 Service. (The minimal lethal dose is in this case, unlike Behring's 

 minimal lethal dose, measured not against 20 gram mice, but against 

 350 gram guinea-pigs.) 



In the actual standardization of tetanus antitoxin, as in that of diph- 

 theria antitoxin, the L+ dose of toxin is employed. The L+ dose is, 

 however, in this case, defined as the smallest quantity of tetanus toxin 

 that will neutralize one-tenth of an immunity unit, plus a quantity of 

 toxin sufficient to kill a 350 gram guinea-pig in just four days. At the 

 Hygienic Laboratory at Washington, a standard toxin and antitoxin 

 are preserved under special conditions, and standard toxin and anti- 

 toxin, arbitrary in their first establishment, are kept constant by being 

 measured against each other from time to time. In measuring the anti- 

 toxic serum thus preserved, at the Hygienic Laboratory, a mixture of 

 one-tenth of a unit of antitoxin and 100 minimal lethal doses of the 

 standard toxin must contain just enough free poison to kill the guinea- 

 pig in four days. This L+ dose of the standard toxin is given out to 

 those interested commercially orotherwise in the production of antitoxin. 



In measuring an unknown antitoxic serum against this L+ dose of 

 toxin, a large number of mixtures are made, each containing the 

 L^. dose of the toxin and varying quantities of the antitoxin. Dilu- 

 tions must always be made with 0.85 per cent salt solution and the 

 total quantity injected into the animals should always be brought up to 



1 Rosenau and Anderson, Pub. Health and Mar. Hosp. Serv. U. S., Hyg. Lab. Bull. 

 43, 1908. 



