962 CONTROL OF BIOLOGICAL PRODUCTS 



TUBERCULIN 



The tests to be mentioned apply to old tuberculin (O.T.) only. There are no satis- 

 factory tests for the various modifications of tuberculin. The tests to be preferred 

 for this preparation, whether intended for diagnostic or therapeutic application, are 

 based on the fact that guinea pigs infected with tuberculosis some weeks earlier react 

 in a specific manner to a subsequent injection of tuberculin. Serological tests have 

 also been employed but are generally considered less satisfactory than those utilizing 

 living animals. 



The method longest in use was devised by Koch and depends upon the death of 

 tuberculous guinea pigs when given a sufficient dose of tuberculin at a suitable time 

 after infection. A modification of this method as applied to official standardization 

 by the Bureau of Animal Industry, Department of Agriculture, was described origi- 

 nally by Schroeder and Brett.' 



According to Lewis and Aronson,^ the procedure employed more recently is as 

 follows: 



A carefully measured minute quantity of tuberculous material, obtained from a tubercu- 

 lous guinea pig, is injected into a series of animals. About three weeks after infection, the 

 sensitiveness of the guinea pigs to tuberculin is determined by giving i guinea pig daily an 

 intra-abdominal injection of 0.375 gm. O.T. per 500 gm. body weight. When the sensitive- 

 ness of the animals has reached a degree at which this dose causes death within twenty-four 

 hours, 2 guinea pigs are injected daily with 0.25 gm. O.T. per 500 gm. weight. The day after 

 it is found that this dose kills i at least of the 2 guinea pigs within twenty-four hours, 6 

 guinea pigs are injected, each with 0.25 gm. O.T. per 500 gm. weight, and, if 4 or more pigs 

 die in twenty-four hours, the remaining infected pigs are regarded as being sufficiently sensi- 

 tive for determining the potency of the tuberculin of unknown strength. When the guinea 

 pigs have reached the desired sensitiveness, as determined by the method just described, 

 they are then divided into groups of six. One group is used to each sample of tuberculin the 

 potency of which is to be determined. Each animal of the group receives an intra-abdominal 

 injection of 0.25 gm. O.T. to be tested for 500 gm. body weight. As a control, another group 

 of 6 guinea pigs is injected, each with an equal amount of the standard tuberculin, and 2 

 non -tuberculous guinea pigs are injected with the tuberculin which is being tested, in order 

 to make certain that the tuberculin contains no excessive amount of toxic material and no 

 viable tubercle bacilli. 



In order to pass the test, it is required that the tuberculin should, within twenty-four 

 hours, kill at least half as many of the test guinea pigs as are killed in the control group 

 injected with the standard tuberculin, and that the animals should show, at autopsy, the 

 characteristic lesions found when tuberculous animals succumb to lethal doses of tuberculin. 



Lewis and Aronson modified this method by determining the dose of tuberculin 

 necessary to evoke a cutaneous allergic reaction in tuberculous guinea pigs. The au- 

 thors give the following instructions for the performance of the titration by the method 

 they proposed : 



The reaction to the intracutaneous injection of tuberculin is very definite on the guinea 

 pigs, and one may easily use this method to ascertain the potency of tuberculin by injecting 

 in adjacent places equal amounts of a tuberculin of unknown potency and a standard product. 



■ Schroeder, E. C, and Brett, G. W.: J. A. Vet. M.A. (7th new ser.), 54, 357. 1919. 

 ^ Lewis, Paul A., and Aronson, Joseph D.: Am. Rev. Tiiherc, 7, 404-11. 1923. 



