METHOD FOR PRODUCTION OF TETANUS TOXIN 337 



Though no comparative tests have been made with toxin 

 produced under the usual anaerobic conditions, the toxin pro- 

 duced by the Hygienic Laboratory method showed a some- 

 what unexpected stabihty. For example: 



Lot 25. 



The test was made on a small amount of toxin, about 30 cc, 

 kept in an ordinary test tube in the ice-box and protected from 

 the air only by a shallow covering of albolene. 



From the above, it will be seen that our method for growing 

 the tetanus bacillus for toxin varies very slightly from that of 

 the Hygienic Laboratory. Instead of beef, we use market 

 veal which has given almost invariably a toxin of 25^00? even 

 going as high as 40^100 occasionally. It is interesting to note 

 that with the bob veal, that is, veal under the legal age limit for 

 selhng, which has always given a more highly potent diphtheria 

 toxin than the older veal, we have rarely obtained a tetanus 

 toxin above 157000 whereas the market veal used as control has 

 produced a toxin of 2X000 strength. ^ 



Anderson recommends that the preliminary cultures should 

 be carried on from one to three weeks, by daily transfers, but 

 with only three generations in glucose broth before inoculating 

 the toxin flasks, we have obtained a toxicity of 1-40,000 show- 

 ing that fewer generations may be sufficient. 



Here it might be well to state that we obtained different re- 

 sults from those recorded by other writers in regard to the growth 

 in the preliminary cultures. Anderson and Leake found that 

 a good growth is obtained in the first generation in 48 hours, 

 in 24 hours in the second and third transplants, and in 16 hours 



^ Furthermore, by inoculating the glucose broth directly with a melted semi- 

 solid agar culture of B. tetani, we have secured a toxin of over - in strength, 



25,000 



the control test which had been inoculated from preliminary broth cultures 

 giving the same degree of toxicity. Confirmatory tests are being made along 

 this line. 



