298 PATHOGENIC BACTERIA. 



The Preparation of the Toxin. The method employed 

 by Roux and others at the present time was first sug- 

 gested by Fernbach, and consists in growing the most 

 virulent bacilli obtainable in alkaline bouillon exposed 

 in a thin layer to the passage of a current of air. 



The cultures are allowed to grow for three or four 

 weeks at a temperature of 37 C., with a stream of 

 moist air constantly passing over them. After the given 

 time has passed, it will be found that the acidity prima- 

 rily produced by the bacillus gives place to a much more 

 intense alkalinity than originally existed. The acme of 

 the toxin-production seems to keep pace with this alka- 

 line production. When "ripe," 0.4 per cent, of trikresol 

 is added to the cultures, which are then filtered through 

 porcelain. If the toxin must be kept before using, it is 

 best to preserve it unfiltered, as it deteriorates more rap- 

 idly after filtration. Unfiltered toxin causes too much 

 local irritation. If the bacillus employed was virulent 

 and the conditions of culture were favorable, the filtered 

 culture should be so toxic that o. i c.cm. would be fatal to 

 a 5OO-gram guinea-pig in twenty-four hours (Roux). Even 

 under the most favorable circumstances it is difficult to 

 obtain a toxin which will kill in less than thirty hours. 



The experience of the author with Fernbach' s appara- 

 tus has not been satisfactory. The passing current of air 

 is a frequent source of contamination to the culture, and 

 requires great care. In the end it is questionable whether 

 the toxin thus produced is better than that obtained from 

 an ordinary flask exposing a large surface to the air. 



Park and Williams did an elaborate work upon the 

 production of diphtheria toxin. 1 They found that 

 4 'toxin of sufficient strength to kill a 4oo-gram guinea- 

 pig in three days and a half in a dose of 0.025 c.cm., 

 developed in suitable bouillon, contained in ordinary 

 Erlenmeycr flasks, within a period of twenty-four hours, 

 ich bouillon the toxin reached its greatest strength 

 in four to seven days (0.005 c.cm. killing a 5oo-gram 



1 Jour, of Exper. Med. t vol. i., No. I, Jan., 1896, p. 164. 



