TOXIN AND ANTITOXIN 105 



In 1894 Calmette 3 and Physalix and Bertrand 4 had indepen- 

 dently succeeded in obtaining an antitoxin against snake poison. In 

 the course of further study of these bodies Calmette 5 determined 

 that the venoms of certain varieties of snakes, the naja and cobra, 

 would remain potent even when subjected to 100 C. for a very 

 short time. In contrast to this the antitoxins to these poisons were 

 destroyed at much lower temperatures. Now when mixtures of 

 the two substances, so proportioned that their injection into ani- 

 mals was innocuous, were heated to 68 C. for considerable 

 periods, toxic properties again became evident, a demonstration that 

 the toxin had not been destroyed, but had remained neutral only in 

 the presence of the intact antitoxins. These experiments were con- 

 firmed by Wassermann, 6 who found that similar conditions pre- 

 vailed in the combination between pyocyaneus toxin and antitoxin. 



The nitration experiments of Martin and Cherry 7 are not con- 

 vincing since they may be taken as indicating either neutralization 

 or toxin destruction. These workers subjected mixtures of snake 

 poison and its specific antitoxin to filtration through gelatin filters, 

 under pressure. Under the experimental conditions thus estab- 

 lished the presumably smaller toxin molecule was allowed to pass 

 through the filter while the larger antitoxin molecule was held back. 

 They showed that if filtered soon after the ingredients have been put 

 together most of the toxin still passes through, but that, as this inter- 

 val is prolonged, less and less comes through, presumably because of 

 the union of the smaller toxin to the larger antitoxin molecule. The 

 chief value of these experiments lies in their proof of the element of 

 time as an important factor in the toxin-antitoxin union. 



In his experiments on snake venom just recorded, Calmette in- 

 terpreted the restitution of toxicity after the heating of neutral mix- 

 tures of cobra neurotoxin and its antitoxin as evidence "qu'il ne 

 s'etait pas forme aucune combinaison de ces deux substances ou que 

 la combinaison realisee etait, au moins, tres instable." Later experi- 

 ments of Martin and Cherry seemed for a time to contradict this con- 

 clusion. Observations by them, analogous to those of Calmette, but 

 carried out with the poison of an Australian snake, seemed to indi- 

 cate that when the toxin and antitoxin were allowed to remain to- 

 gether for a sufficiently long time no restitution of toxicity could be 

 obtained by heating. Apparently the application of heat to such mix- 

 tures merely prevented the further union of antitoxin with any toxin 

 that was not yet bound at the time that the heat was applied. Accord- 



3 Calmette. Compt. rend, de la soc. de biol, 1894. 



4 Physalix and Bertrand. Compt. rend, de la soc. de biol., 1894. 



5 Calmette. Ann. Past., 1895. 



6 Wassermann. Zeitschr. f. Hyg., 22, 1896. 



7 Martin and Cherry. Proc. of the Royal Soc., Vol. 63, 1898. 



