TETANUS. 237 



grown upon feebly alkaline glucose-bouillon contains the 

 specific tetanus-toxin in an exceedingly active state. Often 

 as little as 0.000005 cu - cm - f sucn a filtrate will suffice 

 to cause death in a white mouse. The toxin is destroyed 

 in solution by exposure to a temperature of 65 C. (149 

 F.) for five minutes, and it is gradually enfeebled in 

 the thermostat. In a cold compartment (refrigerator) and 

 protected from light it retains its toxicity unchanged for 

 months. For the preservation of such a toxic filtrate it is 

 advisable to add glycerin in equal amount, or carbolic acid 

 in proportion of 0.5 per cent, and it may then be employed 

 for months in the same way as the solution of a chemic 

 poison of known strength. 



Efforts have not been wanting to isolate the toxin in a 

 pure state. Brieger obtained from cultures two basic sub- 

 stances tetanin and tetanotoxin both of which caused 

 death in animals, preceded by manifestations like those of 

 tetanus. Neither, however, can have any important bear- 

 ing upon the occurrence of the disease, as relatively large 

 amounts were required in order to induce disease in animals. 

 The actual tetanus-toxin, however, must, in accordance 

 with what is known regarding the toxicity of the bouillon- 

 filtrate, be active in minimal amounts. Brieger and Frankel 

 have, according to their method, by precipitation with 

 alcohol, obtained a toxalbumin that is much more active. 

 Even this albuminous powder does not represent the 

 tetanus-toxin chemically pure ; the toxic substance is 

 merely adherent to it. 



The attempts to obtain the virus in its chemic purity 

 have been pursued unremittingly, and it has been obtained 

 in a quite concentrated form. Reference was made in the 

 general section (p. 30) to the investigations of Brieger and 

 Boer, from which it appears that the toxin of the tetanus- 

 bacillus is not an albuminous substance at all. These 

 investigators precipitated the tetanus-toxin by the original 

 toxalbumin-method of Brieger and Frankel with the aid 

 of ammonium sulphate, and then by means of a weak 

 solution of mercuric chlorid (from 10 to 20 cu. cm. of 

 a 0.05 per cent, solution to 10 cu. cm. of tetanus-toxin 

 fluid) precipitated the redissolved toxin. The carefully 

 washed precipitate is well rinsed with water, and decom- 

 posed by successive treatment with ammonium carbonate, 

 ammonium phosphate, and ammonium sulphate. Brieger 



