PROTECTIVE INOCULATIONS. 315 
40° C., and then heated to 102° C., it still retained its antitoxic 
potency. 
Ehrlich, Kossel, and Wassermann (1894) have made experiments 
upon goats, which they found very susceptible to the action of the 
diphtheria poison. Sterilized cultures were first injected in gradually 
increasing amounts, and later virulent cultures. In this way they ob- 
tained a serum which has a value sixty times that of Behring’s “nor- 
mal serum.” In a subsequent communication (1894) Wassermann 
gives an account of his experiments with the milk of immunized 
goats, which contains the antitoxin in considerable quantity, and 
from which it was obtained in a concentrated form by the following 
method: The milk is obtained in sterilized vessels and twenty cubic 
centimetres of normal hydrochloric acid are added to each litre; a 
sufficient quantity of rennet is then added to coagulate the casein, 
and this is separated from the liquid, which is then shaken up with 
chloroform for some time. The liquid is now allowed to stand in 
order that the butter, which has been dissolved by the chloroform, 
may sink to the bottom. The clear liquid is then decanted and the 
antitoxin precipitated from it by means of ammonium sulphate (thirty 
to thirty-three per cent). The precipitate is rapidly dried upon 
porous porcelain plates, in vacuo, and then dissolved in water in the 
proportion of ten parts for one hundred of milk first employed—a 
concentration to one-tenth. Of this solution 0.125 cubic centimetre 
was found to neutralize 0.9 cubic centimetre of a toxin which killed 
guinea-pigs weighing five hundred grammes in the dose of 0.1 cubic 
centimetre. This toxin was an old bouillon culture of the diphtheria 
bacillus to which 0.5 per cent of carbolic acid had been added to 
preserve it. In a communieation of the same date Ehrlich and Was- 
sermann report that they have for some time had a cow immunized 
to such a degree that one cubic centimetre of its milk protects guinea- 
pigs from the fatal effects of 0.9 cubic centimetre of the above-men- 
tioned toxin. The antitoxic value of the milk of an immunized cow 
or goat, as compared with that of its blood, is estimated by Ehrlich 
and Wassermann as from 1:15 to 1:30—usually about 1:20. 
Aronson, in testing his antitoxin, uses a bouillon culture of the 
diphtheria bacillus two and one-half months old, which he preserves 
by the addition of 0.3 per cent of trikresol. He finds that the im- 
munity which results from injections of the antitoxin is established 
at once; that it is not accompanied by any reaction or symptom of 
sickness; and that it is of comparatively short duration. 
As a result of extended experiments made at the Pasteur Institute 
in Paris, Roux has perfected the following method for the production 
