PROTECTIVE INOCULATIONS. 359 



occurred in a dialyzer placed in running water. But by shaking up 

 the dry powder in chloroform the heavy salt sank to the bottom and 

 the purified antitoxin floated on the surface and could be recovered by 

 skimming it off. The powder thus obtained consisted of a mixture of 

 various substances, including the antitoxin, and when obtained from 

 milk having an antitoxic value of 90,000 it was found to have a value 

 of 25,000,000 immunization units. By further purification a still 

 higher value was obtained (55,000,000). In experiments on mice a 

 dose ten thousand times as great as was necessary to produce immu- 

 nity proved to exercise a curative power i.e., a dose of 0.02 gramme 

 for a mouse weighing twenty grammes saved it from being killed by 

 double the minimum fatal dose of the tetanus toxin, after tetanic 

 symptoms had been developed. 



Reference has been made to the production of immunity by the 

 use of cultures made in thymus bouillon. This was made known 

 through the experiments of Brieger, Kitasato, and Wassermann 

 (1892). The thymus bouillon is made from the thymus glands of 

 calves, which are chopped fine in a hash machine and covered with 

 an equal volume of distilled water. The mixture is stirred for some 

 time and then placed in an ice chest for twelve hours; the liquid is 

 then obtained by filtration through gauze with pressure by means of 

 a flesh-press machine. A turbid, slimy fluid is thus obtained, which 

 is diluted with an equal volume of water and made slightly alkaline 

 by the addition of soda solution. It is then sterilized at 100 C. for 

 fifteen minutes. As a result of this the liquid has a grayish-brown 

 color, and some large flocculi in suspension, which are removed by 

 passing it through fine linen. The fluid is then of a milky opal- 

 escence. It is next placed in test tubes and again sterilized. The 

 tetanus bacillus when cultivated in this medium does not form spores, 

 and the toxic potency of the culture is very much reduced 1 : 5,000 to 

 1 : 3,000 of the toxic potency manifested by cultures of the same bacil- 

 lus in ordinary media. Inoculations with cultures in thymus bouillon 

 were found to kill mice in the dose of 0.5 cubic centimetre, while 

 smaller amounts failed to kill and caused the animals to be immune. 

 A culture in ordinary bouillon was fatal to mice in the dose of 0.001 

 cubic centimetre. 



Experiments on rabbits (thirty -five) gave a uniformly successful 

 result in immunizing these animals. Immunity was established in 

 the course of two weeks, and the blood serum of these animals tested 

 on mice showed an antitoxic value of 1,000. 



Reference has already been made to the earlier researches of the 

 Italian investigators, Tizzoni and Cattani. These have been followed 



