280 PROTECTIVE INOCULATIONS. 



chimiques) . These experiments were commenced in 1881. The 

 authors named say : 



" In repeating the experiments of Toussaint upon anthrax blood which 

 had been heated, we made several observations which convinced us that it is 

 possible to confer immunity against anthrax upon sheep by injecting under 

 their skin anthrax blood which does not contain any living bacilli." 



While immunity was produced in this way, Roux and Chamber- 

 land remark that the sheep which had received a comparatively large 

 dose were quite sick when subsequently inoculated with a virulent 

 culture, and the immunity acquired was less reliable than that ob- 

 tained by Pasteur's method with two vaccines of different degrees of 

 attenuation. 



In an investigation made by Hankin, in the laboratory of Professor 

 Koch at the Hygienic Institute of Berlin, the results of which are given 

 in a preliminary account published in the British Medical Journal 

 (October 12th, 1889), the important fact was ascertained that immunity 

 may be produced in susceptible animals by inoculating them with an 

 " albumose " isolated from anthrax cultures. Hankin gives the fol- 

 lowing account of his method of obtaining this immunizing proteid 

 from anthrax cultures : 



" In the course of my process of preparation it is precipitated from its so- 

 lution by the addition of a large bulk of absolute alcohol, and well .washed 

 in this liquid to free it from ptomaines ; it is well known that all such sub- 

 stances are soluble in alcohol. It is then filtered off and dried ; then it is re- 

 dissolved and filtered through a .Chamberland filter. A rough estimate of 

 the percentage of albumose present in the clear solution thus obtained is 

 made colorimetrically by means of the biuret reaction and a peptone solution 

 of known strength." 



"In one experiment four rabbits (Nos. 23 to 26) were inoculated subcuta- 

 neously with virulent anthrax spores. No. 26 served as a control and died 

 in about forty hours. The other three rabbits had the albumose solution in- 

 jected into the ear-vein at the same time. Nos. 24 and 25 each had about 

 the five-millionth of their body-weight, while No. 23 had only the ten- mil- 

 lionth of its body-weight of albumose. No. 25 died in less than forty-eight 

 hours, but Nos. 23 and 24 survived. Ten days later Professor Koch kindly 

 reinoculated these two rabbits for me with very virulent anthrax from an 

 agar-agar culture. Their temperature has remained normal since then, and 

 they are now alive and well a fortnight after this operation. I have also suc- 

 ceeded in producing immunity in mice against attenuated anthrax." 



In a paper published in the Proceedings of the Koyal Society in 

 1890, Dr. Sidney Martin has given an account of his researches relat- 

 ing to " The Chemical Products of the Growth of Bacillus Anthracis, 

 and their Physiological Action. 7 ' In his experiments the cultures 

 were maintained for from ten to fifteen days, and the bacilli were then 

 removed by filtering through a Chamberland filter. The filtrate was 

 found to contain : 



