362 PROTECTIVE INOCULATIONS. 



are redissolved by hot water containing sodium chloride and very 

 dilute potash solution. The elementary analysis gave N 5.90 per 

 cent, C 35.19 per cent, and H 7.02 per cent. The results obtained 

 are believed to show that the active substance present in the lymph 

 is a toxalbumin. In experiments made with Koch's lymph in Pas- 

 teur's laboratory by Bardach, a very decided elevation of temperature 

 was produced in tuberculous guinea-pigs by the subcutaneous injection 

 of 0.1 gramme, and a fatal result by the injection of 0.2 to 0.5 gramme. 

 In man a decided febrile reaction is produced in tuberculous patients 

 by very much smaller doses 0.001 cubic centimetre. 



Hammerschlag, in his chemical researches, found that the tubercle 

 bacillus yields a larger proportion of substances soluble in alcohol and 

 ether than any other bacilli tested (twenty-seven per cent). The alco- 

 holic extract contains fat, lecithin, and a toxic substance which pro- 

 duces convulsions in rabbits and guinea-pigs. The portion insoluble 

 in alcohol and ether contains cellulose and an albuminoid substance. 

 No ptomains were found, but a toxalbumin was isolated, which 

 caused an elevation of temperature in rabbits of 1 to 2 C., lasting 

 for a day or two. 



Koch (1891) has given a full account of his method of prepari-ng 

 crude tuberculin, and also the process by which he obtains from this 

 a tuberculin which appears to be pure, or nearly so. To obtain con- 

 siderable quantities of the crude product the tubercle bacillus is culti- 

 vated in an infusion of calves' flesh, or of beef extract to which one 

 per cent of peptone and four or five per cent of glycerin have been 

 added. This culture liquid must be made slightly alkaline, and it 

 is placed in flasks with a flat bottom, which should not be more than 

 half filled thirty to fifty cubic centimetres. The inoculation is made 

 upon the surface with small masses from a culture upon blood serum 

 or glycerin agar. By accident Koch discovered that these masses 

 floating upon the surface give rise to an abundant development, and 

 to the formation of a tolerably thick and dry white layer, which finally 

 covers the entire surface. At the end of six to eight weeks develop- 

 ment ceases, and the layer after a time sinks to the bottom, breaking 

 up meanwhile into fragments. These cultures, after their purity has 

 been tested by a microscopical examination, are poured into a suitable 

 vessel and evaporated to one- tenth the original volume over a water 

 bath. The liquid is then filtered through procelain. The crude tu- 

 berculin obtained by this process contains from forty to fifty per cent 

 of glycerin, and consequently is not a suitable medium for the develop- 

 ment of saprophy tic bacteria, if they should by accident be introduced 

 into it. It keeps well and preserves its activity indefinitely. 



