BACILLUS OF TUBERCULOSIS. 289 
bacilli themselves. It also contains all the albuminoid 
and other materials originally contained in the bouillon 
which have remained unaffected by the activities of the 
bacilli. There are two preparations known respec- 
tively as the old and the new tuberculin. 
Old tuberculin is prepared as follows: The tubercle 
bacillus is cultivated in an infusion of calf’s flesh, or of 
beef flesh, or extract to which 1 per cent. of peptone 
and 4 to 5 per cent. of glycerin have been added, the 
culture liquid being slightly alkaline. The inoculation 
is made upon the surface from a piece of very thin 
pellicle from a young bouillon culture, or, if the bou- 
illon culture is unobtainable, with small masses from a 
culture on glycerin-agar. These masses, floating on 
the surface, give rise in from three to six weeks, accord- 
ing to the rapidity with which the culture grows, to an 
abundant development and to the formation of a toler- 
ably thick and dry, white crumpled layer, which finally 
covers the entire surface. At the end of four to eight 
weeks development ceases, and the layer after a time 
sinks to the bottom. Fully developed cultures, after 
having been tested for purity by a microscopical exami- 
nation, are passed into a suitable vessel and evaporated 
to one-tenth of their original bulk over a water-bath 
at a temperature of 70° to 80° C. The liquid is then 
filtered through chemically pure sterilized filter-paper. 
The crude tuberculin thus obtained contains 40 to 50 per 
cent. of glycerin and keeps well, retaining its activity 
- indefinitely. 
The method of treatment and the results obtained 
from the old tuberculin have been described recently 
by Koch briefly as follows: After each injection, which 
should be large enough to cause a slight but not a great 
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