Toxic Products 337 



Pursuing the idea of fragmenting the bacilli, or treating 

 them chemically to increase their solubility, Koch found 

 that a 10 per cent, sodium hydrate solution yielded an 

 alkaline extract of the bacillus, which, when injected into 

 animals, produced effects similar to those following the 

 administration of tuberculin, except that they were more 

 brief in duration and more constant in result ; but the dis- 

 advantage of abscess-formation following the injections re- 

 mained. The fluid, when filtered, possessed the properties 

 of tuberculin. 



Mechanical fragmentation of bacilli had been employed 

 by Klebs in his studies of antiphthisin and tuberculocidin, 

 and Koch now used it with advantage. He pulverized living, 

 virulent, but perfectly dry bacilli in an agate mortar, in 

 order to liberate the toxic substance from its protecting 

 envelop of fatty acid, triturating only very small quantities 

 of the bacteria at a time. 



Having thus reduced the bacilli to fragments, he removed 

 them from the mortar, placed them in distilled water, 

 washed them, and collected the fragments by centrif- 

 ugation, as a muddy residuum at the bottom of an opal- 

 escent, clear fluid. For convenience he named the clear 

 fluid TO; the sediment, TR. TO was found to contain 

 tuberculin. In order to separate the essential poison of the 

 bacteria as perfectly as possible from the irritating tuber- 

 culin, the TR fragments were again dried perfectly, trit- 

 urated once more, re-collected in fresh distilled water, and 

 recentrifugated. After the second centrifugation micro- 

 scopic examination showed that the bacillary fragments 

 had not yet been resolved into a uniform mass, for when 

 TO was subjected to staining with carbol-fuchsin and 

 methylene-blue it was found to exhibit a blue reaction, 

 w r hile in TR a cloudy violet reaction was obtained. 



The addition of 50 per cent, of glycerin had no effect 

 upon TO, but caused a cloudy white deposit to be thrown 

 down from TR. This last reaction showed that TR con- 

 tained fragments of the bacilli insoluble in glycerin. 



In making the TR preparation Koch advises the use 

 of a fresh, highly virulent culture not too old. It must 

 be perfectly dried in a vacuum exsiccator, and the tritu- 

 ration, in order to be thorough, should not be done upon 

 more than 100 mg. of the bacilli at a time. A satisfactory 

 separation of the TR from TO is said only to occur when 



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