724 Tuberculosis 



except that they were more brief in duration and more_ constant in result; but 

 the disadvantage of abscess formation following the injections remained. The 

 fluid, when filtered, possessed the properties of tuberculin. 



Mechanical fragmentation of bacilli had been employed by Klebs in his studies 

 of antiphthisin and tuherculocidin, 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 envelope of fatty acid, triturat- 

 ing only very small quantities of the bacteria at a time. 



Having thus reduced the bacilli to fragments, he removed them from the mor- 

 tar, placed them in distilled water, washed them, and collected them by cen- 

 trifugation, as a muddy residuum at the bottom of an opalescent, clear fluid. 

 For convenience he named the clear fluid TO; the sediment, TR. TO was found 

 to contain tuberculin. In ord6r to separate the essential poison of the bacteria 

 as perfectly as possible from the irritating tuberculin, the TR fragments were 

 again dried perfectly, triturated once more, re-collected in fresh distilled^ water, 

 and recentrifugated. After the second centrifugation microscopic examination 

 showed that the badllary 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, while in TR a cloudy violet reaction 

 was obtained. 



The addition of so per cent, of glycerin had no effect upon TO, but caused a 

 doudy white deposit to be thrown down from TR. This last reaction showed 

 that TR contained 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 

 trituration, in order to be thorough, should not be done upon more than loo mg. 

 of the bacilli at a time. A satisfactory separation of the TR from TO is said to 

 occur only when the perfectly clear TO takes up at least so per cent, of the solid 

 substance, as otherwise the quantity of TO in the final preparation is so great as 

 to produce undesirable reactions. 



The fluid is best preserved by the addition of 20 per cent, of glycerin, which does 

 not injure the TR and prevents its decomposition. 



The finished fluid contains 10 mg. of solid constituents to the cubic centimeter, 

 and before administration should be diluted with physiologic salt solution (not 

 solutions of carbolic acid). When administering the remedy to man the injec- 

 tions are made with a hypodermic syringe into the tissues of the back. The 

 beginning dose is J^oo mg., rapidly increased to 20 mg., the injections being made 

 daily. 



Experiment showed that TR had decided immunizing powers. 

 Injected into tuberculous animals in too large a dose it produces 

 a reaction, but its immunizing effects were entirely independent of 

 the reaction. Koch's aim in using this preparation in the therapeutic 

 treatment of tuberculosis was to produce immunity against the 

 tubercle baciUus without reactions by gradual but rapid increase of 

 the dose. In so large a number of cases did Koch produce immunity 

 to tuberculosis by the administration of TR, that he believes it 

 proved beyond a doubt that his observations are correct. 



By proper administration of the TR he was able to render guinea- 

 pigs so completely immune that they were able to withstand inocula- 

 tion with virulent bacilU. The point of inoculation presents no 

 change when the remedy is administered; and the neighboring lymph- 

 glands are generally normal, or when slightly swollen contain no 

 bacilli. 



In speaking of his experiments upon guinea-pigs, Koch says: 



, "I have, in general, got the impression in these experiments that full immuni- 

 zation sets in two or three weeks after the use of large doses. A cure in tubercu- 



