CULTIVATION OF TUBERCLE BACILLUS. 265 



ments which are often present ought to be selected ; dried films 

 are then prepared in the usual way and stained by the Ziehl- 

 Neelsen method (p. 104). In the case of urine or other fluids a 

 deposit should first be obtained by centrifugalising a quantity 

 in a test-tube, or by allowing the fluids to stand in a tall glass 

 vessel (an ordinary burette is very convenient). Film prepara- 

 tions are then made with the deposit and treated as before. If 

 a negative result is obtained in a suspected case, repeated ex- 

 amination should be undertaken. To avoid risk of contamina- 

 tion with the smegma bacillus the meatus of the urethra should 

 be cleansed and the urine first passed should be rejected, or the 

 urine may be drawn off with a sterile catheter. As stated above 

 it is only exceptionally that difficulty will arise to the experienced 

 observer from this cause. (For points to be attended to, vide 



P- 257-) 



(2) Inoculation. The guinea-pig is the most suitable ani- 

 mal. If the material to be tested is a fluid it is injected subcu- 

 taneously or into the peritoneum ; if solid or semi-solid it is 

 placed in a small pocket in the skin or it may be thoroughly 

 broken up in sterile water or other fluid and the emulsion 

 injected. By this method, material in which no tubercle bacilli 

 can be found microscopically may sometimes be shown to be 

 tubercular. 



(3) Cultivation. Owing to the difficulties this is usually 

 quite impracticable as a means of diagnosis, and it is also un- 

 necessary. The best method to obtain pure cultures is to pro- 

 duce tuberculosis in a guinea-pig by inoculation with tubercular 

 material, and to kill the animal after a period of four or five 

 weeks. Then with portions of a tubercular organ, e.g. the spleen, 

 tubes of solidified blood serum (preferably that of the dog, ob- 

 tained aseptically and coagulated by an exposure of three hours 

 to a temperature of 75 C. in the slanting position) are to be 

 inoculated under the strictest aseptic precautions. The por- 

 tions of tissue should be moderately small, and should be gently 

 rubbed over the surface of the medium and brought to rest. 

 The tubes are then to be incubated at 37 C. for a week or 

 ten days, when the pieces of tissue are again to be rubbed over 

 the surface of the medium, turned over, and once more incu- 

 bated for a similar period of time, when growth may possibly 

 be seen in isolated spots on the surface of the serum or ex- 



