Isolation 721 



isms infrequently grow quite well, or can be used for the 

 inoculation of guinea-pigs. 



The most certain method of obtaining a culture of the 

 tubercle bacillus from sputum, pus, etc., is to inoculate a 

 guinea-pig, allow artificial tuberculosis to develop, and make 

 cultures from one of the tuberculous lesions. 



To make such an inoculation with material such as 

 sputum, in which there are many associated micro-organisms 

 that may destroy the guinea-pig from septicemia, Koch ad- 

 vised the following method, with which he never experienced 

 an unfavorable result. 



With a sharp-pointed pair of scissors a snip about J cm. 

 long is made in the skin of the belly- wall. Into this the 

 points of the scissors are thrust, between the skin and the 

 muscles for at least i cm., and the scissors opened and 

 closed so as to make a broad subcutaneous pocket. Into 

 this pocket the needle of the hypodermic syringe contain- 

 ing the injection, or the slender glass point of a pipet con- 

 taining it, is introduced, a drop of fluid expressed and gently 

 rubbed about beneath the skin. When the inoculating in- 

 strument is withdrawn, the mouth of the pocket is left open. 

 A slight suppuration usually occurs and carries out the or- 

 ganisms of wound infection, while the tubercle bacilli are 

 detained and carried to the inguinal nodes, which usually en- 

 large during the first ten days. The guinea-pigs usually die 

 about the twenty-first day after infection. 



The guinea-pig is permitted to live until examination shows 

 the inguinal glands are well enlarged, and toward the middle 

 of the third week is chloroformed to death. The exterior of 

 the body is then wet with i : 1000 solution of bichlorid of 

 mercury and the animal stretched out, belly up, and tacked 

 to a board or tied to an autopsy tray. The skin is ripped up 

 and turned back. The exposed abdominal muscles are now 

 washed with bichlorid solution and a piece of gauze wrung 

 out of the solution temporarily laid on to absorb the excess. 

 With fresh sterile forceps and scissors the abdominal wall is 

 next laid open and fastened back. With the same instru- 

 ments or, preferably, with fresh sterile instruments the 

 spleen, which should be large and full of tubercles, is 

 drawn forward and, one after another, bits the size of a 

 pea cut or torn off and immediately dropped upon the sur- 

 face of appropriate culture-media in appropriate tubes. The 

 fragments of tissue from the spleen of the tuberculous 

 46 



