ANAEROBIC CULTURES 71 



desired, for greater safety, be taken with the forceps, held 

 in the flame for a second or two, and pushed while 

 burning into the tube, and this procedure must always 

 be adopted if the plug be dropped or brush against 

 anything. If the tubes have to be kept for any length 

 of time, especially in the bloodheat incubator, each 

 should be capped with a rubber cap, tinfoil, or gutta- 

 percha tissue which has been soaked in 1-500 corrosive 

 sublimate solution. 



Anaerobic cultures.- Many organisms refuse to grow in 

 the presence of free oxygen, and various expedients have 

 to be adopted to exclude or remove it. The simplest of 

 all is to make the cultivation in a deep stab in glucose 

 agar or gelatin Narrow test-tubes filled three parts full 

 with the medium are best, and immediately before the 

 inoculation they should be placed upright in a beaker of 

 water, boiled for five minutes, and then cooled and 

 solidified in cold water. The object of this is to soften 

 the medium so that it does not split, as a dry medium 

 will, when the needle is plunged into it ; moreover, the 

 needle track closes up more readily, and the dissolved 

 oxygen is expelled. The tubes being cool, the inoculation 

 is made with a long thin wire, either straight or with a 

 closed loop at the end. It is inoculated and plunged 

 steadily into the centre of the medium, nearly to the 

 bottom, rotated, and then withdrawn, and the wool plug 

 is replaced and singed. The tube is then carefully heated 

 at the upper border of the medium so as to melt this 

 slightly and seal the puncture, and a well- fitting rubber cap 

 is applied while the tube is hot. The heating expels a 

 portion of the air, and, with a well-fitting cap, creates a 

 negative pressure within the tube, so that the residual 

 oxygen is not so readily absorbed, or the tubes may be 

 placed in a Buchner apparatus (see below). The tubes are 

 placed in the incubator at a suitable temperature, and 



