212 BACTERIOLOGY 



been expelled. The drawn-out portions of the tubes can 

 then be sealed in the gas-flame without fear of an explosion. 

 The protruding end of the rubber stopper is then painted 

 around with melted parafRn and the tube rolled in the way 

 given for ordinary Esmarch tubes. A tube thus prepared 

 and containing growing colonies is shown in Fig. 38, B. 



The development that now occurs is in an atmosphere of 

 hydrogen, all oxygen having been expelled. During the 

 operation the tube containing the liquefied gelatin should 

 be kept in a water-bath at a temperature sufficiently high 

 to prevent its solidifying, and at the same time not high 

 enough to kill the organisms with which it has been inocu- 

 lated. 



One of the obstacles to the successful performance of this 

 method is the bubbling of the gelatin, the foam from which 

 will often fill the exit-tube and sometimes be forced from it. 

 This may be obviated by reversing the order of proceeding, 

 viz.: roll the Esmarch tube in the ordinary way with the 

 organisms to be studied, using a relatively small amount of 

 gelatin, so as to have as thin a layer as possible when it is 

 rolled. Then replace the cotton plug with the sterilized 

 rubber stopper carrying the glass tubes through which the 

 hydrogen is to be passed, and allow the hydrogen to flow 

 through as in the method first given. The gas now passes 

 over the gelatin instead of through it, and consequently no 

 bubbling results. In all other respects the procedure is the 

 same as that given by Frankel. 



Method of Kitasato and Weil. — For favoring anaerobic 

 conditions Kitasato and Weil have suggested the addition 

 to the culture-media of some strong reducing-agent. They 

 recommend formic acid or sodium formate in 0.3 to 0.5 per 

 cent.; glucose in 1.5 to 2 per cent.; or blue litmus tincture 

 in 5 per cent, by volume. This is, of course, in addition to 



