ANAEROBIC CULTURE IN LIQUID MEDIA. 



Cultures of Anaerobes in Liquid Media. It is necessary to 

 employ such in order to obtain the toxic products of the growth 

 of anaerobes. Glucose 

 broth is most convenient. 

 It is placed either (i) in a 

 conical flask with a lateral 

 opening and a perforated 

 india-rubber stopper, 

 through which a bent glass 

 tube passes, as in Fig. 27, a, 

 by which hydrogen may be 

 delivered, or (2) in a conical 

 flask with a rubber stopper a 



furnished with two holes, as ,?,,,, 



a. rlask lor anaerobes in liquid media. Lateral 



in Fig. 27, b, thrOUgh a tube nozzle and stopper fitted for hydrogen supply, b. A stop- 

 r . per arranged for a flask without lateral nozzle. 



in one of which hydrogen is 



delivered, while through the tube in the other the gas escapes. 

 The inner end of the gas delivery tube must in either case be 

 below the surface of the liquid ; the inner end of the lateral 

 nozzle in the one case, "and the inner end of the escape tube in 

 the other, must of course be above the surface in the liquid. 

 The single tube in the one case and the two tubes in the other 

 ought to be partially drawn out in a flame to facilitate subsequent 

 complete sealing. The ends of the tubes through which the gas 

 is to pass are previously protected by pieces of cotton wool tied 

 on them. It is well previously to place in the tube, through 

 which the hydrogen is to be delivered, a little plug of cotton 

 wool. The flask being thus prepared, it is sterilised by methods 

 B (2) or B (3). On cooling it is ready for inoculation. In the 

 case of the flask with the lateral nozzle, the cotton-wool covering 

 having been momentarily removed a wire charged with the 

 organism is passed down to the bouillon. In the other kind of 

 flask the stopper must be removed for an instant to admit the 

 wire. The flask is then connected with the hydrogen apparatus 

 by means of a short piece of sterile india-rubber tubing, and 

 hydrogen is passed through for half an hour. In the case of 

 flask (i), the lateral nozzle is plugged with molten paraffin 

 covered with alternate layers of cotton wool and paraffin, the 

 whole being tightly bound on with string. The entrance tube 

 is now completely drawn off in the flame before being discon- 



