82 CULTIVATION OF ANAEROBIC BACTERIA. 



test tube for about half an inch, as shown at b, Fig. 53. The space 

 above the cork is then filled with melted sealing wax, which I have 

 found to prevent leakage better than paraffin, which contracts upon 

 cooling. The test tube is inverted while hydrogen is passed through 

 the tube c, and by reason of its levity the gas quickly passes through 

 the cotton air filter and displaces the oxygen in the test tube (Fig. 

 54). After allowing the gas to flow for a few minutes the outlet 

 tube is first sealed in a flame and then the inlet tube. As the cotton 

 filter is interposed between the rubber stopper and the culture mate- 

 rial, no special precautions need be taken for the sterilization of the 

 rubber cork and the glass tubes which it carries. 



FIG. 54. 



FIG. 55. 



This method is more convenient than that previously described, 

 and the only objection to it is that the oxygen is not completely re- 

 moved from the film of solid gelatin or agar attached to the walls of 

 the test tube. But by passing the hydrogen for a long time it would 

 seem that by diffusion the oxygen remaining in this thin layer 

 would be gotten rid of. At all events, this method will serve for all 

 except the very strict aiiaerobics. 



Method of Esmarcli. The following method has been proposed 

 by Esmarch : Three roll tubes are made in the usual way, and into 

 these liquid gelatin, that is nearly cooled to the point of becoming 

 solid, is poured. This fills the tube without melting the layer of 



