84 CULTIVATION OF ANAEROBIC BACTERIA. 
test tube for about half an inch, as shown at 6b, 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. 
Fie. 54. Fie. 55. 
This method is more convenient than that previously described 
and the only objection to it is that the oxygen is not completely ae: 
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 anaérobics. 
Method of Esmarch.—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 
