CULTIVATION OF ANAEROBIC BACTERIA. 79 



St-emberg prepares three Esmarch roll cultures. The cotton 

 plug is pushed down into the tube for a short distance. The 

 end of the tube is closed with a soft-rubber stopper carrying 

 two glass tubes. This stopper is pushed below the level of 

 the tube half an inch, and the space is filled with melted seal- 

 ing-wax. The test-tube is inverted and hydrogen gas passed 

 in through one of the tubes. The gas diffuses through the 

 cotton plug into that portion of the tube containing the cult- 

 ure. After a few minutes the outlet tube is sealed and then 

 the inlet tube. It is not necessary to sterilize the rubber 

 stopper, as the cotton plug is interposed between it and the 

 culture. 



Ravenel places the culture in an air-tight chamber par- 

 tially filled with pyrogallic acid and a 10 per cent, solution 

 of caustic potash. The air is exhausted with an air-pump 

 and the chamber is then hermetically sealed. The chamber 

 contains an upright shelf-stand on which Petri dishes are 

 placed. Any oxygen left in the chamber, or which enters, is 

 absorbed by the pyrogallic acid. 



Koch covers the surface of a glass plate containing the cult- 

 ure with a mica plate, thus excluding oxygen. 



Botkin uses a large bell-jar, which is stood in a dish con- 

 taining liquid paraffin. Two rubber tubes are carried through 

 the paraffin into the jar. Hydrogen gas is then sent into the 

 jar through one of the tubes, and when only pure hydrogen 

 escapes from the other the tubes are withdrawn, the solidify- 

 ing paraffin closing the holes so that no oxygen can enter the 

 jar. It is advisable to put a little pyrogallic acid into the 

 jar in case of leakage. 



Roux suggests still another method. The medium is lique- 

 fied and inoculated. While still liquid it is drawn into a 

 long small-calibred piece of glass tubing, the ends of which 

 have been slightly drawn. When the tube is full the ends 

 are sealed. When the colonies have developed, the tube is 

 broken with a file or diamond and the culture transplanted. 



Any of these methods is subject to such modification as 

 may be desired by the individual investigator. 



