212 The Cultivation of Anaerobic Bacteria 



merit continued favor, and at the present time this method 

 is not employed. 



5. Exclusion of Atmospheric Oxygen by Means of 

 Various Physical Principles and Mechanical Devices. 



This has appealed to the ingenuity of many experimenters, 

 and many means of accomplishing the atmospheric exclusion 

 have been tried with success. 



The most simple plan is that of Hesse, who made a deep 

 puncture in recently boiled and rapidly cooled gelatin or 

 agar-agar, then covered the surface of the medium with 

 sterile oil (Fig. 56). The so-called "shake culture" is an- 



Fig. 55. Buchner's method of 

 making anaerobic cultures. 



Fig. 56. Hesse's method of mak- 

 ing anaerobic cultures. 



other very simple method, suggested by Liborius and Hesse. 

 The medium to be inoculated, contained in a well-filled tube 

 or flask, is boiled to displace the contained air, cooled so as 

 no longer to endanger the introduced bacteria, then inocu- 

 lated, the inoculated bacteria being distributed by gently 

 shaking. On cooling, the medium "sets," the organisms 

 below the surface remaining under anaerobic conditions. 



Kitasato first used paraffin as a covering for the inoculated 

 medium, his recommendation having recently been revived 

 and made successful use of by Park for the cultivation of the 

 tetanus bacillus. The paraffin floats upon the surface of the 



