Exclusion of Atmospheric Oxygen 233 





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 

 medium, melts during sterilization, but does not mix with 

 it, and "sets" when cool. The inoculation is to be made 

 while the culture medium is warm, after boiling and before 

 the paraffin sets. 



Koch studied the colonies of anaerobic organisms by culti- 

 vating them upon a film of gelatin 

 covered by a thin sheet of steril- 

 ized mica, by which the air was 

 excluded. 



Salomonsen has made use of a 

 pipet for making anaerobic cul- 

 tures. It is made of a glass tube 

 a few millimeters in diameter, 

 drawn out to a point at each 

 end. The inoculated gelatin or 

 agar-agar is drawn in while lique- 

 fied and the ends sealed. The 

 tube, of course, contains no air, 

 and perfect anaerobiosis results. 



In the chapter upon "Tetanus " 

 I describe a method of cultivating 

 anaerobic organisms that gave ex- 

 cellent results, though its applica- 

 tion is limited, as it applies only 

 to broth cultures and best to large 

 quantities. 



Theobald Smith has found the 

 fermentation-tube and various 

 modifications of it excellently well 

 adapted to the growth of anae- 

 robes, which, of course, grow only 

 in the closed limb. 



Hens' eggs have been used for 

 anaerobic cultures, and in them 

 the tetanus bacillus grows remark- 

 ably well. Conditions of anaero- 

 biosis are, however, not perfect, 



as can be shown by the behavior of the egg itself. If oxygen 

 be completely shut out by oiling or varnishing the shell, 

 a fertile egg will not develop. 



Figs. 59, 60. Wright's 

 method of making anaero- 

 bic cultures in fluid media 

 (Mallory and Wright). 



