78 "PURE MIXED CULTURES" OF AMCEBA 



complex mixture of chemical substances produced 

 by the accompanying growth of a large variety of 

 different bacteria in the same culture, and subse- 

 quently to isolate the amcebas or their cysts from 

 the presence of all other living organisms. 



At the outset we were aware of the difficult 

 nature of the task from reference to the extensive 

 literature on the subject. Many observers are 

 stated to have obtained pure mixed cultures by 

 various means, but so far as we are aware the 

 result has not always been verified by a bacterio- 

 logical analysis, and in our opinion no culture 

 could be accepted as pure without this pre- 

 cautionary measure. The preparation of a pure 

 mixed culture involves two distinct processes, viz. 

 (1) the separation from its fellows of a single 

 individual amoeba, and (2) its subsequent culti- 

 vation with a pure strain of a bacterium. The 

 former is a simple matter and can be carried out 

 in ten minutes ; the latter is most difficult and has 

 occupied our attention for four months. The 

 two stages will be described separately. 



1. THE SEPARATION FROM ITS FELLOWS OF A 

 SINGLE AMOEBA CYST OR LIVING AMCEBA 



We have found it possible to pick off a single 

 amoeba cyst from a culture by means of a fine 

 capillary tube, and to place it on the surface of 

 a sterile agar plate. This has been done on three 

 occasions, but only on one of them was the 

 amoeba cyst free from all bacteria. At other times, 

 although apparently only one cyst was drawn into 

 the tube, it could not be recovered, probably 



