88 "PURE MIXED CULTURES" OF AMCEBA 



days, and, if all bacteria are killed, wash away the 

 acid, and plant the cysts under sterile precautions 

 with a pure culture of the particular bacillus you 

 select. 



If any difficulty occurs at this point, an analysis 

 may be necessary to determine what bacteria have 

 escaped the action of the germicide, so that by 

 plating out it may be removed. To decide if the 

 culture is pure, treat amosba cysts in culture with 

 a gelatin-liquefying bacillus with the germicide, 

 and transfer to a pure culture of a non-liquefying 

 organism. 



Some Notes on Various Means employed by Other 

 Authors and Ourselves in the Separation of 

 Amcebce or their Cysts from Accompanying 

 Bacteria in the Preparation of Pure Mixed 

 Cultures. 



A. MECHANICAL MEANS 



(1) " Picking off" a single cyst. It is possible to 

 accomplish this free from all living bacteria, but its 

 success is largely dependent on good fortune, and 

 pure cultures made in this way should be carefully 

 examined. One cannot say that, because a cyst 

 looks free from bacteria, it really is so ; as we have 

 many times examined such cysts by the jelly 

 method of staining, and found bacteria, which were 

 invisible when unstained, closely adhering to the 

 edges and to the surface of the cyst. 



The difficulty in separating a single cyst by 

 picking it off is due to the fact that, if the jelly 

 surface is dry, the cyst is firmly adherent (in old 

 cultures almost imbedded), and will not enter the 



