TECHNIQUES 345 



stentors become abundant, the test tube can be emptied into a 

 culture jar which is carried forward as described. 



Alternatively, the test tube with its clone of stentors can be 

 emptied into an Ehrlenmeyer flask, plugged with cotton and fed 

 by repeated additions of food organisms, sub-culturing when the 

 flask becomes filled. Growing the food organisms separately 

 prevents over-nutrification and is therefore recommended for 

 developing clones as well as for producing very abundant and clean 

 cultures for biochemical studies. 



To obtain concentrated animals one can gently shake the flask 

 cultures to loosen stentors attached to the sides and pour the 

 contents into graduate cylinders; for at first the oxygen will be 

 uniform throughout and the stentors (at least coeruleus) will rapidly 

 sink to the bottom in mass and the overlying fluid can be decanted. 

 If it is now desired to free these animals from most of the food 

 organisms one may take advantage of the speed with which most 

 races of coeruleus swim away from the light — or perhaps the reverse 

 in the case of green polymorphus and niger. Whiteley introduces 

 the concentrated animals at the lighted end of a large, horizontal, 

 covered tube with both ends bent upward and filled with Millipore 

 filtered medium (Fig. 96E). Stentors soon migrate to the lighted 

 end, leaving the slower bacteria and food organisms behind, and 

 are promptly removed for study. 



Other methods which have successfully been employed for the 

 cultivation of stentors will now be reviewed. First we have to 

 consider the basic fluid medium. Distilled w^ater is not used 

 because it is injurious and tap water is avoided because it picks up 

 metals in the pipes and may be chlorinated. Natural waters from 

 ponds and lakes are preferred. They may be freed from contaminat- 

 ing organisms either by previous boiling or by fine-filtering — the 

 latter is recommended. These waters will contain dissolved sub- 

 stances natural to the Stentor habitat, but many investigators 

 recommend the addition of a mixture of inorganic salts. (I attempt 

 to supply these along with organic materials in the added milk.) 

 Peters (1904) in particular emphasized the importance of the salt 

 content of the medium and suggested the following mixture, 

 figures representing moles of the salts: CaCU (0-0005), K2HPO4 

 (0-00015), NaNOa (0-00015), and MgS04 (0-00015). This solution 



