The Technique of Culturinq Amoeba protcus. 243 



the water. When the test-tube culture is well established, the 

 contents may be transferred to a larger vessel,* but more than one 

 attempt may have to be made before success is obtained. It is 

 unwise, however, to reject as useless any culture started in this 

 way until it has been given a year's trial, as it frequently happens 

 (especially if the inoculation be made in the summer or autumn, 

 when they are most plentiful) that the amceba? proceed to 

 " encyst," and signs of success may not be evident for six or nine 

 months. 



The somewhat tedious method described above, especially if 

 the amoebai be derived from a variety of sources, results in the 

 formation of a steady culture. If, in addition, the culture has been 

 reinoculated at different times of the year, the resulting lahoratory 

 culture ivill possess the advantage of yielding continuous sujjplies 

 throughout the season, an advantage not possessed by a successful 

 culture derived from one gathering only. Such a culture will be 

 dormant during regularly recurring periods. 



Another method of culturing out the amoebse from a natural 

 source is to collect water- weeds and debris from a likely place, e.g. 

 ditches, slow-running streams, ponds, etc., in autunm, and to store 

 these in a quantity of water during the winter. As the weeds die 

 down they give rise to an abundance of life. Eventually a mass 

 of debris is formed which may yield amoebae without further 

 treatment, or which, when added to fresh water and wheat, 

 eventually yields sufficient to start a laboratory culture. It is 

 often necessary to sub-culture from this, by the methods already 

 described, to get rid of oligochsete and other pests which are likely 

 to be present also. 



Some Exigencies and How to Meet Them. 



1. The wheat may be largely used up to produce a too luxuriant 

 algal growth. In this case it is well to start a new culture 

 carrying oif as many amoebse as possible, discarding the algaB. 



2. From time to time it is a good plan gently to pour off the 

 top water from a culture and to add fresh aerated water. Where 

 it is safe to use tap water the culture may from time to time be 

 placed under a dripping tap. This revives the amoeba?. Theo- 

 retically the green weeds ought to oxygenate the water, but prac- 

 tically, on account no doubt of various metabolic products, the 

 cultures, even when well (stocked with green algte, are wonderfully 

 revived by a change of water. 



3. Oscillaria of various species are troublesome. Where they 

 cannot be removed the culture should be frequently well stirred. 



4. Hydra, especially Hydra vulgaris, is useful in keeping 

 down crustaceans, if they be present. 



* See list of things required. 



