INTERACTIONS OF PROTOPLASMIC MASSES. Ill 



utes and then suddenly displaced by means of a glass filament. 

 This procedure would frequently result in a large mass of proto- 

 plasm being torn from the organism. 



(b) Culture. 



Hollow-ground >lides, each containing two concavities, were 

 used as receptacle- in cultivating the organisms. Seven drops of 

 the desired culture medium were put into each concavity and the 

 animals were transferred to them. After this the slides were 

 labelled and placed in transparent glass moist chambers which 

 were kept before a window, unless otherwise stated. 



At first the method described by Jennings ('16) for Dirfln^ia 

 was used in preparing the culture medium, i.e., a quantity of 

 ooze from ;i pond in \\hich the organisms were found to be living 

 was shaken up in water, this was allowed to settle, and then the 

 water was decanted oft and used. Although other investigators 

 have obtained successful results by using culture media prepared 

 in tlii> way, several objections to its use in these experiments 

 were evident: first, fresh material was hard to obtain, since no 

 ponds were near the laboratory in which the work was done; 

 secondly, a constant culture medium was not insured, for the 

 plant and animal life in a given j>ond vary constantly; and 

 finally, it was necessary ever to be on guard against possible con- 

 tamination with wild specimens and injurious chemical agents. 

 After encountering these difficulties for two months the- above 

 method was discarded in favor of a hay infusion culture medium 

 which dispensed with all of the objections mentioned, and in 

 which the organisms thrived even better. It was prepared as 

 follows: 10 grams of clean timothy hay was placed in a clean 

 beaker containing 250 c.c. of neutral distilled water. This was 

 boiled slowly for five minutes, then strained through two thick- 

 nesses of cheese cloth, after which it was .-tc.ivd. in quantities of 

 about 3 cc., in small sterile test tubes. Tin- tube- were then 

 plugged with cotton and placed in boiling water for fifteen min- 

 utes. Two days later they were subject i-d to boiling water again 

 for the same period of time, in order to kill any }>. u-ria which 

 might have resisted the first sterili/ation by being in the spore . 

 Mage. The liquid in the tubes constituted what wa< known as 

 stock solution; it would keep for month^ wit IK mt deteriorating 



