Holophryidae 101 



four or five generations are usually produced at room temperature 

 (21 C.) in every 24-hour period, and at least two Paramecia are needed 

 for a single Didinium to attain its full growth prior to division. At higher 

 temperatures the voraciousness and rapidity of reproduction of Didinium 

 are astonishing, in that as many as nine generations may be produced 

 within 24 hours, and the task of supplying Paramecia to large cultures 

 becomes overwhelming. 



When Didinia are needed for class use or for research purposes over a 

 period of days or weeks, they are therefore best maintained in small 

 stock cultures, made up, for example, in watch glasses. In the prepara- 

 tion of these cultures chemically clean watch glasses are filled with the 

 desired amount of spring water or pond water (filtered or boiled to re- 

 move or kill Entomostraca) . Or they may be filled with 0.01% modified 

 Knop solution or with diluted hay infusion from a Paramecium culture. 

 Paramecia are then concentrated with the centrifuge and transferred to 

 the watch glasses, after which a few specimens of Didinium are added. A 

 temperature of about 20 C. is the optimum for the growth of Didinium. 



The Paramecia should react normally soon after being transferred 

 to the watch glasses. A medium that is injurious to Paramecium is un- 

 suitable for the growth of Didinium, and, conversely, any fluid medium 

 in which Paramecium exhibits normal behavior and retains its normal 

 cell configuration is usually favorable for Didinium. Occasionally spring 

 water and pond water have a slightly unfavorable hydrogen-ion concen- 

 tration which needs to be corrected by the addition of about 5 cc. of 

 phosphate buffer mixture of pH 6.8 to each 100 cc. of water. The range 

 of hydrogen-ion concentration which active specimens of Didinium can 

 tolerate is considerable. It varies from pH 5.0 to pH 9.6, but the 

 optimum lies between pH 6.8 and pH 7.2. 



Modified Knop solution of suitable concentration (about 0.01%) may 

 be prepared conveniently from the usual three stock solutions: namely, 

 10% Ca(N0 3 ) 2 , 5% KNO3 and, 5% MgS0 4 . 7 H 2 0. A 1% solution is 

 first made up by adding the following amounts of the stock solutions to 

 150 cc. distilled water: 10 cc. Ca(N0 3 ) 2 , 7-5 cc. KN0 3 and 7.5 cc. 

 MgSO.j. Then a 0.01% solution is prepared by adding 1 cc. of the 1% 

 solution to 99 cc. of distilled water. Finally, 5 cc. of NaOH-KH 2 P0 4 

 buffer mixture of pH 6.8 is added to each 100 cc. of 0.01% solution. 



This solution is often preferable to spring water or pond water for 

 maintaining cultures for experimental purposes, in that its chemical 

 composition is known and constant. It has an added advantage in that 

 it is unfavorable for the growth of bacteria. 



Hay infusion, either from a flourishing Paramecium culture or from 

 an old, declining culture, usually has a favorable hydrogen-ion concentra- 

 tion, but it is sometimes too concentrated, and in the preparation of 



NAVY PRO IFTT m 



