32 CULTURE METHODS AND TECHNIQUE 



Collodion for the dialyzing sacks is prepared by dissolving five 

 grams of guncotton in 100 cc. of a solvent consisting of equal 

 parts of absolute alcohol and sulfuric ether. The sacks are prefer- 

 ably prepared by the test tube method, and convenient tubes will 

 measure about 30 by 200 mm., with lip. The tubes must be thor- 

 oughly cleaned (a final rinsing with ether being desirable) and 

 dried. The tube is held in a slanting position and gradually re- 

 volved as about 50 cc. of the collodion is slowly poured in, and 

 thus an even roll, or coating throughout, with no bubbles, should 

 be effected. A second coating is obtained by similarly revolving 

 the tube as the surplus collodion is poured out. The tubes are 

 then supported upright, mouth downward, and the drip at the lip 

 removed. They should then be dried rapidly in a draft at a 

 window or preferably under an electric fan, or by exhaust, con- 

 stantly revolving each tube meanwhile to maintain even distribution 

 of the collodion. When dry, first free the edge of the collodion 

 from the tube with a scalpel and then immerse the tube in a vessel 

 of water so that as the sack is made free water will pass in between 

 the collodion film and the glass, and thus the removal of the film 

 may be readily effected. 



The silicate mixture is put in these tubes and they are tied 

 securely with a rubber cord and suspended in water over night. 

 Running tap water may be commonly employed, but for more 

 accurate work it will be necessary to use changes of distilled water. 

 The dialyzed liquid should react neutral to litmus and should show 

 only a trace of chlorides with silver nitrate. 



The nutrient solution employed may be that mentioned on page 

 26, except that the concentration is five times as strong, as pre- 

 viously indicated.' It is now necessary to boil both the silicate 

 preparation and the nutrient solution a few minutes to remove air. 

 Then cool down to room temperature, mix and stir, and put into 

 the separatory funnel to facilitate pipetting into tubes or other 

 vessels to be employed in the work. The silicate in the desired 

 vessels is then autoclaved for about fifteen minutes. This should 

 insure thorough solidification. Slanting tubes may also be pre- 

 pared. It may be necessary for the operator to vary the concen- 

 tration of the salts, or to experiment with small quantities when 

 using unusual proportions of mineral salts. 



