PRESERVATION OF CULTURES 55 



surrounding the tubes is raised to approximately —5° C. 1 by supplying 

 additional methyl cellosolve that has been precooled to approximately 0° C. 

 The bath is maintained at this level continuously until the preparations 

 appear thoroughly dry. In drying, the serum contracts slightly to form a 

 well-defined chalky pellet. When the pellets are apparently dry, the tubes 

 are raised above the bath and evacuation is continued for one-half to three- 

 quarters of an hour at room temperature to insure as complete removal of 

 water as possible, after which time they are sealed off with a gas-oxygen 

 torch (fig. 14 A). On the following day each tube is tested for the presence 

 of a good vacuum by means of a high-frequency, spark coil tester, and only 

 those tubes which show such a vacuum are retained. Tubes not maintain- 

 ing a satisfactory vacuum are very rarely encountered in actual practice. 

 Quadruplicate tubes are regularly made for each stock culture, and the 

 finished preparations are stored in a refrigerator. 



In recultivating the molds, the tubes are marked with a file scratch, sur- 

 face-sterilized, and the tube broken inside a wrapping of sterile cotton. 

 The content, which is in the form of a well-formed pellet (fig. 13 B), is then 

 dissolved in 1 to 2 cc. of sterile broth or water. This is streaked on agar 

 plates and colonies are allowed to develop. New isolations can be made 

 within a period of a few days. It is possible, of course, to go directly from 

 the lyophile tubes into flasks or other cultures used in actual experiments, 

 but generally speaking, much larger quantities of material would need to 

 be processed. 



The feasibility of preserving molds in lyophile form over long periods 

 has by no means been proved, but results to date are very encouraging. 

 Should it be found that spores of molds, like bacterial cells, can be kept 

 viable by this method for many years, it will prove ideal as a means of pre- 



1 Wickerham and Andreasen in 1942 governed the temperature at which the sus- 

 pension was dried by adjusting the level of the tubes above a bath which was kept 

 at a very low temperature. Subsequent to this, Wickerham developed the procedure 

 outlined above. 



contained and portable unit of sixty tube capacity (designed by Dr. L. J. Wickerham) 

 which utilizes a column of anhydrous calcium sulphate ("drierite") to collect water 

 vapor removed from drying preparations. Ai and B\, Glass manifolds; A 2 and B 2 , 

 Thermometers; A 3 , Dewar flask containing water vapor trap immersed in CO2 

 ice and methyl cellosolve; B u Column of drierite; A^ and B b , Freezing bath con- 

 taining CO2 ice and methyl cellosolve in which preparations are immersed for 

 quick freezing and subsequent temperature control; Ae, and Be, Vacuum pump; At, 

 Vacuum guage (mounted at opposite end of apparatus shown in figure B) ; As and B s , 

 Gas-oxygen torch for sealing off dried preparations; B 9 , Terminals on which finished 

 tubes are mounted to be tested for presence of good vacuum by means of a high fre- 

 quency spark coil tester (not shown); Bio, Oxygen tank; B u , Screw lift for raising 

 and lowering manifold and attached tubes (In figure A , manifold is raised and lowered 

 manually and locked into position by means of a wing bolt). 



