MAINTENANCE AND PRESERVATION OF CULTURES 105 



as an absorbent or desiccating medium for preserving cultures. In the 

 first case soil that is suitably pulverized and screened is placed in culture 

 tubes, and enough water or very dilute culture medium is added to raise 

 it to about 60 per cent of its maximum water-holding capacity. It must 

 be autoclaved at least 1 hour^ and tested for sterility before use. After 

 the culture has sufficiently grown, it is simply allow^ed to become air dry 

 and is then suitably sealed and stored at room temperature or refrigerated. 



If soil is to be used merely as an absorbent, it is first air dried, then 

 tubed and autoclaved, and finally heated in a drying oven until moisture- 

 free, after which the tubes are stored in a desiccator until needed. They 

 are then inoculated with a broth culture or with a cell suspension washed 

 from an agar slope, as described under ''LyophiHzation." For preparing 

 a cell suspension a 2 per cent solution of peptone is useful. A 0.05- to 

 0.1-ml portion of this suspension is placed with a pipet on 1 ml of soil, and 

 the tubes are returned to a desiccator until dry or may be tightly stop- 

 pered if the soil already appears dry. 



A variation using silica gel granules as an absorbent has been proposed 

 by Dr. J. Lederberg, of the University of Wisconsin. Silica gel of the 

 grade known as 40, 6-16 mesh is used, and 1-1.5 g is placed in culture 

 tubes about 75 by 8 mm, which are then plugged and baked in an oven at 

 170°C for 2 hr. They are cooled and stored in a desiccator until needed. 

 A cell suspension in 2 per cent peptone is prepared and pipetted on to the 

 granules as described above. The tubes may then be hermetically 

 sealed, without evacuating, or stoppered and stored in a desiccator. The 

 method is still in the trial stage but has given promise of being successfully 

 adaptable. 



Other Desiccants 



Pieces of thread and strips of filter paper have long been used as absorb- 

 ents of spore suspensions of both aerobes and anaerobes. After immer- 

 sion in a spore suspension, followed by air drying, these materials can be 

 stored in glassine envelopes or culture tubes for many months with satis- 

 factory retention of viability. 



Deep Freezing 



The storage temperatures indicated in the follomng section for holding 

 cultures of various bacteria between routine transfers are only low enough 

 to retard or suppress normal growth; they are not necessarily the mini- 

 mum temperatures at which the cultures will survive. Temperatures 



1 Some technicians advise much more drastic sterilization, up to 2 hr at a time and 

 repeated three times at 1-day intervals. Excessive heating of soil should be avoided, 

 however, because it breaks down the organic components and may release toxic 

 products. We consider 1 hx at 15 lb as generally enough. 



