18 



A. KOHN AND M. LION 



due to freezing of E. coli, as indicated by lysozyme sensitivity, 

 showed that the cells are lysed by lysozyme immediately upon 

 thawing to an extent many times greater than that of cells 

 incubated at 37° for a short period of time after thawing (Fig, 

 2)13. It could further be shown that following this immediate 



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20 40 60 80 100 120 

 Minutes after thawing 



Fig. 2. Loss of sensitivity to lysis by lysozyme of E. coli at different times 

 after thawing. At times indicated by arrows 30 jug/ml of lysozyme was 

 added. Dotted line indicates control lysis up to the time of addition of 

 lysozyme. Each determination made in a separate tube from the same 

 culture. Optical density measured at 540 mjn/(TQf.'^). 



recovery after thawing, considered to be osmotic or mechanical, 

 there was a period of metabolic recovery which could be 

 inhibited by starvation or metabolic poisons such as KCN, but 

 not by chloramphenicol or 2,4-dinitrophenol (Fig. 3). 



Immediate damage to cells by freezing may be prevented by 

 adding to the bacterial suspension 'protective colloids' such as 

 skim milk. The explanation offered for the action of these 

 colloids is that they prevent the death of bacteria by crushing 

 due to extracellular ice when the inter-crystallic water films are 

 thin. Colloids increase the thickness of these films and thus 



