POLLARD AND KENNEDY 



\0,000r \00,000r 



139 



1,000,000 



~b.000.000r 



Q 



z: 



O" 



z: 



O: 



ClipS 



98% 



75% 



8% 



0.07% 



Fig. 2. A schematic representation of the statistical analysis of radiation damage. It is 

 supposed that to escape damage a microsomal particle must escape an ionization in its 

 structure, or even the diffusion of an active agent from ionization to its structure. The 

 probability of escape can be seen to be clearly dependent on the particle size and the amount 

 of ionization. 



be a migration by diffusion of the energy release to the extent of 30 A. This 

 little spread of energy has been indicated by a dotted line around the ioniza- 

 tion. It can be seen that the chance of escape at 10,000 r is very good, the cal- 

 culated figure being 98 per cent. In the second region the microsomal particle 

 has been given an increase of 30 A in radius to allow for the possibility of 

 migration of energy, and now each ionization has to be considered as a single 

 dot. The chance of escape is now less but still high; it is estimated at 75 per 

 cent. With 1,000,000 r it is only 8 per cent, and with 3,000,000 r the chance of 

 escape is very remote indeed, and it is expressed by the unlikely figure of 0.07 

 per cent. 



The experimental method, therefore, is to expose bacterial cultures held in 

 a static condition by being chilled to 4° C to X rays, to permit them to warm 

 up, and then to add labeled phosphate or sulfate to them and observe the up- 

 take of radioactivity as compared with the uptake of the unirradiated prepara- 

 tion. The ratio of the two is then compared with the chance of escape, and 

 deductions regarding the size of the particle are made from this probability. 



EXPERIMENTAL METHOD 



The experiments with phosphate are complicated by the fact that very com- 

 monly phosphate buffer is used in minimal media. This means that a large 

 excess of cold phosphate is present, rendering uptake somewhat difficult. 

 Accordingly the procedure we evolved was to grow the bacteria overnight in 

 the C minimal medium as described by Roberts, Abelson, Bolton, Britten, and 

 Cowie [7] and then transfer for a 90-minute growth period to Tris buffered 

 medium containing 80 mg of phosphate/1. The 90 minutes permitted the bac- 

 teria to establish themselves satisfactorily in the new medium; they were 

 then chilled and irradiated with a 250-kv X-ray machine at approximately 

 3000 r/minute. Subsequent to irradiation both the control and the irradiated 

 sample were warmed in a 37° bath and an appropriate number of counts of 

 radioactive phosphate was added to each one. The bacteria were sampled at 



