30 BACTERIOPHAGES 



petri dish; (2) the time required for plating each sample is less 

 than 30 seconds; (3) the plaque size is larger than that given by 

 the spreading method ; and (4) the efficiency of plating is often 

 higher. The agar layer method has permitted accurate kinetic 

 study of reactions that were too rapid to be followed by other 

 methods of phage assay, and has been indispensable in genetic 

 studies relying on recognition of different types of plaque. 



2. Dilution End-Points 



The dilution end-point method consists in finding the smallest 

 amount of the phage preparation that will bring about lysis of a 

 growing culture of susceptible bacteria. Subject to the assump- 

 tion that one phage particle will cause lysis, this method measures 

 the number of particles. However, if the adsorption is poor or 

 phage multiplication slow, lysis may occur only when a consider- 

 able number of phage particles are inoculated, and so the phage 

 population will be grossly underestimated. In such cases the 

 estimate can be improved by testing for presence of phage in all 

 tubes in which lysis failed. 



This method can be made quite precise by multiplying the 

 number of tubes tested at the limiting dilution. The phage 

 preparation is diluted to the point where each sample should 

 contain about one phage particle. Then 50 young cultures of 

 the host bacterium are each inoculated with a sample of the 

 diluted phage preparation and incubated. Each tube is then 

 checked for lysis or for presence of phage. If the test for infec- 

 tive particles is reliable, the proportion of cultures in which 

 phage multiplication did not occur will be equal to e~", from 

 which n, the mean number of infective particles per sample, can 

 be computed. The measurement fails unless roughly half but 

 not more of the tubes tested prove to contain phage, which means 

 that the method is too laborious for routine assays. It is in- 

 dispensable, however, in certain types of experiment, and the 

 student of phage should be familiar with the Poisson distribu- 

 tion, of which the computation mentioned is an application. 

 The accuracy of experimental results obtained by this method 



