CHAPTER III 



ENUMERATION OF BACTERIOPHAGE 

 PARTICLES 



Quantitative work with bacteriophages depends on simple 

 and accurate means of counting the particles. It is essential to 

 be able to determine relative numbers of phage particles in 

 different samples with precision and it is often important to be 

 able to measure the absolute number of particles. 



Three principal assay methods have been used: (7) plaque 

 counts on nutrient agar plates seeded with phage-susceptible 

 bacteria, (2) dilution end-points using lysis of fluid bacterial 

 cultures as an indicator for presence of phage, and (3) measure- 

 ments of the length of time required for lysis of a standard fluid 

 bacterial culture. 



1. Plaque Counts 



Only the first method mentioned is generally useful. The 

 method was originally described by d'Herelle in 1917. An ap- 

 propriate dilution of a phage preparation is mixed with a con- 

 centrated suspension of susceptible bacteria and an aliquot of 

 the mixture is spread on the surface of an agar plate. On in- 

 cubation the bacteria grow as a film, spotted with circular clear 

 areas or plaques produced by the lytic action of the bacterio- 

 phage. D'Herelle found that large amounts of phage could be 

 recovered from the plaques, but that none was present in the 

 unlysed areas of the plate. Since the number of plaques ob- 

 tained was directly proportional to the amount of phage prepara- 

 tion spread on the plate, the plaques could be understood as 

 phage colonies containing the descendents of single phage 

 particles. It does not follow, however, that every phage particle 



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