APPENDIX 451 



agar layer should be allowed to harden with the Petri dish rest- 

 ing on a leveled sheet of plate glass, to insure uniform distribu- 

 tion of the virus plaques over the plate surface. If the melted 

 soft agar is held at 46 ° C. for much over an hour the agar will 

 start to gel and poor plates will result. Neither bacteria nor 

 viruses seem to be harmed by a short stay at 46 ° C. 



Advantages of the agar layer method are that the host bacteria 

 and virus particles may be more uniformly distributed over the 

 surface of the plate than by the spreading technique and a 

 larger volume of virus may be plated. The greater porosity of 

 the soft agar layer permits more rapid diffusion of the phage 

 particles and development of larger plaques than are obtained 

 by the spreading technique, and hence variations in plaque 

 morphology may be more readily studied. The chief disadvan- 

 tage is that melted agar must be maintained at 46 ° C. until 

 used. Use of this method has tremendously facilitated the 

 plating of bacterial viruses since it is more rapid than the spread- 

 ing technique. 



The dark field colony counter (Spencer Optical Co.) is a good 

 device for observation and counting of plaques, since the plaques 

 appear as dark holes in the brightly illuminated layer of bac- 

 terial growth. An automatically recording counting apparatus 

 described by Varney (1935) marks each plaque as it is counted 

 and at the same time operates an electric recorder. 



Efficiency of Plating 



The efficiency of plating may be defined as that proportion 

 of viable phage particles which actually form plaques when 

 plated. It depends on the ability of the phage particles to be- 

 come adsorbed to a suitable host cell and on the ability of these 

 infected cells to liberate more phage again, thus initiating a chain 

 reaction which results in a plaque. The plaque continues to 

 increase in size until the stationary phase of bacterial growth is 

 reached, at which point infected cells no longer liberate virus. 

 A phage particle which does not become absorbed to a host 



