A. H. DOERMANN 647 



the addition of one part in ten of a high titer T6 phage filtrate (concentration of T6 

 in lysing medium was ca. 4 X 10^ particles per ml.) and cyanide brought to a final 

 concentration of 0.01 M. Specially designed experiments showed that at this concentra- 

 tion the cyanide does not inactivate free phage particles, nor does the amount which 

 reaches the plate affect titration by interference with plaque development. 



T6 was used as the lysing phage because in several experiments it proved to be a 

 more effective lysing agent than any of the other T phages tested. Since only single 

 stocks of the phages were compared in the early experiments, the superiority of T6 

 over the other phages may have been due to a difference in the particular stock used, 

 and not to an inherent difference among the phages. In fact, later experiments with 

 different T6 stocks showed marked differences in lysing efficiency, and phage titer 

 proved to be a poor criterion of lysing ability. The experiments described here were 

 made with T6 stocks selected for their ability to induce lysis from without. The se- 

 lections were made on the basis of nephelometric comparisons. 



Platings were made in agar layer (0.7 per cent agar) poured over nutrient agar plates 

 (1.3 per cent agar), and in order to assay T4 in the presence of high titer T6, the in- 

 dicator strain, B/6, was used. B/6 is completely resistant to T6 (no host range mutants 

 have so far been found which will lyse the strain used here) and gives full efficiency 

 of plating (compared to B) with T4. 



EXPERIMENTAL 



Experiments with the Stayidard Lysing Medium. — The experimental procedure used 

 consisted essentially of a one-step growth experiment (5) with certain modifications. 

 B/r/1 cells in the exponential growth phase were concentrated by centrifugation to 

 about 10^ cells per ml. To these concentrated bacteria T4r48 was added and this ad- 

 sorption mixture was incubated for 1 to 2 minutes with aeration, allowing at least 80 

 per cent of the phage to be adsorbed to the bacteria. Then a 40-fold or larger dilution 

 was made into growth medium containing anti-T4 rabbit serum. The serum inactivated 

 most of the residual unadsorbed phage. After several minutes' incubation in the serum 

 tube, a further dilution was made to reduce the serum concentration to one of relative 

 inactivity. The resulting culture will be referred to as the source culture (SC). The 

 entire experiment was carried out with the infected bacteria from SC. The titer of 

 infected B/r/l in this tube was approximately 10^ cells per ml. 



Simultaneously with the dilution into the tube containing serum, another dilution 

 from the adsorption tube was made. From the latter an estimation of the unadsorbed 

 phage was made by assaying the supernatant after sedimentation of the cells. This 

 step permits calculation of the multiplicity of infection (5). 



From SC a further dilution of 1:20 was made at some time before the end of the 

 latent period. The resulting culture, containing approximately 5 X 10' infected bac- 

 teria per ml., was used for determining the normal end of the latent period and for 

 estimating the average yield of phage per infected cell. It will be called the control 

 growth tube (GT). In addition, a number of precisely timed 20-fold dilutions were made 

 from SC into lysing medium. These were titrated after they had been incubated in 

 the lysing medium for 30 minutes or longer. Serial platings from the lysing medium 

 cultures over a longer period of time have shown that the phage titer remained constant 

 after 30 minutes' incubation. The titer calculated from these platings, divided by the 



77 



