118 ALICK ISAACS 



showed 1 infective dose of virus for every 25 to 50 cells, and between 50 and 

 90 % of this amount was recoverable throughout the lag period. It seems 

 important to repeat these experiments with seed virus which does not show 

 the enhancement effect. Overman and Tamm (1957) incubated very large 

 doses of virus (18 X 10 6 infective doses) with pieces of chorioallantoic 

 membrane in vitro. The amount of virus taken up by the cells was not meas- 

 ured but there was only a slight decline, roughly 2-fold, between the third 

 and eighth hours of incubation in the amount of virus recoverable from ground 

 membrane pieces. 



Shaffer and Enders (1939) noted that 1| hours after inoculating 120 

 infective doses of herpes simplex virus on the chick chorioallantoic membrane, 

 only 10 doses could be recovered from ground membranes. However, further 

 experiments with antiserum made it appear that only a small proportion of 

 the virus had been taken up by the membrane in that time. Scott et al. (1953) 

 carried out similar experiments but with a much larger inoculum, 8 X 10 4 

 infective doses. The recovery of virus from ground membranes after 1, 4, and 

 G hours' incubation was usually less than 10 infective doses, but the uptake of 

 virus during this time was not measured. In further experiments of the same 

 kind, Modi and Tobin (1954) obtained a recovery of 1 to 10 % of the inoculum 

 in the interval 1 to 6 hours after inoculation. In this case the inoculum was 

 between 2.10 6 and 5.10 8 infective doses, but the amount absorbed is not known. 

 Wildy (1954) carried out a series of most ingenious experiments to investigate 

 the question of a possible eclipse phase for this virus. He inoculated eggs with 

 100 infective miits of virus and at different time intervals eggs were rotated 

 through 120°. The effect of this was that the bubble of air forming the 

 artificial air sac was carried around to a new area of the membrane, taking 

 along with it any virus which had not previously been fixed to the membrane. 

 In this way it was found that the total amount of inoculated virus was 

 partitioned between the two areas, 90 % of the inoculum being fixed to the 

 first area in 90 minutes. The total amount of virus which could be recovered 

 from membranes and overlying fluids declined gradually and was about 33 % 

 of the original amount after 4 to 6 hours. In this experiment the inoculum was 

 suspended in broth containing gelatin; after the period of incubation at 

 37°C, the eggs were chilled in order to solidify the gelatin and thus facilitate 

 harvesting of all the virus in the overlying fluid. Since the gelatin was found 

 experimentally to delay fixation of virus by the membrane, it is probable 

 that part of the 33 % of the virus recovered between 4 and 6 hours represented 

 virus not yet fixed to the membrane. Gostling and Bedson (1956) infected 

 trypsinized chick embryo cell suspensions by contact with herpes virus 

 overnight at 4°C. Under these conditions the uptake of virus was found to be 

 only about 1 %, or less, of the inoculum. When such infected cell suspensions 

 were incubated at 37°C, the level of infective virus recoverable from the cells 



