116 ALICK ISAACS 



appears that only 3.2 %, on the average, of the amount of infective virus 

 taken up could be recovered from the membranes during the lag period. 

 Although a number of different techniques of extraction were used, the 

 recovery of virus was not improved. 



Schafer and Munk (1952) inoculated very large doses (10 9 — 10 10 LD 50 ) of 

 fowl plague virus into de-embryonated eggs, washed out the inoculum with 

 buffer 30-105 minutes afterwards, and then measured the infectivity of 

 homogenized membranes. During the lag period, the infective titer was 10~ 5 

 to 10~ 6 . This represents 0.01 % of the virus inoculum, but it is not known how 

 much of the inoculum was actually taken up by the cells. 



Granoff (1955) inoculated 10 7 - 4 LD 50 of Newcastle disease virus (NDV) 

 into the allantoic cavity of chick embryos and calculated that 80 % of the 

 seed was adsorbed after 2 hours (this figure would be an overestimate if a 

 significant degree of virus inactivation occurs during 2 hours' incubation at 

 37°C). However, less than 1 % of the inoculated virus could be detected in 

 ground membrane extracts at this time. More recently, Kubin et at. (1957) 

 studied the growth of NDV in monolayers of chick embryo lung epithelium. 

 The technique was similar to that used by Rubin et al. (1955) for western 

 equine encephalitis virus. During the lag period, the number of virus infective 

 doses recoverable by freezing and thawing the cells was about 1 % of the 

 number of cell yielders (or infective centers). This proportion remained 

 constant throughout the lag period. 



Rubin (1955) studied the development of the Rous sarcoma virus in 

 suspensions of tumor cells. By inoculating cells onto the chick chorioallantoic 

 membrane, it was shown that the number of cells capable of initiating pocks 

 was about 1/8 (range of 1/1.9 to 1/17 in different experiments) of the total 

 number in the suspensions. These cells release virus at a slow rate during 

 incubation at 37 °C, but when the cells were frozen and thawed three times 

 before incubation (a procedure which of itself did not reduce virus infectivity) 

 the amount of virus released corresponded to one infective dose for 250 cells. 

 On the average, therefore, the recovery of virus during the lag period was 

 about 3 virus particles for every 100 infected cells. 



The recoveries found for influenza and Rous sarcoma viruses during the lag 

 period are slightly higher than the level of about 1 % (or less) of the virus 

 taken up, which was found for the remaining viruses. However, at least for 

 influenza viruses, this estimate is probably too high, as will be described in 

 Section II, C. 



4. Larger Viruses 



A number of recent studies bear on the recovery of vaccinia and herpes 

 simplex viruses from infected chorioallantoic membranes during the lag 

 period, and, in general, the recoveries found have been higher than those 



