324 H. B. ANDERVONT 



mesodermal and ectodermal lesions in the membrane, and established the 

 feasibility of using the discrete lesions for virus titration. Groupe et al. 

 (1957a), with a refined procedure, procured a correlation between the number 

 of lesions and the quantity of virus used to infect the membrane. 



Prince (1958a) recognized that the variability between embryos in their 

 responses to infection with the Eous virus was a major deterrent to the use of 

 Keogh's technique for quantitative assays of the virus, and has done much to 

 make it practical. He tested embryos from 9 stocks of chickens for suscepti- 

 bility to the virus and found a wide variation in the responses of their 

 membranes. Embryos from one inbred line of chickens were highly susceptible 

 and proved to be satisfactory for quantitative assay. Prince (1958b) then used 

 the most susceptible and most resistant lines of chickens to ascertain the 

 nature of the susceptibility to the virus. Sera from adult chickens were 

 tested for neutralizing antibodies to the virus; it was found that sera from 

 chickens whose embryos were most susceptible contained the most antibody. 

 This led to a test for antibodies in membranes from the susceptible embryos 

 because about 10 % of these were resistant to the virus: membranes from 

 susceptible and resistant embryos showed no essential differences in their 

 neutralizing activities. These results indicated that humoral antibodies were 

 not responsible for the resistance of the membranes. When embryos procured 

 by reciprocal matings between susceptible and resistant chickens were tested 

 with the virus, their membranes proved to be equally susceptible. It was 

 concluded that genetic factors, and not humoral antibodies, were responsible 

 for the resistance of the chorioallantoic membrane to the virus. 



Rubin (1957) applied the technique in conjunction with tissue cultures and 

 other methods to perform a series of interesting studies on cell- virus relation- 

 ships. He confirmed Keogh's findings that the number of lesions produced 

 upon the chorioallantoic membrane was dependent upon the concentration 

 of virus in the inoculum, and he interpreted this as suggesting that each 

 tumor on the membrane was caused by a single virus particle. Tissue cultures 

 of Rous sarcoma cells were assayed for the virus by exposing membranes to 

 the supernatant fluids. By means of this procedure it was determined that 

 the rate of virus production per cell was low but remained constant for a 

 considerable time because of the intimate relationship between cell and virus. 

 This indicated that, in contrast to bacterial viruses, the Rous sarcoma virus 

 was produced slowly by most sarcoma cells but these cells produced virus for a 

 long time. Through determinations of the amounts of intracellular and extra- 

 cellular virus, Rubin concluded that the average infectious virus particle 

 stayed within the cell for one-half hour after its completion. Also, the rate of 

 virus production correlated with an increase in cell size and, because this 

 increase took place only in the cytoplasm of the cell, it was inferred that 

 virus formation took place within the cytoplasm. 



