140 - 
Time after infection (hr) 
Figure 4.1-A. Evaluation of the replication potential of AvlCFl in HeLa 
cells by measurement of production of infectious virus. HeLa cells were 
grown to 80% confluency in Dulbecco's modified Eagle medium, 10% fetal calf 
serum, 1% glutamine, 100 U/ml penicillin, 100 /ig/ml streptomycin and in- 
fected with AvlCFl or Ad-dl327 at a multiplicity of infection (MOI) of 60 
plaque forming units (pfu) per cell. After 90 minutes ("0" time), 24 hr, 48 
hr, or 72 hr, cells and media were collected and all cells were recovered 
and freeze- thawed 5 times to release intracellular adenovirus. Serial 
dilutions of the freeze-thaw lysate were prepared and used to infect 293 
cells grown to 95% confluency. Ninety minutes after infection, the media 
was replaced with soft agarose overlay media (Appendix 1; Green et al., 
1967) and the cells were incubated at 37°C, 5% C0 2 in a humidified atmo- 
sphere. After two weeks, the plates were evaluated for the presence of 
adenoviral plaques. The titer of adenovirus was calculated from the number 
of plaques at a given dilution and the dilution factor. 
[ 690 ] 
Recombinant DNA Research, Volume 16 
