In a second set of experiments searching for replication-competent virus (Table 
6), approximately 10 8 BHK cells were infected with packaged SFV-lacZ/Helper 
2 virus (+ activation), at MOI of 3-4 and 60. After 16 hours of incubation, the 
cell culture medium was titered for PFU. Altogether, 1.34xl0 10 activated 
particles were examined. Despite the potential amplification of any replication- 
competent virus, no PFU were detected. 
Table 6 
Test for Replication-Competent Virus 
Using SFV-lacZ/Helper 2 
Quantity of 
Virus (IU)* 
MOI 
Number of Cells 
Infected 
PFU Observed 15 
-Act 
+Act 
6x 10 8 
3 
2x 10 8 
0 
0 
8 x 10 8 
4 
2 x 10 8 
0 
0 
1 x 10 8 
60 
1 x 10 8 
0 
0 
1 x 10 8 
60 
1 x 10 8 
0 
0 
*Titre determined after activatino, by IF. 
b +Act, with protease activation; -act, without activation. 
An important question is how few replication-competent virus were detectable 
with this assay and whether the high moi of 60 might have caused interference, 
thereby masking the detection of low levels of replication-competent virus. 
The possibility of interference was first examined (Table 7) by scoring for lacZ- 
positive BHK cells infected with one or 1000 IU of packaged SFV3 lacZ/Helper 
1 virus, alone or together with packaged SFV-hTR/Helper 2 virus (a recombinant 
clone of the human transferrin receptor), at MOI ranging from three to 30. No 
significant interference was detected, even at an MOI of 30. 
[454] 
Recombinant DNA Research, Volume 18 
