Figure 2.
Rescue of Fr-MLV (env ΔRBD) infection by soluble RBD on 293 mCAT1 cells. (A) In each well of a 6-well plate, 5 × 105 human 293-derived cells expressing the Fr-MLV receptor, mCAT1, were exposed to Fr-MLV (env ΔRBD) containing E. coli lacZ in the presence of increasing concentrations of purified Fr-RBD (0–400 nM). A 1:10 dilution of viral supernatant (1 ml/well), RBD, and polybrene (8 μg/ml) was added to cells simultaneously, and the plates were incubated overnight at 37°C before refeeding with fresh media. After an additional 24 h of incubation, cells were assayed for acquired β-galactosidase activity. The experiments were performed in triplicate, and standard errors are indicated. (B) Six-well plates of 293 mCAT1 cells were exposed to 1 ml per well of Fr-MLV (env ΔRBD) for 3 h on ice. Virus-containing media were then replaced with fresh media on half of the wells, and RBD (450 nM) was either added or not to each well. After 48 h, cells were fixed and stained, and β-galactosidase-positive cells were counted. The experiments were performed in triplicate, and standard errors are indicated.