FIG. 5.

Cell binding and internalization of fluorescent virus. (A) Binding at 4°C of density gradient-purified SAD ΔP(eGFP-P) to the cell surface. BSR cells were incubated for 2 h at 4°C with virus, fixed, and counterstained with propidium iodide. Small green dots (<1 μm) were detectable at the cell surface (top row). Neutralization with anti-G antibodies prior to incubation with cells resulted in larger aggregates (>1 μm [middle row]) that were still able to attach. Analysis of multiple sections of cells revealed that fluorescent signals were detectable exclusively at the cell surface. No comparable signals were obtained in the absence of virus (bottom row). (B) Internalization of bound SAD ΔP virus particles into large vesicular structures was observed after subsequent temperature shift to 37°C (top row). Most neutralized virus (anti-G antibodies) remained at the cell surface; however, some virus aggregates were also internalized (middle row). No green fluorescence was observed in cells not infected with SAD ΔP(eGFP-P) virus (bottom row). All laser scans shown in the figure were made at identical parameter settings.