FIG. 7.

Subcellular distribution of S glycoprotein in group-specific ORF deletion viruses. Cultures of Vero cells were inoculated with wild-type Urbani SARS-CoV, icSARS-CoV, or icSARS-CoV ΔORF3a. At 12 h, the infected cells were fixed in ice-cold 100% methanol overnight, washed with PBS, and blocked in PBS containing 5% BSA. Cells were washed and the coverslips incubated with mouse anti-S or ORF3a antibody diluted 1:1,000 in PBS containing 1% BSA, 0.05% NP-40, and 2% normal goat serum. Secondary antibody was fluorescein isothiocyanate-conjugated anti-mouse antibody diluted 1:1,000 in PBS containing 1% BSA, 0.05% NP-40, and 2% normal goat serum. Coverslips were washed, mounted, and visualized with a Nikon Microphot FXA Upright fluorescence microscope.