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. 2007 Oct 3;81(24):13889–13903. doi: 10.1128/JVI.01231-07

FIG. 1.

FIG. 1.

Endocytosis of gB during infection assayed by biotinylation. (A) Biotinylation of gB was assayed in 143B cells with noncleavable biotin 8 h after infection with KOS. Samples were immunoprecipitated with a polyclonal anti-gB antibody and then separated by sodium dodecyl sulfate-7% polyacrylamide gel electrophoresis under nonreducing conditions, and biotinylated glycoproteins were revealed with streptavidin-HRP by Western blotting (lanes 1 to 4). After stripping, the membrane was revealed with a monoclonal anti-gB antibody (lanes 5 to 8). Lanes: 1, 2, 5, and 6, uninfected control cells; lanes 3, 4, 7, and 8, infected cells; lanes 1, 3, 5, and 7, nonbiotinylated; lanes 2, 4, 6, and 8, biotinylated. (B) 143B cells were biotinylated with Sulfo-NHS-SS-cleavable biotin 8 h after infection with KOS and left for 4 h for infection to proceed. After treatment of the cells with GSH, lysed samples were gB immunoprecipitated and visualized with streptavidin-HRP (lane 3). As controls, cells were gB immunoprecipitated immediately after the biotinylation step, without (lane 1) or after (lane 2) GSH treatment. Lane 4 shows the gB Western blot of lane 3 after stripping.