Fig. 7.
gL− virions also show little conformation change in gB. (a) NMuMG cells were incubated (2 h, 4 °C) with WT (3 p.f.u. per cell) or gL− (50 p.f.u. per cell for equivalent binding) MuHV-4, then washed with PBS and either fixed immediately or first further incubated (2 h, 37 °C) to allow virion endocytosis. The cells were stained for pre-fusion gB with mAb BN-1A7 (IgG2a) or for post-fusion gB with mAb MG-1A12 (IgG2a) (both green), for LAMP-1 (red) and with DAPI (blue). Co-localization appears as yellow. Equivalent data were obtained in three further experiments. (b) Cells and viruses were incubated as in (a), then antibody binding was detected with an alkaline phosphatase-conjugated IgG2a-specific secondary antibody, p-nitrophenylphosphate substrate and A405. For each condition, the A405 was normalized to the value obtained at 4 °C. The bars show mean±sem values from six wells. After incubation at 37 °C, the WT BN-1A7 signal was reduced significantly relative to gL− (P<10−9 by Student’s t-test) and the WT MG-1A12 signal was increased significantly (P<10−5). Equivalent data were obtained in a repeat experiment.