Mutant PrP misfolding and aggregation are necessary to impair VSV-G transport.A, HeLa cells were transfected with plasmids encoding WT, M128V, PG14, or PG14ΔHC PrP. After 24 h at 35 °C cells were fixed, permeabilized, stained with polyclonal anti-GM130 and monoclonal anti-PrP 12B2 antibodies followed by Alexa Fluor-conjugated anti-IgG secondary antibodies. Cells were viewed with green excitation/emission settings to detect VSV-G and blue excitation/emission settings to detect GM130. Scale bar 10 μm. B, the normalized fluorescent density of VSV-G in the Golgi was measured and expressed as fold change over vector-transfected cells. Data are the mean ± SD of 20 to 40 cells. ∗∗∗∗p < 0.0001 versus vector by one-way ANOVA, Tukey’s post-hoc test.