VSV-G accumulates in the Golgi of HeLa cells expressing mutant PrPs.A, HeLa cells were transfected with plasmids encoding WT, PG14, CJD or FFI PrP, and VSV-G-EGFP fusion protein. 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, percentages of cells accumulating VSV-G in the Golgi. Data are the mean ± SD of three independent experiments. ∗∗p < 0.01, ∗∗∗p < 0.001 versus vector, and #p < 0.05, ##p < 0.01 versus WT by one-way ANOVA, Tukey’s post-hoc test. C, the normalized fluorescent density of VSV-G in the Golgi was measured in PrP-expressing cells and expressed as fold change over vector-transfected cells. Each bar indicates the mean ± SD of 65 to 80 cells from three independent experiments; ∗∗∗∗p < 0.0001 versus vector and WT by one-way ANOVA, Tukey’s post-hoc test.