Low concentration of digitonin selectively permeabilizes the plasma membrane. (A and B) HaCaT cells were grown for 24 h and then fixed and permeabilized with either 1 mg/ml (A) or 5 μg/ml (B) digitonin. Lamp1, cytosol-facing late endosome marker Lamp1; calnexin (Calnx), luminal endoplasmic reticulum marker; DAPI, nuclear marker. (C and D) HaCaT cells were grown for 24 h and then fixed and permeabilized with either 0.5% Triton X-100 or 5 μg/ml digitonin. p230, cytosol-facing TGN marker; TGN46, luminal TGN marker; DAPI, nuclear marker. (E to H) HaCaT cells were infected with HPV16 pseudovirus for 18 h in the presence of 1 μM bafilomycin A1. Cells were fixed, permeabilized using 5 μg/ml digitonin, and stained using L2-specific antibodies (K4, K1, and 33L2-1) or an L1-specific antibody (33L1-7) following the Click-iT reaction. Colors are coded as indicated by the labels on the figure.