(A) Immunostaining of JFH-1 infected cells for NS5A (green) and endogenous OSBP (red) with DAPI nuclear counterstaining (blue). Manders’ coefficients: M1=0.60 (fraction of NS5A overlapping OSBP); M2=0.07 (fraction of OSBP overlapping NS5A). Bar, 10 μm.
(B) Uninfected or HCV-infected Huh7.5.1 cell homogenates (H) were centrifuged to prepare a crude membrane pellet and a “water soluble” supernatant (WS). The pellet was treated with cold 1% NP-40 and spun again; the “detergent soluble” (DS) fraction was removed and detergent-resistant membranes were fractionated on a density gradient. Fractions (numbered in order from light to heavy) were analyzed by immunoblotting for the indicated proteins.
(C) Immunoblots of homogenates from uninfected or HCV-infected Huh7.5.1 cells, treated with or without cold 1% NP-40 followed by treatment with or without proteinase K (PK).