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Fig. 5. Subcellular localization of wild-type and FLAG-tagged human and bovine PrP in BHK cells transfected with recombinant Semliki-Forest virus RNAs. The subcellular location was determined by immunofluoroscence microscopy employing the mAb anti-PrP 3B5 and a secondary Ab (goat anti-mouse IgG) conjugated with indocarbocyanine (Cy3). Immunostaining of non-permeabilized BHK cells (A-F, left panels) transfected with (A) SFV-1 RNA (without any insert), (B) recombinant SFV-huPrP1-253 RNA, (C) recombinant SFV-huPrP22FLAG RNA, (D) recombinant SFV-huPrP227FLAG RNA, (E) recombinant SFV-boPrP1-264 RNA and (F) recombinant SFV-boPrP239FLAG RNA. (A-F, right panels) Intracellular immunostaining of permeabilized, transfected BHK cells, respectively. (magnification A-F: x630).