Goat anti-HIV-1 gp160 (MRC ADP 72 408/5104); rabbit anti-HIV-1 p24 (Gag) (ARP 432); donkey anti-goat IgG Cy3 or FITC; and donkey anti-rabbit IgG Alexa 647 (Life Technologies, Oslo, Norway) were used for immunofluorescence assays. The goat anti-human IgG-FITC (Fc-specific) antibodies (ab 97224, abcam, Cambridge, UK) were used for live-cell staining. Goat anti-HIV-1 gp120 (Bio-Rad 5000–0557, Bio-Rad, Watford, UK); goat anti-HIV-1 p24 (Gag) (Bio-Rad 4999–9007, Bio-Rad, Watford, UK); and mouse monoclonal anti-goat/sheep IgG–AP GT34 (Sigma, Saint Louis, MO, USA) were used for Western blotting. The anti-HIV-1 Env human monoclonal antibodies (MAbs) PG9, PGT128, CAP256 VRC26.08, and VRC01 were expressed in FreeStyle 293F cells (Life Technologies, Oslo, Norway) using the PEIMAX transfection reagent (Polysciences, Warrington, PA, USA). The monoclonal antibodies were purified from cell-free supernatants after 6 days using protein A affinity chromatography [39 (link)].
MBDK and HeLa cells (supplied by ATCC) were grown in Dulbecco’s Modified Eagle Medium (DMEM) (high glucose), plus L-glutamine (Lonza), plus 10% of fetal calf serum, plus 10% of penicillin-streptomycin (Pen-Strep) (both from Gibco).
MBDK and HeLa cells (supplied by ATCC) were grown in Dulbecco’s Modified Eagle Medium (DMEM) (high glucose), plus L-glutamine (Lonza), plus 10% of fetal calf serum, plus 10% of penicillin-streptomycin (Pen-Strep) (both from Gibco).
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