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. 2010 Mar 24;84(11):5687–5694. doi: 10.1128/JVI.02583-09

FIG. 2.

FIG. 2.

Characterization of ASLV EnvA mutants. (A) Incorporation into MLV pseudovirions. Equal volumes of MLV pseudovirions bearing the indicated EnvA mutants were run on duplicate gels and transferred to nitrocellulose. One immunoblot was probed with anti-A-tail IgG (gp37) and visualized with horseradish peroxidase (HRP)-conjugated anti-rabbit IgG (p30). The other gel was probed with anti-MLV capsid and visualized with HRP-conjugated anti-rat IgG. The bands were quantitated using a Rosenman Pixel Quantification plug-in for Adobe Photoshop. The numbers reported are for EnvA/gag ratios. (B) PNGase treatment of WTA, D432A, and F439A. Virions were treated with PNGase (+) or mock treated (−), resolved by SDS-PAGE, and transferred to nitrocellulose, and the gp37 band was probed as described for panel A. Bars, glycosylated gp37; *, deglycosylated gp37. (C) Immunoprecipitation of sTva with WTA and F439A pseudovirions. Biotinylated sTva was incubated with virions in 1% NP-40 lysis buffer, immunoprecipitated with anti-A tail IgG bound to protein A-conjugated beads, resolved by SDS-PAGE, transferred to nitrocellulose, and probed with HRP-conjugated streptavidin. Mock, no virus added.