Figure 11.
Effect of ankyrin expression on Gag protein levels in HIV-1-infected SupT1 cells. Control and ankyrin-expressing SupT1 cell lines were infected with HIV-1NL4-3 inoculum at 10 MOI for 16 h at 37°C. After washing with serum-free medium, cells were resuspended in prewarmed medium containing 500 μg/mL hygromycin B and 10% FCS, and seeded into 6-well plates. Cells were harvested and lysed at day 9 and 11 pi. Cell lysates were fractionated into nuclear pellet, membrane compartment and cytosol, and each fraction assayed for CAp24 content by ELISA, and Gag proteins by SDS-PAGE and Western blot (WB) analysis. Protein concentration in all samples was normalized to 10 μg/mL. (A), ELISA of whole cell lysates (WCL); (B), ELISA of membrane/particulate fractions. Data presented are mean from triplicate experiments, with error bars indicating the standard error to mean. (C, D), SDS-PAGE and WB analysis of WCL. Samples were taken at day 11 pi, and analyzed using anti-CAp24 (C) and anti-MAp17 mAb (D).