FIG. 1.

Characterization of MLV(HBV) pseudotype virus. (A) Protein expression in virion and cell lysates. 293T cells were transfected with pHIT112, pHIT60, and pCMV-L with or without pCMV-S. Virus particles were partially purified from culture supernatants by pelleting through 20% sucrose. Cell lysates and lysed virus pellets from the supernatant (sup.) were analyzed by Western blotting with anti-HBs antibody (top) or anti-MLV Gag antibody (bottom). Gp41 and p39 are L-HBsAg; gp27 and p24 are S-HBsAg. p30 and Pr65 are Gag and its precursor protein, respectively. (B) Immunoprecipitation of MLV(HBV) virus. MLV(HBV) and MLV(−) pseudotype viruses were harvested from the culture supernatant and concentrated by centrifugation at 26,000 rpm in an SW28 rotor for 90 min. After resuspension in NTE buffer (10 mM Tris-HCl [pH 7.4], 150 mM NaCl, 1 mM EDTA) with the control serum (Ctrl) or the polyclonal antibody (Ab) against HBsAg, the virus was incubated overnight at 4°C. The antibody-bound virus particles were pulled down by protein G beads. The virus particles were eluted by boiling in sample buffer and immunoblotted with polyclonal antibody against MLV Gag (p30) protein.