FIG. 11.

Association of H2 and A28. (A) BS-C-1 cells were infected with vA28-HAi/H2-V5 in the absence or presence of IPTG, and virions lacking A28-HA and containing A28-HA, respectively, were purified by sedimentation through two sucrose cushions and a sucrose gradient. The virions were incubated with NP-40 or NP-40 plus dithiothreitol and centrifuged to separate the soluble membrane and insoluble core fractions (34). The H2-V5 and A28-HA proteins were then detected by Western blotting using MAbs to the epitope tags conjugated to peroxidase. (B) BS-C-1 cells were infected with vA28-HA/H2-V5 or vA28-HA for 20 h, and the cells were lysed with 1% NP-40 in PBS. The detergent-soluble fraction was incubated with agarose beads to remove proteins that bind nonspecifically and then with anti-V5 MAb or anti-HA MAb immobilized on agarose beads. The bound proteins were analyzed by Western blotting with anti-HA or anti-V5 MAb conjugated to peroxidase.