(A–C) Detection of p21 in HIV-1 PICs. CMK (restrictive) and H9 (permissive) cells were infected with HIV-1_IIIB and cytoplasmic PICs isolated at 7 hours after infection. Viral matrix protein is a component of PICs (22, 23). (A) Crude cytoplasmic PICs were isolated from CMK cells. Antibodies against p21 or HIV-1 Gag protein matrix (MA) (50) recovered similar levels of integrase (IN) (lanes 1 and 2). Purified integrase served as a marker (lane 3). Mock-infected cell control is shown in lane 4. (B) Analyses following PIC purification by spin column chromatography. Antibodies against p21 coimmunoprecipitated p21 with integrase in CMK cells (lane 4). Antibodies against p21 failed to recover IN when CMK cells were treated with p21 siRNA (lane 6) but not when cells were treated with mutated p21-siRNA (lane 7) (15). Isotype controls IgG3k (for MA) and IgG1 (for p21) failed to recover IN (lanes 2 and 8). Purified integrase served as a marker (lane 1). (C) Anti-MA antibodies coimmunoprecipitated p21 in PICs made from CMK cells (lane 1) but not from H9 cells (lane 2). p21-containing nuclear extract served as a marker (lane 5).