CD9 is associated with MHC class II and CD38 on human monocytes. Monocytes were surface-labeled with biotin before lysis in NP-40 (A) or CHAPS (B-D). Immunoprecipitations were then performed with the HLA-DR (D1.12), CD38 (IB4), or isotype control mAbs. After electrophoresis under nonreducing conditions and transfer to a nitrocellulose membrane, the precipitated material was revealed by chemiluminescence. (C) The HLA-DR, CD38, and CD9 molecules were immunoprecipitated, from unlabeled monocytes, with the specific D1.12, IB4, and PHN 200 mAbs, respectively. Immunoprecipitates were analyzed by Western blot with biotinlabeled CD38 (Leu 17), CD9 (Syb.1), or anti–HLA-DR (DA6.147) mAbs. (D) Monocyte lysates were precleared twice in a period of 24 hours with CD38 (IB4) or isotype control mAb together with protein G-sepharose. HLA-DR and CD38 were then immunoprecipitated and Western blotted with biotin-labeled CD38 (Leu 17), CD9 (Syb.1), or anti–HLA-DR (DA6.147) mAbs. Supernatants of CD38 and HLA-DR precipitates obtained after Ig or CD38 preclearing were blotted with an antiactin mAb and show that equal quantities of proteins were used for immunoprecipitations (not shown). WB indicates Western blot; IP, immunoprecipitation.