FIG. 10.

Detection of endogenous p235 in 3T3-L1 adipocytes (A) and expressed epitope-tagged p235 in COS-7 cells (B) by immunoprecipitation and Western blotting. (A) Proteins from 3T3-L1 lysates were immunoprecipitated (IP) on preimmune (pre-imm) or anti-p235 antibodies (R6953) as described in Materials and Methods. After five washes in RIPA buffer, the immunoprecipitated proteins were resolved by SDS-PAGE, transferred to nitrocellulose membranes, and immunoblotted with anti-p235 peptide antiserum (R6953). (B) COS-7 cells were transfected with pCMV5-HA-p235s cDNA or with the empty vector as indicated. Cell lysates were immunoprecipitated with the anti-p235 or anti-HA antiserum. The lysates of transfected cells, together with HA and p235 immunoprecipitates, were sequentially Western blotted with anti-HA and anti-p235 antisera as indicated.