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. Author manuscript; available in PMC: 2009 Dec 4.
Published in final edited form as: Exp Cell Res. 2008 Dec 3;315(3):542–555. doi: 10.1016/j.yexcr.2008.11.011

Fig. 1.

Fig. 1

A complex of AKAP350A with CCAR1 and caprin-1. (A) Immunostaining of HepG2 cells with rabbit polyclonal antibodies against AKAP350A show co-localization at the Golgi apparatus with marker of cis-Golgi gm130 as well as diffuse cytosolic staining. Bar = 10 μm. (B) AKAP350 in subcellular fractions of HepG2. Confluent HepG2 cells were homogenized and fractionated by serial spins and western blots were probed for AKAP350 and gm130. 3,000g membranes were enriched for gm130. (C) Cell lysates (L) were immunoisolated with non-specific rabbit IgG (C) or anti-AKAP350A (a350A) and preparations were blotted for both AKAP350 and RIIa. (D) Verification of proteomics data by immunoprecipitation followed by western blot. Lysates of HepG2 cells were immunoprecipitated with anti-AKAP350A and CCAR1 antibody or pre-immune rabbit serum. Eluates of immunoprecipitations (IPs) corresponding 50 mg of starting material and 10 mg of starting lysate were resolved on the same SDS-PAGE gel and probed for AKAP350A, CCAR1, caprin-1 and a-tubulin (as a control for specific immunoprecipitation). CCAR1 and caprin-1 were both immunoisolated with AKAP350A-specific antibody. Caprin-1 as well as a fraction of AKAP350A were recovered with anti-CCAR1 antibody. All immunoisolations were free of a-tubulin-contamination.

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