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Fig. 2. MIM-B protein is widely expressed and is not downregulated in metastatic cell lines. (A) Characterisation of anti-MIM-B antibody. (Left) Lysates from mouse brain (L) were used to immunoprecipitate endogenous MIM-B using pre-immune serum (PI) or affinity-purified anti-MIM-B antibody (Pur) and analysed by immunoblotting with anti-MIM-B antibody. (Right) Lysates from COS-7 cells transfected with constructs expressing myc-MIM-B, myc-MIM-B {Delta}IMD ({Delta}IMD) or myc-IMD (see Fig. 1) were analysed by immunoblotting with anti-MIM-B antibody and reprobed with anti-myc antibody. Positions of molecular size markers are indicated in kDa. (B) Tissue distribution of MIM-B. 50 µg lysates from various mouse tissues were separated on SDS-PAGE and analysed by immunoblotting with anti-MIM-B antibody. SK, skeletal. (C) MIM-B expression is not downregulated in metastatic cell lines. Expression of endogenous MIM-B was studied by immunoblotting with anti-MIM-B antibody in non-metastatic (-) or metastatic (+) cell lines from bladder (top), prostate or breast (bottom). Tubulin was subsequently probed as a loading control. This result is representative of three independent experiments.