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Fig. 1. Western blot analysis of KSR expression in adult mouse tissues and cell lines. A, testing the specificity of the antibody. COS cells were transfected with a vector expressing myc-Ksr or with the vector control, and protein lysates were prepared and analyzed with the KSR antibody. The antibody detects a single protein species of Mr 108,000 only in the cells transfected with the myc-Ksr vector. B, analysis of adult mouse tissues. Western blots were prepared using 20 µg of tissue lysate and analyzed with the anti-KSR rat monoclonal antibody (top panel). To control for protein loading, the Western blots were analyzed with an anti-vinculin mouse monoclonal antibody (bottom panel). Only the faster migrating isoform (B-KSR1) is detected in cerebral cortex and cerebellum, and only the slower migrating isoform (KSR1) is detected in bladder and lung. The ovary expresses both isoforms. C, analysis of three commonly used cell lines. Western blots were prepared using 20 µg of cell lysate and analyzed with the anti-KSR rat monoclonal antibody (top panel) and an anti-vinculin mouse monoclonal antibody (bottom panel). Expression of KSR1 is only detected in ES cells.