Western blot analysis was performed on whole cell extracts (30 µg lysate) of A-431 (Lane 1), Hep G2 (Lane 2), MCF7 (Lane 3), Jurkat (lane 4), HEK-293 (lane 5) and Panc-1 (lane 6). The blots were probed with Anti-IRS1 Rabbit Polyclonal Antibody (Product # PA1-1057, 1:250 dilution) and detected by chemiluminescence using Goat anti-Rabbit IgG (Heavy Chain) Superclonal™ Secondary Antibody, HRP conjugate (Product # A27036, 0.4 µg/mL, 1:2500 dilution). A 131 kDa band corresponding to IRS1 was observed across cell lines tested. Known quantity of protein samples were electrophoresed using Novex® NuPAGE® 4-12 % Bis-Tris gel (Product # NP0321BOX), XCell SureLock™ Electrophoresis System (Product # EI0002) and Novex® Sharp Pre-Stained Protein Standard (Product # LC5800). Resolved proteins were then transferred onto a nitrocellulose membrane with iBlot® 2 Dry Blotting System (Product # IB21001). The membrane was probed with the relevant primary and secondary Antibody following blocking with 5 % skimmed milk. Chemiluminescent detection was performed using Pierce™ ECL Western Blotting Substrate (Product # 32106).