Western blot analysis was performed on whole cell extracts (30ug) of A-431 (Lane 1), HCT 116 (Lane 2), HEK-293 (Lane 3), HeLa (Lane 4), SH-SY5Y (Lane 5), NIH/3T3 (Lane 6) and PC-12 (Lane 7). The blots were probed with 0014-02 (2ug/mL) and detected by chemiluminescence using Goat anti-Rabbit IgG (H+L) Secondary Antibody, HRP conjµgate (0.4ug/mL, 1:2500 dilution). A ~ 28kD band corresponding to 14-3-3 theta was observed across cell lines tested. Known quantity of protein samples were electrophoresed using 4-12% Bis-Tris gel, Electrophoresis System and Novex® Sharp Pre-Stained Protein Standard. Resolved proteins were then transferred onto a nitrocellulose membrane with iBlot® 2 Dry Blotting System. 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.

Western blot analysis of HeLa cells using 0014-02. Lane 1: 1ug/mL, Lane 2: 0.5ug/mL, Lane 3: 0.1ug/mL