Western blot was performed using Anti-BID Recombinant Rabbit Monoclonal Antibody (2) (Product # MA5-29050) and a 20 kDa band corresponding to BID was observed across cell lines and tissue extracts tested and decreased upon Staurosporine and also with Actinomycin D along with TNF Alpha treatment. The expression of BID is reported to be low in Mouse Brain compared to Mouse Spleen and Mouse Thymus. Fig.a shows whole cell extracts (30 µg lysate) of A-431 (Lane 1), A-431 treated with Staurosporine (1uM for 8 hr) (Lane 2) and A-431 treated with Actinomycin along with TNF Alpha (5ug/ml and 10ng/ml for 5 hr) (Lane 3), Fig.b shows whole cell extracts (30 µg lysate) of Hep G2 (Lane 1), Jurkat (Lane 2), MOLT-4 (Lane 3), HeLa (Lane 4), PC-3 (Lane 5), SH-SY5Y (Lane 6) and tissue extracts of Mouse Spleen (Lane 7), Mouse Thymus (Lane 8) and Mouse Brain (Lane 9) were electrophoresed using Novex® NuPAGE® 12 % Bis-Tris gel (Product # NP0342BOX). Resolved proteins were then transferred onto a nitrocellulose membrane (Product # IB23001) by iBlot® 2 Dry Blotting System (Product # IB21001). The blot was probed with the primary antibody (1:1000 dilution) and detected by chemiluminescence Goat anti-Rabbit IgG (H+L) Superclonal™ Recombinant Secondary Antibody, HRP (Product # A27036, 1:4000 dilution) using the iBright FL 1000 (Product # A32752). Chemiluminescent detection was performed using Novex® ECL Chemiluminescent Substrate Reagent Kit (Product # WP20005).