Western Blot Analysis of Apoptosis Markers

Western blot analyses were performed as previously described [33 (link)]. Briefly, cells were washed twice with PBS and then lysed via sonication in lysis buffer (Intron, Seoul, Korea). The samples were separated on 10–15% SDS-PAGE gels, and then transferred to nitrocellulose membranes (Protran BA83; Whatman). Immunoblotting was performed using the following primary antibodies and dilutions: anti-Capase 8 (1:1000; Cell Signaling, Danvers, MA, USA), anti-Caspase 3 (1:1000; Cell Signaling, Danvers, MA, USA), anti-Caspase 7 (1:1000; Cell Signaling, Danvers, MA, USA), anti-cleaved Caspase 7 (1:1000; Cell Signaling, Danvers, MA, USA), PARP (1:1000; Cell Signaling, Danvers, MA, USA), BCL2 (1:1000; Cell Signaling, Danvers, MA, USA), DR4 (1:1000; Abcam, Cambridge, MA), DR5 (1:000; Abcam, Cambridge, MA, USA) and anti-beta actin (1:10000; Sigma, St. Louis, MO, USA). Horseradish peroxidase-labeled rabbit anti-mouse (Abcam, diluted 1:5000, Cambridge, MA, USA) and goat anti-rabbit (Santa Cruz, diluted 1:2000, Finnell Street, Dallas, TX, USA) secondary antibodies were used. The proteins were visualized using an ECL detection system (Ab Frontier, Seoul, Korea).