Rabbit anti caspase 8

Protein was extracted on ice from embryos at E10.5 using RIPA buffer. Protein concentrations were determined using the Bradford assay. Western blot was performed using conventional methods, with 50-μg protein run per sample on NuPAGE 4–12% Bis-Tris gel (Life technologies), followed by transfer to PVDF membranes (XCell II Blot Module, Invitrogen). The primary antibodies were rabbit anti-caspase-8 (1:500, Proteintech, Chicago, USA) and anti-β-TUBULIN (1:10,000, Proteintech, Chicago, USA). After incubation with secondary antibody (1:10,000, DAKO), blots were developed using ECL Prime (GE Healthcare Life Sciences) or ECL Western Blotting Substrate (Promega). ImageJ was used to detect densitometry. The results were normalized using β-TUBULIN loading control. Independent experiments were carried out three to four times per sample.

The following antibodies were used: rabbit anti-CXCR2 (Abcam, USA), mouse anti-Ki67 (BD Biosciences, USA), rabbit anti-Akt, rabbit anti-phospho-Akt (Ser473), rabbit anti-ERK1/2, mouse anti-phospho-ERK1/2 and PARP (Cell Signaling Technologies, USA), mouse anti-GAPDH, rabbit anti-Caspase-8, rabbit anti-Bax and Bcl-2 (ProteinTech, USA), and rabbit anti-VEGF and NFкB (Santa Cruz), mouse anti-α-tubulin (Sigma Aldrich). G31P is prepared as described previously [21 (link), 22 ].