Anti wstf

GBM cells were lysed in RIPA Lysis and Extraction Buffer (Thermo Fisher Scientific), and acid protein kit (Thermo Fisher Scientific) was applied to detect the protein concentrations in the cellular lysates. The lysates were separated by SDS-PAGE, electro- transferred onto PVDF membrane (Millipore, Bedford, MA, USA) and blocked in 5% BSA. Following overnight probe with the primary antibodies: anti-WSTF (1:2000, Cell Signaling, Beverly, MA, USA), anti-Bax (1:3000, Cell Signaling), anti-cleaved caspase- 3 (1:3500, Cell Signaling), anti-AKT, anti-p-AKT and anti-β- actin (1:4000, Cell Signaling), the membranes were incubated with corresponding horseradish peroxidase-labeled secondary antibody (1:5000; Cell Signaling). The immunoreactivities were detected by enhanced chemiluminescence (KeyGen, Nanjin, China).

For western blot analysis, whole-cell lysates were separated by SDS-PAGE and transferred onto PVDF membranes using Criterion TGX gels and Trans-Blot Turbo Midi PVDF Transfer Packs (Bio-Rad, Berkeley, CA, USA). The blots were then incubated with specific primary antibodies as follows: anti-CHK1 (no. 2360), anti-CHK2 (no. 6334), anti-phospho-CHK1 Ser317 (no. 12302), anti-phospho-CHK2 Thr68 (no. 2197), anti-BRCA1 (no. 9010), anti-phospho-BRCA1 Ser1524 (no. 9009), anti-TFII-I (no. 4562) and anti-WSTF (no. 2152) all purchased from Cell Signaling Technology, Inc. The primary anti-RCF2 antibody (no. ab174271) was purchased from Abcam (Cambridge, UK).