Hrp conjugated mouse anti β actin

Tumours were lysed in RIPA buffer and 500 μg of lysate were used for immunoprecipitation of VEGFR2. The following antibodies were used in this study: rabbit anti-VEGFR2 (55B11, Cell Signalling; 2479), HRP-conjugated anti-phosphotyrosine (4G10^®, Millipore; 16-105), goat anti-Albumin (Abcam; ab19194), and HRP-conjugated mouse anti-β-actin (Santa Cruz; sc-47778). For a quantitative analysis the membranes were scanned with a fluorescence scanner and the signal strength was determined by using ImageQuant^® software. Full-size images are presented in Supplementary Fig. 10.

Human lung tissue samples used for Western blot to assess BIM were obtained through the LTRC. Samples were homogenized in mammalian protein extraction reagent (Thermo Scientific) supplemented with protease and phosphatase inhibitors (Roche). Western blotting was performed with an SDS-PAGE electrophoresis system (Bio-Rad), on a 4%–20% Tris gel (Bio-Rad) in Tris running buffer, blotted to a PVDF membrane (Bio-Rad) and probed with primary antibodies, including rabbit anti–phosphorylated-p53 (Serine15) (1:1000, Cell Signaling Technologies, 9284), rabbit anti-p53 (1:1000, Cell Signaling Technologies, 9282), rabbit anti-BIM (1:1000, Cell Signaling Technologies, 2933), mouse anti-BRCA1 (1:100, Santa Cruz Biotechnology, sc-6954), rabbit anti-Vinculin (1:10,000, Abcam, ab219649), and HRP-conjugated mouse anti–β-actin (1:5000, Santa Cruz Biotechnology, sc-47778). After washing in TBS with 0.01% Tween, blots were incubated with HRP-conjugated anti-rabbit (1:5000, Cell Signaling Technology, 7074) or HRP-conjugated anti-mouse (1:5000, Cell Signaling Technology 7076) where appropriate, and signal was detected by autoradiography with ECL Supersignal West Femto maximum sensitivity substrate (Thermo Fisher Scientific). The detection of bands was carried out on a ChemiDoc MP imaging system (Bio-Rad Laboratories), and densitometry was performed using ImageJ software (NIH).