Rabbit anti rad51 h 92 antibody

U2OS cells were grown on coverslips or microlaser dishes, fixed with 4% paraformaldehyde for 15 min at room temperature and permeabilized in 0.2% Triton X-100 for 10 min at room temperature. Treated cells were then incubated with primary antibodies for 1 h at 37 °C, washed and incubated with secondary antibody for 1 h at 37 °C. The slides were mounted in Vectashield mounting medium with DAPI (Vector Laboratories). Confocal images were acquired on Leica TCS SP5 (Leica). Image analysis was carried out with SP5 image software. The following antibodies were used for immunofluorescence: rabbit anti-Rad51 (H-92) antibody (SantaCruz, sc-8349), rabbit anti-Mre11 antibody (Novus, NB100-142), anti-MDC antibody (Bethyl, bl3423), anti-53BP1 antibody (SantaCruz, sc-22760), anti-Rap80 antibody (Bethyl, A300-763A), anti-ATM p1981 antibody (Abcam, ab36810), FITC-conjugated anti-BrdU antibody (BD biosciences), mouse anti-RPA antibody (Sigma). Anti-mouse IgG-Cy3 antibody (Sigma, c2181), anti-rabbit IgG-FITC antibody (Sigma, f7512), anti-rabbit IgG-Cy3 antibody (Sigma, and C2306), anti-mouse IgG-FITC antibody (Sigma, F6257).

Whole-cell extract prepared in RIPA buffer (20 mM Tris-HCl, pH 7.4, 20% glycerol, 0.5% NP-40, 1 mM MgCl₂, 0.5 M NaCl, 1 mM EDTA, 1 mM EGTA and protease inhibitors) or immunoprecipitates were separated on SDS-PAGE, transferred to PVDF membrane, incubated with the indicated antibodies and detected by ECL Western Blotting Detection Kit (BD Bioscience). The following antibodies were used for western blotting and immunoprecipitation: rabbit anti-Ago2/elF2C2 antibody (Abcam, ab32381), mouse Ago1 antibody (Milipore, 04-083), mouse Ago 3 antibody (Novus, L021V1), mouse Ago4 antibody (Milipore, 05-967), mouse anti-Dicer antibody (Abcam, ab14601), rabbit anti-Rad51(H-92) antibody (SantaCruz, sc-8349), anti-ATM antibody (Abcam, ab81292), rabbit anti-Mre11 antibody (Novus, NB100-142), mouse anti-RPA antibody (Abcam, ab2175), mouse anti-Ku70 antibody (SantaCruz, sc-5309), mouse anti-β-tubulin antibody (Sigma, T5293), rabbit anti-Myc antibody (Abcam, ab9106), mouse anti-MDC1 antibody (Abcam, ab50003), rabbit anti-53BP1 antibody (Bethyl, A300-272A), mouse anti-GFP antibody (CWBIO, CW0258), anti-mouse IgG-HRP antibody (Dakocytomation, p0161) and anti-rabbit IgG-HRP antibody (Dakocytomation, p0448).