Anti btg1

Cells were mixed with lysis buffer (50 mM Tris [pH 7.5], 1% NP-40, 10 mM NaCl, 20 μg/mL aprotinin, 20 μg/mL leupeptin, and 1 mM phenylmethylsulphonyl fluoride) and were placed on ice for 20 min. The lysates were assessed for protein concentration, and 100 μg of each protein sample was resolved by 12% SDS-PAGE and electroblotted onto nitrocellulose membranes (GE Healthcare, London, UK). After a 1-h incubation in blocking solution [5% non-fat milk in Tris-buffered saline with Tween 20 (TBS-T)], the membranes were exposed to primary antibodies (anti-BTG1, 1:200, polyclonal; Abcam, Cambridge, MA, USA, and β-actin, 1:5,000; Santa Cruz Biotechnology, Santa Cruz, CA, USA) overnight at 4°C. The blots were washed three times in TBS-T and incubated with horseradish peroxidase- conjugated secondary antibody (Cell Signaling Technology, Beverly, MA, USA) for 1 h at room temperature. Protein bands were visualized using enhanced chemiluminescence reagent (iNtRON Biotechnology, Seongnam, Gyeonggi-do, Republic of Korea).