Las 1000 uv mini imager

Western blot analysis was performed as previously described [48 (link)]. Protein samples were prepared by sample homogenization in radio immunoprecipitation assay (RIPA) buffer (1 % Nonidet P40, 0.5 % sodium deoxycholate, 0.1 % sodium dodecyl sulfate-polyacrylamide in PBS) [49 (link)]. Equal amounts (20 μg / lane) of protein were subjected to a 10–20 % sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) gradient and then transferred to Polyvinylidene-Fluoride membranes (Scientific Research Special, USA). Membranes were blocked for 1 h with 5 % nonfat dry milk in 0.1 % Tween–20 in PBS and then incubated overnight at 4 °C with AKR1B1 antibody (ABGENT, CA, USA) at 1:1000 dilution. Following 3 washes with PBS containing TBST (Tris and Tween Solution Buffer), the membranes were incubated with HRP (horseradish peroxidase)-labeled anti-mouse immunoglobulin G. The specific band was visualized with BeyoECLPlus (Santa Cruz Biotechnology, TX, USA). The density of each band was analyzed using a LAS-1000 UV mini imager (Fuji Film, Tokyo, Japan).

Western blotting analysis was performed as previously described^{21 (link)}. The primary antibodies were rabbit antibodies against GFP (1:500 dilution, AB3080; Chemicon), human β-actin (1:2,500, ab8227; Abcam), mouse antibodies against Flag M2 (1:500, F3165; Sigma), human CAF-1 p150 (1:1,000, ab24746; Abcam). The secondary antibodies were horseradish peroxidase-conjugated donkey anti-rabbit IgG (1:2,500, NA934; Amersham Biosciences) and sheep anti-mouse IgG (1:7,500, NA931; Amersham Biosciences). ECL Plus reagents were from Amersham Biosciences. Chemiluminescence was detected using an LAS1000UV mini imager and quantified using the Image Gauge software (Fuji Film).