Ab92473

Total proteins were extracted from transfected or non-transfected cells, loaded (50 μg per lane) on 10% sodium dodecyl sulfate (SDS)-polyacrylamide gels, and transferred onto PVDF membranes (Thermo Fisher Scientific). The membranes were blocked for 2 h at 37°C with 5% nonfat milk in Tris-buffered saline with Tween 20 (TBST) and then incubated overnight at 4°C with the following primary antibodies: CDK1 (ab133327, Abcam, Cambridge, UK), cyclin B1 (ab18250; Abcam), cyclin A2 (18202-1-AP; Proteintech, Wuhan, China), MLH1 (ab92312; Abcam), PMS2 (ab110638; Abcam), MSH2 (ab92473; Abcam), MSH6 (ab92471; Abcam) and GAPDH (T0004; Affinity, Changzhou, China). Then, the membranes were incubated with an HRP-conjugated secondary antibody for 1 h at 37°C and covered with ECL luminescence reagent (Perkin-Elmer Inc., Waltham, MA, USA). GAPDH was used as the internal control.

The collected cells were lysed in ice-cold RIPA buffer, and protein lysates were then quantified with a BCA Protein Assay Kit. Western blotting was carried out as described previously [24 (link)]. The following antibodies were purchased, as indicated: a primary antibody against hMSH2 (ab92473, Abcam, Cambridge, UK), β-Actin (ab8226, Abcam, Cambridge, UK), and an anti-rabbit secondary antibody (Rockland, Gilbertsville, PA, USA). β-Actin was employed as a loading control. Immunoreactive proteins were detected by an Odyssey infrared imaging system (LI-COR Biosciences, Lincoln, NE, USA), and band intensities were quantified using ImageJ.