Western Blotting Technique for Protein Detection

A standard Western blotting technique was performed. Briefly, the membranes were blocked using phosphate-buffered saline (PBS) containing 1% casein (Bio-Rad) and then probed with a 1:1,000 dilution of a primary antibody overnight at 4°C. The appropriate secondary antibody was used at a 1:10,000 dilution and incubated at room temperature for 1 h. Signal was detected with ECL+ (GE Healthcare), and the film was developed by a film processor (GE Healthcare). The antibodies used for Western blotting were mouse anti-Myc (Cell Signaling), rabbit anti-actin (Cell Signaling), rabbit anti-GST (Cell Signaling) antibodies and the custom-made EhAgo2-2 polyclonal antibodies in rabbit (28 (link)).