KKU-213 cells were treated with CMA3 at indicated concentrations and times. Protein lysate was prepared as described elsewhere (29 (link)). Protein concentrations were determined by the bicinchoninic acid protein assay (Thermo Fisher Scientific, Inc.). A total of 20 µg of protein was separated by SDS-PAGE (10-15%) and transferred to a PVDF membrane (GE Healthcare Japan). The membrane was blocked with 5% skim milk in Tris-buffered saline containing 0.1% Tween-20 (TBST) at room temperature for 30 min and was incubated with specific primary antibodies (1:1,000) at 4°C overnight. Membranes were washed with TBST 3 times prior to the incubation with the corresponding HRP-conjugated secondary antibody (1:2,000) for 1 h at room temperature. Signals were detected using Chemi-Lumi One Super reagents (Nacalai Tesque, Inc.) and visualized by ImageQuant LAS 4000 system (GE Healthcare Japan). The quantitative analyses of blots were quantified by Image J (30 (link)). Hsc70 was used as an internal control.