2.5 Sodium dodecyl sulfate- polyacrylamide gel electrophoresis (SDS-PAGE) and immunoblotting
The purity of the isolated αs1-CN and contamination of isolated β-CN was confirmed by SDS-PAGE and immunoblotting. The gel (15% polyacrylamide) was stained with Coomassie Brilliant Blue R-250 (FUJIFILM Wako Pure Chemical Corporation, Osaka, Japan). For immunoblotting, proteins were transferred to polyvinylidene difluoride membranes. After blocking (1% w/v bovine serum albumin in tris-buffered saline), αs1-CN was detected by anti-α-casein mouse monoclonal antibody and peroxidase-conjugated anti-mouse IgG.