Western blot analysis of extracts of various cell lines, using Phospho-TAK1-S412 antibody (TA378278) at 1:1000 dilution.C2C12 cells were treated by TGF-β (10 ng/ml) at 37℃ for 30 minutes.C6 cells were treated by Calyculin A (100 nM) at 37℃ for 30 minutes after serum-starvation overnight.|Secondary antibody: HRP Goat Anti-Rabbit IgG (H+L) at 1:10000 dilution.|Lysates/proteins: 25ug per lane.|Blocking buffer: 3% nonfat dry milk in TBST.|Detection: ECL Enhanced Kit .|Exposure time: 120s.