Western blot analysis of Phospho-MYL9 (T18 + S19) on different lysates with Rabbit anti-Phospho-MYL9 (T18 + S19) antibody (HA751447) at 1/5,000 dilution. Lane 1: PC-12 cell lysate Lane 2: PC-12 treated with 100nM Calyculin A for 30 minutes cell lysate Lane 3: C2C12 cell lysate Lane 4: C2C12 starved overnight then add 100nM Calyculin A for 30 minutes then treated with 100ng/mL insulin for 20 minutes cell lysate Lane 5: C2C12 starved overnight then add 100nM Calyculin A for 30 minutes then treated with 100ng/mL insulin for 20 minutes cell lysate, then the membrane treated with λpp for 1 hour Lysates/proteins at 20 µg/Lane. Predicted band size: 20 kDa Observed band size: 19 kDa Exposure time: 30 seconds; ECL: K1801; 4-20% SDS-PAGE gel. Proteins were transferred to a PVDF membrane and blocked with 5% NFDM/TBST for 1 hour at room temperature. The primary antibody (HA751447) at 1/5,000 dilution was used in primary antibody dilution (K1803) at 4℃ overnight. Goat Anti-Rabbit IgG - HRP Secondary Antibody (HA1001) at 1/50,000 dilution was used for 1 hour at room temperature.