Western Blot Analysis

Briefly, the total protein was collected from H9c2 cells and separated on sodium dodecyl sulphate-polyacrylamide gel electrophoresis (SDS-PAGE) gels. Subsequently, the proteins were electrotransferred to polyvinylidene difluoride membranes (Millipore, Billerica, MA, USA) and incubated overnight (4°C) with primary antibodies (Table 1). After that, the membranes were washed and incubated with appropriate secondary antibodies for 2 hours (37°C). Protein bands were developed using an enhanced chemiluminescence (ECL) reagent (Wanleibio, Shenyang, China), and the band intensity was quantified using Gel-Pro Analyzer Software (Media Cybernetics, Bethesda, MD, USA). β-actin served as an internal loading control. The relative protein level was determined as the ratio of the gray value of the target band to that of the β-actin band.

Primary Antibodies Used in Western Blot