2.9. Western blotting

Total proteins were extracted from tissue and cells, and the proteins in the sample were separated using SDS‐polyacrylamide gel electrophoresis. The separated proteins were then transferred to a polyvinylidene fluoride (PVDF) membrane. The PVDF membrane was immersed in blocking solution and shaken at room temperature for 1 hour. Then, the membrane was incubated with anti‐NMP1 primary antibodies(1:1000; Proteintech, Rosemont, IL, USA) for 2 hours at room temperature. Then, the membrane was incubated with secondary antibodies [horseradish oxidase‐labelled goat anti‐rabbit IgG (H + L); 1:5000] and was shaken at room temperature for 1 hour. After washing, the membrane was incubated with the developer solution and exposed to an X‐ray film to visualize the immunoreactive proteins.