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Wiegman CH, Michaeloudes C, Haji G, et al

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After denaturing in a boiling water bath, the proteins were separated by SDSpolyacrylamide gel electrophoresis (SDSPAGE), transferred to a PVDF membrane, blocked with 5% milk in TBST for 30 min at room temperature, and then incubated with primary antibody at 4 C overnight with shaking