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After blocking with 5% BSA for 1 h, the membranes were incubated with anti--SMA (1:1,000 dilution, Abcam, USA) anti-Nrf2 (1:1,000 dilution, Proteintech Group, Rosemont, IL, USA), anti-HO-1 (1:1,000 dilution, Proteintech Group, Rosemont, IL, USA), anti-GPX4 (1:1,000 dilution, Abmart, Shanghai, China), anti-xCT (1:1,000 dilution, Shanghai, China), anti--tubulin antibodies (1:1,000 dilution, Cell Signaling Technology, Danvers, MA, USA) or anti-Lamin-B1 antibodies (1:10,000 dilution, Proteintech Group, Rosemont, IL, USA) at 4 C overnight, followed by horseradish peroxidase-linked anti-mouse (1:1,000 dilution, Cell Signaling Technology, Danvers, MA, USA) or anti-rabbit secondary antibodies (1:2,000 dilution, Cell Signaling Technology, Danvers, MA, USA) at 37 C for 1 h

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However, there are drawbacks to using this in evaluation of NACs effectiveness on hangovers