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When pretreating the cells with N -ethylmaleimide (NEM, 20 mM), a trapping reagent of thiol species, for 0.5 h, the signals from the cytoplasmic regions are decreased in both green and yellow channels (Figure 7DF)

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These results were validated using fluorescent miscroscopy and immunocytochemical staining ( Figure 4B ), which showed that CB-839 induced the accumulation of LC3 puncta in HG-3 and MEC-1 cells