This protocol describes how to visualize, detect, and analyze redox signals (oxidative bursts) at the ER-mitochondrial interface. It uses drug-inducible crosslinking to target the genetically encoded glutathione redox sensor Grx1roGFP2 to organellar contact sites to measure local redox changes associated with transient depolarizations of the mitochondrial membrane potential (flickers). The strategy allows imaging of the oxidized to reduced glutathione ratio (GSSG:GSH) in subcellular regions below the diffraction limit with good temporal resolution and minimum phototoxicity. Moreover, the strategy also applies to diverse parameters including pH, H2O2, and Ca2+.
Booth, David M.; Várnai, Péter; Joseph, Suresh K; and Hajnóczky, György, "Fluorescence imaging detection of nanodomain redox signaling events at organellar contacts" (2022). Department of Pathology, Anatomy, and Cell Biology Faculty Papers. Paper 342.
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