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A FRET-based fluorescence turn-on probe is designed, which employs a sydnonimine as the linker to match specific fluorophore and quencher pairs and releases the fluorescence after the “click-and-release” reaction. Furthermore, we realized selective fluorescence labeling by exploiting the mutual orthogonality between sydnonimine–DIBAC and tetrazine–1,3-Cp cycloaddition pairs.

Graphical abstract: Fluorogenic sydnonimine probes for orthogonal labeling

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