Fluorogenic Red to Near-Infrared Tetrazine–Cyanine Probes for Bioorthogonal Organelle Membrane Imaging and Spatiotemporal Disruption

Organelle membranes are fundamental interfaces that enable specialized cellular functions. Achieving organelle membrane labeling with small-molecule fluorescent probes remains highly desirable yet challenging. Here, we report a robust bioorthogonal platform based on fluorogenic tetrazine-cyanine (TBCy) probes, constructed by integrating tetrazine into a tunable BF 2 -bridged cyanine (BCy) scaffold, to develop a generalizable organelle membrane labeling strategy. The TBCy probes exhibit outstanding fluorogenicity, robust photostability, red-to-near-infrared absorption/emission, and charge neutrality. Coupling with rationally designed organelle membrane-targeted probes, the biocompatible TBCy variants enable no-wash, high-specificity labeling of lysosomal, mitochondrial, and endoplasmic reticulum membranes. This capability facilitates multicolor super-resolution STED imaging and the real-time tracking of their dynamics. Moreover, through bioorthogonal activation, photosensitized TBCy allows spatiotemporally controlled disruption of the lysosomal membrane under light irradiation, leading to lysosomal membrane damage-triggered cell death. This work not only provides a versatile toolbox for organelle membrane function elucidation and manipulation but also expands the design and application scope of fluorogenic tetrazine probes in precise bioimaging and therapy.