Supramolecular ‘catch-and-release’ strategy for bioorthogonal fluorogenic imaging across the visible spectrum

Fluorogenic probes that unmask fluorescence signals in response to a bioorthogonal reaction are a powerful new addition to biological imaging. They can provide significantly reduced background fluorescence and minimize non-specific signals, potentially allowing real-time high-contrast imaging without washing out excess fluorophores. While diverse classes of highly refined synthetic fluorophores are readily available now, their integration into a bioorthogonal fluorogenic scheme still necessitates another level of extensive design efforts and customized structural alterations to optimize quenching mechanisms for each given fluorophore scaffold. Herein, we present an easy-to-implement and highly generalizable supramolecular ‘catch-and-release’ strategy for generating an efficient bioorthogonal fluorogenic response from essentially any readily available fluorophores without further structural alterations. We designed this distinct strategy based on the macrocyclic cucurbit[7]uril (CB7) host, where a fluorogenic response is achieved by programming a guest displacement reaction from the macrocycle cavity. We used this strategy to rapidly generate fluorogenic probes across the visible spectrum from structurally diverse classes of fluorophore scaffolds, including coumarin, bodipy, rhodamine, and cyanine. These probes were applied to no-wash fluorogenic imaging of various target molecules in live cells and tissue with minimal background and no appreciable non-specific signal. Notably, the orthogonal reactivity profile of the system allowed us to pair this host-guest fluorogenic probe with the covalently clickable fluorogenic probe to achieve high-contrast super-resolution and multiplexed fluorogenic imaging in cells and tissue.