In vivo NIR-to-NIR imaging with bispidine-lanthanide(iii) complexes

Two-photon luminescent bioprobes have emerged as powerful tools for in cellulo and in vivo imaging. Here, we report a family of bispidine-based chelators L^1a, L^1b and L^1c bearing push-pull two-photon antennas for efficient sensitization of lanthanide(iii) ions. The ligands and their complexes are readily accessible in good yields of 16-22% and 39-87%, respectively. The resulting complexes [EuL^1a]⁺, [SmL^1a]⁺, [DyL^1b]⁺, [TbL^1b]⁺ and [YbL^1c]⁺ display two-photon excitability combined with high quantum yields, long luminescence lifetimes, and high brightness, making them highly suitable for bioimaging applications. Importantly, the probes [EuL^1a]⁺, [SmL^1a]⁺, [DyL^1b]⁺, and [TbL^1b]⁺ show no apparent nonspecific binding in cellulo and can be readily washed out, indicating minimal interaction with cellular membranes or the intracellular matrix. Furthermore, [EuL^1a]⁺ and [YbL^1c]⁺ were successfully applied for in vivo imaging in zebrafish larvae, confirming their non-specificity and demonstrating their potential for angiographic imaging. The vascular system was clearly visualized, with sharp visible emission observed for [EuL^1a]⁺ and a completely dark background obtained for [YbL^1c]⁺. These results establish bispidine-based lanthanide(iii) complexes as robust and versatile lanthanide(iii) luminescent probes for biological imaging.