Rational Design of Tunable Near-Infrared Oxazine Probe with Large Stokes Shift for Leucine Aminopeptidase Detection and Imaging

Near-Infrared (NIR) fluorescence imaging with the advantages of deep tissue penetration and minimum background, has been widely employed and developed in the study of biological applications. However, small Stokes shifts, difficulty in optical tuning, and pH sensitivity are still the major limitations faced by current NIR dyes. To solve these problems, we rationally designed a pH insensitive amino-tunable NIR oxazine fluorophore DQF-NH2, which exhibited large Stokes shift (125 nm) accompanied with NIR excitation/emission due to the introduction an asymmetrical alternating vibronic feature. By benefiting from the excellent photophysical properties of DQF-NH2, we have successfully constructed the probe DQF-NH2-LAP with the ability to detect endogenous LAP. Bioimaging assays demonstrated that DQF-NH2-LAP can not only effectively detect LAP in living cells, but also was successfully applied to image tumor tissue in vivo. We anticipate that the functionalizable dye DQF-NH2 may be a potential new NIR dye platform with an optically tunable group for the development of future desirable probes for bioimaging. A novel amino-tunable near-infrared oxazine dye (DQF-NH2) with large Stokes shift (125 nm) has been designed by a molecular integration strategy of optically tunable groups and unsymmetric oxazine fluorophore. As an example of its application, we use this NIR dye to develop a highly sensitive fluorescent probe DQF-NH2-LAP for detecting and imaging leucine aminopeptidase in living cells and in vivo.image