Quantitatively imaging the spatiotemporal distribution of biological events in living organisms is essential to understand fundamental biological processes. Self-calibrating ratiometric fluorescent probes enable accurate and reliable imaging and sensing, but conventional probes using wavelength of 400-900 nm suffer from extremely low resolution for in vivo application due to the disastrous photon scattering and tissue autofluorescence background. Here, we develop a NIR-IIb (1500-1700 nm) emissive nanoprobe for high-resolution ratiometric fluorescence imaging in vivo. The obtained nanoprobe shows fast ratiometric response to hypochlorous acid (HOCl) with a detection limit down to 500 nM, through an absorption competition-induced emission (ACIE) bioimaging system between lanthanide-based downconversion nanoparticles and Cy7.5 fluorophores. Additionally, we demonstrate the superior spatial resolution of 1550 nm to a penetration depth of 3.5 mm in a scattering tissue phantom, which is 7.1-fold and 2.1-fold higher than that of 1064 and 1344 nm, respectively. With this nanoprobe, clear anatomical structures of lymphatic inflammation in ratiometric channel are observed with a precise resolution of ∼477 μm. This study will motivate the further research on the development of NIR-II probes for high-resolution biosensing in vivo.
Keywords: NIR-II; bioimaging; biosensing; nanoprobe; ratiometric fluorescence.