Fluorescence-amplified nanocrystals in the second near-infrared window for in vivo real-time dynamic multiplexed imaging

Optical imaging in the second near-infrared window (NIR-II, 1,000–1,700 nm) holds great promise for non-invasive in vivo detection. However, real-time dynamic multiplexed imaging remains challenging due to the lack of available fluorescence probes and multiplexing techniques in the ideal NIR-IIb (1,500–1,700 nm) ‘deep-tissue-transparent’ sub-window. Here we report on thulium-based cubic-phase downshifting nanoparticles (α-TmNPs) with 1,632 nm fluorescence amplification. This strategy was also validated for the fluorescence enhancement of nanoparticles doped with NIR-II Er 3+ (α-ErNPs) or Ho 3+ (α-HoNPs). In parallel, we developed a simultaneous dual-channel imaging system with high spatiotemporal synchronization and accuracy. The NIR-IIb α-TmNPs and α-ErNPs facilitated the non-invasive real-time dynamic multiplexed imaging of cerebrovascular vasomotion activity and the single-cell-level neutrophil behaviour in mouse subcutaneous tissue and ischaemic stroke model. Lanthanide downshifting nanoparticles with tunable emissions in the NIR-IIb sub-window (1,500–1,700 nm) region are ideal for deep-tissue imaging. Biofunctionalized core–shell, cubic-phase thulium-based nanoprobes show the non-invasive imaging of murine cerebral vasculature and the tracking of single immune cells and their extravasation in an inflammatory microenvironment.