Publications

Intravital fluorescence imaging of vasculature morphology and dynamics in the brain and in tumors with large penetration depth and high signal-to-background ratio (SBR) is highly desirable for the study and theranostics of vascular-related diseases and cancers. Herein, a highly bright fluorophore (BTPETQ) with long-wavelength absorption and aggregation-induced near-infrared (NIR) emission (maximum at ≈700 nm) is designed for intravital two-photon fluorescence (2PF) imaging of a mouse brain and tumor vasculatures under NIR-II light (1200 nm) excitation. BTPETQ dots fabricated via nanoprecipitation show uniform size of around 42 nm and a high quantum yield of 19 ± 1% in aqueous media. The 2PF imaging of the mouse brain vasculatures labeled by BTPETQ dots reveals a 3D blood vessel network with an ultradeep depth of 924 μm. In addition, BTPETQ dots show enhanced 2PF in tumor vasculatures due to their unique leaky structures, which facilitates the differentiation of normal blood vessels from tumor vessels with high SBR in deep tumor tissues. Moreover, the extravasation and accumulation of BTPETQ dots in deep tumor (more than 900 μm) is visualized under NIR-II excitation. This study highlights the importance of developing NIR-II light excitable efficient NIR fluorophores for in vivo deep tissue and high contrast tumor imaging.

Two-photon fluorescence imaging allows in vivo study of biological structures and activities in deep tissues, in which bright fluorophores with high photostability and good biocompatibility are highly desirable. Herein, a small-molecule fluorogen with aggregation-induced emission (AIEgen) is complexed with fetal bovine serum (FBS) proteins to develop a protein-sized AIEgen–protein hybrid nanocomposite (TPEPy-FBS) with bright far-red/near-infrared (NIR) emission, excellent photostability, and low phototoxicity for deep and high-resolution in vivo two-photon brain vasculature imaging. Upon complexation with FBS, the fluorescence of TPEPy is greatly intensified and a sixfold enhancement is observed with 10% FBS in aqueous media. The yielded TPEPy-FBS shows good physical stability in aqueous media and the phototoxicity of TPEPy is dramatically inhibited after complexation with FBS. Moreover, TPEPy-FBS exhibits bright two-photon fluorescence in far-red/NIR region and good photostability upon femtosecond laser excitation, which facilitates high performance in vivo imaging. A large imaging depth of 656 μm is obtained in brain vasculature network imaging with a high signal-to-background ratio of 234, where a small blood capillary of 1.05 μm can be resolved at an imaging depth of 656 μm. Highlighted is a simple and versatile strategy to develop efficient two-photon probes for in vivo biological imaging.