Wan, YingpengYingpengWanGao, YijianYijianGaoChen, Yu‐NengYu‐NengChenLee, Ka‐WaiKa‐WaiLeeWang, Hao‐WenHao‐WenWangTsai, Ya‐JieYa‐JieTsaiChen, WeilongWeilongChenLi, ShengliangShengliangLiWong, Ken‐TsungKen‐TsungWongLee, Chun‐SingChun‐SingLee2025-09-172025-09-172025-08-29https://scholars.lib.ntu.edu.tw/handle/123456789/732174Near-infrared-II (NIR-II) fluorescence imaging has transformed biomedical imaging by providing low autofluorescence, deep tissue penetration, and superior spatial resolution, essential for precise in vivo biological visualization. However, developing NIR-II-emitting molecular fluorophores with high brightness remains challenging due to the difficulty in simultaneously achieving high absorption coefficients and excellent fluorescence emission in aggregated states. This study addresses these challenges at both the molecular and aggregate levels through a series of donor-acceptor-donor (D-A-D) configured small molecules. The initial twisted molecule, DTTD, exhibits low absorption and significant fluorescence quenching upon aggregation. To mitigate these issues, DMTTD is synthesized with a planarized backbone by locking the π bridge with the phenylene ring of donor, resulting in a higher absorption coefficient and reduced quenching due to its rigid structure and bulky donors. Further enhancement is achieved by incorporating branched alkyl chains into DMTTD for wrapping the molecular skeleton, creating DETTD, which maintains a high absorption coefficient and improved resistance to quenching in aggregated states. DETTD nanoparticles (NPs) with high NIR-II brightness of 117.5 M−1 cm−1 enabled high-resolution imaging of mice vasculature and gastrointestinal tract. This study establishes a general paradigm for simultaneously mitigating fluorescence quenching and improving absorption coefficients to achieve bright NIR-II fluorophores.enanti-quenchingdonor-acceptor-donor dyehigh brightnessNIR-II fluorescence imaging[SDGs]SDG3journal article10.1002/adma.202510386