NIR-II fluorescence imaging demonstrates significant advantages in biological imaging with its high signal-to-background ratio （SBR） and deep tissue penetration， showing broad application prospects in biomedical fields. The classification of NIR-II imaging windowsfacilitates the optimization of imaging processes. Among these， the 1400-1500 nm imaging window benefits from its unique water absorption characteristics， enabling effective suppression of scattering background and achieving high-contrast imaging. This study systematically evaluates the imaging potential of the 1400-1500 nm window through simulation studies and in vivo experiments. To advance the clinical translation of fluorescence imaging in the 1400-1500 nm window， indocyanine green （ICG）， an organic small-molecule dye approved by the U.S. Food and Drug Administration （FDA）， was employed as the fluorescent probe. Utilizing its extended fluorescence emission tail in the NIR-II region， high-contrast and high-resolution imaging of mouse vasculature and intestinal structures was achieved in the 1400-1500 nm window. Furthermore， in combination with methylene blue （MB）， another FDA-approved agent， high-quality dual-channel NIR-II imaging was successfully implemented enabling precise localization of blood vessels and lymph nodes in mice. This research further explores the unique advantages of the 1400-1500 nm imaging window in biological imaging and its clinical application potential. It also provides valuable references for the clinical translation of NIR-II fluorescence imaging.