A Dual-Channel NIR Fluorescent Probe for Self-Calibrated Ratiometric Imaging and In Vivo Discrimination of Malignant Tumors via Alkaline Phosphatase Activity

). Encapsulation within lipid nanomicelles facilitates efficient cellular uptake and affords excellent specificity with minimal cytotoxicity. The dual-channel platform successfully distinguished ALP-high tumor cells (HeLa and HepG2) from ALP-low melanoma cells (B16F10) and normal cells (NIH3T3) by confocal imaging. In xenograft mouse models bearing HeLa or B16F10 tumors, ratiometric fluorescence imaging revealed a significantly higher signal ratio in HeLa tumors, enabling clear discrimination based on endogenous ALP activity. The probe exhibited selective activation and prolonged retention in ALP-overexpressing HeLa tumors, underscoring its potential for precise in vivo tumor imaging and enzymatic activity profiling. This work establishes a generalizable strategy for high-fidelity ratiometric enzymatic sensing and provides a useful tool for cancer diagnostics and microenvironmental analysis.