In Vivo NIR-II Ratiometric Imaging of Peroxynitrite for Tracking Redox Dynamics in Tumor Immunochemotherapy

Peroxynitrite (ONOO^-) is a key mediator of redox imbalance in inflammation and cancer, with in vivo monitoring of its dynamic behavior still posing a major challenge. Here, we developed a BD@PEG nanoprobe that enabled dual-excitation NIR-II ratiometric imaging with superior biocompatibility, quantitative fidelity, and ONOO^- selectivity. When applied to a drug-induced liver injury model, BD@PEG-based NIR-II ratiometric imaging allowed real-time monitoring of hepatic ONOO^- dynamics, directly correlating oxidative/nitrosative stress to histopathological alterations, and demonstrating significant attenuation under glutathione treatment. In a CT26 tumor model, the NIR-II ratiometric imaging strategy revealed distinct ONOO^- modulation in response to different therapeutic interventions. Furthermore, two-plex imaging by integrating NIR-IIb oxyhemoglobin saturation analysis afforded concurrent assessment of nitrosative stress and vascular oxygenation. We found that the combination of chemotherapy and immunotherapy induced synergistic ONOO^- production, enhanced immune cell infiltration, and achieved superior antitumor efficacy. Overall, this study established a versatile imaging platform for dynamic redox monitoring and oxygenation assessment in vivo, providing mechanistic insight into ONOO^--mediated pathology and a translational tool for optimizing therapeutic interventions in cancer and inflammation.