A Dual-Cascade Activatable Photoacoustic Sensor for Tracking Iron(II) and Reactive Oxygen Species during Ferroptosis

Ferroptosis is an iron-dependent form of programmed cell death driven by lipid peroxidation and is closely associated with various diseases. Conventional fluorescence imaging of ferroptosis suffers from limited tissue penetration and specificity. Photoacoustic (PA) imaging overcomes these limitations by coupling optical contrast with centimeter-scale penetration and high spatial resolution, but most existing PA probes respond to single biomarkers and are prone to off-target activation by nonferroptotic oxidative stress. Here, we report the design and synthesis of a manganese-based Prussian blue analogue (MnPB) nanosensor for monitoring ferroptosis. The nanosensor operates via a cascade reaction with two key biomarkers, Fe 2+ and H 2 O 2, enabling a differential PA signal readout. The designed nanosensor first reacts with Fe 2+ through ion exchange, which enhances the PA signal at 800 nm. Subsequently, the resulting intermediate product is oxidized by H 2 O 2, leading to a significant amplification of the PA signal at 700 nm. This dual wavelength activation enables a differential signal readout that transitions from negative to positive, functioning as a specific “AND” logic gate for the codetection of both markers. This nanosensor exhibits excellent stability, biocompatibility, and high selectivity against various biological interferents. In both cellular and 4T1 tumor-bearing mouse models, the MnPB nanosensor enables precise visualization of Fe 2+ and ROS dynamics during ferroptosis in vivo. This work presents a novel and robust strategy for the dynamic, noninvasive monitoring of ferroptosis in vivo, offering a powerful tool for advancing our understanding of tumor biology and evaluating the efficacy of ferroptosis-based cancer therapies.