Apt-Nanogel-Kit for Real-Time Quantitative Monitoring of the Released H 2 O 2 from Living Cells and Point-of-Care Application

Hydrogen peroxide (H₂O₂) is a key redox signaling molecule that plays a vital regulatory role in diverse physiological processes. However, existing tools for near-real-time H₂O₂ detection are often constrained by limited applicability in dynamic biological environments. To address these challenges, we developed a point-of-care aptamer-functionalized nanozyme-hydrogel kit (Apt-ngel-kit) that integrates nanozyme-based homogeneous catalytic fluorescence with digital signal processing, enabling highly sensitive, quantitative, and near-real-time monitoring of in situ H₂O₂ secretion. The system leverages the exceptional enzymatic activity of ferromanganese silicate nanozymes (FMSN), which were immobilized in a polyacrylamide hydrogel coloaded with the catalytic substrate o-Phenylenediamine (OPD) to create a homogeneous, stable, and reproducible catalytic nanoplatform. Furthermore, to detect the near-real-time secretion of H₂O₂ within cells, we conjugated Apt to the nanogel surface to improve the biocompatibility of the Apt-ngel-kit. It is noteworthy that this system not only achieves an ultralow in situ detection limit of 8.32 × 10^-23 M, but also quantifies H₂O₂ released by individual cells and distinguishes the secretion characteristics between cancer patients and healthy individuals, demonstrating its potential as a rapid and effective tool for early disease diagnosis.