Dual-Readout Cerium Oxide Nanoparticle Paper-Based Sensors for Redox Profiling via Hydrogen Peroxide Detection in 2D and 3D Chronic Lymphocytic Leukemia Models

Reactive oxygen species, particularly hydrogen peroxide, play a key role in cancer biology and disease progression. However, reliable tools for monitoring H₂O₂ dynamics in complex cellular models remain limited, particularly those combining analytical performance with accessibility and operational simplicity. To address this need, a low-cost paper-based analytical device (PAD) was developed for dual readouts, integrating electrochemical (ePAD) and smartphone-assisted optical/colorimetric (oPAD) formats. Both approaches employ cerium oxide nanoparticles (nCe) as nanozymes for H₂O₂ detection. With high analytical performance in both electrochemical (3 nM–500 µM linear range, LOD 0.4 nM) and optical (1–1000 µM linear range, LOD 0.3 µM) modes, covering physiologically relevant H₂O₂ concentrations, the dual-readout strategy enables independent confirmation of H₂O₂ levels in highly complex biological systems under rapid analysis (<1 min), requiring only 5 µL of sample and exhibiting excellent operational stability. Validation in 2D cultures and 3D CLL co-culture systems demonstrates the PAD capabilities to monitor redox dynamics in tumour microenvironment models. These results highlight the potential of integrating dual-readout PADs for reliable redox analysis in biologically relevant systems, paving the way for decentralized redox sensing and broader applications in biomedical research.