Prussian blue electrocatalyst in bioanalysis: From oxidase biosensors to peroxidase kinetics assays

Prussian blue (PB) has evolved as the most advantageous electrocatalyst for preparing H₂O₂ biosensors due to its exceptional high sensitivity and specificity, which are vital to minimizing the interferences in complex biological samples. However, the widespread adoption of PB-based bioanalysis has been severely restricted by its inherent instability in neutral and alkaline environments, compromising the measurement accuracy and reliability. This review provides an overview of the strategies used to improve PB stability in bioanalysis, such as integration with carbon nanomaterials, polymer encapsulation (e.g., Nafion and chitosan), and transition metal ion (e.g., Ni, Co)-doping. Furthermore, we comprehensively cover the bioanalytical applications of PB: from traditional oxidase-based biosensors to emerging high-impact applications enabled by the enhanced PB stability. These includ real-time H₂O₂ quantification in tumors and brain tissue, electrochemical immunosensing, and kinetics assays for peroxidases and peroxygenases acting on insoluble substrates.