Electrochemical biosensor based on cellulose nanocrystals functionalized with peptide-module probes for sensitive...

The high transmissibility and rapid evolution of the influenza virus have contributed to its global spread. Consequently, there is an urgent need for versatile and advanced approaches that enable rapid biosensor development for effective influenza virus detection and control. In this study, we developed a sensitive electrochemical biosensor in which a fusion of cellulose-binding modules and virus-binding peptide (CBM31-VBP1) was site-specifically immobilized on a cellulose nanocrystal (CNC)-coated electrode. The CBM31-VBP1/CNC system enhanced electrical conductivity, porosity, electroactive surface area, and active site availability for H5N1 recognition. The sensing mechanism operated via signal suppression, with the reduction in peak current driven by specific interactions between immobilized virus-binding peptides on the CNC-coated Au electrode and viral hemagglutinin (HA) proteins. The biosensor enabled sensitive detection of H5N1 hemagglutinin (HA), achieving a limit of detection (LOD) of 3.16 nM. The CBM31-guided immobilization strategy established a stable and oriented biointerface that facilitated reliable virus recognition. In addition, the biosensor demonstrated excellent storage stability, maintaining activity for 14 days at 4 °C. Moreover, low concentrations of H5N1 HA in virus transport medium were reliably detected. These findings highlight the CBM31-VBP1/CNC platform as a promising biointerface engineering strategy for sensitive and accurate detection of influenza virus, with potential to expand point-of-care diagnostic applications to diverse diseases.