Biomimetic isolation of affinity peptides for electrochemical detection of influenza virus antigen

In this study, we developed an electrochemical biosensor coated with a peptide that can detect the protein hemagglutinin (HA) present in the envelope of the influenza virus (H5N1). Using phage display, high-affinity peptide-displaying phage particles (GHPHYNNPSLQL) with specific binding abilities for HA were identified, which were evaluated using enzyme-linked immunosorbent assays (ELISAs). Based on the molecular structure identified by the functional studies, six newly designed peptides were chemically synthesized, and their binding interactions with HA proteins were characterized by atomic force microscopy (AFM), scanning electron microscopy (SEM), isothermal titration calorimetry (ITC), and surface plasmon resonance spectroscopy (SPR). Square wave voltammetry (SWV) revealed the presence of a linear relationship between the change in current and concentration of HA protein. From these observations, we selected IFA BP1 peptide as a potential molecular receptor and confirmed that the limit of detection (LOD) was found to be 3.26 nM. In addition, our electrochemical sensor successfully detected HA spiked in human plasma samples. Furthermore, good results for reproducibility and recovery using this biosensor were observed within a range of 31.2–500 nM concentrations and 82.3–111.5%, respectively. Importantly, the levels of HA proteins in the samples of human plasma measured using this sensor system corresponded well with those determined using a commercial ELISA method and also offered advantages such as low cost of fabrication, faster analysis, and use of lower sample volumes, suggesting that it could be applied for rapid and effective detection of the influenza virus antigen.