Fabrication of electrochemical biosensor consisted of multi-functional DNA structure/porous au nanoparticle for avian influenza virus (H5N1) in chicken serum

Abstract

Avian influenza virus (AIV) is one of the most harmful pathogens to living things due to its fast infection, various mutations, and dangerous symptoms. In this study, we fabricated a label-free AIV H5N1 biosensor composed of multi-functional DNA structure on a porous Au nanoparticles (pAuNPs) fabricated electrode using the electrochemical (EC) technique. As a multi-functional bioprobe, the DNA 3 way-junction (3WJ) was introduced. Each fragment of DNA 3WJ was rolled to recognition part (hemagglutinin (HA) protein detection aptamer), EC signal generation part (horseradish peroxidase (HRP)-mimicked DNAzyme), and immobilization part (Thiol group). Each fragment was assembled in order to form the DNA 3WJ for AI detection and the assembled structure was confirmed by native-tris boric acid magnesium polyacrylamide gel electrophoresis (TBM-PAGE). Moreover, in order to increase the electrochemical signal sensitivity, pAuNPs were synthesized. The property of pAuNPs was investigated by field emission scanning electron microscopy (FE-SEM), Transmission electron microscopy (TEM), Ultraviolet–visible (UV-VIS) spectroscopy and zeta potential analysis. The DNA 3WJ on pAuNPs-modified Au electrode was then prepared using the layer-by-layer (LbL) assembly method. FE-SEM and atomic force microscopy (AFM) were used to investigate the surface morphology. Cyclic voltammetry (CV) was carried out to confirm the HA protein binding to DNA 3WJ-modified electrode. Moreover, The HA protein can be detected 1 pM in HEPES solution and 1 pM in diluted-chicken serum, respectively. The present study showed label-free, simple fabrication, and easy-to-tailor detection elements for AIV. The present biosensor can be a powerful candidate for various virus detection platforms.