Direct immunosensing of avian influenza A virus in whole blood using hybrid nanocomposites

Abstract

A sandwich-based electrochemical immunosensor was designed for detection of avian influenza virus (AIV) strains H5N1 and H4N6. This sensor was developed using gold-graphene nanocomposites, immobilized viral antibodies, and CdTe quantum dot electrochemical tagging. The nanocomposites were formed by the simultaneous reduction of a gold salt and graphene using hydroquinone as the reducing agent, thus producing non-spherical gold nanoparticles on graphene sheets. Viral antibodies were immobilized on nanocomposites and CdTe quantum dots through N-(3-dimethylaminopropyl)-N'-ethylcarbodiimide and N-hydroxysuccinimide chemistry. Cyclic voltammetry studies were used to validate the detection of H5N1 surface protein and H4N6 inactivated virus. The immunosensor detected H5 protein in phosphate buffer solution (pH 7.4) with a limit of detection (LOD) of 10 fg/mL and a linear detection range was established for 10 ng/mL to 10 pg/mL. The biosensor detected H4N6 in three parts diluted whole chicken blood with a LOD of 1.28x10-7 hemagglutinating units (HAU). Commercial ELISA testing for H5N1 and H4N6 showed limits of detection of 10 ng/mL and 0.128 HAU, respectively. The sensor showed 106-fold increased detection of H4N6 virus in blood in comparison to its commercial ELISA kit counterpart. The developed immunosensor effectively change the way avian influenza is detected, monitored, and controlled; transforming time-consuming reactive methods, into rapid predictive technology.