An amperometric biosensor for specific detection of glycated hemoglobin based on recombinant engineered fructosyl peptide oxidase.

Here, we present a specific biosensor based on the detection of glycated hemoglobin (HbA1c) proteolytic digestion product, fructosyl valyl histidine (Fru-ValHis). A recombinant engineered fructosyl peptide oxidase (FPOX) enzyme with improved specificity was immobilized on the electrode surface modified by chitosan (CHIT), graphene oxide (GO) and gold nanoparticles (AuNPs). The biosensor exhibited a linear response toward different concentrations of Fru-ValHis ranging from 0.1 to 2 mM with a sensitivity of 8.45 µA mM-1 cm-2. Detection limit of the current biosensor for Fru-ValHis was 0.3 µM as the lowest quantity required giving a signal to a background. Analytical recovery of added Fru-ValHis in whole blood was 95.1-98.35% for FPOX/AuNPs/GO/CHIT/FTO electrode. For Fru-ValHis determination by FPOX-AuNPs-GO-CHIT/FTO electrode, within-run coefficient of variation (CV) was between 1.3% and 2.4% and between run CV was between 2.1% and 3.5%. A significant change in electron transfer resistance after the incubation of FPOX-modified electrode with Fru-ValHis was observed, while no response was achieved with control, indicating specific measurement of Fru-ValHis. Moreover, designed biosensor measured HbA1c in human blood samples and the results were well agreed with that obtained with NORUDIA™ N HbA1c diagnostic kit. Overall, suitable specificity of the engineered FPOX made the bioelectrode responded well to the Fru-ValHis level, which demonstrates a promising application for specific detection of HbA1c biomarker.