Impedimetric Genosensor Based on Controllable Pi–Pi Functionalization of Zirconia Decorated Graphene Nanoflakes for the Detection of Epidermal Growth Factor Receptor Exon-19 Mutation

Abstract

A disposable genosensor enhanced with 1-pyrenebutyric-acid-N-hydroxysuccinimide-ester (PY) -functionalized-zirconia-graphene nanocomposite has been fabricated for the detection of exon-19 mutation in epidermal growth factor receptor (EGFR). A green approach using solvent-exfoliation and solvothermal synthesis was employed to prepare the PY-functionalized-zirconia-graphene nanocomposite. The fabrication steps of the genosensor were evaluated by electrochemical impedance spectroscopy (EIS) and cyclic voltammetry (CV). Both tests advocated that the capture-DNA designed to detect the wild-type sequence of exon-19 was effectively immobilized on the electrode surface via a stable amide bond with the biolinker. The response of the sensor was amplified by optimizing the ratio of PY, the concentration and the immobilization time of the capture-DNA, as well as the hybridization time and temperature of the capture-target DNA. Under optimized sensing conditions, the wild-type DNA can be quantified from 5 nM to $1~\mu \text{M}$ , with an R2 = 0.99 and a limit of detection (LOD) of 1.7 nM. The sensor exhibited a reproducibility of 8.3% relative standard deviation (RSD), and satisfactory stability after 8 days of storage. The fabricated genosensor is disposable, label-free, and cost-effective, making it advantageous for clinical applications. This work not only demonstrated a reliable technique for the detection of lung cancer-associated DNA mutation but also offered a promising portable platform for the detection of other genetic mutational diseases in general.