Facile and in situ production of reduced graphene oxide nanosheets paste electrode via electrochemical exfoliation of carbon paste electrode for fabricating a VEGF165 tumor marker aptasensor

The sensitive detection of trace amounts of vascular endothelial growth factor (VEGF165) in samples holds significant potential for clinical cancer diagnosis across various cancer types. This work introduces a simple, quick, and economical electrochemical method for exfoliating a graphite paste electrode (CPE) to directly synthesize a reduced graphene oxide nanosheet paste electrode (r-GONPE) for the determination of VEGF165. Thionine (Th) was used as a redox probe due to its strong interaction with the graphene surface. The functionalization of r-GONPE by thionine improves its stability while preserving the intrinsic properties of r-GONPE. Au nanoparticles were electrodeposited on the r-GONPE/Th electrode to stabilize the SH-aptamer by self-assembly for selective interaction and to enhance the speed of electron transfer. FESEM images confirmed the formation of a fixed, stable, very thin, and large few-layer reduced graphene oxide nanosheet on the outer surface of the CPE. The surface area of the bare CPE increased significantly after electrochemical exfoliation of the surface graphite layers (from 0.059 cm² for CPE to 0.281 cm² for r-GONPE). Electrical characterization showed that the conversion of graphite to graphene resulted in a five-fold increase in peak height and a decrease in peak separation by nearly 30 mV. The selective interaction of the VEGF165 with the r-GONPE/Th/nano-Au electrode modified with SH-aptamer reduced the oxidation peak current of thionine in the DPV signal. Thus, a wide linear calibration ranges from 5.0 pM to 320 pM and a low LOD of 1.03 pM (using the 3σ/S equation) were obtained. The aptasensor demonstrated consistent stability and strong selectivity against typical interferences when detecting picomolar concentrations of VEGF165. It also showed high accuracy and speed in detecting VEGF165 in human serum samples. Additionally, this aptasensor is advantageous due to its cost-effectiveness and simple fabrication process compared to traditional methods that use complex nanocomposites for signal enhancement.