Physical absorption vs covalent binding of graphene oxide on glassy carbon electrode towards a robust aptasensor for ratiometric electrochemical detection of vascular endothelial growth factor (VEGF) in serum

Achieving a biosensing interface with desirable performance is essential to a successful biosensor. In this paper, we developed a robust aptasensor by modifying methylene blue loaded graphene oxide (GO/MB) and ferrocene-labeled aptamer onto glassy carbon (GC) electrodes to realize the dual electrochemical signal mode ratiometric quantification of vascular endothelial growth factor (VEGF) in serum. For comparison, two sensing interfaces based on the covalent attachment (GC-ph-GO/MB-streptavidin-aptamer) and physical absorption GC-GO/MB-streptavidin-aptamer) of GO/MB to GC electrodes were fabricated, respectively. Redox molecule ferrocene labeled structure-switching aptamers having affinity to VEGF were used as the recognition probes providing electrochemical signal readout with the presence of the analyte VEGF. Larger amount of aptamers was observed on the GC-GO/MB-streptavidin-aptamer than that on GC-ph-GO/MB-streptavidin-aptamer while having the similar stability, which contributed to the higher sensitivity of GC-GO/MB-streptavidin-aptamer. The linear range of GC-GO/MB-streptavidin-aptamer and GC-ph-GO/MB-streptavidin-aptamer for detection of VEGF was 10–500 pg mL⁻¹ and 20–500 pg mL⁻¹, respectively. The GC-GO/MB-streptavidin-aptamer surface was able to realize the dual-signal mode ratiometric quantification of VEGF with wider linear range comparing with a single-signal mode detection. This electrochemical apatasensor provides a robust universal biosensing platform for sensitive ratiometric detection of a spectrum of analytes in real time.