ABSTRACT
An immunosensor for label-free electrochemical detection of MiniChromosome Maintenance Protein 5, MCM5, a protein overexpressed in cervical cancer, based on a gold electrode is reported. The electrode was first modified with a submonolayer (capture layer) of 11-mercaptoundecanoic acid (11-MUA) and then activated with N-(3-dimethylaminopropyl)-N-ethylcarbodiimide hydrochloride (EDC) and N-hydroxysuccinimide (NHS) to immobilize the capture antibody. The change in electrode surface properties (wettability) during the formation of the 11-MUA layers was determined using the static water contact angle (WCA). The binding of MCM5 antigens to the capture antibody was monitored by cyclic voltammetry (CV) and electrochemical impedance spectroscopy (EIS) using 5 mM [Fe(CN)6]3-/4- in 0.1 M LiClO4(aq) as an electroactive probe. AC Impedance was used to measure charge transfer resistance (Rct), which reflects impeded electron transfer when the antigen is bound to the antibody functionalized surface. After exposing the antibody-functionalized surface to MCM5 antigens, Rct increases linearly with the logarithmic value of MCM5 antigen concentration, with a linear dynamic range of 10-6 to 10-11 g/mL, a correlation coefficient of 0.99, and a detection limit of 2.9 pM (10-11 g/mL). This excellent sensitivity was achieved with simple preparation steps and minimal reagent consumption, without the need for complicated procedures such as enzymatic amplification, fluorescent labeling, or nanoparticle modification.