Local electrochemical impedance spectroscopy for the evaluation of labelled antibodies

Abstract

Electrochemical impedance spectroscopy (EIS) is a powerful, non-destructive and informative technique, which has been successfully applied for the characterization of GOx-based biosensor surfaces [1]. However, conventional EIS based techniques represent only averaged response of the entire electrochemical system. In order to get more advanced mapping of the electrochemical system, scanning electrochemical microscopy (SECM) merged with EIS (SEIM) could be applied. In the SEIM based technique localized impedance measurements could be performed in the frequency range when the surface of interest is scanned by an ultramicroelectrode (UME). The result of SEIM could be visualized by mapping one of the calculated parameters, e.g. charge transfer resistance or double layer capacitance as a function of 3D coordinates [2,3]. The detection of antigen and antibody interaction at the localized surface areas can be performed by SECM [4]. SECM is a better technique than spectroscopy and ellipsometry, which can be used for the determination/evaluation of proteins on the surface but do not indicate or map their exact location. In this research, SEIM was applied for the experiments on: i) a gold wafer; ii) a gold wafer modified with a self-assembling monolayer (SAM) made of 11-mercaptoundecanoic acid and iii) a gold wafer modified with SAM and enzyme-labelled antibodies. The efficiency of covalent enzyme-labelled antibodies immobilization and location of these conjugates on the surface were evaluated applying the most suitable equivalent electrical circuits (EEC). The thickness and ion-permeability of formed protein layer was evaluated by the calculation of charge transfer resistance (RCT) in selected EEC. RCT increased together with every added modification layer.