Gold Array Based Electrochemical Immunobiosensor For Detection Of Escherichia Coli O157:H7

A novel electrochemical immunbiosensor was developed for rapid detection of Escherichia coli O157:H7 based on gold electrode array. The immobilization of antibodies onto the electrodes was carried out by a monolayer of 16-mercaptohexadecanoic acid (MHDA), a long-chain carboxylic acid-terminated alkanethiol, self-assembled on gold electrodes. The monolayer was activated by l-ethyl-3-(3-dimethylaminopropyl) carbodiimide (EDC) and N-hydroxysuccinimide (NHS) to generate a stable acyl amino ester intermediate. Antibodies were then immobilized after aminolysis of the NHS adduct. The binding of target bacteria onto the immobilized antibodies changed the surface characteristics of gold electrodes, which can be detected by electrochemical measurements.The electrochemical impedance spectroscopy and cyclic voltammetric method were employed after each step of surface modification in pH 7.4 PBS buffer containing a redox probe of [Fe(CN)6]^3-/4-. The effect of self-assembled monolayers (SAMs) and the bacteria layers on the electrochemical properties of the electrodes were quantitatively analyzed in terms of constant phase element (CPE) and electron transfer resistance using proper equivalent circuit. It was demonstrated that these layers act as barriers for the electron transfer between the electrode surface and the redox species in the solution, resulting in most significant increase in the electron transfer resistance compared to CPE. The immunobiosensor could detect the target bacteria with detection limit of 2.0 × 10² CFU/ml. A linear response in the electron transfer resistance was found between 2.0 × 10² 2.0 × 10⁷ CFU/ml. The biosensor could detect six different concentrations of bacteria at the same time, which shortened the detection time in a large scale.