An electrochemical immunosensor that can operate in biological fluids: applications for the detection of anti-Glycosylated Hemoglobin (anti-HbA1c) Antibodies

摘要

Electrochemical Immunosensors have been a promising approach in diagnostics. However, they have to overcome the enduring challenge of biofouling when applied in biological fluids. The research presented here therefore focuses on developing a simple, straightforward and efficient methodology of fabricating an electrochemical immunosensor capable of functioning in complex matrices. Towards its applications, the important anti-glycosylated hemoglobin antibody will be used as a sample analyte. The immunosensor developed here, consists an electrode, antifouling layer and sensing gold nanoparticles. The key question was how thick the organic layer can be to achieve efficient electron transfer across the electrode-insulator-nanoparticle (NP) construct. Thus, electron transfer process of the electrode-insulator-NP constructs was first studied in the context of a recent theory proposed by Chazalviel and Allongue. A precise tuning of the thickness of the organic film, using the electrodeposition of poly(ethylenediamine) on glassy carbon electrodes, followed by the study of its effect on the efficiency of the electron transfer in NP-constructs, verified the theory and demonstrated a transition from thickness-independent to thickness-dependent electron transfer as the layer thickness exceeds a certain threshold of 20 A. Also, another study showed that the organic film must be able to form uniformly continuous films to fully exploit the potential of electrode-insulator-NP constructs. ethylene glycol, was found to be an appropriate organic layer to modify the gold surfaces and fabricating the immunosensor. The entire fabrication process was performed in a straightforward method. Cyclic voltammetry for the modified gold electrodes showed a close to ideal surface-bound electrochemistry for ferrocene with an E-half of +277 mV (vs. Ag|AgCl) which falls in an ideal potential window for the stability of the construct. The sensing surface demonstrated great antifouling and selectivity properties toward different antibodies and proteins, and an excellent capability of functioning in blood was observed. Detection of anti-HbA1c antibody as a sample analyte in solution was assessed using enzyme-linked immunosorbent assay and square wave voltammetry where a good sensitivity to detect the antibody with a concentration range of 0.2 to 2.0 ug/ml was revealed.