Immunosensor Based On Carbon Nanomaterials And Electrochemiluminescence Resonance Energy Transfer For Mouse IgG Detection

Electrochemiluminescence (ECL) is an effective trace analytical technology due to it integration of high sensitivity and good temporal/spatial controllability. As a quantitative measurement method for antigen or antibody, electrochemiluminescence immunoassay (ECLIA) is based on electrochemiluminescence signal change before and after immunoreactions. ECL immunosensor is a novel biosensor based on specific antigen-antibody interactions, which combines the high sensitivity of sensor technology with high specificity of immunoreaction. Since ECL immunosensor shows some remarkable features such as low background, high sensitivity, wide dynamic range, good temporal/spatial controllability, simple instrumentation and rapidity, it has been developed into an important detection mean for antibody, antigen and hapten with different molecular size.Similar to the fluorescence energy transfer, the excited ECL luminophor can act as an energy donor and transfer its energy to energy acceptor. Among all the traditional ECL system, Ru(bpy)32+ and its derivatives are the most widely used ones due to its outstanding characteristics, such as good water solubility, high ECL efficiency, good chemical stability and electrochemical renewability. In this thesis, a novel disposable ECL immunosensor was developed, in which Ru(bpy)32+ acted as energy donor in the ECL energy transfer system.Amorphous carbon nanoparticles (ACNPs) showing highly quenching efficience were prepared from candle soot with a very simple protocol. In this work, it was found that ACNPs can efficiently quench the ECL emission from Ru(bpy)32+. Thus this ECL quenching system was adopted to establish a facile, low-cost immunosensor for a model analyte (MIgG). First, ACNPs were linked with antigens to obtain ACNP -antigen conjugate. Then, the immunoreaction between the Ru(bpy)32+-labeled antibody and the ACNP-antigen conjugate bridged the Ru(bpy)32+ donor and the ACNPs acceptor, which led to quenching of ECL emission from Ru(bpy)32+ via energy transfer. In the presence of the target antigen, this antigen competed with the ACNP-antigen for the immobilized Ru(bpy)32+-labeled antibody, thus led to decreased occurrence of the ECL quenching. This ACNPs-based ECL quenching system was successfully applied to detect mouse IgG. The method showed a linear range of 0.50-400 ng/mL and a detection limit of 0.35 ng/mL. The proposed immunosensor showed a much lower detection limit compared with ELISA assay(3.2μg/mL). The stability, the specificity and the recovery of the proposed immunosensor were also acceptable. Compared with other approaches using luminescent materials as the acceptors, the present protocol does not suffer from overlap of the emission spectra of donor and acceptor. The proposed method showed its advantages of simple fabrication, low cost, high sensitivity, suitability for point-of-care test and field analysis.