Study On Biosensors Based On Novel Nanocomposite Films

Electrochemical biosensors plays an important role in food, pharmaceutical and clinical fields because of the advantages such as good accuracy, fast analysis, high sensitivity and simple detection process. With the rapid progress in nanotechnology, nanomaterials have become the subject of intense investigation in the fabrication of electrochemical biosensors. Thus, in this thesis, some different types of electrochemical biosensors were constructed, and the application of some nanomaterials in electrochemical biosensors and the fabrication of sensing interface were investigated.A nonenzymatic electrochemical sensor based on silver nanowires (Ag NWs) and a nonenzymatic electrochemical sensor base on gold-silver bimetallic nanotubes (Au-Ag NTs) were constructed. Uniform Ag NWs were synthesized by a polyol process, whose average diameter was about100nm. The Au-Ag NTs with hollow tubular structures were synthesized by substitution reaction using the Ag NWs as the templates. The electrode modified with Ag NWs and chitosan (CS) film could detect hydrogen peroxide in the linear range of0.008～1.35mmol L-1. with a detection limit of2μmol L-1(S/N=3). The hydrogen peroxide in milk samples were assessed by the proposed sensor, and the recovery was in the range of91.7%～107.9%. The electrode modified with Au-Ag NTs/CS film could detect ascorbic acid in the linear range of0.005～2.000mmol L-1, with a detection limit of2μmol L-1(S/N=3). The ascorbic acid in vitamin tablets were detected by the prepared sensor, and the results were consistent with the labeled values. The proposed nonenzymatic sensors showed acceptable reproducibility and good long-term stability.A bienzyme biosensor for the detection of cholesterol was developed based on the horseradish peroxidase/cholesterol oxidase (HRP/ChOx) bienzyme system. The (PAH-MCNTs-GNPs/HRP)m/(PAH-MCNTs-GNPs/ChOx)n multilayer film was fabricated via layer-by-layer (LBL) assembly of positively charged poly(allylamine hydrochloride)-multiwalled carbon nanotubes-gold nanoparticles (PAH-MCNTs-GNPs) and negatively charged enzymes. The fabrication process of the bienzyme multilayer film was characterized by scanning electron microscopy (SEM) and electrochemical impedance spectroscopy (EIS). The mechanism of the detection of cholesterol was confirmed by cyclic voltammetry (CV). The comparison experiments showed that the presence of HRP in the bienzyme system could significantly enhance the response signal, meanwhile, the MCNTs and GNPs with excellent electron transfer rate could also improve the response signal. Under the optimal conditions, the bienzyme biosensor could detect cholesterol in the linear range of0.18～11mmol L-1,with a detection limit of0.02mmol L-1(S/N=3). Human serum samples were assayed by the prepared bienzyme biosensor, and the recovery was in the range of92.2%～98.5%, indicating the proposed bienzyme biosensor could be further applied to clinical detection of cholesterol.A new electrochemical immunosensor was constructed. Firstly, the uniform nanomultilayer film was fabricated via LBL assembly of positively charged poly(diallyldimethylammonium chloride)-multiwalled carbon nanotubes carbon nanotubes (PDCNTs) nanocomposite and negatively charged poly(sodium-p-styrene-sulfonate)(PSS). Subsequently, gold nanoclusters were electrodeposited on the electrode to immobilize antibodies. The fabrication process of the immunosensor were characterized by SEM, CV and EIS. Under the optimal conditions, the proposed immunosensor could detect carcinoembryonic antigen (CEA) in two linear ranges from0.1to2.0ng mL-1and from2.0to160.0ng mL-1, with a detection limit of0.06ng mL-1. The comparison experiments showed that the MCNTs could promoted the electron transfer and increased the amount of immobilized anti-CEA. The prepared immunosensor showed regeneration ability, good long-term stability and acceptable selectivity, and could be used to detect serum CEA levels.A simple and controllable one-step electrodeposition method for the preparation of a chitosan-carbon nanotubes-gold nanoparlicles (CS-CNTs-GNPs) nanocomposite film was used to fabricate an electrochemical immunosensor. The antibodies were immobilized on the electrode modified with CS-CNTs-GNPs nanocomposite film. The porous three-dimensional CS-CNTs-GNPs nanocomposite film, which offered a large specific surface area for immobilization of antibodies, exhibited improved conductivity, high stability and good biocompatibility. The fabrication process of the immunosensor were characterized by EIS and CV. Under the optimal conditions, the proposed immunosensor could detect CEA in two linear ranges from0.1to2.0ng mL-1and from2.0to200.0ng mL-1. with a detection limit of0.04ng mL-1. The immunosensor based on CS-CNTs-GNPs nanocomposite film as the antibody immobilization matrix could exhibited showed regeneration ability, good long-term stability and acceptable selectivity. Human serum samples were assayed by the prepared immunosensor and and ELISA,and the results were acceptable.