Application Of Functional Magnetic Nanoparticles In Immunoassay And Study Of Biosensor

There are six chapters in the paper.In the first chapter, the magnetic polymer microsphere, which included the significance, structure, characteristic and prospect, was reviewed. The preparative methods of magnetic polymer microspheres were described in detail. In addition, the application of magnetic polymer microspheres, which included magnetism target medicament, enzyme immobilizing, cell separating, immunoassay, DNA separating etc, was also systemically reviewed.In the second chapter, on the basis of the former work, we studied optimum conditions preparation for magnetic polymer microsphere. The nano-magnetic polymer carboxyl microsphere and amino microsphere were prepared successfully by dispersion polymerization. Moreover characteristics of magnetic polymer microsphere were verified. The morphology of the magnetic polymer microsphere was examined by using transmission electron microscopy (TEM) and scanning electron microscopy (SEM). The magnetic measurement was carried out at a physical property system. The functional group of the microsphere was confirmed by Fourier transform infrared (FTIR) spectroscopy. The fluorescent experiment was studied on the magnetic polymer microsphere connected with antibody labeled fluorescent substance. The magnetic polymer microsphere showed the following characteristics, such as super paramagnetism, narrow distribution of diameter, bounding to bioactive substance effectively.In the third chapter, the magnetic polymer microsphere was applied to immunoassay. In the former section of the chapter, the enzyme-linked Immunovoltammetry was used to determine antigen with magnetic polymer microsphere. The surface of the nano-magnetic polymer microsphere covalently bound with antibody, and then captured antigen and secondary antibody labeled with horseradish peroxide enzyme. The microsphere with sandwich complex was separated magnetically from sample solution and the enzyme labeled at the complex catalyzed the substrate adding to reaction, the electroactive product was detected by using differential pulse voltammetry (DPV). Thisnew analytical assay offered the promise for rapid, low-cost, good separate efficiency and high sensitivity. The detection limit of antigen was found to be 0.06ng/mL. The sensitivity was much higher than that of enzyme-linked immunosorbnent assay (ELISA) used in diagnostic analysis. In the latter section of the chapter, the magnetic polymer microsphere was used to prepare electrochemistry biosensor. At first, the surface of the nano-magnetic polymer microsphere covalently bound with antibody, and then captured antigen and secondary antibody labeled with electroactive substance, The microsphere with sandwich complex was separated magnetically from sample solution and was attracted to gold electrode surface by magnetic field. The electroactive substance labeled at the complex was detected directly by using differential pulse voltammetry. This new analytical assay offered the promise for rapid, low-cost and high sensitivity. The determination was carried out directly no substrate adding to solution. Moreover, because magnetic polymer microsphere had superparamagnetic property and was dispersed into solution after magnetic field disappear and then the electrode surface was renewed immediately. The analytical method was fit for on-line assay.In the forth chapter, a sensitive microgravimetric biosensor had been developed by using secondary antibody labeled with magnetic nano-microspheres, which could be combined with antigen and antibody on the gold electrode of a quartz crystal microbalance. These modified nano-microspheres had been used as the amplified label for microgravimetric signal because their heavy mass and the sensitivity of detection had been enhanced greatly.In the fifth chapter, a sensitive electrochemical biosensor was developed for the detection of nucleic acid hybridization and sequence-specific DNAs. The biosensor was based on a graphite electrode modified ssDNA using 2.3.5-triphenyl-tetrazolium chloride as hybridization indicator. It showed high electroactivity and low background current in electrochemical measurements and high selectivity to bind to dsDNAs on the electrode surface. The detection limit of the biosensor for DNA was 6.0 X 10'" mol/L in differential pulse, voltammetry.In the last chapter, some neurocompounds were investigated by using the electrodemodified with silk fibroin membrane. The silk fibroin protein membrane was an amphoteric ion-exchange membrane composed of both weak acidic and weak basic groups. The membrane with isoelectric point of pH 4.5, the modified electrode was negatively charged in pH > 4.5 solutions, rejected anions and attracted cations. In pH < 4.5 solutions, the electrode charged positively, rejected cations and attracted anions. The pH-responsive charge recognition of the modified electrode was investigated for some neurocompounds. A fibroin carbon fiber electrode had been used successfully for in-vivo determination.