Preparation, Sers Property And Application Of Polymer/Silver-NPs Composite Microspheres

Surface enhanced Raman spectroscopy (SERS) holds great potential for analytic application due to its fingerprint-like and ultra-highly sensitive property, espically SERS has been subjected to intensive attention in biomolecular analysis because of the advantages of extremely stable signal and high spectral revolution. Biological analysis based on SERS-tags has wide applications in biomedical research, food safety monitoring, medical diagnostics, forensic analysis, cultural relic identification and so on. The sensitivity of SERS mainly depends on the morphology and size of SERS substrates. Based on the research background, this thesis has focused on the preparation of highly active SERS substrates and their application in biological analysis area. The detailed results of each part are listed as following:(1) A facile, efficient and environmentally friendly route for preparation of PSA/Ag-NPs composite microspheres was developed. Firstly, a series of poly (styrene-co-acrylic acid)(PSA) microspheres with different contents of-COOH were prepared by surfactant-free emulsion polymerization using different ratios of styrene (St) and acrylic acid (AA) as monomers. Secondly, PSA microspheres served as the supporting template and PVP acted as a reducer and stabilizer to obtain Ag nanoparticles (Ag-NPs) coated PSA composite microspheres (PSA/Ag-NPs) via in situ reduction of AgNO3by PVP in aqueous solution. The effect of carboxyl group amount and reaction temperature on surface coverage and morphology of Ag-NPs immobilized on PSA microspheres was systematically studied. The results reveal that the bigger-COOH amount on the surface of microspheres and higher reaction temperature was in favor of the increase of Ag-NPs coverage, but too high reaction temperature led to the formation of silver nanoparticle agglomerates on the microsphere surface and a great amount of colloidal silver nanoparticles in the solution.(2) PSA/Ag-NPs composite microspheres were used as SERS substrates to detect trace melamine in solution. At first, their SERS activity was evaluated using4-ABT as the model molecule. The result shows that the SERS activity of PSA/Ag-NPs composite microspheres increases with the increase of the Ag-NPs coverage on the surface of the PSA/Ag-NPs composite microspheres, and the composite microspheres with the uniform and complete coverage show the highest SERS activity with the detection limit down to10-9M for4-ABT. Potential application of the PSA/Ag-NPs composite microspheres as SERS substrates was demonstrated with the detection of melamine, and the detection limit was about10-3M. Chemical noises from PVP and other impurities were observed, which were attributed mainly to the competitive adsorption of PVP on the surfaces of Ag-NPs. After tetrahydrofuran (THF) washing of the PSA/Ag-NPs that removed the PVP and other residuals, the signal/noise levels of SERS were greatly improved and the detection limit of melamine was increased to be10-7M.(3) A class of SERS encoded core-shell microspheres (SERS-tags) was synthesized. The SERS-tags are composed of PSA/Ag-NPs microspheres as SERS-active substrates, Raman active molecules as coding for specific analyte, and silica shell for protecting the Ag-NPs and the Raman molecules from the exterior chemical and biological interferences and conjugation with biomolecules. The experimental results show that:(a) the SERS-tags exhibited high chemical stability and could resist to the etching of strong acid;(b) the SERS-tags at different concentrations produced consistent Raman signals under the identical measuring conditions and a highly linear relationship between SERS signal intensities and concentrations of nanoparticle SERS-tags with an R2=0.989was obtained;(c) At least four different tags (a four "color" system) were quantitatively differentiated in a mixture of SERS-tags indicating an excellent multiplexing potential of SERS-tags. The silica surfaces of SERS-tags were further conjugated with probe DNA to prepare SERS-probes. The detection of single-stranded oligonucleotide (ssDNA) targets was successfully accomplished using SERS-probes in a chip-based sandwich hybridization assay in a mixed ssDNA target solution. Therefore, the as-prepared SERS-probes are suitable for high specific detection of biomolecules with high sensitivity and the remarkable multiplexing capability associated with the SERS method.(4) We demonstrated a strategy for the detection of target DNA based on SERS-tags. Firstly, the magnetite colloidal nanocrystal clusters (MCNCs) with high saturation magnetization value were synthesized via a solvothermal reaction, and then they were coated with high biocompatible poly (acrylic acid)(PAA) to obtain MCNCs/PAA core/sell microspheres with fast magnetic responsiveness and excellent biocompatibility. A proof-of-concept sandwich hybridization assay for DNA detection was performed utilizing DNA-functionalized SERS-tags as probes and magnetic nanoparticles as suspended capture substrates and a separating tool, respectively. The quantitative detection of target DNA strands showed a well-defined linear correlation between SERS signal intensity and concentrations of target DNA strands, and the limit of detection was as low as10-11M. Multiplexed detection was further successfully performed with simultaneous spectral identification of up to three different kinds of DNA targets in an assay in light of the unique spectroscopic fingerprints of each type of SERS-tags. So the as-developed strategy holds significant promise for specific detection of biomolecules with high sensitivity and exquisite multiplexing capability.