An Immunochromatographic Assay Based On Surface-enhanced Raman Scattering(SERS) Using Noble Metal Nanoparticles

Surface-enhanced Raman scattering(SERS) has been widely used in biological studies owing to its nondestructive to samples and high sensitivity; while immunochromatographic assay(ICA) is widely used in the on-site detection in food safety and toxicant because its user-friendly operation, low cost and fairly good shelf life. In this study, we combined SERS with ICA and developed a novel analytical method named SERS-ICA. The thesis includes three parts as follows:1. A novel ultrasensitive competitive immunochromatographic assay(ICA) based on surface-enhanced Raman scattering(SERS) for direct detection of 3- Amino-5- methylmorpholino-2-oxazolidinone(AMOZ, the metabolite of a nitrofuran) in tissue and urine sampleswas was developed in which the monoclonal antibody(m Ab) against AMOZ and the Raman reporter [4-mercapto-benzoic acid(MBA)] dual labelled colloidal GNPs(e.g. Ab-Au-MBA) was prepared and used as an immune-probe. The assay was completed within 15 min. After ICA procedures, the specific Raman scattering intensity of MBA on the test line was measured for quantitative detection of AMOZ. The values of IC50 and limit of detection(LOD) of the assay for AMOZ were 0.04 ng m L-1and 0.28 pg m L-1, respectively. There was no cross-reactivity(CR) of the assay with other three nitrofurans and five compounds. The tissue and urine samples were spiked with AMOZ and the recoveries of AMOZ were in range of 87.8–105.9% with the relative standard deviations(RSD) of 1.4–9.0%(n = 3). The results demonstrated that the proposed SERS-based ICA was able to directly detect AMOZ with high sensitivity, specificity, stability, precision and accuracy.2. To detect phenylethanolamine A(PA, a new emerging β-agonist) by the proposed SERS-ICA, we successfully prepared AuMBA@Ag-Ab [e.g. the polyclonal antibody of PA labeled Au@Ag core-shell nanoparticles(NPs) sandwiched with a Raman reporter(4-mercaptobenzoic acid, MBA)] and used it as a novel immune-probe due to the higher SERS intensity of MBA on bimetallic core/shell compared to that of Au nanoparticles. The assay was completed within 15 min. The IC50 and limit of detection(LOD) values of the ICA for PA detection were 0.06 ng m L-1 and 0.32 pg m L-1, respectively, which were 1-3 orders of magnitude lower than those obtained by other immunoassays, indicating the ultrasensitivity of this ICA. There was no cross-reactivity(CR) of the assay with another three β-agonists(ractopamine, clenbuterol and salbutamol), suggesting high specificity of the SERS-based ICA. A spiking experiment revealed that the recoveries of PA from pig urine samples were in range of 99.9-101.2% with relative standard deviations(RSDs) of 3.6-5.8%. The results demonstrated that this SERS-based ICA was able to quantitatively detect PA in urine samples with high sensitivity, specificity, precision, and accuracy and might be a powerful method for the analysis of other target analytes in the food area.3. After the study of SERS-ICA based on above two nanoparticles(e.g. Au nanoparticles and Au@Ag core/shell nanoparticles), other novel nanoparticles which might show strong Raman signal enhanced effect were investigated. We prepared two kinds of nonspherical nanoparticles, e.g. Au nanorods and the nanocubes with camber concave surface, and explored the preparation conditions and SERS property of these nanoparticles in the use of SERS-ICA.