Studies And Applications Of Surface-enhanced Raman Spectroscopy Based Immunochromatographic Assay(SERS-ICA)

Because the million times of scattering signal enhancement by rough noble metal surface, Surface-enhanced Raman spectroscopy (SERS) can provide a nondestructive and ultrasensitive detection down to the possibility of single molecule level and has been one of the most sensitive analytical methods so far. SERS based Immunoassay (SERSIA) has attracted increasing attentions because of its high sensitivity arose from SERS and high selectivity based on the specific reaction between antibody and antigen. However, the application of SERSIA is largely limited by their time-consuming and tedious procedure, low reproducibility as well as the high cost. These problems include Ⅰ:The preparation of the substrates and the performance of the SERSIA are complicated because multiple processes such as blocking and washing are involved. Ⅱ:Due to non-uniform distribution of "hot spots" on the substrate, it is very difficult to get the reproducible SERS signal. Ⅲ: The targets were limited for macromolecules like protein or bacteria and cells, few works has been done for small molecules. Ⅳ:Purchase and maintenance of the instrucments are costly and substrates like gold film are very expensive. The shortcomings mentioned above severely hinder the normal SERS based immunoassays to be as a feasible methodology.In this study, we have investigated some kinds of substrate materials and proposed a rapid, low cost, simple and ultra-sensitive analytical method which integrates SERS with immunochromatographic assay (ICA) for the detection of two analytes based on NC membrane. The study was focused on the following aspects:①Magnetic bead@Au nanoparticle, noble metal SERS substrate, concave glass slides and NC membrace were selected as SERSIA substrate materials and combined with Raman dye labeled immuno-gold nanoprobes for the detection of CL. Among of them, Magnetic bead@Au nanoparticles were hard to prepared, the pretreatment of concave glass slides was tedious, and both of them were hard to achieve reproducible SERS signals; gold substrate was very expensive; whereas NC membrane was simple and convenient, the Raman signals were well distributed because of the labeled immuno-Gold nanoprobes were highly concentrated in the test line (T line) area which was only150μm width.②Combianing SERS with ICA, we established a new kind of immunoassay called SERS-ICA and successfully applied to detecte clenbuterol using portable Raman analyzer. The IC50was0.01ng mL-1, the LOD was0.06pg ml/-1, the results of specificity and reproducibility indicated that the proposed SERS-ICA would be potential as a powerfully rapid, simple and ultra-sensitive analytical method for the determination of various target analytes.③To test the proposed SERS-ICA for the detection of macromolecular compound, we established a sandwich SERS-ICA for carcinoembryonic antigen (CEA). The linear range of determination was from3to150ng mL-1, and the limit of detection (LOD) was50times lower than that achieved by traditional ICA. The results of specificity has also been discussed.