SERS-Based Multi Protein Detection Using Gold-Patterned Microarray Chip

Abstract

Immunoassay is a simple and steadfast method to quantify the unknown sample using antibody-antigen interaction. It has been widely used for biomedical diagnosis, environmental monitoring, and so forth. For example, enzyme linked immunosorbent assay (ELISA) method is commonly used to identify and quantify the target molecule due to its specificity and simplicity. But broad spectral bandwidth and high limit of detection hindered the sensitivity, reproducibility and multiplicity. Recently, surface enhanced Raman scattering (SERS)-based immunoassay platform has been draw great attention in research field. Its high degree of detection sensitivity and potential for simultaneous multi-analyte detection makes it better than conventional ELISA, because of its 100 times narrower spectrum bandwidth and up to 1014 times enhancement of the signal. This study describes the modifications made by SERS method in the field of immunoassay technology, which was completely ruled by ELISA in the previous decades. Here we report a conceptually new optical detection method to simultaneously quantify multiple biomarkers on a single surface. For this purpose we used a SERS-based detection technique with a gold-patterned microarray chip. The gold-patterned microarray chip was designed to use it as an immunoassay template together with multiple Raman probes. The gold surface was modified with 11-mercaptoundecanoic acid by self-assembled monolayer (SAM) method. In addition, hollow gold nanospheres (HGNs) were synthesized and used as highly sensitive and reproducible SERS probe for multiplex biomarker detection. To examine correlation between antibodies, sandwich ELISA was performed against two different origin sets of IgGs, and results were compared with SERS measurements. For SERS detection, human and rabbit monoclonal antibodies (McAb) were immobilized on the gold surface and different concentrations of multi-antigens were captured simultaneously. Then different Raman dye-embedded HGNs, conjugated with human and rabbit polyclonal antibodies (PcAb) were binded with specific antigens by antigen-antibody interaction, and sandwich type immunocomplexes were formed on the gold-patterned microarray wells and different intensity signals were generated because of the different concentrations of antigens. The concentration of biomarkers was evaluated by using the SERS method. Here, selected animal origin of antibodies prevents the cross reactivity and nonspecific interactions. When the antibody immobilized on SERS nanoprobes and antigens are completely match with each other, specific Raman bands will appear. This study fulfills the current needs of sensitivity and quantitation in simultaneous multi-biomarker assay. This SERS-based immunoassay technique is expected to be an effective bio-detection tool which can be applied to a fast and sensitive sensing of multiple biomarkers.