Surface-Enhanced\nRaman Spectroscopy-Based, Homogeneous,\nMultiplexed Immunoassay with Antibody-Fragments-Decorated Gold Nanoparticles

Abstract

We report the development of a novel\nsingle-step, multiplexed,\nhomogeneous immunoassay platform for sensitive detection of protein\ntargets based on our realization of high surface-enhanced raman spectroscopy\n(SERS) signal enhancement by controlled assembly of SERS nanoparticles.\nAn essential design of this platform is the use of gold nanoparticles\nor nanorods codecorated with specially reduced antibody half-fragments,\nnonfluorescent Raman-active dyes, and passivating proteins as the\nSERS nanoparticles. These nanoparticles offer a facile approach to\naccomplish orientational immobilization of antibodies, minimized interparticle\ndistance, multicolor Raman fingerprint coding, low fluorescence background,\nas well as excellent biocompatibility and stability. Through sandwiched\nantibody–antigen interactions, controlled assembly of SERS\nnanoparticles is realized with a strong SERS signal achieved via plasmonic\ncoupling, creating an immunoassay platform for rapid, sensitive, multiplexed\nquantification of proteins. This platform is demonstrated for reproducible\nquantification of three cytokines, interferon gamma, interleukin-2,\nand tumor necrosis factor alpha, with large signal-to-noise ratio.\nIt is also successfully applied to multiplexed cytokine analysis for\nT cell secretion studies in complicated biological samples. The developed\nSERS immunoassay platform may create a simple but valuable tool for\nfacilitating accurate validation and early detection of disease biomarkers\nas well as for point-of-care tests in clinical diagnostics.