Gold nanoparticles (GNPs) are well-documented for their size and surface chemistry-dependent electronic and optical properties that are extensively utilized to develop highly sensitive immunoassays. GNP-based immuno-polymerase chain reaction (immuno-PCR) is especially interesting due to the facile loading of biomolecules on the surface of GNP probes and has been utilized to develop analyte-specific assays. In this study, the role of size and surface chemistry of GNPs is explored in detail to develop a highly sensitive and reproducible immuno-PCR assay for specific detection of biotinylated analytes. Our results indicate that smaller-sized gold nanoparticles outperform the larger ones in terms of their sensitivity in immuno-PCR assay and show superior loading of proteins and oligonucleotides on the surface of nanoparticles. Furthermore, the role of different macromolecular stabilizers (such as polyethylene glycol (PEG), bovine serum albumin (BSA), and PEGylated BSA) was compared to optimize the loading of biomolecules and to improve the signal-to-noise ratio of GNP probes. mPEG-BSA-functionalized GNP probes of 15 nm were found to be highly sensitive at low concentrations of analytes and significantly (~ 30 fold) improve the limit of detection of analytes in comparison with ELISA assay.
Keywords: GNP-based immuno-PCR; High sensitivity; Macromolecule stabilizers; Reproducibility; Size-dependent loading of biomolecules.
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