ABSTRACT
Two double-cysteine mutants of a small protein judiciously modified so that the cysteines appear at axially opposite sides of the native fold were prepared such that different axes were defined in the two mutants. Upon reduction, the disulfide bonds are broken, and the proteins act as bifunctional ligands toward Ag nanoparticles, encouraging their assembly into nanoparticle dimers and small aggregates such that, when excited with laser light, the proteins are automatically located at electromagnetic hot spots within the aggregates. Because the protein molecules are small (~2.3 nm) and because the electromagnetic energy at a hot spot tends to increase as the size of the interparticle gap decreases, this nanoparticle-protein-nanoparticle geometry significantly enhances the Raman emission at the metallic surface. Exploiting this effect, we have recorded surface-enhanced Raman spectra (SERS) of the proteins at near-single-molecule level. The observed SERS spectra were dominated by the vibrations of molecular groups near the anchor points of the proteins.
ABSTRACT
We report a simple strategy for placing analyte molecules in hot spots between closely spaced nanowires leading to intense SERS enhancement. The results are highly reproducible from experiment to experiment likely because of the regularity of the SERS substrate, which consists of highly ordered and regular silver nanowires fabricated in porous aluminum oxide. Because the silver nanowires are sealed in the pores of PAO, this system is potentially immune to contamination until it is ready for use, at which point the alumina matrix is etched, thereby allowing the silver nanowires to collapse into bundles and form hot spots in the region of close contact between the nanowires, trapping the analyte in those junctions.
