Raman and Fluorescence Spectroscopy of Single Molecules and Metal Particles

Doctoral thesis, 2002

Technological and conceptual advances have led to the realization of single-molecule spectroscopy. This permit detailed investigations of heterogeneous systems in fields ranging from condensed matter physics to biochemistry. It was recently discovered that surface-enhanced Raman scattering (SERS) can be used to study single molecules. A part of this thesis deals with single-molecule SERS measurements of heme-proteins adsorbed on small Ag-particle aggregates. The SERS signal exhibits temporal and spectral fluctuations, a phenomenon that appears to be characteristic of single-molecule optical spectroscopy at room temperature. The enormous SERS sensitivity can be understood in the framework of electromagnetic interparticle coupling. Various metal particles were explored as SERS substrates. Electron beam lithography was used to make substrates consisting of particles with different shapes, sizes, and inter-particle distances. Given the precise control of fabrication, these substrates are suitable for quantitative analysis. For many sensor applications, however, cheap and fast metal deposition in an aqueous environment is desirable. A simple and robust synthesis route of noble metal nanoparticles for optical sensing based on laser-controlled growth and deposition is demonstrated. We further examined the irreversible and highly heterogeneous photochemical transformation of aromatic precursors to amorphous carbon that occurs on aggregated and immobilized Ag nanocrystals, using SERS. The results highlight the importance of surface-enhanced photochemistry in experiments which rely on high electromagnetic fields near metal nanoparticles. We also observed photo-induced effects on hemoglobin in red blood cells. The implementation of a through-the-objective total internal reflection (TIR) microscope is described. This type of microscopy permits single-molecule fluorescence imaging because of the low fluorescence background and the high detection efficiency inherent to the technique. The fluorescence emission from single dye-molecules, immobilized in polymer films, was measured and the sensitivity of the TIR set-up was compared to the epi-illumination geometry.