Time-Gated Raman Micro-Spectroscopy for Rejection of Fluorescence Background in Biological Specimens.

摘要

For the purpose of my masters thesis I investigate different methods to retrieve Raman spectra from overwhelming fluorescence background in chemical and biological samples. Raman spectroscopy has been a focus of intense research as a tool for addressing biomedical problems for the past several years. For instance, Raman spectra have been shown to provide diagnostic value for discriminating between cancerous and non-cancerous cells and tissues. It has also been used to classify and quantify bacteria and bacterial cultures and to assess bone health. However, Raman scattering has a small cross section and while most biological samples have significant fluorescence background signals. Although several methods have been proposed to eliminate the fluorescence background, none of these methods address the more fundamental problem of signal-to-noise that arises when trying to detect a small signal on a much larger background. An ultra-fast switch can temporally separate the Raman signal from the slower evolving fluorescence background when pulsed excitation light sources are used. If the laser pulse duration is much shorter than the fluorescence lifetime, the instantaneous occurring Raman scattered light can be temporally separated. Ultra-fast switches operating in the picosecond time scale are the most common all-optical, meaning that a gate is opened and closed by a light-matter interaction. However, the need for high power laser systems drive these gates remains unsolved. The resultant high light intensities on the sample have been a major problem that has been addressed by the methods described here. In this work, several optical experiments were set up to perform measurements involving a Sagnac interferometer as well as a polarization maintaining photonic crystal fiber based optical Kerr switch. Furthermore, a collinear Kerr gate with carbon disulfide as a nonlinear material was developed and tested. This setup operates with 3 orders-of-magnitude less power than previously reported systems and by careful paying attention to the power conservation, all-optical switching with a one picosecond gating time and 5% peak gating efficiency was achieved. Using this system, measurements on a sample composed of perylene dye dissolved in toluene and the stem of a Jasminum multiflorum were performed. In both cases, a high signal-to-noise spectrum of the high-wavenumber region of the spectrum was recorded in the presence of an overwhelming fluorescence background.