Stimulated Raman Scattering Microspectral Imaging Noise And Signal Processing

Raman spectrum is the change of the wavelength（frequency）of scattered light caused by the energy exchange between light and the affected molecule in the process of inelastic collision.The detection technology based on Raman spectroscopy has the advantages of accuracy,non-destructive,no marking,simple sample preparation and low interference of water.The combination of Raman spectrum and microscopic imaging can obtain the Raman spectrum of specific band while obtaining the cell microscopic image,and deeply analyze the cell life activities in the way of"spectrum combination".Because the spontaneous Raman signal is very weak,it is often limited in the applications that need to obtain the spectrum quickly.Stimulated Raman scattering（SRS）is produced by the coupling of the photoelectric field of strong laser with the excitation of electrons in atoms,the vibration in molecules or the lattice in crystals.It has strong stimulated characteristics,and its signal is at least 4 orders of magnitude higher than that of spontaneous Raman scattering.Greatly improve the acquisition speed of Raman spectrum information,and realize the spectral scanning imaging of video rate for biological tissues such as cells.SRS technology can obtain the Raman spectrum of biochemical substances in the corresponding region while obtaining the cell image,so as to realize the analysis of sample structure and composition.On the basis of summarizing the development status of stimulated Raman and other related spectroscopy technologies,the main research carried out in this paper is as follows:Assisted in the construction of SRS spectral imaging system,and compiled spectral acquisition and scanning imaging software;The influence of modulation depth of acousto optic modulator on SRS signal strength and signal-to-noise ratio and its mechanism are studied.In the process of SRS spectral imaging,when the residence time of each pixel is fixed,the influence of the time constant and order of phase-locked amplifier on the signal-to-noise ratio of SRS image is studied,and the setting method of relevant parameters is given.Using this system,the SRS spectra of dimethyl sulfoxide and the hyperspectral SRS spectra of the mixed solution of PMMA and PS spheres were obtained.SRS images of lipids in a single leukemia cell were obtained under the condition of optimal signal-to-noise ratio.The relevant research results prove to some extent that the software and hardware system can obtain a spectral resolution close to 26cm^-1 and a spatial resolution close to245nm in the wavenumber range of 2800-3100 cm^-1.The work of this paper lays a foundation for the next research on single-cell SRS spectral imaging and label free detection of cancer cells.