Study On Super Resolution Microscopy

With the rapid development of life sciences,human have studied into single cells,subcellular and molecular levels.However,the resolution of conventional optical microscope has been limited by diffraction limit.Light diffractions duing to the fluctuation occurs.So the beam can not focus to a point but an Airy spot.The size of Airy spot should be approximately half the wavelength of visible light.200 nm is the theoretical resolution limit of conventional optical microscopy.Now improving spatial resolution of optical microscopes have become a key issues in the microscopy field.In the recent years,with the development of modern measurement technology and physicists of the technological innovations,far-field optical microscopes have made revolutionary progress and break the diffraction limit to the nanometer scale.They can study the internal structure of cells directly on the single molecule level.Stochastic optical reconstruction microscopy(STORM)and Stimulated Emission Depletion microscopy(STED)are two kinds of super resolution microscopy with their own unique characteristics.The main contents of this paper includes the following sections:1.We study the original data obtained by stochastic optical reconstruction microscopy system,comparing the current location algorithm of simple molecules,designing a 3D single PSF algorithm based on Gaussian fitting.Images processing was simulated by MATLAB.Then we optimiz the original CPU serial processing to CPU parallel processing,improving imaging speed by four times,and improving the temporal resolution of STORM.2.We designed a 2PE-STED microscopy system,including synchronization system and modulation systems of excite and STED illuminating sourse.We introduced a bragg cell in the system and declined the illumination three orders of magnitude for the sample with low input power,simple operation,low cost and without pulse-synchronous,and improve fluorescent yeild 15-25 times.We apply the system for the observation of ATTO 452 marked on Chinese hamster ovary(CHO)cells of clathrin.Then we got a promissing result.