Fluorescence Emission Difference Microscopy And System

Far-field super-resolution fluorescence microscopy has been widely recognized as an efficient method for biological and medical observations and investigations, because of its high resolution, real-time observation, minimal perturbation, and convenient operation. On the basis of Fluorescence Emission Difference (FED) theory, we proposed several effective methods to further improve the performance of the existing FED, including increasing the imaging speed--this project built up a high-speed FED system which has been demonstrated to achieve a resolution of 150nm that is beyond the diffraction barrier, under a real-time imaging speed of lfps; improving the imaging quality---this project simulated the deterioration which negative sidelobes brought to FED images and proposed a solution by extending the point spread function (PSF) of the solid spot; enhancing the spatial resolution---this project built up the first FED system based on cylindrical vector beams in China and achieved a spatial resolution of 110nm which is beyond the diffraction barrier; realizing isotropic imaging FED---this project analyzed and simulated the isotropic three-dimensional hollow spot and built up a spatial light modulator FED system.In this thesis, I first briefly introduced some popular super-resolution techniques, especially the proposal and development of FED and its current research situation. Then the main concept of FED, related techniques and implementation approaches were elaborated. After that a high-speed FED system was built and its performance was then testified through imaging kinds of samples. Aside of imaging speed, the next few chapters introduced several efficient ways to further enhance the function of FED system including eliminating the deformation by matching the PSFs of solid spots and hollow spots with beam modulation, enhancing the resolution and image quality by extending the solid spot and shrinking the hollow spot with cylindrical vector beams, simplifying the system and realizing isotropic imaging with SLM. Finally I concluded the work in this project and pointed out the possible directions of future research.