Astigmatism imaging is widely used to encode the 3D position of fluorophore in single-particle tracking and super-resolution localization microscopy. Here, we present a fast and precise localization algorithm based on gradient fitting to decode the 3D subpixel position of the fluorophore. This algorithm determines the center of the emitter by finding the position with the best-fit gradient direction distribution to the measured point spread function (PSF), and can retrieve the 3D subpixel position of the emitter in a single iteration. Through numerical simulation and experiments with mammalian cells, we demonstrate that our algorithm yields comparable localization precision to the traditional iterative Gaussian function fitting (GF) based method, while exhibits over two orders-of-magnitude faster execution speed. Our algorithm is a promising online reconstruction method for 3D super-resolution microscopy.
Hongqiang Ma, Jianquan Xu, Jingyi Jin, Ying Gao, Li Lan, and Yang Liu, "Fast and precise 3D fluorophore localization by gradient fitting," Proc. SPIE 9714, Single Molecule Spectroscopy and Superresolution Imaging IX, 97140G (Presented at SPIE BiOS: February 14, 2016; Published: 1 March 2016); https://doi.org/10.1117/12.2211731.
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