3D localization of high particle density images using sparse recovery

Abstract

If particles are too close in space, their images may be overlapped when they are observed with microscopes because of diffraction limitation, which makes them difficult to be distinguished or localized. This limitation also affects the efficiency of localization of those single-particle-localization microcopies, such as stochastic optical reconstruction microscopy (STORM) and (fluorescence) photoactivated localization microscopy [(F)PALM]. In this work, we developed a 3D sparse recovery (3D-SR) method, with the aim of localizing particles with high density in three dimensions, which cannot be resolved using original STROM or (F)PALM. A cylindrical lens was introduced to a traditional wide-field microscope in order to form the 3D point spread function for 3D-SR. The performance of the 3D-SR method was evaluated using simulation. Simulated results demonstrated that, even for particle densities as high as ${4\mathrm{\mu m}}^{-2}$ on a transversal projection, particles could still be localized with high accuracy. The standard deviations were found to be 25.59 nm along the transverse direction and 50.42 nm along the axial direction. Compared with the existing 3D localization methods used in high particle density cases, such as 3D-DAOSTORM, 3D-SR allows a higher activated fluorophore density per frame.

© 2015 Optical Society of America

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