Optical sectioning microscopy can provide highly detailed three dimensional (3D) images of biological samples. However, it requires acquisition of many images per volume, and is therefore time consuming, and may not be suitable for live cell 3D imaging. We propose the use of the modified Gerchberg-Saxton phase retrieval algorithm to enable full 3D imaging of gold nanoparticles tagged sample using only two images. The reconstructed field is free space propagated to all other focus planes using post processing, and the 2D z-stack is merged to create a 3D image of the sample with high fidelity. Because we propose to apply the phase retrieving on nano particles, the regular ambiguities typical to the Gerchberg-Saxton algorithm, are eliminated. The proposed concept is then further enhanced also for tracking of single fluorescent particles within a three dimensional (3D) cellular environment based on image processing algorithms that can significantly increases localization accuracy of the 3D point spread function in respect to regular Gaussian fitting. All proposed concepts are validated both on simulated data as well as experimentally.
Tali Ilovitsh, Asaf Ilovitsh, Aryeh M. Weiss, Rinat Meir, and Zeev Zalevsky, "Phase retrieval and 3D imaging in gold nanoparticles based fluorescence microscopy (Conference Presentation)," Proc. SPIE 10074, Quantitative Phase Imaging III, 1007406 (Presented at SPIE BiOS: January 29, 2017; Published: 24 April 2017); https://doi.org/10.1117/12.2251587.5380600157001.
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