Single shot, three-dimensional fluorescence microscopy with a spatially rotating point spread function

Author(s):

Zhaojun Wang and Yanan Cai and Yansheng Liang and Xing Zhou and Shaohui Yan and Dan Dan and Piero R. Bianco and Ming Lei and Baoli Yao

Abstract:

“A wide-field fluorescence microscope with a double-helix point spread function (PSF) is constructed to obtain the specimen’s three-dimensional distribution with a single snapshot. Spiral-phase-based computer-generated holograms (CGHs) are adopted to make the depth-of-field of the microscope adjustable. The impact of system aberrations on the double-helix PSF at high numerical aperture is analyzed to reveal the necessity of the aberration correction. A modified cepstrum-based reconstruction scheme is promoted in accordance with properties of the new double-helix PSF. The extended depth-of-field images and the corresponding depth maps for both a simulated sample and a tilted section slice of bovine pulmonary artery endothelial (BPAE) cells are recovered, respectively, verifying that the depth-of-field is properly extended and the depth of the specimen can be estimated at a precision of 23.4nm. This three-dimensional fluorescence microscope with a framerate-rank time resolution is suitable for studying the fast developing process of thin and sparsely distributed micron-scale cells in extended depth-of-field.”

Link to Publications Page

Publication: Biomedical Optics Express

Issue/Year/DOI: Biomedical Optics Express, Vol. 8, Issue 12, (2017)
DOI: 10.1364/BOE.8.005493

Fast label-free microscopy technique for 3D dynamic quantitative imaging of living cells

Author(s):

José A. Rodrigo, Juan M. Soto, and Tatiana Alieva

Abstract:

“The refractive index (RI) is an important optical characteristic that is often exploited in label-free microscopy for analysis of biological objects. A technique for 3D RI reconstruction of living cells has to be fast enough to capture the cell dynamics and preferably needs to be compatible with standard wide-field microscopes. To solve this challenging problem, we present a technique that provides fast measurement and processing of data required for real-time 3D visualization of the object RI. Specifically, the 3D RI is reconstructed from the measurement of bright-field intensity images, axially scanned by a high-speed focus tunable lens mounted in front of a sCMOS camera, by using a direct deconvolution approach designed for partially coherent light microscopy in the non-paraxial regime. Both the measurement system and the partially coherent illumination, that provides optical sectioning and speckle-noise suppression, enable compatibility with wide-field microscopes resulting in a competitive and affordable alternative to the current holographic laser microscopes. Our experimental demonstrations show video-rate 3D RI visualization of living bacteria both freely swimming and optically manipulated by using freestyle laser traps allowing for their trapping and transport along 3D trajectories. These results prove that is possible to conduct simultaneous 4D label-free quantitative imaging and optical manipulation of living cells, which is promising for the study of the cell biophysics and biology.”

Link to Publications Page

Publication: Optics Express

Issue/Year/DOI: Optics Express Volume 8, Issue 12
DOI: 10.1364/BOE.8.005507