Light-sheet microscopy with isotropic, sub-micron resolution and solvent-independent large-scale imaging

Chakraborty, Tonmoy; Driscoll, Meghan; Murphy, Malea M.; Roudot, Philippe; Chang, Bo-Jui; Vora, Saumya; Wong, Wen Mai; Nielson, Cara D.; Zhang, Hua; Zhemkov, Vladimir; Hiremath, Chitkale; Cruz, Estanislao Daniel De La; Bezprozvanny, Ilya; Zhao, Hu; Tomer, Raju; Heintzmann, Rainer; Meeks, Julian P.; Marciano, Denise K.; Morrison, Sean J.; Danuser, Gaudenz; Dean, Kevin M.; Fiolka, Reto

doi:10.1101/605493

Cited by 6 publications

(4 citation statements)

References 39 publications

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“…Furthermore, we demonstrate a method for dual-view registration and deconvolution of images after remapping into an inverted RGB colorspace to simulate the appearance of brightfield H&E images. It should be mentioned that isotropic resolution can also be achieved by axially swept light sheet microscopy (ASLM) without the need of deconvolution [37,38]. ASLM leverages axial sweeping of a scanned light sheet, with only signals excited by the diffraction-limited beam waist area being collected via an electronic confocal slit detection scheme, and with the axial resolution being mainly dependent on how thin the illumination beam waist is.…”

Section: Discussionmentioning

confidence: 99%

Enhanced resolution 3D digital cytology and pathology with dual-view inverted selective plane illumination microscopy

Brown

2019

Biomed. Opt. Express

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The current gold-standard histopathology for tissue analysis is destructive, time consuming, and limited to 2D slices. Light sheet microscopy has emerged as a powerful tool for 3D imaging of tissue biospecimens with its fast speed and low photo-damage, but usually with worse axial resolution and complicated configuration for sample imaging. Here, we utilized inverted selective plane illumination microscopy for easy sample mounting and imaging, and dual-view imaging and deconvolution to overcome the anisotropic resolution. We have rendered 3D images of fresh cytology cell blocks and millimeter-to centimetersized fixed tissue samples with high resolution in both lateral and axial directions. More accurate cellular quantification, higher image sharpness, and more image details have been achieved with the dual-view method compared with single-view imaging.

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Section: Discussionmentioning

confidence: 99%

Enhanced resolution 3D digital cytology and pathology with dual-view inverted selective plane illumination microscopy

Brown

2019

Biomed. Opt. Express

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“…Furthermore, several post-acquisition image processing techniques have been made accessible to microscopists in open source software platforms such as MATLAB and Im-ageJ (also known as Fiji). Image deblurring is performed via deconvolution using an optical system point spread function (PSF), obtained from imaging diffraction-limited beads embedded in agar-or resin-based embedding media [85,91]. Isotropic image contrast across an attenuated FOV is restored using intensity thresholding techniques such as histogram equalization or nonlinear haze removal [60].…”

Section: Light Sheet Modality Considerationsmentioning

confidence: 99%

Development of Planar Illumination Strategies for Solving Mysteries in the Sub-Cellular Realm

Teranikar

Lim

Ijaseun

et al. 2022

IJMS

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Optical microscopy has vastly expanded the frontiers of structural and functional biology, due to the non-invasive probing of dynamic volumes in vivo. However, traditional widefield microscopy illuminating the entire field of view (FOV) is adversely affected by out-of-focus light scatter. Consequently, standard upright or inverted microscopes are inept in sampling diffraction-limited volumes smaller than the optical system’s point spread function (PSF). Over the last few decades, several planar and structured (sinusoidal) illumination modalities have offered unprecedented access to sub-cellular organelles and 4D (3D + time) image acquisition. Furthermore, these optical sectioning systems remain unaffected by the size of biological samples, providing high signal-to-noise (SNR) ratios for objective lenses (OLs) with long working distances (WDs). This review aims to guide biologists regarding planar illumination strategies, capable of harnessing sub-micron spatial resolution with a millimeter depth of penetration.

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“…Among these methods, the SLM is the most practical beam shaper, because it offers a versatile application for beam shaping as well as beam aberration correction and beam aperture controlling simultaneously with the tiling technique [15]. All of these approaches were successfully applied to the basic configuration of the SPIM with two or three objective lenses [10,11,13,14,[16][17][18][19][20][21] to imaging a small transparent specimen or cleared fixed sample, but are not optimized for high resolution whole functional imaging of an opaque live sample. This analysis revealed the temporal sequence of dynamic MyoII-GFP localization in distinct morphogenetic movements during gastrulation in the Drosophila embryo.…”

Section: Introductionmentioning

confidence: 99%

Multiview tiling light sheet microscopy for 3D high resolution live imaging

Aakhte

Mueller

2021

Preprint

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Light sheet or selective plane illumination microscopy (SPIM) is ideally suited for in toto imaging of living specimens at high temporal-spatial resolution. In SPIM, the light scattering that occurs during imaging of opaque specimens brings about limitations in terms of resolution and the imaging field of view. To ameliorate this shortcoming, the illumination beam can be engineered into a highly confined light sheet over a large field of view and multi-view imaging can be performed by applying multiple lenses combined with mechanical rotation of the sample. Here, we present a Multiview tiling SPIM (MT-SPIM) that combines the Multi-view SPIM (M-SPIM) with a confined, multi-tiled light sheet. The MT-SPIM provides high-resolution, robust and rotation-free imaging of living specimens. We applied the MT-SPIM to image nuclei and Myosin II from the cellular to subcellular spatial scale in early Drosophila embryogenesis. We show that the MT-SPIM improves the axial-resolution relative to the conventional M-SPIM by a factor of two. We further demonstrate that this axial resolution enhancement improves the automated segmentation of Myosin II distribution and of nuclear volumes and shapes.

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Light-sheet microscopy with isotropic, sub-micron resolution and solvent-independent large-scale imaging

Cited by 6 publications

References 39 publications

Enhanced resolution 3D digital cytology and pathology with dual-view inverted selective plane illumination microscopy

Enhanced resolution 3D digital cytology and pathology with dual-view inverted selective plane illumination microscopy

Development of Planar Illumination Strategies for Solving Mysteries in the Sub-Cellular Realm

Multiview tiling light sheet microscopy for 3D high resolution live imaging

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