scholarly journals Sub-voxel light-sheet microscopy for high-resolution, high-throughput volumetric imaging of large biomedical specimens

2018 ◽  
Author(s):  
Peng Fei ◽  
Jun Nie ◽  
Juhyun Lee ◽  
Yichen Ding ◽  
Shuoran Li ◽  
...  
Keyword(s):  
High Resolution ◽  
High Throughput ◽  
Light Sheet ◽  
Large Field ◽  
Mouse Heart ◽  
Processing Unit ◽  
Intact Mouse ◽  
Volumetric Imaging ◽  
Light Sheet Microscopy ◽  
Wide Range

A key challenge when imaging whole biomedical specimens is how to quickly obtain massive cellular information over a large field of view (FOV). Here, we report a sub-voxel light-sheet microscopy (SLSM) method enabling high-throughput volumetric imaging of mesoscale specimens at cellular-resolution. A non-axial, continuous scanning strategy is used to rapidly acquire a stack of large-FOV images with three-dimensional (3-D) nanoscale shifts encoded. Then by adopting a sub-voxel-resolving procedure, the SLSM method models these low-resolution, cross-correlated images in the spatial domain and iteratively recovers a 3-D image with improved resolution throughout the sample. This technique can surpass the optical limit of a conventional light-sheet microscope by more than three times, with high acquisition speeds of gigavoxels per minute. As demonstrated by quick reconstruction (minutes to hours) of various samples, e.g., 3-D cultured cells, an intact mouse heart, mouse brain, and live zebrafish embryo, the SLSM method presents a high-throughput way to circumvent the tradeoff between intoto mapping of large-scale tissue (>100 mm3) and isotropic imaging of single-cell (~1-μm resolution). It also eliminates the need of complicated mechanical stitching or precisely modulated illumination, using a simple light-sheet setup and fast graphics-processing-unit (GPU)-based computation to achieve high-throughput, high-resolution 3-D microscopy, which could be tailored for a wide range of biomedical applications in pathology, histology, neuroscience, etc.

scholarly journals Subvoxel light-sheet microscopy for high-resolution high-throughput volumetric imaging of large biomedical specimens

2019 ◽  
Vol 1 (01) ◽  
pp. 1 ◽  
Author(s):  
Peng Fei ◽  
Jun Nie ◽  
Juhyun Lee ◽  
Yichen Ding ◽  
Shuoran Li ◽  
...  
Keyword(s):  
High Resolution ◽  
High Throughput ◽  
Light Sheet ◽  
Volumetric Imaging ◽  
Light Sheet Microscopy

scholarly journals A hybrid open-top light-sheet microscope for multi-scale imaging of cleared tissues

2021 ◽  
Author(s):  
Adam Glaser ◽  
Kevin Bishop ◽  
Lindsey Barner ◽  
Etsuo Susaki ◽  
Shimpei Kubota ◽  
...  
Keyword(s):  
Hybrid System ◽  
High Throughput ◽  
Light Sheet ◽  
Refractive Indices ◽  
Volumetric Imaging ◽  
Tissue Clearing ◽  
Sample Geometry ◽  
Multi Scale ◽  
Light Sheet Microscopy ◽  
User Friendly

Abstract Light-sheet microscopy has emerged as the preferred means for high-throughput volumetric imaging of cleared tissues. However, there is a need for a user-friendly system that can address imaging applications with varied requirements in terms of resolution (mesoscopic to sub-micrometer), sample geometry (size, shape, and number), and compatibility with tissue-clearing protocols and sample holders of various refractive indices. We present a ‘hybrid’ system that combines a novel non-orthogonal dual-objective and conventional (orthogonal) open-top light-sheet architecture for versatile multi-scale volumetric imaging.

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

Development ◽  
2021 ◽  
Author(s):  
Mostafa Aakhte ◽  
H.-Arno J. Müller
Keyword(s):  
High Resolution ◽  
Myosin Ii ◽  
Resolution Enhancement ◽  
Light Sheet ◽  
Field Of View ◽  
Large Field ◽  
Axial Resolution ◽  
Light Sheet Microscopy ◽  
Drosophila Embryogenesis ◽  
Mechanical Rotation

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.

scholarly journals High-Resolution, Large Field-of-View, and Multi-View Single Objective Light-Sheet Microscopy

2020 ◽  
Author(s):  
Bin Yang ◽  
Alfred Millett-Sikking ◽  
Merlin Lange ◽  
Ahmet Can Solak ◽  
Hirofumi Kobayashi ◽  
...  
Keyword(s):  
High Resolution ◽  
Light Sheet ◽  
Field Of View ◽  
Large Field ◽  
Sample Rotation ◽  
Light Sheet Microscopy ◽  
Spatio Temporal ◽  
Large Living ◽  
Single Objective ◽  
Imaging Speed

Light-sheet microscopy has become the preferred method for long-term imaging of large living samples because of its low photo-invasiveness and good optical sectioning capabilities. Unfortunately, refraction and scattering often pose obstacles to light-sheet propagation and limit imaging depth. This is typically addressed by imaging multiple complementary views to obtain high and uniform image quality throughout the sample. However, multi-view imaging often requires complex multi-objective configurations that complicate sample mounting, or sample rotation that decreases imaging speed. Recent developments in single-objective light-sheet microscopy have shown that it is possible to achieve high spatio-temporal resolution with a single objective for both illumination and detection. Here we describe a single-objective light-sheet microscope that achieves: (i) high-resolution and large field-of-view imaging via a custom remote focusing objective; (ii) simpler design and ergonomics by remote placement of coverslips; (iii) fast volumetric imaging by means of light-sheet stabilised stage scanning – a novel scanning modality that extends the imaging volume without compromising imaging speed nor quality; (iv) multi-view imaging by means of dual orthogonal light-sheet illumination. Finally, we demonstrate the speed, field of view and resolution of our novel instrument by imaging zebrafish tail development.

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

2021 ◽  
Author(s):  
Mostafa Aakhte ◽  
Hans-Arno J Mueller
Keyword(s):  
High Resolution ◽  
Myosin Ii ◽  
Resolution Enhancement ◽  
Light Sheet ◽  
Field Of View ◽  
Large Field ◽  
Axial Resolution ◽  
Light Sheet Microscopy ◽  
Drosophila Embryogenesis ◽  
Mechanical Rotation

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.

scholarly journals A hybrid open-top light-sheet microscope for multi-scale imaging of cleared tissues

2020 ◽  
Author(s):  
Adam K. Glaser ◽  
Kevin W. Bishop ◽  
Lindsey A. Barner ◽  
Robert B. Serafin ◽  
Jonathan T.C. Liu
Keyword(s):  
Hybrid System ◽  
High Throughput ◽  
Light Sheet ◽  
Refractive Indices ◽  
Volumetric Imaging ◽  
Tissue Clearing ◽  
Sample Geometry ◽  
Multi Scale ◽  
Light Sheet Microscopy ◽  
User Friendly

AbstractLight-sheet microscopy has emerged as the preferred means for high-throughput volumetric imaging of cleared tissues. However, there is a need for a user-friendly system that can address diverse imaging applications with varied requirements in terms of resolution (mesoscopic to sub-micron), sample geometry (size, shape, and number), and compatibility with tissue-clearing protocols of different refractive indices. We present a hybrid system that combines a novel non-orthogonal dual-objective and conventional open-top light-sheet architecture for highly versatile multi-scale volumetric imaging.One sentence summaryGlaser et al. describe a hybrid open-top light-sheet microscope to image cleared tissues at mesoscopic to sub-micron resolution and depths of up to 1 cm.

scholarly journals A Versatile Oblique Plane Microscope for Large-Scale and High-Resolution Imaging of Subcellular Dynamics

2020 ◽  
Author(s):  
Etai Sapoznik ◽  
Bo-Jui Chang ◽  
Jaewon Huh ◽  
Robert J. Ju ◽  
Evgenia V. Azarova ◽  
...  
Keyword(s):  
High Resolution ◽  
Numerical Aperture ◽  
Light Sheet ◽  
Membrane Dynamics ◽  
Field Of View ◽  
Large Field ◽  
Endoplasmic Reticulum Membrane ◽  
High Resolution Imaging ◽  
Light Sheet Microscopy ◽  
Resolution Imaging

AbstractWe present an Oblique Plane Microscope that uses a bespoke glass-tipped tertiary objective to improve the resolution, field of view, and usability over previous variants. Owing to its high numerical aperture optics, this microscope achieves lateral and axial resolutions that are comparable to the square illumination mode of Lattice Light-Sheet Microscopy, but in a user friendly and versatile format. Given this performance, we demonstrate high-resolution imaging of clathrin-mediated endocytosis, vimentin, the endoplasmic reticulum, membrane dynamics, and Natural Killer-mediated cytotoxicity. Furthermore, we image biological phenomena that would be otherwise challenging or impossible to perform in a traditional light-sheet microscope geometry, including cell migration through confined spaces within a microfluidic device, subcellular photoactivation of Rac1, diffusion of cytoplasmic rheological tracers at a volumetric rate of 14 Hz, and large field of view imaging of neurons, developing embryos, and centimeter-scale tissue sections.

scholarly journals Converting lateral scanning into axial focusing to speed up 3D microscopy

2020 ◽  
Author(s):  
Tonmoy Chakraborty ◽  
Bo-Jui Chang ◽  
Stephan Daetwyler ◽  
Etai Sapoznik ◽  
Bingying Chen ◽  
...  
Keyword(s):  
High Resolution ◽  
Spherical Aberration ◽  
Optical Design ◽  
Image Plane ◽  
Light Sheet ◽  
Volumetric Imaging ◽  
Light Sheet Microscopy ◽  
Scanning Rate ◽  
Order Of Magnitude ◽  
Axial Scanning

AbstractIn optical microscopy, the slow axial scanning rate of the objective or the sample has traditionally limited the speed of 3D volumetric imaging. Recently, by conjugating either a movable-mirror to the image plane or an electrotuneable lens (ETL) to the back-focal plane respectively, rapid axial scanning has been achieved. However, mechanical actuation of a mirror limits axial scanning rate (usually only 10-100 Hz for piezoelectric or voice coil based actuators), while ETLs introduce spherical and higher order aberrations, thereby preventing high-resolution imaging. Here, we introduce a novel optical design that can transform a lateral-scan motion into a spherical-aberration-free, high-resolution, rapid axial scan. Using a galvanometric mirror, we scan a laser beam laterally in a remote-focusing arm, which is then back-reflected from different heights of a mirror in image space. We characterize the optical performance of this remote focusing technique and use it to accelerate axially swept light-sheet microscopy (ASLM) by one order of magnitude, allowing the quantification of rapid vesicular dynamics in 3D.

scholarly journals Large-field high-resolution two-photon digital scanned light-sheet microscopy

Cell Research ◽  
2014 ◽  
Vol 25 (2) ◽  
pp. 254-257 ◽  
Author(s):  
Weijian Zong ◽  
Jia Zhao ◽  
Xuanyang Chen ◽  
Yuan Lin ◽  
Huixia Ren ◽  
...  
Keyword(s):  
High Resolution ◽  
Light Sheet ◽  
Large Field ◽  
Two Photon ◽  
Light Sheet Microscopy

scholarly journals A versatile oblique plane microscope for large-scale and high-resolution imaging of subcellular dynamics

eLife ◽  
2020 ◽  
Vol 9 ◽  
Author(s):  
Etai Sapoznik ◽  
Bo-Jui Chang ◽  
Jaewon Huh ◽  
Robert J Ju ◽  
Evgenia V Azarova ◽  
...  
Keyword(s):  
High Resolution ◽  
Numerical Aperture ◽  
Light Sheet ◽  
Membrane Dynamics ◽  
Field Of View ◽  
Large Field ◽  
Endoplasmic Reticulum Membrane ◽  
High Resolution Imaging ◽  
Light Sheet Microscopy ◽  
Resolution Imaging

We present an oblique plane microscope (OPM) that uses a bespoke glass-tipped tertiary objective to improve the resolution, field of view, and usability over previous variants. Owing to its high numerical aperture optics, this microscope achieves lateral and axial resolutions that are comparable to the square illumination mode of lattice light-sheet microscopy, but in a user friendly and versatile format. Given this performance, we demonstrate high-resolution imaging of clathrin-mediated endocytosis, vimentin, the endoplasmic reticulum, membrane dynamics, and Natural Killer-mediated cytotoxicity. Furthermore, we image biological phenomena that would be otherwise challenging or impossible to perform in a traditional light-sheet microscope geometry, including cell migration through confined spaces within a microfluidic device, subcellular photoactivation of Rac1, diffusion of cytoplasmic rheological tracers at a volumetric rate of 14 Hz, and large field of view imaging of neurons, developing embryos, and centimeter-scale tissue sections.

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