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

2019 ◽  
Author(s):  
Tonmoy Chakraborty ◽  
Meghan Driscoll ◽  
Malea Murphy ◽  
Philippe Roudot ◽  
Bo-Jui Chang ◽  
...  
Keyword(s):  
Dendritic Spines ◽  
Large Scale ◽  
Light Sheet ◽  
Automated Detection ◽  
Field Of View ◽  
Large Field ◽  
Axial Resolution ◽  
Optical Sectioning ◽  
Isotropic Resolution ◽  
Light Sheet Microscopy

AbstractWe present cleared tissue Axially Swept Light-Sheet Microscopy (ctASLM), which achieves sub-micron isotropic resolution, high optical sectioning capability, and large field of view imaging (870×870 μm2) over a broad range of immersion media. ctASLM can image live, expanded, and both aqueous and organic chemically cleared tissue preparations and provides 2- to 5-fold better axial resolution than confocal or other reported cleared tissue light-sheet microscopes. We image millimeter-sized tissues with sub-micron 3D resolution, which enabled us to perform automated detection of cells and subcellular features such as dendritic spines.

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 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 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.

Enhancement of image quality in planar Airy light-sheet microscopy via subtraction method

2021 ◽  
Author(s):  
Suhui Deng ◽  
Liusong Yuan ◽  
Peiwei Cheng ◽  
Yuhao Wang ◽  
Mingping Liu
Keyword(s):  
Image Quality ◽  
Light Sheet ◽  
Large Field ◽  
Subtraction Method ◽  
Optical Sectioning ◽  
Two Photon ◽  
Light Sheet Microscopy ◽  
Airy Beam ◽  
Photon Excitation ◽  
Distribution Uniformity

Abstract The use of propagation-invariant Airy beams enables a light-sheet microscopy with a large field-of-view. Without relying upon two-photon excitation or deconvolution-based processing to eliminate out-of focus blur caused by the side lobes, here, we present how the subtraction method is applied to enhance the image quality in digital scanned light-sheet microscopy with Airy beam. In the proposed method, planar Airy beam with the symmetric transversal structure is used to excite the sample. A hollow Airy beam with zero intensity at the focal plane is created, which is mainly used to excite the out-of-focus signal. By scanning the sample twice with the normal planar Airy beam and the hollow Airy beam, digital post-processing of the obtained images by subtraction allows for the rejection of out-of-focus blur and improves the optical sectioning, the axial resolution and the intensity distribution uniformity of the light-sheet microscopy.

scholarly journals Light-sheet microscopy with digital Fourier analysis measures transport properties over large field-of-view

2016 ◽  
Vol 24 (18) ◽  
pp. 20881 ◽  
Author(s):  
Devynn M. Wulstein ◽  
Kathryn E. Regan ◽  
Rae M. Robertson-Anderson ◽  
Ryan McGorty
Keyword(s):  
Fourier Analysis ◽  
Transport Properties ◽  
Light Sheet ◽  
Field Of View ◽  
Large Field ◽  
Light Sheet Microscopy

scholarly journals Light-Sheet Microscopy for Surface Topography Measurements and Quantitative Analysis

Sensors ◽  
2020 ◽  
Vol 20 (10) ◽  
pp. 2842 ◽  
Author(s):  
Zhanpeng Xu ◽  
Erik Forsberg ◽  
Yang Guo ◽  
Fuhong Cai ◽  
Sailing He
Keyword(s):  
Surface Topography ◽  
Spatial Resolution ◽  
Scattered Light ◽  
Three Dimensional ◽  
Light Sheet ◽  
Field Of View ◽  
Large Field ◽  
Linear Calibration ◽  
Industrial Sample ◽  
Light Sheet Microscopy

A novel light-sheet microscopy (LSM) system that uses the laser triangulation method to quantitatively calculate and analyze the surface topography of opaque samples is discussed. A spatial resolution of at least 10 μm in z-direction, 10 μm in x-direction and 25 μm in y-direction with a large field-of-view (FOV) is achieved. A set of sample measurements that verify the system′s functionality in various applications are presented. The system has a simple mechanical structure, such that the spatial resolution is easily improved by replacement of the objective, and a linear calibration formula, which enables convenient system calibration. As implemented, the system has strong potential for, e.g., industrial sample line inspections, however, since the method utilizes reflected/scattered light, it also has the potential for three-dimensional analysis of translucent and layered structures.

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 Light-sheet engineering using the Field Synthesis theorem

2019 ◽  
Author(s):  
Bo-Jui Chang ◽  
Reto Fiolka
Keyword(s):  
Focal Plane ◽  
Light Sheet ◽  
Field Of View ◽  
Resolving Power ◽  
Square Lattice ◽  
Axial Resolution ◽  
Spatiotemporal Resolution ◽  
Light Sheet Microscopy ◽  
Line Beam ◽  
The Cost

AbstractRecent advances in light-sheet microscopy have enabled sensitive imaging with high spatiotemporal resolution. However, the creation of thin light-sheets for high axial resolution is challenging, as the thickness of the sheet, field of view and confinement of the excitation need to be carefully balanced. Some of the thinnest light-sheets created so far have found little practical use as they excite too much out-of-focus fluorescence. In contrast, the most commonly used lightsheet for subcellular imaging, the square lattice, has excellent excitation confinement at the cost of lower axial resolving power. Here we leverage the recently discovered Field Synthesis theorem to create light-sheets where thickness and illumination confinement can be continuously tuned. Explicitly, we scan a line beam across a portion of an annulus mask on the back focal plane of the illumination objective, which we call it C-light-sheets. We experimentally characterize these light-sheets and their application on biological samples.

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.

scholarly journals Using Stage- and Slit-Scanning to Improve Contrast and Optical Sectioning in Dual-View Inverted Light Sheet Microscopy (diSPIM)

2016 ◽  
Vol 231 (1) ◽  
pp. 26-39 ◽  
Author(s):  
Abhishek Kumar ◽  
Ryan Christensen ◽  
Min Guo ◽  
Panos Chandris ◽  
William Duncan ◽  
...  
Keyword(s):  
Light Sheet ◽  
Optical Sectioning ◽  
Light Sheet Microscopy ◽  
Slit Scanning
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