scholarly journals Enhanced field-of-view high-resolution lattice light sheet microscopy with phased array beam formers

2021 ◽  
Author(s):  
Jeremy Witzens ◽  
Marc Spehr ◽  
Pol Van Dorpe ◽  
Niels Verellen
Keyword(s):  
High Resolution ◽  
Phased Array ◽  
Light Sheet ◽  
Field Of View ◽  
Light Sheet Microscopy

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 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 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.

Rapid high resolution 3D imaging of expanded biological specimens with lattice light sheet microscopy

Methods ◽  
2020 ◽  
Vol 174 ◽  
pp. 11-19 ◽  
Author(s):  
Yun-Chi Tsai ◽  
Wei-Chun Tang ◽  
Christine Siok Lan Low ◽  
Yen-Ting Liu ◽  
Jyun-Sian Wu ◽  
...  
Keyword(s):  
High Resolution ◽  
3D Imaging ◽  
Light Sheet ◽  
Light Sheet Microscopy

scholarly journals Reflective imaging improves resolution, speed, and collection efficiency in light sheet microscopy

2017 ◽  
Author(s):  
Yicong Wu ◽  
Abhishek Kumar ◽  
Corey Smith ◽  
Evan Ardiel ◽  
Panagiotis Chandris ◽  
...  
Keyword(s):  
High Resolution ◽  
High Speed ◽  
Collection Efficiency ◽  
Single Cells ◽  
Numerical Aperture ◽  
Light Sheet ◽  
Three Dimensions ◽  
Spatiotemporal Resolution ◽  
Light Sheet Microscopy ◽  
Simultaneous Acquisition

AbstractLight-sheet fluorescence microscopy (LSFM) enables high-speed, high-resolution, gentle imaging of live biological specimens over extended periods. Here we describe a technique that improves the spatiotemporal resolution and collection efficiency of LSFM without modifying the underlying microscope. By imaging samples on reflective coverslips, we enable simultaneous collection of multiple views, obtaining 4 complementary views in 250 ms, half the period it would otherwise take to collect only two views in symmetric dual-view selective plane illumination microscopy (diSPIM). We also report a modified deconvolution algorithm that removes the associated epifluorescence contamination and fuses all views for resolution recovery. Furthermore, we enhance spatial resolution (to < 300 nm in all three dimensions) by applying our method to a new asymmetric diSPIM, permitting simultaneous acquisition of two high-resolution views otherwise difficult to obtain due to steric constraints at high numerical aperture (NA). We demonstrate the broad applicability of our method in a variety of samples of moderate (< 50 μm) thickness, studying mitochondrial, membrane, Golgi, and microtubule dynamics in single cells and calcium activity in nematode embryos.

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