scholarly journals Converting lateral scanning into axial focusing to speed up three-dimensional microscopy

2020 ◽  
Vol 9 (1) ◽  
Author(s):  
Tonmoy Chakraborty ◽  
Bingying Chen ◽  
Stephan Daetwyler ◽  
Bo-Jui Chang ◽  
Oliver Vanderpoorten ◽  
...  
Keyword(s):  
High Resolution ◽  
Spherical Aberration ◽  
Optical Design ◽  
Light Sheet ◽  
Three Dimensions ◽  
High Resolution Imaging ◽  
Light Sheet Microscopy ◽  
Scanning Rate ◽  
Resolution Imaging ◽  
Axial Scanning

Abstract In optical microscopy, the slow axial scanning rate of the objective or the sample has traditionally limited the speed of volumetric imaging. Recently, by conjugating either a movable mirror to the image plane in a remote-focusing geometry or an electrically tuneable lens (ETL) to the back focal plane, rapid axial scanning has been achieved. However, mechanical actuation of a mirror limits the axial scanning rate (usually only 10–100 Hz for piezoelectric or voice coil-based actuators), while ETLs introduce spherical and higher-order aberrations that prevent high-resolution imaging. In an effort to overcome these limitations, we introduce a novel optical design that transforms a lateral-scan motion into a spherical aberration-free axial scan that can be used for high-resolution imaging. 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 the image space. We characterize the optical performance of this remote-focusing technique and use it to accelerate axially swept light-sheet microscopy by an order of magnitude, allowing the quantification of rapid vesicular dynamics in three dimensions. We also demonstrate resonant remote focusing at 12 kHz with a two-photon raster-scanning microscope, which allows rapid imaging of brain tissues and zebrafish cardiac dynamics with diffraction-limited resolution.

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.

Adaptive light-sheet microscopy for long-term, high-resolution imaging in living organisms

2016 ◽  
Vol 34 (12) ◽  
pp. 1267-1278 ◽  
Author(s):  
Loïc A Royer ◽  
William C Lemon ◽  
Raghav K Chhetri ◽  
Yinan Wan ◽  
Michael Coleman ◽  
...  
Keyword(s):  
High Resolution ◽  
Light Sheet ◽  
High Resolution Imaging ◽  
Light Sheet Microscopy ◽  
Resolution Imaging ◽  
Living Organisms

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.

Optical design of the High Resolution Imaging Channel of SIMBIO-SYS: publisher’s note

2019 ◽  
Vol 58 (19) ◽  
pp. 5310
Author(s):  
Michele Zusi ◽  
Riccardo Paolinetti ◽  
Vincenzo Della Corte ◽  
Gabriella Marra ◽  
Marco Baroni ◽  
...  
Keyword(s):  
High Resolution ◽  
Optical Design ◽  
High Resolution Imaging ◽  
Resolution Imaging

scholarly journals Light sheet theta microscopy for rapid high-resolution imaging of large biological samples

BMC Biology ◽  
2018 ◽  
Vol 16 (1) ◽  
Author(s):  
Bianca Migliori ◽  
Malika S. Datta ◽  
Christophe Dupre ◽  
Mehmet C. Apak ◽  
Shoh Asano ◽  
...  
Keyword(s):  
High Resolution ◽  
Biological Samples ◽  
Light Sheet ◽  
High Resolution Imaging ◽  
Resolution Imaging

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.

Optical design of the High Resolution Imaging Channel of SIMBIO-SYS

2019 ◽  
Vol 58 (15) ◽  
pp. 4059 ◽  
Author(s):  
Michele Zusi ◽  
Riccardo Paolinetti ◽  
Vincenzo Della Corte ◽  
Gabriella Marra ◽  
Marco Baroni ◽  
...  
Keyword(s):  
High Resolution ◽  
Optical Design ◽  
High Resolution Imaging ◽  
Resolution Imaging
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