scholarly journals Minutes-timescale 3D isotropic imaging of entire organs at subcellular resolution by content-aware compressed-sensing light-sheet microscopy

2019 ◽  
Author(s):  
Chunyu Fang ◽  
Tingting Chu ◽  
Tingting Yu ◽  
Yujie Huang ◽  
Yusha Li ◽  
...  
Keyword(s):  
Compressed Sensing ◽  
Spatial Resolution ◽  
Light Sheet ◽  
System Level ◽  
Projection Neurons ◽  
Long Distance ◽  
Light Sheet Microscopy ◽  
Macro Scale ◽  
Subcellular Resolution ◽  
Content Aware

AbstractInstant 3D imaging of entire organs and organisms at cellular resolution is a recurring challenge in life science. Here we report on a computational light-sheet microscopy able to achieve minute-timescale mapping of entire macro-scale organs at high spatial resolution, thereby overcoming the throughput limit of current 3D microscopy implementations. Through combining a dual-side confocally-scanned Bessel light-sheet illumination which provides thinner-and-wider optical sectioning of deep tissues, with a content-aware compressed sensing (CACS) computation pipeline which further improves the contrast and resolution based on a single acquisition, our method yields 3D images with high, isotropic spatial resolution and rapid acquisition improved by two-orders of magnitude. We demonstrate the imaging of whole brain (∼400 mm3), entire gastrocnemius and tibialis muscles (∼200 mm3) of mouse at subcellular resolution (0.5-μm isovoxel) and ultra-high throughput of 5∼10 minutes per sample. Various system-level cellular analyses, such as mapping cell populations at different brain sub-regions, tracing long-distance projection neurons over the entire brain, and calculating neuromuscular junction occupancy across whole muscle, were also readily enabled by our method.

scholarly journals Minutes-timescale 3D isotropic imaging of entire organs at subcellular resolution by content-aware compressed-sensing light-sheet microscopy

2021 ◽  
Vol 12 (1) ◽  
Author(s):  
Chunyu Fang ◽  
Tingting Yu ◽  
Tingting Chu ◽  
Wenyang Feng ◽  
Fang Zhao ◽  
...  
Keyword(s):  
Compressed Sensing ◽  
Light Sheet ◽  
System Level ◽  
Projection Neurons ◽  
Long Distance ◽  
Rapid Acquisition ◽  
Light Sheet Microscopy ◽  
Macro Scale ◽  
Subcellular Resolution ◽  
Content Aware

AbstractRapid 3D imaging of entire organs and organisms at cellular resolution is a recurring challenge in life science. Here we report on a computational light-sheet microscopy able to achieve minute-timescale high-resolution mapping of entire macro-scale organs. Through combining a dual-side confocally-scanned Bessel light-sheet illumination which provides thinner-and-wider optical sectioning of deep tissues, with a content-aware compressed sensing (CACS) computation pipeline which further improves the contrast and resolution based on a single acquisition, our approach yields 3D images with high, isotropic spatial resolution and rapid acquisition over two-order-of-magnitude faster than conventional 3D microscopy implementations. We demonstrate the imaging of whole brain (~400 mm3), entire gastrocnemius and tibialis muscles (~200 mm3) of mouse at ultra-high throughput of 5~10 min per sample and post-improved subcellular resolution of ~ 1.5 μm (0.5-μm iso-voxel size). Various system-level cellular analyses, such as mapping cell populations at different brain sub-regions, tracing long-distance projection neurons over the entire brain, and calculating neuromuscular junction occupancy across whole muscle, are also readily accomplished by our method.

scholarly journals Simultaneous multiview capture and fusion improves spatial resolution in wide-field and light-sheet microscopy

Optica ◽  
2016 ◽  
Vol 3 (8) ◽  
pp. 897 ◽  
Author(s):  
Yicong Wu ◽  
Panagiotis Chandris ◽  
Peter W. Winter ◽  
Edward Y. Kim ◽  
Valentin Jaumouillé ◽  
...  
Keyword(s):  
Spatial Resolution ◽  
Light Sheet ◽  
Wide Field ◽  
Light Sheet Microscopy

scholarly journals Multimodal Magnetic Resonance Histology and Light Sheet Imaging for Quantitative Neurogenetics of the Mouse

2021 ◽  
Author(s):  
G. Allan Johnson ◽  
Gary Cofer ◽  
James Cook ◽  
James Gee ◽  
Adam Hall ◽  
...  
Keyword(s):  
Magnetic Resonance ◽  
Spatial Resolution ◽  
Mouse Brain ◽  
Angular Resolution ◽  
Imaging Techniques ◽  
Brain Connectivity ◽  
Light Sheet ◽  
Whole Brain ◽  
Light Sheet Microscopy

Paul Lauterbur closed his seminal paper on MRI with the statement that "zeugmatographic (imaging) techniques should find many useful applications in studies of the internal structures, states and composition of microscopic objects" {Lauterbur, 1973 #967}. Magnetic resonance microscopy was subsequently demonstrated in 1986 by three groups{Aguayo, 1986 #968}{Eccles, 1986 #969}{Johnson, 1986 #970}. The application of MRI to the study of tissue structure, i.e. magnetic resonance histology (MRH) was suggested in 1993 {Johnson, 1993 #957}. MRH, while based on the same physical principals as MRI is something fundamentally different than the clinical exams which are typically limited to voxel dimensions of ~ 1 mm3. Preclinical imaging systems can acquire images with voxels ~ 1000 times smaller. The MR histology images presented here have been acquired at yet another factor of 1000 increase in spatial resolution. Figure S1 in the supplement shows a comparison of a state-of-the-art fractional anisotropy images of a C57 mouse brain in vivo @ 150 um resolution (voxel volume of 3.3 x10-3 mm3) with the atlas we have generated for this work at 15 um spatial resolution (voxel volume of 3.3 x 10-6 mm3). In previous work, we have demonstrated the utility of MR histology in neurogenetics at spatial/angular resolution of 45 um /46 angles {Wang N, 2020 #972}. At this spatial/angular resolution it is possible to map whole brain connectivity with high correspondence to retroviral tracers {Calabrese, 2015 #895}. But the MRH derived connectomes can be derived in less than a day where the retroviral tracer studies require months/years {Oh, 2014 #971}. The resolution index (angular samples/voxel volume) for this previous work was >500,000 {Johnson, 2018 #894}. Figure S2 shows a comparison between that previous work and the new atlas presented in this paper with a resolution index of 32 million. Light sheet microscopy (LSM) has undergone similar rapid evolution over the last 20 years. The invention of tissue clearing, advances in immuno histochemistry and development of selective plane illumination microscopy (SPIM) now make it possible to acquire whole mouse brain images at submicron spatial resolution with a vast array of cell specific markers{Ueda, 2020 #974}{Park, 2018 #953}{Murray, 2015 #952}{Gao, 2014 #973}. And these advantages can be realized in scan times of < 6hrs. The major limitation from these studies is the distortion in the tissue from dissection from the cranium, swelling from clearing and staining, and tissue damage from handling. We report here the merger of these two methods: 1. MRH with the brain in the skull to provide accurate geometry, cytoarchitectural measures using scalar imaging metrics and whole brain connectivity at 15 um isotropic spatial resolution with super resolution track density images @ 5 um isotropic resolution; 2. whole brain multichannel LSM @ 1.8x1.8x4.0 um; 3. a big image data infrastructure that enables label mapping from the atlas to the MR image, geometric correction to the light sheet data, label mapping to the light sheet volumes and quantitative extraction of regional cell density. These methods make it possible to generate a comprehensive collection of image derived phenotypes (IDP) of cells and circuits covering the whole mouse brain with throughput that can be scaled for quantitative neurogenetics.

scholarly journals Fast, multicolor 3-D imaging of brain organoids with a new single-objective two-photon virtual light-sheet microscope

2018 ◽  
Author(s):  
Irina Rakotoson ◽  
Brigitte Delhomme ◽  
Philippe Djian ◽  
Andreas Deeg ◽  
Maia Brunstein ◽  
...  
Keyword(s):  
Spatial Resolution ◽  
Three Dimensional ◽  
Numerical Aperture ◽  
Light Sheet ◽  
Confocal Microscope ◽  
Large Field ◽  
Excitation Light ◽  
Two Photon ◽  
Light Sheet Microscopy ◽  
Functional Features

ABSTRACTHuman inducible pluripotent stem cells (hiPSCs) hold a large potential for disease modeling. hiPSC-derived human astrocyte and neuronal cultures permit investigations of neural signaling pathways with subcellular resolution. Combinatorial cultures, and three-dimensional (3-D) embryonic bodies enlarge the scope of investigations to multi-cellular phenomena. A the highest level of complexity, brain organoids that – in many aspects – recapitulate anatomical and functional features of the developing brain permit the study of developmental and morphological aspects of human disease. An ideal microscope for 3-D tissue imaging at these different scales would combine features from both confocal laser-scanning and light-sheet microscopes: a micrometric optical sectioning capacity and sub-micrometric spatial resolution, a large field of view and high frame rate, and a low degree of invasiveness, i.e., ideally, a better photon efficiency than that of a confocal microscope. In the present work, we describe such an instrument that belongs to the class of two-photon (2P) light-sheet microsocpes. Its particularity is that – unlike existing two- or three-lens designs – it is using a single, low-magnification, high-numerical aperture objective for the generation and scanning of a virtual light sheet. The microscope builds on a modified Nipkow-Petran spinning-disk scheme for achieving wide-field excitation. However, unlike the common Yokogawa design that uses a tandem disk, our concept combines micro lenses, dichroic mirrors and detection pinholes on a single disk. This design, advantageous for 2P excitation circumvents problems arising with the tandem disk from the large wavelength-difference between the infrared excitation light and visible fluorescence. 2P fluorescence excited in by the light sheet is collected by the same objective and imaged onto a fast sCMOS camera. We demonstrate three-dimensional imaging of TO-PRO3-stained embryonic bodies and of brain organoids, under control conditions and after rapid (partial) transparisation with triethanolamine and /ormamide (RTF) and compare the performance of our instrument to that of a confocal microscope having a similar numerical aperture. 2P-virtual light-sheet microscopy permits one order of magnitude faster imaging, affords less photobleaching and permits better depth penetration than a confocal microscope with similar spatial resolution.

scholarly journals High-efficiency, large-area lattice light-sheet generation by dielectric metasurfaces

Nanophotonics ◽  
2020 ◽  
Vol 9 (12) ◽  
pp. 4043-4051
Author(s):  
Fenghua Shi ◽  
Jing Wen ◽  
Dangyuan Lei
Keyword(s):  
Spatial Resolution ◽  
Temporal Resolution ◽  
High Efficiency ◽  
Numerical Aperture ◽  
Light Sheet ◽  
Field Of View ◽  
Objective Lens ◽  
High Temporal Resolution ◽  
Light Sheet Microscopy ◽  
Illumination Efficiency

AbstractLattice light-sheet microscopy (LLSM) was developed for long-term live-cell imaging with ultra-fine three-dimensional (3D) spatial resolution, high temporal resolution, and low photo-toxicity by illuminating the sample with a thin lattice-like light-sheet. Currently available schemes for generating thin lattice light-sheets often require complex optical designs. Meanwhile, limited by the bulky objective lens and optical components, the light throughput of existing LLSM systems is rather low. To circumvent the above problems, we utilize a dielectric metasurface of a single footprint to replace the conventional illumination modules used in the conventional LLSM and generate a lattice light-sheet with a ~3-fold broader illumination area and a significantly leveraged illumination efficiency, which consequently leads to a larger field of view with a higher temporal resolution at no extra cost of the spatial resolution. We demonstrate that the metasurface can manipulate spatial frequencies of an input laser beam in orthogonal directions independently to break the trade-off between the field of view and illumination efficiency of the lattice light-sheet. Compared to the conventional LLSM, our metasurface module serving as an ultra-compact illumination component for LLSM at an ease will potentially enable a finer spatial resolution with a larger numerical-aperture detection objective lens.

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 System level analyses of motor-related neural activities in larval Drosophila

2018 ◽  
Author(s):  
Youngteak Yoon ◽  
Jeonghyuk Park ◽  
Atsushi Taniguchi ◽  
Hiroshi Kohsaka ◽  
Ken Nakae ◽  
...  
Keyword(s):  
Animal Movement ◽  
Large Population ◽  
Light Sheet ◽  
Time Frame ◽  
Frame Rate ◽  
System Level ◽  
Nuclear Staining ◽  
Fictive Locomotion ◽  
Light Sheet Microscopy ◽  
Neural Activities

ABSTRACTThe way in which the central nervous system (CNS) governs animal movement is complex and difficult to solve solely by the analyses of muscle movement patterns. We tackle this problem by observing the activity of a large population of neurons in the CNS of larval Drosophila. We focused on two major behaviors of the larvae, forward and backward locomotion, and analyzed the neuronal activity related to these behaviors during fictive locomotion that spontaneously occurs in the isolated CNS. We expressed genetically-encoded calcium indicator, GCaMP, and a nuclear marker in all neurons and used digital scanned light-sheet microscopy to record neural activities in the entire ventral nerve cord at a fast frame rate. We developed image processing tools that automatically detect the cell position based on the nuclear staining and allocate the activity signals to each detected cell. We also applied a machine learning-based method that we developed recently to assign motor status in each time frame. Based on these methods, we find cells whose activity is biased to forward versus backward locomotion and vice versa. In particular, we identified a group of neurons near the boundary of subesophageal zone (SEZ) and thoracic neuromeres, which are strongly active during an early phase of backward but not forward fictive locomotion. Our experimental procedure and computational pipeline enable systematic identification of neurons to show characteristic motor activities in larval Drosophila.

scholarly journals Imaging plant germline differentiation within Arabidopsis flowers by light sheet microscopy

eLife ◽  
2020 ◽  
Vol 9 ◽  
Author(s):  
Sona Valuchova ◽  
Pavlina Mikulkova ◽  
Jana Pecinkova ◽  
Jana Klimova ◽  
Michal Krumnikl ◽  
...  
Keyword(s):  
Higher Plants ◽  
Three Dimensional ◽  
Cell Lineage ◽  
Light Sheet ◽  
Three Dimensional Model ◽  
Light Sheet Microscopy ◽  
Meiotic Chromosomes ◽  
Short Period ◽  
Germline Differentiation ◽  
Subcellular Resolution

In higher plants, germline differentiation occurs during a relatively short period within developing flowers. Understanding of the mechanisms that govern germline differentiation lags behind other plant developmental processes. This is largely because the germline is restricted to relatively few cells buried deep within floral tissues, which makes them difficult to study. To overcome this limitation, we have developed a methodology for live imaging of the germ cell lineage within floral organs of Arabidopsis using light sheet fluorescence microscopy. We have established reporter lines, cultivation conditions, and imaging protocols for high-resolution microscopy of developing flowers continuously for up to several days. We used multiview imagining to reconstruct a three-dimensional model of a flower at subcellular resolution. We demonstrate the power of this approach by capturing male and female meiosis, asymmetric pollen division, movement of meiotic chromosomes, and unusual restitution mitosis in tapetum cells. This method will enable new avenues of research into plant sexual reproduction.

Sign in / Sign up

Export Citation Format

Share Document