Browse
Submit Data
Databases
API
Help

Dataset Information

10 Views

0 Connections

0 Citations

0 Reanalyses

0 Downloads

Omics score: 0

Combined iDISCO and CUBIC tissue clearing and lightsheet microscopy for in toto analysis of the adult mouse ovary†.

ABSTRACT: At any given time, the ovary contains a number of follicles in distinct growth stages, each with a set of identifying characteristics. Although follicle counting and staging using histological stains on paraffin-embedded ovary sections has been the gold standard in assessing ovarian health in fertility studies, the final counts rely on extrapolation factors that diverge greatly among studies. These methods also limit our ability to investigate spatial aspects of ovary organization. Recent advances in optical tissue clearing and lightsheet microscopy have permitted comprehensive analysis of intact tissues. In this study, we set out to determine the best clearing and imaging methods to generate 3D images of the complete adult mouse ovary that could be used for accurate assessments of ovarian follicles. We found that a combination of iDISCO and CUBIC was the best method to clear the immunostained ovary. Using lightsheet microscopy, we generated 3D images of the intact ovary and performed qualitative assessments of follicles at all stages of development. This study is an important step toward developing quantitative computational models that allow rapid and accurate assessments of growing and quiescent primordial follicles, and to investigate the integrity of extrinsic ovarian components including vascular and neuronal networks.

SUBMITTER: McKey J

PROVIDER: S-EPMC7186783 | biostudies-literature |

REPOSITORIES: biostudies-literature

ACCESS DATA

Json Xml

Similar Datasets

Rapid single-wavelength lightsheet localization microscopy for clarified tissue.

Project description:Optical super-resolution microscopy allows nanoscale imaging of protein molecules in intact biological tissues. However, it is still challenging to perform large volume super-resolution imaging for entire animal organs. Here we develop a single-wavelength Bessel lightsheet method, optimized for refractive-index matching with clarified specimens to overcome the aberrations encountered in imaging thick tissues. Using spontaneous blinking fluorophores to label proteins of interest, we resolve the morphology of most, if not all, dopaminergic neurons in the whole adult brain (3.64 × 107 µm3) of Drosophila melanogaster at the nanometer scale with high imaging speed (436 µm3 per second) for localization. Quantitative single-molecule localization reveals the subcellular distribution of a monoamine transporter protein in the axons of a single, identified serotonergic Dorsal Paired Medial (DPM) neuron. Large datasets are obtained from imaging one brain per day to provide a robust statistical analysis of these imaging data.

| S-EPMC6800451 | biostudies-literature

Tissue Clearing and Light Sheet Microscopy: Imaging the Unsectioned Adult Zebra Finch Brain at Cellular Resolution.

Project description:The inherent complexity of brain tissue, with brain cells intertwining locally and projecting to distant regions, has made three-dimensional visualization of intact brains a highly desirable but challenging task in neuroscience. The natural opaqueness of tissue has traditionally limited researchers to techniques short of single cell resolution such as computer tomography or magnetic resonance imaging. By contrast, techniques with single-cell resolution required mechanical slicing into thin sections, which entails tissue distortions that severely hinder accurate reconstruction of large volumes. Recent developments in tissue clearing and light sheet microscopy have made it possible to investigate large volumes at micrometer resolution. The value of tissue clearing has been shown in a variety of tissue types and animal models. However, its potential for examining the songbird brain remains unexplored. Songbirds are an established model system for the study of vocal learning and sensorimotor control. They share with humans the capacity to adapt vocalizations based on auditory input. Song learning and production are controlled in songbirds by the song system, which forms a network of interconnected discrete brain nuclei. Here, we use the CUBIC and iDISCO+ protocols for clearing adult songbird brain tissue. Combined with light sheet imaging, we show the potential of tissue clearing for the investigation of connectivity between song nuclei, as well as for neuroanatomy and brain vasculature studies.

| S-EPMC6382697 | biostudies-literature

Protocol for Imaging and Analysis of Mouse Tumor Models with CUBIC Tissue Clearing.

Project description:Tissue-clearing technologies have developed rapidly in the past decade, especially for use in neuroscience research. We previously reported that CUBIC, which is one tissue-clearing method, is useful for applications in cancer research. CUBIC cancer analysis can be used to detect cancer metastasis with single-cell resolution at whole mouse body/organ level. This approach can also analyze the tumor characteristics with high-quality 3D images. Here, we describe a detailed CUBIC cancer protocol from tissue clearing, capturing 3D images and post-immunohistochemistry. For complete details on the use and execution of this protocol, please refer to Kubota et al. (2017).

| S-EPMC7757565 | biostudies-literature

Three-Dimensional Study of Alzheimer's Disease Hallmarks Using the iDISCO Clearing Method.

Project description:Amyloidosis is a major problem in over one hundred diseases, including Alzheimer's disease (AD). Using the iDISCO visualization method involving targeted molecular labeling, tissue clearing, and light-sheet microscopy, we studied plaque formation in the intact AD mouse brain at up to 27 months of age. We visualized amyloid plaques in 3D together with tau, microglia, and vasculature. Volume imaging coupled to automated detection and mapping enables precise and fast quantification of plaques within the entire intact mouse brain. The present methodology is also applicable to analysis of frozen human brain samples without specialized preservation. Remarkably, amyloid plaques in human brain tissues showed greater 3D complexity and surprisingly large three-dimensional amyloid patterns, or TAPs. The ability to visualize amyloid in 3D, especially in the context of their micro-environment, and the discovery of large TAPs may have important scientific and medical implications.

| S-EPMC5040352 | biostudies-literature

Three-dimensional evaluation of murine ovarian follicles using a modified CUBIC tissue clearing method.

Project description:BackgroundRecently, we demonstrated the three-dimensional (3D) localization of murine trophoblast giant cells in the pregnant uterus using a modified Clear Unobstructed Brain Imaging Cocktails and Computational analysis (CUBIC) tissue-clearing method and hybrid construct consisting of the cytomegalovirus enhancer fused to the chicken beta-actin promoter (CAG) conjugated enhanced green fluorescent protein (EGFP) transgenic mice. In this study, we applied this method to obtain a transparent whole-image of the ovary and observed the 3D localization of individual oocytes in the developing follicles.MethodsOvarian samples were obtained from EGFP transgenic mice and subjected to nuclear staining with propidium iodide (PI) and CUBIC treatment. The detection of double fluorescence signals (green and red) and subsequent reconstruction of 3D images of the whole ovary were performed by light-sheet microscopy and computer programs, respectively.ResultsThe ovary became transparent using the CUBIC method and each nucleus of the follicle component cells was uniformly fluoro-stained by PI perfusion. In contrast, EGFP signals were strong in oocytes, whereas those of surrounding granulosa cells were faint. These signal differences in EGFP expression among oocytes, granulosa cells, and theca-interstitial cells produce well-contrasted images of the growing follicles, providing clear information of the 3D localization of individual oocytes.ConclusionThese results indicate that this procedure is one of the effective approaches to analyze the 3D structure of follicles in the whole ovary.

| S-EPMC6091003 | biostudies-literature

Passive Clearing and 3D Lightsheet Imaging of the Intact and Injured Spinal Cord in Mice.

Project description:The spinal cord contains a diverse array of sensory and motor circuits that are essential for normal function. Spinal cord injury (SCI) permanently disrupts neural circuits through initial mechanical damage, as well as a cascade of secondary injury events that further expand the spinal cord lesion, resulting in permanent paralysis. Tissue clearing and 3D imaging have recently emerged as promising techniques to improve our understanding of the complex neural circuitry of the spinal cord and the changes that result from damage due to SCI. However, the application of this technology for studying the intact and injured spinal cord remains limited. Here, we optimized the passive CLARITY technique (PACT) to obtain gentle and efficient clearing of the murine spinal cord without the need for specialized equipment. We demonstrate that PACT clearing enables 3D imaging of multiple fluorescent labels in the spinal cord to assess molecularly defined neuronal populations, acute inflammation, long-term tissue damage, and cell transplantation. Collectively, these procedures provide a framework for expanding the utility of tissue clearing to enhance the study of spinal cord neural circuits, as well as cellular- and tissue-level changes that occur following SCI.

| S-EPMC8366232 | biostudies-literature

Visualizing Ocular Morphogenesis by Lightsheet Microscopy Using rx3:GFP Transgenic Zebrafish.

Project description:Vertebrate eye development is a complex process that begins near the end of embryo gastrulation and requires the precise coordination of cell migration, proliferation, and differentiation. Time-lapse imagining offers unique insight to the behavior of cells during eye development because it allows us to visualize oculogenesis in vivo. Zebrafish are an excellent model to visualize this process due to their highly conserved vertebrate eye and their ability to develop rapidly and externally while remaining optically transparent. Time-lapse imaging studies of zebrafish eye development are greatly facilitated by use of the transgenic zebrafish line Tg(rx3:GFP). In the developing forebrain, rx3:GFP expression marks the cells of the single eye field, and GFP continues to be expressed as the eye field evaginates to form an optic vesicle, which then invaginates to form an optic cup. High resolution time lapse imaging of rx3:GFP expression, therefore, allows us to track the eye primordium through time as it develops into the retina. Lightsheet microscopy is an ideal method to image ocular morphogenesis over time due to its ability to penetrate thicker samples for fluorescent imaging, minimize photobleaching and phototoxicity, and image at a high speed. Here, a protocol is provided for time-lapse imaging of ocular morphogenesis using a commercially available lightsheet microscope and an image processing workstation to analyze the resulting data. This protocol details the procedures for embryo anesthesia, embedding in low melting temperature agarose, suspension in the imaging chamber, setting up the imaging parameters, and finally analyzing the imaging data using image analysis software. The resulting dataset can provide valuable insights into the process of ocular morphogenesis, as well as perturbations to this process as a result of genetic mutation, exposure to pharmacological agents, or other experimental manipulations.

| S-EPMC8274632 | biostudies-literature

Optical clearing of archive-compatible paraffin embedded tissue for multiphoton microscopy.

Project description:Standard histopathology techniques (including paraffin embedding) are incompatible with thick tissue multiphoton imaging, and standard clearing techniques on those specimens destroy some molecular information. We demonstrate multiphoton imaging in specimens prepared according to standard histopathology techniques. This permits unlabeled 3-dimensional histology on archival tissue banks, which is of great value in evaluating prognostic indicators.

| S-EPMC3493235 | biostudies-other

Whole-Brain Analysis of Cells and Circuits by Tissue Clearing and Light-Sheet Microscopy.

Project description:In this photo essay, we present a sampling of technologies from laboratories at the forefront of whole-brain clearing and imaging for high-resolution analysis of cell populations and neuronal circuits. The data presented here were provided for the eponymous Mini-Symposium presented at the Society for Neuroscience's 2018 annual meeting.

| S-EPMC6706004 | biostudies-literature

Quantifying three-dimensional rodent retina vascular development using optical tissue clearing and light-sheet microscopy.

Project description:Retinal vasculature develops in a highly orchestrated three-dimensional (3-D) sequence. The stages of retinal vascularization are highly susceptible to oxygen perturbations. We demonstrate that optical tissue clearing of intact rat retinas and light-sheet microscopy provides rapid 3-D characterization of vascular complexity during retinal development. Compared with flat mount preparations that dissect the retina and primarily image the outermost vascular layers, intact cleared retinas imaged using light-sheet fluorescence microscopy display changes in the 3-D retinal vasculature rapidly without the need for point scanning techniques. Using a severe model of retinal vascular disruption, we demonstrate that a simple metric based on Sholl analysis captures the vascular changes observed during retinal development in 3-D. Taken together, these results provide a methodology for rapidly quantifying the 3-D development of the entire rodent retinal vasculature.

| S-EPMC5514054 | biostudies-literature

OmicsDI is part of the ELIXIR infrastructure

OmicsDI is an Elixir interoperability service. Learn more ›

Tweets

OmicsDI Databases

PRIDE
PeptideAtlas
MassIVE
JPOST Repository
Physiome Model Repository

EGA
EVA
ENA
LINCS
PAXDB
Cell Collective

MetaboLights
Metabolomics Workbench
MetabolomeExpress
GNPS
BioModels
FAIRDOMHub

ArrayExpress
dbGaP
ExpressionAtlas
GEO
NODE

Information

Databases
Help
API
Contact us
Code on GitHub
Terms of use
Submit Data