scholarly journals Multi-fluorescence high-resolution episcopic microscopy (MF-HREM) for three dimensional imaging of adult murine organs

2020 ◽  
Author(s):  
Claire Walsh ◽  
Natalie Holroyd ◽  
Eoin Finnerty ◽  
Sean G. Ryan ◽  
Paul W. Sweeney ◽  
...  
Keyword(s):  
High Resolution ◽  
3D Imaging ◽  
Three Dimensional ◽  
Image Resolution ◽  
Biological Research ◽  
Wide Field ◽  
Vascular Networks ◽  
Optical Sectioning ◽  
Trade Offs ◽  
Xenograft Models

AbstractThree-dimensional microscopy of large biological samples (>0.5 cm3) is transforming biological research. Many existing techniques require trade-offs between image resolution and sample size, require clearing or use optical sectioning. These factors complicate the implementation of large volume 3D imaging. Here we present Multi-fluorescent High Resolution Episcopic Microscopy (MF-HREM) which allows 3D imaging of large samples without the need for clearing or optical sectioning.MF-HREM uses serial-sectioning and block-facing wide-field fluorescence, without the need for tissue clearing or optical sectioning. We detail developments in sample processing including stain penetration, resin embedding and imaging. In addition, we describe image post-processing methods needed to segment and further quantify these data. Finally, we demonstrate the wide applicability of MF-HREM by: 1) quantifying adult mouse glomeruli. 2) identifying injected cells and vascular networks in tumour xenograft models; 3) quantifying vascular networks and white matter track orientation in mouse brain.

Wide-angle, high-resolution three-dimensional (3D) imaging using non-mechanical beam steering

2021 ◽  
Author(s):  
Christopher L. Hoy ◽  
Jay Stockley ◽  
Kelly Kluttz ◽  
Doug McKnight ◽  
Lance Hosting ◽  
...  
Keyword(s):  
High Resolution ◽  
3D Imaging ◽  
Beam Steering ◽  
Three Dimensional ◽  
Wide Angle

scholarly journals Three-Dimensional High-Resolution Digital Inline Hologram Reconstruction with a Volumetric Deconvolution Method

Sensors ◽  
2018 ◽  
Vol 18 (9) ◽  
pp. 2918 ◽  
Author(s):  
Junseong Eom ◽  
Sangjun Moon
Keyword(s):  
High Resolution ◽  
Whole Blood ◽  
Tissue Sample ◽  
Three Dimensional ◽  
Kidney Tissue ◽  
Wide Field ◽  
Deconvolution Technique ◽  
Micron Size ◽  
Volumetric Images ◽  
Focus Image

The digital in-line holographic microscope (DIHM) was developed for a 2D imaging technology and has recently been adapted to 3D imaging methods, providing new approaches to obtaining volumetric images with both a high resolution and wide field-of-view (FOV), which allows the physical limitations to be overcome. However, during the sectioning process of 3D image generation, the out-of-focus image of the object becomes a significant impediment to obtaining evident 3D features in the 2D sectioning plane of a thick biological sample. Based on phase retrieved high-resolution holographic imaging and a 3D deconvolution technique, we demonstrate that a high-resolution 3D volumetric image, which significantly reduces wave-front reconstruction and out-of-focus artifacts, can be achieved. The results show a 3D volumetric image that is more finely focused compared to a conventional 3D stacked image from 2D reconstructed images in relation to micron-size polystyrene beads, a whole blood smear, and a kidney tissue sample. We believe that this technology can be applicable for medical-grade images of smeared whole blood or an optically cleared tissue sample for mobile phytological microscopy and laser sectioning microscopy.

3D Microscopy using Confocal Microscopy

1996 ◽  
Vol 54 ◽  
pp. 270-271
Author(s):  
Ernst H. K. Stelzer ◽  
Steffen Lindek
Keyword(s):  
Incident Light ◽  
Three Dimensional ◽  
Numerical Aperture ◽  
Biological Research ◽  
Half Maximum ◽  
Optical Sectioning ◽  
Observation Volume ◽  
Spread Function ◽  
Relationship Of ◽  
The Relationship

The importance of confocal fluorescence microscopy in modem biological research results from its optical sectioning capability, which allows the three-dimensional analysis of thick specimens. This property is due to the combination of a point-like light source and a point-like detector, which restrict the illumination and detection volumes, respectively. Only the volume that is illuminated and detected is relevant to the confocal observation volume. The smaller it is, the better is the resolution. The performance of a confocal microscope is thus primarily specified by the spatial extent of the confocal point spread function (PSF). The extent can be estimated, e.g., by the volume enclosed by the isosurface at half maximum of the PSF (VHM – volume at half maximum).The relationship of the parameters that determine the lateral resolution of a microscope has been described by Ernst Abbé. The diameter of a light spot in the focal plane Δx is proportional to the wavelength λ of the incident light and inversely proportional to the numerical aperture of the optical system (N.A. = n, ∙ sin α).

A processing work-flow for measuring erythrocytes velocity in extended vascular networks from wide field high-resolution optical imaging data

NeuroImage ◽  
2012 ◽  
Vol 59 (3) ◽  
pp. 2569-2588 ◽  
Author(s):  
Thomas Deneux ◽  
Sylvain Takerkart ◽  
Amiram Grinvald ◽  
Guillaume S. Masson ◽  
Ivo Vanzetta
Keyword(s):  
High Resolution ◽  
Optical Imaging ◽  
Work Flow ◽  
Wide Field ◽  
Imaging Data ◽  
Vascular Networks

Three Dimensional Imaging of Microelectronic Devices Using a CrossBeam FIB

2004 ◽  
Author(s):  
Eric Lifshin ◽  
James Evertsen ◽  
Edward Principe ◽  
John Friel
Keyword(s):  
High Resolution ◽  
3D Imaging ◽  
Ion Beam ◽  
Three Dimensional ◽  
Serial Sections ◽  
Microelectronic Devices ◽  
3D Microscopy ◽  
Internal Structures ◽  
Insight Into

Abstract Increased insight into the internal structure of microelectronic devices can be achieved through the use of three dimensional (3D) imaging based on image stacks of serial sections obtained with a combined electron and ion beam (CrossBeam) FIB. This study describes how such data can be collected and presented, some of the factors that need to be optimized to get the best images, and the limitations of the method. It can be viewed as a first step in the emerging area of high resolution 3D microscopy, a technique that can lead to more accurate characterization of the shapes of internal structures and their interconnectivity at the nanoscale.

scholarly journals Quantitative morphometric measurements using site selective image cytometry of intact tissue

2008 ◽  
Vol 6 (suppl_1) ◽  
Author(s):  
Hyuk-Sang Kwon ◽  
Yoon Sung Nam ◽  
Dominika M Wiktor-Brown ◽  
Bevin P Engelward ◽  
Peter T.C So
Keyword(s):  
High Resolution ◽  
Three Dimensional ◽  
Image Cytometry ◽  
Regions Of Interest ◽  
Wide Field ◽  
Detailed Measurement ◽  
Cell Clusters ◽  
Two Photon ◽  
Recombinant Cell ◽  
Site Selective

Site selective two-photon tissue image cytometry has previously been successfully applied to measure the number of rare cells in three-dimensional tissue specimens up to cubic millimetres in size. However, the extension of this approach for high-throughput quantification of cellular morphological states has not been demonstrated. In this paper, we report the use of site-selective tissue image cytometry for the study of homologous recombination (HR) events during cell division in the pancreas of transgenic mice. Since HRs are rare events, recombinant cells distribute sparsely inside the organ. A detailed measurement throughout the whole tissue is thus not practical. Instead, the site selective two-photon tissue cytometer incorporates a low magnification, wide field, one-photon imaging subsystem that rapidly identifies regions of interest containing recombinant cell clusters. Subsequently, high-resolution three-dimensional assays based on two-photon microscopy can be performed only in these regions of interest. We further show that three-dimensional morphology extraction algorithms can be used to analyse the resultant high-resolution two-photon image stacks providing information not only on the frequency and the distribution of these recombinant cell clusters and their constituent cells, but also on their morphology.

scholarly journals Three-dimensional microscale hanging drop arrays with geometric control for drug screening and live tissue imaging

2021 ◽  
Vol 7 (17) ◽  
pp. eabc1323
Author(s):  
A. Ganguli ◽  
A. Mostafa ◽  
C. Saavedra ◽  
Y. Kim ◽  
P. Le ◽  
...  
Keyword(s):  
High Resolution ◽  
Real Time ◽  
Drug Screening ◽  
Three Dimensional ◽  
3D Culture ◽  
Geometric Control ◽  
Hanging Drop ◽  
Design Elements ◽  
Trade Offs ◽  
On Chip

Existing three-dimensional (3D) culture techniques are limited by trade-offs between throughput, capacity for high-resolution imaging in living state, and geometric control. Here, we introduce a modular microscale hanging drop culture where simple design elements allow high replicates for drug screening, direct on-chip real-time or high-resolution confocal microscopy, and geometric control in 3D. Thousands of spheroids can be formed on our microchip in a single step and without any selective pressure from specific matrices. Microchip cultures from human LN229 glioblastoma and patient-derived mouse xenograft cells retained genomic alterations of originating tumors based on mate pair sequencing. We measured response to drugs over time with real-time microscopy on-chip. Last, by engineering droplets to form predetermined geometric shapes, we were able to manipulate the geometry of cultured cell masses. These outcomes can enable broad applications in advancing personalized medicine for cancer and drug discovery, tissue engineering, and stem cell research.

scholarly journals Three-dimensional nanoscale imaging by plasmonic Brownian microscopy

Nanophotonics ◽  
2017 ◽  
Vol 7 (2) ◽  
pp. 489-495 ◽  
Author(s):  
Anna Labno ◽  
Christopher Gladden ◽  
Jeongmin Kim ◽  
Dylan Lu ◽  
Xiaobo Yin ◽  
...  
Keyword(s):  
3D Imaging ◽  
Optical Tomography ◽  
Three Dimensional ◽  
Optical Resolution ◽  
Super Resolution ◽  
Scanning Speed ◽  
Large Field ◽  
Brownian Diffusion ◽  
Wide Field ◽  
Unlabeled Sample

AbstractThree-dimensional (3D) imaging at the nanoscale is a key to understanding of nanomaterials and complex systems. While scanning probe microscopy (SPM) has been the workhorse of nanoscale metrology, its slow scanning speed by a single probe tip can limit the application of SPM to wide-field imaging of 3D complex nanostructures. Both electron microscopy and optical tomography allow 3D imaging, but are limited to the use in vacuum environment due to electron scattering and to optical resolution in micron scales, respectively. Here we demonstrate plasmonic Brownian microscopy (PBM) as a way to improve the imaging speed of SPM. Unlike photonic force microscopy where a single trapped particle is used for a serial scanning, PBM utilizes a massive number of plasmonic nanoparticles (NPs) under Brownian diffusion in solution to scan in parallel around the unlabeled sample object. The motion of NPs under an evanescent field is three-dimensionally localized to reconstruct the super-resolution topology of 3D dielectric objects. Our method allows high throughput imaging of complex 3D structures over a large field of view, even with internal structures such as cavities that cannot be accessed by conventional mechanical tips in SPM.

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