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5D imaging approaches reveal the formation of distinct intracellular cAMP spatial gradients 24

5D imaging approaches reveal the formation of distinct intra...

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conference on Three-dimensional and multidimensional microscopy - image acquisition and processing XXIV

作者： Rich, Thomas C. Annamdevula, Naga Trinh, Kenny Britain, Andrea L. Mayes, Samuel A. Griswold, John R. Deal, Joshua Hoffman, Chase West, Savannah Leavesley, Silas J. Univ S Alabama Pharmacol Mobile AL 36688 USA Univ S Alabama Ctr Lung Biol Mobile AL 36688 USA Univ S Alabama Basic Med Sci Grad Program Mobile AL 36688 USA Univ S Alabama Chem & Biomol Engn Mobile AL 36688 USA Univ S Alabama Biomed Sci Mobile AL 36688 USA

ISBN: (数字)9781510605824

ISBN: (纸本)9781510605817;9781510605824

Cyclic AMP (cAMP) is a ubiquitous second messenger known to differentially regulate many cellular functions. Several lines of evidence suggest that the distribution of cAMP within cells is not uniform. However, to date, no studies have measured the kinetics of 3D cAMP distributions within cells. This is largely due to the low signal-tonoise ratio of FRET-based probes. We previously reported that hyperspectral imaging improves the signal-to-noise ratio of FRET measurements. Here we utilized hyperspectral imaging approaches to measure FRET signals in five dimensions (5D) -three spatial (x, y, z), wavelength (lambda), and time (t) -allowing us to visualize cAMP gradients in pulmonary endothelial cells. cAMP levels were measured using a FRET-based sensor (H188) comprised of a cAMP binding domain sandwiched between FRET donor and acceptor - Turquoise and Venus fluorescent proteins. We observed cAMP gradients in response to 0.1 or 1 mu M isoproterenol, 0.1 or 1 mu M PGE1, or 50 mu M forskolin. Forskolin-and isoproterenol-induced cAMP gradients formed from the apical (high cAMP) to basolateral (low cAMP) face of cells. In contrast, PGE1-induced cAMP gradients originated from both the basolateral and apical faces of cells. Data suggest that 2D (x, y) studies of cAMP compartmentalization may lead to erroneous conclusions about the existence of cAMP gradients, and that 3D (x, y, z) studies are required to assess mechanisms of signaling specificity. Results demonstrate that 5D imaging technologies are powerful tools for measuring biochemical processes in discrete subcellular domains. This work was supported by NIH P01HL066299, R01HL058506, S10RR027535, AHA 16PRE27130004 and the Abraham Mitchell Cancer Research Fund.

关键词： Hyperspectral imaging spectral imaging microscopy FRET Forster resonance energy transfer cAMP second messenger signaling pulmonary endothelium

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PHASE RETRIEVAL FROM 4-dimensional ELECTRON DIFFRACTION DATASETS

PHASE RETRIEVAL FROM 4-DIMENSIONAL ELECTRON DIFFRACTION DATA...

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IEEE International conference on image processing (ICIP)

作者： Friedrich, Thomas Yu, Chu-Ping Verbeek, Johan Pennycook, Timothy Van Aert, Sandra Univ Antwerp Electron Microscopy Mat Sci EMAT Groenenborgerlaan 171 B-2020 Antwerp Belgium Univ Antwerp NANOlab Ctr Excellence Groenenborgerlaan 171 B-2020 Antwerp Belgium

ISBN: (纸本)9781665441155

We present a computational imaging mode for large scale electron microscopy data, which retrieves a complex wave from noisy/sparse intensity recordings using a deep learning approach and subsequently reconstructs an image of the specimen from the Convolutional Neural Network (CNN) predicted exit waves. We demonstrate that an appropriate forward model in combination with open data frameworks can be used to generate large synthetic datasets for training. In combination with augmenting the data with Poisson noise corresponding to varying dose-values, we effectively eliminate overfitting issues. The U-NET[1] based architecture of the CNN is adapted to the task at hand and performs well while maintaining a relatively small size and fast performance. The validity of the approach is confirmed by comparing the reconstruction to well-established methods using simulated, as well as real electron microscopy data. The proposed method is shown to be effective particularly in the low dose range, evident by strong suppression of noise, good spatial resolution, and sensitivity to different atom types, enabling the simultaneous visualisation of light and heavy elements and making different atomic species distinguishable. Since the method acts on a very local scale and is comparatively fast it bears the potential to be used for near-real-time reconstruction during data acquisition.

关键词： phase retrieval inverse problem electron diffraction 4D-STEM CBED

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A Novel 3D Architecture for High Dynamic Range image Sensor and on-Chip Data Compression

A Novel 3D Architecture for High Dynamic Range Image Sensor ...

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conference on Sensors, Cameras, and Systems for Industrial, Scientific, and Consumer Applications xii

作者： Guezzi-Messaoud, F. Dupret, A. Peizerat, A. Blanchard, Y. CEA Leti L3I Lab 17 Rue Martyrs F-38054 Grenoble France Paris Est Univ ESIEE Engn ESYCOM Lab Paris France

ISBN: (纸本)9780819484123

High Dynamic Range (HDR) image sensors aim at having a dynamic over 120dB. Compared to classical architectures this is obtained at the cost of a higher transistor count, thus lower fill factor. Three dimensional integrated circuits (3D-IC) somehow change the constraints, photodiodes and electronics can be stacked on different layers, giving more processing powers without compromising the fill factor. In this paper, we propose an original architecture for a high dynamic 3D image sensor with data reduction obtained by local compression. HDR acquisition is based on a floating point coding shared by a group of pixel (macro-pixel), thus giving also a first level of compression. A second level of compression is performed by using a Discrete Cosine Transform (DCT). With this new concept a good image quality (PSNR of about 40 dB) and a high dynamic range (120 dB) are obtained within a pixel area of 5 mu mx5 mu m.

关键词： High Dynamic Range Compression CMOS image sensor 3D integration floating point coding

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Visualization environment for electron microscopy

Visualization environment for electron microscopy

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Proceedings of the 1997 5th Pacific conference on Computer Graphics and Applications

作者： Boier Martin, Ioana M. Marinescu, Dan C. Indiana Univ South Bend United States

In this paper we describe an environment for visualization and processing of low-dose, low-contrast electron micrographs of biological specimens. We focus on image selection, the first step in the process of reconstruction of the 3D structure of a specimen from its projections. Noise from a variety of sources makes automatic detection of particle positions a difficult task. New image acquisition devices and modern electron microscopy methods require processing and rendering of very large images (50-100 million pixels). We describe techniques for processing large images, algorithms for detecting particle positions on compressed images using the Crosspoint method, and methods for position refinement. EMMA, an interactive visualization environment for experimenting with particle identification methods is presented.

关键词： Three dimensional computer graphics

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A high performance biometric signal and image processing method to reveal blood perfusion towards 3D oxygen saturation mapping

A high performance biometric signal and image processing met...

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conference on Imaging, Manipulation, and Analysis of Biomolecules, Cells, and Tissues xii

作者： Imms, Ryan Hu, Sijung Azorin-Peris, Vicente Trico, Michael Summers, Ron Univ Loughborough Sch Elect Elect & Syst Engn Ashby Rd Loughborough LE11 3TU Leics England Paris Sud Univ Polytech Paris Sud Engn Sch F-91440 Orsay France

ISBN: (纸本)9780819498601

Non-contact imaging photoplethysmography (PPG) is a recent development in the field of physiological data acquisition, currently undergoing a large amount of research to characterize and define the range of its capabilities. Contact-based PPG techniques have been broadly used in clinical scenarios for a number of years to obtain direct information about the degree of oxygen saturation for patients. With the advent of imaging techniques, there is strong potential to enable access to additional information such as multi-dimensional blood perfusion and saturation mapping. The further development of effective opto-physiological monitoring techniques is dependent upon novel modelling techniques coupled with improved sensor design and effective signal processing methodologies. The biometric signal and imaging processing platform (bSIPP) provides a comprehensive set of features for extraction and analysis of recorded iPPG data, enabling direct comparison with other biomedical diagnostic tools such as ECG and EEG. Additionally, utilizing information about the nature of tissue structure has enabled the generation of an engineering model describing the behaviour of light during its travel through the biological tissue. This enables the estimation of the relative oxygen saturation and blood perfusion in different layers of the tissue to be calculated, which has the potential to be a useful diagnostic tool.

关键词： imaging photoplethysmography pulse oximetry oxygen saturation mapping signal acquisition and processing

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Three-dimensional imaging through millimeter thick tissue specimens

Three-dimensional imaging through millimeter thick tissue sp...

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conference on Three-dimensional microscopy - image acquisition and processing IV

作者： Jones, MG Halliwell, M Bull, DR Davies, D Chinyama, C Wells, PNT United Bristol & NHS Trust & Bristol General Hospital (United Kingdom) Univ. of Bristol (United Kingdom) Univ. of Bristol and Southmead Hospital (United Kingdom) Univ. of Bristol and NHS Trust (United Kingdom)

ISBN: (纸本)0819423955

A 780 nm stage-scanned confocal transmission microscope using compact disk (CD) player optics has been constructed and used to study test objects (including scattering effects) and breast tissue specimens (25-1500 mu m thick). Confocal alignment in transmission is achieved using the CD optical mechanism moving coils to scan the objective lens until intensity peaks;the specimen is present but motionless, Intensity variation as a 3D function of mis-alignment of the two foci may allow estimation of optical deterioration caused by the specimen;appropriate confocal apertures and sample spacings can then be selected. Experimental measurements have been supplemented by 2D Monte-Carlo modeling of photon transport for a scattering-limited confocal transmission system. Modeling has characterized: transmission as a function of focal depth in a slab;contrast produced by a range of embedded spherical target radii;and optical mis-alignment. Poor dye perfusion within thick stained specimens results in little visible internal structure, This may be improved using partially phase dependent imaging (congruent to split-detector), sensitive to specimen refractive index variations.

关键词： confocal transmission imaging thick-tissue Monte-Carlo scattering phase-sensitive imaging

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Applying DIP techniques to microscopic biological images

Applying DIP techniques to microscopic biological images

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conference on Nonlinear image processing and Pattern Analysis xii

作者： Araújo, AA Faria, BM Silva, MAR Reis, HJ Univ Fed Minas Gerais Dept Ciencias Computacao Nucl Proc Digital Imagens BR-31270010 Belo Horizonte MG Brazil

ISBN: (纸本)0819439827

This work reports and illustrates the application of enhancement techniques to animal nervous system images from a Laser Scanning Confocal Microscope. images obtained from this equipment are used to help researchers on localizing several organelles and proteins. Different image components of the same tissue sample can be acquired varying the confocal microscope laser bean wavelength. Due to non-ideal acquisition, numerous images contain artifacts, poor distribution of gray levels and unsystematic contrast gradient. Several techniques have been implemented in order to enhance the images, including noise and artifacts reduction, contrast expansion and enhancements on organelles borders, such as emboss and 3D-visualization. A methodology to accurately solve the frequent contrast gradient problem has been implemented. The approach is based on blurring filter, histogram equalization and arithmetic operations. image coloring is another issue. Each of the acquired components must be merged into one single image with its respective color. The final phase of the work consisted of gathering all implemented techniques to elaborate an application that enclosed facilities to automatically open files from confocal file format (.pic format), apply the developed methodologies to enhance the images, build the multicomponent artificial color image and save the results in common formats. This application must deal with large amounts of images easily, providing facilities to batch processing and image indexing and labeling.

关键词： confocal microscopy image processing image editor image enhancement

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Tomographic phase microscopy using optical tweezers 4

Tomographic phase microscopy using optical tweezers

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conference on Advanced microscopy Techniques IV and Neurophotonics II

作者： Habaza, Mor Gilboa, Barak Roichman, Yael Shaked, Natan T. Tel Aviv Univ Dept Biomed Engn IL-69978 Tel Aviv Israel Tel Aviv Univ Dept Exact Sci IL-69978 Tel Aviv Israel

ISBN: (纸本)9781628417012

We review our technique for tomographic phase microscopy with optical tweezers [1]. This tomographic phase microscopy approach enables full 3-D refractive-index reconstruction. Tomographic phase microscopy measures quantitatively the 3-D distribution of refractive-index in biological cells. We integrated our external interferometric module with holographic optical tweezers for obtaining quantitative phase maps of biological samples from a wide range of angles. The close-to-common-path, off-axis interferometric system enables a full-rotation tomographic acquisition of a single cell using holographic optical tweezers for trapping and manipulating with a desired array of traps, while acquiring phase information of a single cell from all different angles and maintaining the native surrounding medium. We experimentally demonstrated two reconstruction algorithms: the filtered back-projection method and the Fourier diffraction method for 3-D refractive index imaging of yeast cells.

关键词： Holography Interference microscopy Tomography Optical tweezers image reconstruction techniques Three dimensional image processing

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Three-dimensional microscopy using a diffraction grating primary objective

Three-dimensional microscopy using a diffraction grating pri...

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International conference on Applications of Photonics Technology

作者： Ditto, TD Aspex Inc New York NY 10013 USA

ISBN: (纸本)081945527X

Three-dimensional magnification of a diffusing surface can be achieved using profilometry based on holographic chirped frequency gratings and laser line projection. The recently patented method being investigated both supplants earlier 3-D microscopy methods using diffraction range finders and also competes favorably with extant confocal microscopy in coverage area, speed of acquisition and unit cost. The geometry of grazing incidence in a diffraction range finder shows an intrinsic anamorphic magnification. A narrow waist of input rays is expanded to the full width of the grating itself. The spatial magnification is simply the ratio of the length of the grating to the waist of the input column of rays. As the column of input rays approaches the grating plane, this magnification ratio approaches infinity. We develop a theory that predicts a limit to resolution as a function of illumination wave length and grating pitch of the primary objective. A simple microscope can achieve near micron resolution over several centimeters of depth in the visible light regime. An empirical demonstration is made using silver halide holograms.

关键词： Diffraction gratings Diffraction Microscopes Grazing incidence Cameras 3D acquisition 3D image processing microscopy Spectral resolution 3D metrology

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Three dimensional simulations of SEM imaging and charging

Three dimensional simulations of SEM imaging and charging

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conference on Metrology, Inspection, and Process Control for Microlithography XV

作者： Grella, L Lorusso, G Niemi, T Chuang, TC Adler, DL KLA Tencor Corp San Jose CA 95134 USA

ISBN: (纸本)0819440302

SEM based CD control and wafer inspection has an increasingly active role in the semiconductor industry. Current design rules require a CD control with a precision in the nanometer range. In order to achieve this precision, a complete model of the image formation mechanism is desirable. For this reason we present a three-dimensional simulation of scanning electron microscope (SEM) images. The simulations include Monte Carlo electron scattering, charging in the substrate and electron ray-tracing in the column. We investigate some specific cases in CD-SEM metrology: We will describe the effect of scan orientation relative to the orientation of the imaged feature on the apparent beam width (ABW), the effect of magnification on contact imaging, and the effect of residue in resist trenches. Our results, regarding these examples, clearly indicate that a fully three-dimensional numerical simulation is needed to obtain an understanding of image formation and resolution limiting factors.

关键词： Monte Carlo methods Scanning electron microscopy 3D image processing 3D modeling image acquisition 3D metrology Computer simulations image resolution Metrology Scattering

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