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Sample records for 3d-structured illumination microscopy

  1. 3D structured illumination microscopy

    NASA Astrophysics Data System (ADS)

    Dougherty, William M.; Goodwin, Paul C.

    2011-03-01

    Three-dimensional structured illumination microscopy achieves double the lateral and axial resolution of wide-field microscopy, using conventional fluorescent dyes, proteins and sample preparation techniques. A three-dimensional interference-fringe pattern excites the fluorescence, filling in the "missing cone" of the wide field optical transfer function, thereby enabling axial (z) discrimination. The pattern acts as a spatial carrier frequency that mixes with the higher spatial frequency components of the image, which usually succumb to the diffraction limit. The fluorescence image encodes the high frequency content as a down-mixed, moiré-like pattern. A series of images is required, wherein the 3D pattern is shifted and rotated, providing down-mixed data for a system of linear equations. Super-resolution is obtained by solving these equations. The speed with which the image series can be obtained can be a problem for the microscopy of living cells. Challenges include pattern-switching speeds, optical efficiency, wavefront quality and fringe contrast, fringe pitch optimization, and polarization issues. We will review some recent developments in 3D-SIM hardware with the goal of super-resolved z-stacks of motile cells.

  2. Quantitative 3D structured illumination microscopy of nuclear structures.

    PubMed

    Kraus, Felix; Miron, Ezequiel; Demmerle, Justin; Chitiashvili, Tsotne; Budco, Alexei; Alle, Quentin; Matsuda, Atsushi; Leonhardt, Heinrich; Schermelleh, Lothar; Markaki, Yolanda

    2017-05-01

    3D structured illumination microscopy (3D-SIM) is the super-resolution technique of choice for multicolor volumetric imaging. Here we provide a validated sample preparation protocol for labeling nuclei of cultured mammalian cells, image acquisition and registration practices, and downstream image analysis of nuclear structures and epigenetic marks. Using immunostaining and replication labeling combined with image segmentation, centroid mapping and nearest-neighbor analyses in open-source environments, 3D maps of nuclear structures are analyzed in individual cells and normalized to fluorescence standards on the nanometer scale. This protocol fills an unmet need for the application of 3D-SIM to the technically challenging nuclear environment, and subsequent quantitative analysis of 3D nuclear structures and epigenetic modifications. In addition, it establishes practical guidelines and open-source solutions using ImageJ/Fiji and the TANGO plugin for high-quality and routinely comparable data generation in immunostaining experiments that apply across model systems. From sample preparation through image analysis, the protocol can be executed within one week.

  3. Fluorescence in situ hybridization applications for super-resolution 3D structured illumination microscopy.

    PubMed

    Markaki, Yolanda; Smeets, Daniel; Cremer, Marion; Schermelleh, Lothar

    2013-01-01

    Fluorescence in situ hybridization on three-dimensionally preserved cells (3D-FISH) is an efficient tool to analyze the subcellular localization and spatial arrangement of targeted DNA sequences and RNA transcripts at the single cell level. 3D reconstructions from serial optical sections obtained by confocal laser scanning microscopy (CLSM) have long been considered the gold standard for 3D-FISH analyses. Recent super-resolution techniques circumvent the diffraction-limit of optical resolution and have defined a new state-of-the-art in bioimaging. Three-dimensional structured illumination microscopy (3D-SIM) represents one of these technologies. Notably, 3D-SIM renders an eightfold improved volumetric resolution over conventional imaging, and allows the simultaneous visualization of differently labeled target structures. These features make this approach highly attractive for the analysis of spatial relations and substructures of nuclear targets that escape detection by conventional light microscopy. Here, we focus on the application of 3D-SIM for the visualization of subnuclear 3D-FISH preparations. In comparison with conventional fluorescence microscopy, the quality of 3D-SIM data is dependent to a much greater extent on the optimal sample preparation, labeling and acquisition conditions. We describe typical problems encountered with super-resolution imaging of in situ hybridizations in mammalian tissue culture cells and provide optimized DNA-/(RNA)-FISH protocols including combinations with immunofluorescence staining (Immuno-FISH) and DNA replication labeling using click chemistry.

  4. Super-resolution imaging of the cytokinetic Z ring in live bacteria using fast 3D-structured illumination microscopy (f3D-SIM).

    PubMed

    Turnbull, Lynne; Strauss, Michael P; Liew, Andrew T F; Monahan, Leigh G; Whitchurch, Cynthia B; Harry, Elizabeth J

    2014-09-29

    Imaging of biological samples using fluorescence microscopy has advanced substantially with new technologies to overcome the resolution barrier of the diffraction of light allowing super-resolution of live samples. There are currently three main types of super-resolution techniques - stimulated emission depletion (STED), single-molecule localization microscopy (including techniques such as PALM, STORM, and GDSIM), and structured illumination microscopy (SIM). While STED and single-molecule localization techniques show the largest increases in resolution, they have been slower to offer increased speeds of image acquisition. Three-dimensional SIM (3D-SIM) is a wide-field fluorescence microscopy technique that offers a number of advantages over both single-molecule localization and STED. Resolution is improved, with typical lateral and axial resolutions of 110 and 280 nm, respectively and depth of sampling of up to 30 µm from the coverslip, allowing for imaging of whole cells. Recent advancements (fast 3D-SIM) in the technology increasing the capture rate of raw images allows for fast capture of biological processes occurring in seconds, while significantly reducing photo-toxicity and photobleaching. Here we describe the use of one such method to image bacterial cells harboring the fluorescently-labelled cytokinetic FtsZ protein to show how cells are analyzed and the type of unique information that this technique can provide.

  5. Super-resolution Imaging of the Cytokinetic Z Ring in Live Bacteria Using Fast 3D-Structured Illumination Microscopy (f3D-SIM)

    PubMed Central

    Liew, Andrew T. F.; Monahan, Leigh G.; Whitchurch, Cynthia B.; Harry, Elizabeth J.

    2014-01-01

    Imaging of biological samples using fluorescence microscopy has advanced substantially with new technologies to overcome the resolution barrier of the diffraction of light allowing super-resolution of live samples. There are currently three main types of super-resolution techniques – stimulated emission depletion (STED), single-molecule localization microscopy (including techniques such as PALM, STORM, and GDSIM), and structured illumination microscopy (SIM). While STED and single-molecule localization techniques show the largest increases in resolution, they have been slower to offer increased speeds of image acquisition. Three-dimensional SIM (3D-SIM) is a wide-field fluorescence microscopy technique that offers a number of advantages over both single-molecule localization and STED. Resolution is improved, with typical lateral and axial resolutions of 110 and 280 nm, respectively and depth of sampling of up to 30 µm from the coverslip, allowing for imaging of whole cells. Recent advancements (fast 3D-SIM) in the technology increasing the capture rate of raw images allows for fast capture of biological processes occurring in seconds, while significantly reducing photo-toxicity and photobleaching. Here we describe the use of one such method to image bacterial cells harboring the fluorescently-labelled cytokinetic FtsZ protein to show how cells are analyzed and the type of unique information that this technique can provide. PMID:25286090

  6. Cytology 3D structure formation based on optical microscopy images

    NASA Astrophysics Data System (ADS)

    Pronichev, A. N.; Polyakov, E. V.; Shabalova, I. P.; Djangirova, T. V.; Zaitsev, S. M.

    2017-01-01

    The article the article is devoted to optimization of the parameters of imaging of biological preparations in optical microscopy using a multispectral camera in visible range of electromagnetic radiation. A model for the image forming of virtual preparations was proposed. The optimum number of layers was determined for the object scan in depth and holistic perception of its switching according to the results of the experiment.

  7. Novel scanning electron microscopy methods for analyzing the 3D structure of the Golgi apparatus.

    PubMed

    Koga, Daisuke; Ushiki, Tatsuo; Watanabe, Tsuyoshi

    2017-01-01

    The structure of the Golgi apparatus has been extensively examined by light and electron microscopy, but details of its three-dimensional (3D) structure have remained unclear because of the technical limitations of conventional microscopy techniques. To overcome this problem, we have developed several novel scanning electron microscopy (SEM) methods for observing the 3D structure of subcellular organelles including the Golgi apparatus: (1) an osmium maceration method that facilitates SEM observation of membranous organelles, including the Golgi apparatus, by selectively removing soluble cytoplasmic proteins, (2) an osmium impregnation/maceration method that combines an osmium impregnation method with the osmium maceration method to determine the polarity of the Golgi apparatus by SEM, (3) a correlative light and SEM method that combines a cryosectioning technique with the osmium maceration method to enable correlation of the immunocytochemical distribution of molecules with the 3D ultrastructure of the Golgi apparatus, and (4) array tomography based on the systematic collection and integration of SEM images of serial ultrathin sections on glass slides for revealing the 3D ultrastructure of the entire Golgi apparatus. Together, the novel SEM techniques listed above can reveal the complete 3D structure of the Golgi apparatus in different cell types.

  8. 3D structure tensor analysis of light microscopy data for validating diffusion MRI

    PubMed Central

    Khan, Ahmad Raza; Cornea, Anda; Leigland, Lindsey A.; Kohama, Steven G.; Jespersen, Sune Nørhøj; Kroenke, Christopher D.

    2015-01-01

    Diffusion magnetic resonance imaging (d-MRI) is a powerful non-invasive and non-destructive technique for characterizing brain tissue on the microscopic scale. However, the lack of validation of d-MRI by independent experimental means poses an obstacle to accurate interpretation of data acquired using this method. Recently, structure tensor analysis has been applied to light microscopy images, and this technique holds promise to be a powerful validation strategy for d-MRI. Advantages of this approach include its similarity to d-MRI in terms of averaging the effects of a large number of cellular structures, and its simplicity, which enables it to be implemented in a high-throughput manner. However, a drawback of previous implementations of this technique arises from it being restricted to 2D. As a result, structure tensor analyses have been limited to tissue sectioned in a direction orthogonal to the direction of interest. Here we describe the analytical framework for extending structure tensor analysis to 3D, and utilize the results to analyze serial image “stacks” acquired with confocal microscopy of rhesus macaque hippocampal tissue. Implementation of 3D structure tensor procedures requires removal of sources of anisotropy introduced in tissue preparation and confocal imaging. This is accomplished with image processing steps to mitigate the effects of anisotropic tissue shrinkage, and the effects of anisotropy in the point spread function (PSF). In order to address the latter confound, we describe procedures for measuring the dependence of PSF anisotropy on distance from the microscope objective within tissue. Prior to microscopy, ex vivo d-MRI measurements performed on the hippocampal tissue revealed three regions of tissue with mutually orthogonal directions of least restricted diffusion that correspond to CA1, alveus and inferior longitudinal fasciculus. We demonstrate the ability of 3D structure tensor analysis to identify structure tensor orientations

  9. Structured line illumination Raman microscopy

    PubMed Central

    Watanabe, Kozue; Palonpon, Almar F.; Smith, Nicholas I.; Chiu, Liang-da; Kasai, Atsushi; Hashimoto, Hitoshi; Kawata, Satoshi; Fujita, Katsumasa

    2015-01-01

    In the last couple of decades, the spatial resolution in optical microscopy has increased to unprecedented levels by exploiting the fluorescence properties of the probe. At about the same time, Raman imaging techniques have emerged as a way to image inherent chemical information in a sample without using fluorescent probes. However, in many applications, the achievable resolution is limited to about half the wavelength of excitation light. Here we report the use of structured illumination to increase the spatial resolution of label-free spontaneous Raman microscopy, generating highly detailed spatial contrast from the ensemble of molecular information in the sample. Using structured line illumination in slit-scanning Raman microscopy, we demonstrate a marked improvement in spatial resolution and show the applicability to a range of samples, including both biological and inorganic chemical component mapping. This technique is expected to contribute towards greater understanding of chemical component distributions in organic and inorganic materials. PMID:26626144

  10. Structured illumination temporal compressive microscopy

    PubMed Central

    Yuan, Xin; Pang, Shuo

    2016-01-01

    We present a compressive video microscope based on structured illumination with incoherent light source. The source-side illumination coding scheme allows the emission photons being collected by the full aperture of the microscope objective, and thus is suitable for the fluorescence readout mode. A 2-step iterative reconstruction algorithm, termed BWISE, has been developed to address the mismatch between the illumination pattern size and the detector pixel size. Image sequences with a temporal compression ratio of 4:1 were demonstrated. PMID:27231586

  11. Improved Interference configuration for structured illumination microscopy

    NASA Astrophysics Data System (ADS)

    Chen, Houkai; Wei, Shibiao; Wu, Xiaojing; Yang, Yong; Zhang, Yuquan; Du, Luping; Liu, Jun; Zhu, Siwei; Yuan, Xiaocong

    2017-02-01

    We present an improved structured illumination configuration for structured illumination microscopy (SIM) based on spatial light modulator. Precise phase shifts and rotation of illumination fringes can be dynamically controlled using a spatial light modulator. The method is different from the conventional illumination configuration that are based on interference of ±1 diffractive order light. The experimental setup requires less optical elements making it compact, reliable, and suitable for integration. The method has been applied in the standing-wave total internal reflection fluorescent microscopy. High lateral resolution of sub-100 nm was achieved in single directional resolution enhancement experiments.

  12. Saturated pattern-illuminated Fourier ptychography microscopy

    NASA Astrophysics Data System (ADS)

    Fang, Yue; Chen, Youhua; Kuang, Cuifang; Xiu, Peng; Liu, Qiulan; Ge, Baoliang; Liu, Xu

    2017-01-01

    We report a series of simulation studies which extends pattern-illuminated Fourier ptychography microscopy by integrating with the nonlinearity arising from saturation of the fluorophore excited state for super-resolution fluorescence imaging. This extended technique, termed Saturated pattern-illuminated Fourier ptychography (SpiFP) microscopy, could achieve a resolution four times that of wide field when the illuminating light intensity approaches the saturation threshold in simulations. Increasing light intensity leads to further resolution enhancement. In order to demonstrate the performance of SpiFP, we make a comparison between SpiFP and saturated structure illumination microscopy in simulations, and prove that the SpiFP exhibits superior robustness to noise, aberration correcting ability, and pattern’s flexibility. Introducing the saturation of the fluorescent emission brings in notable improvements in imaging performance, implying its potential in nanoscale-sized biological observations by wide-field microscopy.

  13. Structured illumination fluorescence Fourier ptychographic microscopy

    NASA Astrophysics Data System (ADS)

    Xiu, Peng; Chen, Youhua; Kuang, Cuifang; Fang, Yue; Wang, Yifan; Fan, Jiannan; Xu, Yingke; Liu, Xu

    2016-12-01

    We apply a Fourier ptychographic algorithm for fluorescent samples using structured illumination. The samples are illuminated with structured light patterns and the raw imaging data using traditional structured illumination fluorescence microscopy (SIM) are acquired. We then extract equivalent oblique illuminated images of fluorescent samples from the SIM images. An optimized Fourier ptychography algorithm is proposed, which ensures the fidelity of the reconstructed the super-resolution results. This method can break the diffraction limit to a resolution of λ/4, and has a better signal-to-noise ratio (SNR) than SIM, especially when the background noise is high.

  14. Bessel light sheet structured illumination microscopy

    NASA Astrophysics Data System (ADS)

    Noshirvani Allahabadi, Golchehr

    Biomedical study researchers using animals to model disease and treatment need fast, deep, noninvasive, and inexpensive multi-channel imaging methods. Traditional fluorescence microscopy meets those criteria to an extent. Specifically, two-photon and confocal microscopy, the two most commonly used methods, are limited in penetration depth, cost, resolution, and field of view. In addition, two-photon microscopy has limited ability in multi-channel imaging. Light sheet microscopy, a fast developing 3D fluorescence imaging method, offers attractive advantages over traditional two-photon and confocal microscopy. Light sheet microscopy is much more applicable for in vivo 3D time-lapsed imaging, owing to its selective illumination of tissue layer, superior speed, low light exposure, high penetration depth, and low levels of photobleaching. However, standard light sheet microscopy using Gaussian beam excitation has two main disadvantages: 1) the field of view (FOV) of light sheet microscopy is limited by the depth of focus of the Gaussian beam. 2) Light-sheet images can be degraded by scattering, which limits the penetration of the excitation beam and blurs emission images in deep tissue layers. While two-sided sheet illumination, which doubles the field of view by illuminating the sample from opposite sides, offers a potential solution, the technique adds complexity and cost to the imaging system. We investigate a new technique to address these limitations: Bessel light sheet microscopy in combination with incoherent nonlinear Structured Illumination Microscopy (SIM). Results demonstrate that, at visible wavelengths, Bessel excitation penetrates up to 250 microns deep in the scattering media with single-side illumination. Bessel light sheet microscope achieves confocal level resolution at a lateral resolution of 0.3 micron and an axial resolution of 1 micron. Incoherent nonlinear SIM further reduces the diffused background in Bessel light sheet images, resulting in

  15. Efficient illumination for microsecond tracking microscopy.

    PubMed

    Dulin, David; Barland, Stephane; Hachair, Xavier; Pedaci, Francesco

    2014-01-01

    The possibility to observe microsecond dynamics at the sub-micron scale, opened by recent technological advances in fast camera sensors, will affect many biophysical studies based on particle tracking in optical microscopy. A main limiting factor for further development of fast video microscopy remains the illumination of the sample, which must deliver sufficient light to the camera to allow microsecond exposure times. Here we systematically compare the main illumination systems employed in holographic tracking microscopy, and we show that a superluminescent diode and a modulated laser diode perform the best in terms of image quality and acquisition speed, respectively. In particular, we show that the simple and inexpensive laser illumination enables less than 1 μs camera exposure time at high magnification on a large field of view without coherence image artifacts, together with a good hologram quality that allows nm-tracking of microscopic beads to be performed. This comparison of sources can guide in choosing the most efficient illumination system with respect to the specific application.

  16. Image Correlation Microscopy for Uniform Illumination

    PubMed Central

    Gaborski, Thomas R.; Sealander, Michael N.; Ehrenberg, Morton; Waugh, Richard E.; McGrath, James L.

    2011-01-01

    Image cross-correlation microscopy (ICM) is a technique that quantifies the motion of fluorescent features in an image by measuring the temporal autocorrelation function decay in a time-lapse image sequence. ICM has traditionally employed laser-scanning microscopes because the technique emerged as an extension of laser-based fluorescence correlation spectroscopy (FCS). In this work, we show that image correlation can also be used to measure fluorescence dynamics in uniform illumination or wide-field imaging systems and we call our new approach uniform illumination image correlation microscopy (UI-ICM). Wide-field microscopy is not only a simpler, less expensive imaging modality, but it offers the capability of greater temporal resolution over laser-scanning systems. In traditional laser-scanning ICM, lateral mobility is calculated from the temporal de-correlation of an image, where the characteristic length is the illuminating laser beam width. In wide-field microscopy, the diffusion length is defined by the feature size using the spatial autocorrelation function (SACF). Correlation function decay in time occurs as an object diffuses from its original position. We show that theoretical and simulated comparisons between Gaussian and uniform features indicate the temporal autocorrelation function (TACF) depends strongly on particle size and not particle shape. In this report, we establish the relationships between the SACF feature size, TACF characteristic time and the diffusion coefficient for UI-ICM using analytical, Monte-Carlo and experimental validation with particle tracking algorithms. Additionally, we demonstrate UI-ICM analysis of adhesion molecule domain aggregation and diffusion on the surface of human neutrophils. PMID:20055917

  17. The evolution of structured illumination microscopy in studies of HIV.

    PubMed

    Marno, Kelly; Al'Zoubi, Lara; Pearson, Matthew; Posch, Markus; McKnight, Áine; Wheeler, Ann P

    2015-10-15

    The resolution limit of conventional light microscopy has proven to be limiting for many biological structures such as viruses including Human immunodeficiency virus (HIV). Individual HIV virions are impossible to study using confocal microscopy as they are well below the 200 nm resolution limit of conventional light microscopes. Structured illumination microscopy (SIM) allows a twofold enhancement in image resolution compared to standard widefield illumination and so provides an excellent tool for study of HIV. Viral capsids (CAs) vary between 110 and 146 nm so this study challenges the performance of SIM microscopes. SIM microscopy was first developed in 2000, commercialised in 2007 and rapidly developed. Here we present the changes in capabilities of the SIM microscopes for study of HIV localisation as the instrumentation for structured illumination microscopy has evolved over the past 8 years.

  18. Selective plane illumination microscopy techniques in developmental biology

    PubMed Central

    Huisken, Jan; Stainier, Didier Y. R.

    2009-01-01

    Summary Selective plane illumination microscopy (SPIM) and other fluorescence microscopy techniques in which a focused sheet of light serves to illuminate the sample have become increasingly popular in developmental studies. Fluorescence light-sheet microscopy bridges the gap in image quality between fluorescence stereomicroscopy and high-resolution imaging of fixed tissue sections. In addition, high depth penetration, low bleaching and high acquisition speeds make light-sheet microscopy ideally suited for extended time-lapse experiments in live embryos. This review compares the benefits and challenges of light-sheet microscopy with established fluorescence microscopy techniques such as confocal microscopy and discusses the different implementations and applications of this easily adaptable technology. PMID:19465594

  19. Comparison of image reconstruction methods for structured illumination microscopy

    NASA Astrophysics Data System (ADS)

    Lukeš, Tomas; Hagen, Guy M.; Křížek, Pavel; Švindrych, Zdeněk.; Fliegel, Karel; Klíma, Miloš

    2014-05-01

    Structured illumination microscopy (SIM) is a recent microscopy technique that enables one to go beyond the diffraction limit using patterned illumination. The high frequency information is encoded through aliasing into the observed image. By acquiring multiple images with different illumination patterns aliased components can be separated and a highresolution image reconstructed. Here we investigate image processing methods that perform the task of high-resolution image reconstruction, namely square-law detection, scaled subtraction, super-resolution SIM (SR-SIM), and Bayesian estimation. The optical sectioning and lateral resolution improvement abilities of these algorithms were tested under various noise level conditions on simulated data and on fluorescence microscopy images of a pollen grain test sample and of a cultured cell stained for the actin cytoskeleton. In order to compare the performance of the algorithms, the following objective criteria were evaluated: Signal to Noise Ratio (SNR), Signal to Background Ratio (SBR), circular average of the power spectral density and the S3 sharpness index. The results show that SR-SIM and Bayesian estimation combine illumination patterned images more effectively and provide better lateral resolution in exchange for more complex image processing. SR-SIM requires one to precisely shift the separated spectral components to their proper positions in reciprocal space. High noise levels in the raw data can cause inaccuracies in the shifts of the spectral components which degrade the super-resolved image. Bayesian estimation has proven to be more robust to changes in noise level and illumination pattern frequency.

  20. Lensless phase microscopy using phase retrieval with multiple illumination wavelengths.

    PubMed

    Bao, Peng; Situ, Guohai; Pedrini, Giancarlo; Osten, Wolfgang

    2012-08-01

    A phase retrieval method for microscopy using multiple illumination wavelengths is proposed. A fast algorithm suitable for calculations with high numerical aperture is used for the iterative retrieval of the object wavefront. The advantages and limitations of the technique are systematically analyzed and demonstrated by both simulation and experimental results.

  1. Restoration of uneven illumination in light sheet microscopy images.

    PubMed

    Uddin, Mohammad Shorif; Lee, Hwee Kuan; Preibisch, Stephan; Tomancak, Pavel

    2011-08-01

    Light microscopy images suffer from poor contrast due to light absorption and scattering by the media. The resulting decay in contrast varies exponentially across the image along the incident light path. Classical space invariant deconvolution approaches, while very effective in deblurring, are not designed for the restoration of uneven illumination in microscopy images. In this article, we present a modified radiative transfer theory approach to solve the contrast degradation problem of light sheet microscopy (LSM) images. We confirmed the effectiveness of our approach through simulation as well as real LSM images.

  2. Structured illumination microscopy of autofluorescent aggregations in human tissue.

    PubMed

    Best, Gerrit; Amberger, Roman; Baddeley, David; Ach, Thomas; Dithmar, Stefan; Heintzmann, Rainer; Cremer, Christoph

    2011-06-01

    Sections from human eye tissue were analyzed with Structured Illumination Microscopy (SIM) using a specially designed microscope setup. In this microscope the structured illumination was generated with a Twyman-Green Interferometer. This SIM technique allowed us to acquire light-optical images of autofluorophore distributions in the tissue with previously unmatched optical resolution. In this work the unique setup of the microscope made possible the application of SIM with three different excitation wavelengths (488, 568 and 647 nm), thus enabling us to gather spectral information about the autofluorescence signal.

  3. Structured illumination microscopy with unknown patterns and a statistical prior

    PubMed Central

    Yeh, Li-Hao; Tian, Lei; Waller, Laura

    2017-01-01

    Structured illumination microscopy (SIM) improves resolution by down-modulating high-frequency information of an object to fit within the passband of the optical system. Generally, the reconstruction process requires prior knowledge of the illumination patterns, which implies a well-calibrated and aberration-free system. Here, we propose a new algorithmic self-calibration strategy for SIM that does not need to know the exact patterns a priori, but only their covariance. The algorithm, termed PE-SIMS, includes a pattern-estimation (PE) step requiring the uniformity of the sum of the illumination patterns and a SIM reconstruction procedure using a statistical prior (SIMS). Additionally, we perform a pixel reassignment process (SIMS-PR) to enhance the reconstruction quality. We achieve 2× better resolution than a conventional widefield microscope, while remaining insensitive to aberration-induced pattern distortion and robust against parameter tuning. PMID:28270977

  4. On illumination schemes for wide-field CARS microscopy.

    PubMed

    Toytman, I; Simanovskii, D; Palanker, D

    2009-04-27

    New system for a wide-field CARS microscopy is demonstrated, including two schemes of non-phase-matching illumination. Several advantages including high Stokes pulse energy, pulse-to-pulse stability and inherent synchronization between pump and Stokes pulses were brought by use of methane-filled Raman converter. Spatial resolution of the system with axially symmetric illumination, 0.5 microm, was found to correspond to diffraction limit of the imaging objective. Selective sensitivity to lipid-rich myelin sheaths in the nerve tissue has been demonstrated and confirmed by comparison with histological samples stained with myelin-specific dye. Single-shot imaging capability of the system has been demonstrated with a speckling-free illumination on a monolayer of 3 microm polystyrene beads.

  5. High-resolution 3D structural and optical analyses of hybrid or composite materials by means of scanning probe microscopy combined with the ultramicrotome technique: an example of application to engineering of liquid crystals doped with fluorescent quantum dots

    NASA Astrophysics Data System (ADS)

    Mochalov, Konstantin E.; Efimov, Anton E.; Bobrovsky, Alexey Yu.; Agapov, Igor I.; Chistyakov, Anton A.; Oleinikov, Vladimir A.; Nabiev, Igor

    2013-05-01

    Combination of nanometer-scale 3D structural analysis with optical characterization of the same material is a challenging task. Its results may be important for nanophotonics, materials science, and quality control. We have developed a new technique for complementary high-resolution structural and optical characterization followed by optical spectroscopic and microscopic measurements accompanied by reconstruction of the 3D structure in the same area of the sample. The 3D structure is reconstructed by combination of ultramicrotomic and SPM techniques allowing the study of the 3D distribution of implanted nanoparticles and their effect on the matrix structure. The combination of scanning probe nanotomography (SPN) and optical microspectroscopy makes it possible to direct estimate how the 3D structural characteristics of materials affect their macroscopic optical properties. The technique developed has been applied to the engineering of materials made from cholesteric liquid crystals and fluorescent quantum dots (QDs). These materials permit photochemical patterning and image recording through the changes in the dissymmetry factor of circular polarization of QD emission. The differences in the polarisation images and morphological characteristics of the liquid crystal matrix have proved to be correlated with the arrangement of the areas of homogeneous distribution and nonhomogeneous clustering of QDs. The reconstruction of the 3D structure of the liquid crystal matrix in the areas of homogeneous QD distribution has shown that QDs embedded into cholesteric liquid crystal matrices do not perturb their periodic planar texture. The combined optical/SPM/ultramicrotome technique will be indispensable for evaluating the effects of inorganic nanoparticles on the organisation of organic and liquid crystal matrices, biomedical materials, cells, and tissues.

  6. Lensfree on-chip tomographic microscopy employing multi-angle illumination and pixel super-resolution.

    PubMed

    Isikman, Serhan O; Bishara, Waheb; Ozcan, Aydogan

    2012-08-16

    Tomographic imaging has been a widely used tool in medicine as it can provide three-dimensional (3D) structural information regarding objects of different size scales. In micrometer and millimeter scales, optical microscopy modalities find increasing use owing to the non-ionizing nature of visible light, and the availability of a rich set of illumination sources (such as lasers and light-emitting-diodes) and detection elements (such as large format CCD and CMOS detector-arrays). Among the recently developed optical tomographic microscopy modalities, one can include optical coherence tomography, optical diffraction tomography, optical projection tomography and light-sheet microscopy. These platforms provide sectional imaging of cells, microorganisms and model animals such as C. elegans, zebrafish and mouse embryos. Existing 3D optical imagers generally have relatively bulky and complex architectures, limiting the availability of these equipments to advanced laboratories, and impeding their integration with lab-on-a-chip platforms and microfluidic chips. To provide an alternative tomographic microscope, we recently developed lensfree optical tomography (LOT) as a high-throughput, compact and cost-effective optical tomography modality. LOT discards the use of lenses and bulky optical components, and instead relies on multi-angle illumination and digital computation to achieve depth-resolved imaging of micro-objects over a large imaging volume. LOT can image biological specimen at a spatial resolution of <1 μm x <1 μm x <3 μm in the x, y and z dimensions, respectively, over a large imaging volume of 15-100 mm(3), and can be particularly useful for lab-on-a-chip platforms.

  7. 3D fluorescence anisotropy imaging using selective plane illumination microscopy

    PubMed Central

    Hedde, Per Niklas; Ranjit, Suman; Gratton, Enrico

    2015-01-01

    Fluorescence anisotropy imaging is a popular method to visualize changes in organization and conformation of biomolecules within cells and tissues. In such an experiment, depolarization effects resulting from differences in orientation, proximity and rotational mobility of fluorescently labeled molecules are probed with high spatial resolution. Fluorescence anisotropy is typically imaged using laser scanning and epifluorescence-based approaches. Unfortunately, those techniques are limited in either axial resolution, image acquisition speed, or by photobleaching. In the last decade, however, selective plane illumination microscopy has emerged as the preferred choice for three-dimensional time lapse imaging combining axial sectioning capability with fast, camera-based image acquisition, and minimal light exposure. We demonstrate how selective plane illumination microscopy can be utilized for three-dimensional fluorescence anisotropy imaging of live cells. We further examined the formation of focal adhesions by three-dimensional time lapse anisotropy imaging of CHO-K1 cells expressing an EGFP-paxillin fusion protein. PMID:26368202

  8. Quantitative sectioning and noise analysis for structured illumination microscopy

    PubMed Central

    Hagen, Nathan; Gao, Liang; Tkaczyk, Tomasz S.

    2011-01-01

    Structured illumination (SI) has long been regarded as a nonquantitative technique for obtaining sectioned microscopic images. Its lack of quantitative results has restricted the use of SI sectioning to qualitative imaging experiments, and has also limited researchers’ ability to compare SI against competing sectioning methods such as confocal microscopy. We show how to modify the standard SI sectioning algorithm to make the technique quantitative, and provide formulas for calculating the noise in the sectioned images. The results indicate that, for an illumination source providing the same spatially-integrated photon flux at the object plane, and for the same effective slice thicknesses, SI sectioning can provide higher SNR images than confocal microscopy for an equivalent setup when the modulation contrast exceeds about 0.09. PMID:22274364

  9. Structured illumination microscopy for vibrational molecular imaging (Conference Presentation)

    NASA Astrophysics Data System (ADS)

    Watanabe, Kozue; Palonpon, Almar F.; Smith, Nicholas I.; Chiu, Liang-da; Kasai, Atsushi; Hashimoto, Hitoshi; Kawata, Satoshi; Fujita, Katsumasa

    2016-09-01

    Raman microscopy is a powerful tool for analytical imaging. The wavelength shift of Raman scattering corresponds to molecular vibrational energy. Therefore, we can access rich chemical information, such as distribution, concentration, and chemical environment of sample molecules. Despite these strengths of Raman microscopy, the spatial resolution has been a limiting factor for many practical applications. In this study, we developed a large-area, high-resolution Raman microscope by utilizing structured illumination microscopy (SIM) to overcome the spatial resolution limit. A structured line-illumination (SLI) Raman microscope was constructed. The structured illumination is introduced along the line direction by the interference of two line-shaped beams. In SIM, the spatial frequency mixing between structured illumination and Raman scattering from the sample allows access to the high spatial frequency information beyond the conventional cut-off. As a result, the FWHM of 40-nm fluorescence particle images showed a clear resolution enhancement in the line direction: 366 nm in LI and 199 nm in SLI microscope. Using the developed microscope, we successfully demonstrated high-resolution Raman imaging of various kinds of specimens, such as few-layer graphene, graphite, mouse brain tissue, and polymer nanoparticles. The high resolution Raman images showed the capability to extract original spectral features from the mixed Raman spectra of a multi-component sample because of the enhanced spatial resolution, which is advantageous in observing complex spectral features. The Raman microscopy technique reported here enables us to see the detailed chemical structures of chemical, biological, and medical samples with a spatial resolution smaller than 200 nm.

  10. Fault Tolerant Algorithm for Structured Illumination Microscopy with Incoherent Light

    NASA Astrophysics Data System (ADS)

    Wang, Kun; Heidingsfelder, Philipp; Gao, Jun; Yu, Liandong; Ott, Peter

    2015-04-01

    In this contribution we present a new algorithm for structured illumination microscopy with incoherent light. Existing algorithms for determining the contrast values of the focal depth response require a high accurate phase shift of the fringe pattern illumination. The presented algorithm, which is robust against inaccurate phase shift of the fringe pattern, reduces significantly the requirements for the phase shift and consequently the costs of the microscope. The new algorithm was tested by a preliminary experiment, whereby the grating was shifted by an elastic guided micro-motion mechanism employing a low-cost stepper motor replacing the conventional expensive piezo drive. The determined focal depth response is very smooth and corresponds very well to the theoretical focal depth response.

  11. Highly Resolved Intravital Striped-illumination Microscopy of Germinal Centers

    PubMed Central

    Andresen, Volker; Sporbert, Anje

    2014-01-01

    Monitoring cellular communication by intravital deep-tissue multi-photon microscopy is the key for understanding the fate of immune cells within thick tissue samples and organs in health and disease. By controlling the scanning pattern in multi-photon microscopy and applying appropriate numerical algorithms, we developed a striped-illumination approach, which enabled us to achieve 3-fold better axial resolution and improved signal-to-noise ratio, i.e. contrast, in more than 100 µm tissue depth within highly scattering tissue of lymphoid organs as compared to standard multi-photon microscopy. The acquisition speed as well as photobleaching and photodamage effects were similar to standard photo-multiplier-based technique, whereas the imaging depth was slightly lower due to the use of field detectors. By using the striped-illumination approach, we are able to observe the dynamics of immune complex deposits on secondary follicular dendritic cells – on the level of a few protein molecules in germinal centers. PMID:24748007

  12. Image recombination transform algorithm for superresolution structured illumination microscopy

    NASA Astrophysics Data System (ADS)

    Zhou, Xing; Lei, Ming; Dan, Dan; Yao, Baoli; Yang, Yanlong; Qian, Jia; Chen, Guangde; Bianco, Piero R.

    2016-09-01

    Structured illumination microscopy (SIM) is an attractive choice for fast superresolution imaging. The generation of structured illumination patterns made by interference of laser beams is broadly employed to obtain high modulation depth of patterns, while the polarizations of the laser beams must be elaborately controlled to guarantee the high contrast of interference intensity, which brings a more complex configuration for the polarization control. The emerging pattern projection strategy is much more compact, but the modulation depth of patterns is deteriorated by the optical transfer function of the optical system, especially in high spatial frequency near the diffraction limit. Therefore, the traditional superresolution reconstruction algorithm for interference-based SIM will suffer from many artifacts in the case of projection-based SIM that possesses a low modulation depth. Here, we propose an alternative reconstruction algorithm based on image recombination transform, which provides an alternative solution to address this problem even in a weak modulation depth. We demonstrated the effectiveness of this algorithm in the multicolor superresolution imaging of bovine pulmonary arterial endothelial cells in our developed projection-based SIM system, which applies a computer controlled digital micromirror device for fast fringe generation and multicolor light-emitting diodes for illumination. The merit of the system incorporated with the proposed algorithm allows for a low excitation intensity fluorescence imaging even less than 1 W/cm2, which is beneficial for the long-term, in vivo superresolved imaging of live cells and tissues.

  13. Wide-field Fourier ptychographic microscopy using laser illumination source

    PubMed Central

    Chung, Jaebum; Lu, Hangwen; Ou, Xiaoze; Zhou, Haojiang; Yang, Changhuei

    2016-01-01

    Fourier ptychographic (FP) microscopy is a coherent imaging method that can synthesize an image with a higher bandwidth using multiple low-bandwidth images captured at different spatial frequency regions. The method’s demand for multiple images drives the need for a brighter illumination scheme and a high-frame-rate camera for a faster acquisition. We report the use of a guided laser beam as an illumination source for an FP microscope. It uses a mirror array and a 2-dimensional scanning Galvo mirror system to provide a sample with plane-wave illuminations at diverse incidence angles. The use of a laser presents speckles in the image capturing process due to reflections between glass surfaces in the system. They appear as slowly varying background fluctuations in the final reconstructed image. We are able to mitigate these artifacts by including a phase image obtained by differential phase contrast (DPC) deconvolution in the FP algorithm. We use a 1-Watt laser configured to provide a collimated beam with 150 mW of power and beam diameter of 1 cm to allow for the total capturing time of 0.96 seconds for 96 raw FPM input images in our system, with the camera sensor’s frame rate being the bottleneck for speed. We demonstrate a factor of 4 resolution improvement using a 0.1 NA objective lens over the full camera field-of-view of 2.7 mm by 1.5 mm. PMID:27896016

  14. Color digital lensless holographic microscopy: laser versus LED illumination.

    PubMed

    Garcia-Sucerquia, Jorge

    2016-08-20

    A comparison of the performance of color digital lensless holographic microscopy (CDLHM) as utilized for illumination of RGB lasers or a super-bright white-light LED with a set of spectral filters is presented. As the use of lasers in CDLHM conceals the possibility of having a compact, lightweight, portable, and low cost microscope, and additionally the limited available laser radiation wavelengths limit a real multispectral imaging microscope, here we present the use of super-bright white-light LED and spectral filters for illuminating the sample. The performance of RGB laser-CDLHM and LED-CDLHM is evaluated on imaging a section of the head of a Drosophila melanogaster fly. This comparison shows that there is trade-off between the spatial resolution of the microscope and the light sources utilized, which can be understood with regard to the coherence properties of the illuminating light. Despite the smaller spatial coherence features of LED-CDLHM in comparison with laser-CDLHM, the former shows promise as a portable RGB digital lensless holographic microscope that could be extended to other wavelengths by the use of different spectral filters.

  15. Effect of distorted illumination waves on coherent diffraction microscopy

    SciTech Connect

    Kohmura, Yoshiki; Nishino, Yoshinori; Ishikawa, Tetsuya; Miao Jianwei

    2005-12-15

    Coherent diffraction microscopy requires a well-defined illumination wave such as a plane wave on a specimen. Experimentally, a small pinhole or a focused beam is often used to reduce the illumination area but they unavoidably distort the waves. The distortion of the illumination wave causes artifacts in the phase retrieval of oversampled diffraction patterns. Using computer simulations, we searched for the conditions where strong artifacts arise by changing the Fresnel number, pinhole size, alignment error and photon statistics. The experimental setup with Fresnel number of around 1 and smaller than 1 realized a small reconstruction error when the pinhole radius is larger than a few times the specimen size. These conditions are suitable for the rotation of specimens for the three-dimensional (3D) observations. Such investigation will have an impact in the design of coherent diffraction microscopes for the 3D characterization of nanoscale materials and biological systems using the third generation synchrotron radiation and future x-ray free-electron lasers.

  16. Inclined selective plane illumination microscopy adaptor for conventional microscopes.

    PubMed

    Cutrale, Francesco; Gratton, Enrico

    2012-11-01

    Driven by the biological sciences, there is an increased need for imaging modalities capable of live cell imaging with high spatial and temporal resolution. To achieve this goal in a comprehensive manner, three-dimensional acquisitions are necessary. Ideal features of a modern microscope system should include high imaging speed, high contrast ratio, low photo-bleaching and photo-toxicity, good resolution in a 3D context, and mosaic acquisition for large samples. Given the importance of collecting data in live sample further increases the technical challenges required to solve these issues. This work presents a practical version of a microscopy method, Selective Plane Illumination Microscopy re-introduced by Huisken et al. (Science2004,305,1007-1009). This method is gaining importance in the biomedical field, but its use is limited by difficulties associated with unconventional microscope design which employs two objectives and a particular kind of sample preparation needed to insert the sample between the objectives. Based on the selective plane illumination principle but with a design similar to the Total Internal Reflection Fluorescence microscope, Dunsby (Dunsby, Opt Express 2008,16,20306-20316) demonstrated the oblique plane microscope (OPM) using a single objective which uses conventional sample preparation protocols. However, the Dunsby instrument was not intended to be part of a commercial microscope. In this work, we describe a system with the advantages of OPM and that can be used as an adaptor to commonly used microscopes, such as IX-71 Olympus, simplifying the construction of the OPM and increasing performance of a conventional microscope. We named our design inclined selective plane illumination microscope (iSPIM).

  17. Fast structured illumination microscopy using rolling shutter cameras

    NASA Astrophysics Data System (ADS)

    Song, Liyan; Lu-Walther, Hui-Wen; Förster, Ronny; Jost, Aurélie; Kielhorn, Martin; Zhou, Jianying; Heintzmann, Rainer

    2016-05-01

    Spatial light modulators (SLM) update in a synchronous manner, whereas the data readout process in fast structured illumination systems is usually done using a rolling shutter camera with asynchronous readout. In structured illumination microscopy (SIM), this leads to synchronization problems causing a speed limit for fast acquisition. In this paper we present a configuration to overcome this limit by exploiting the extremely fast SLM display and dividing it into several segments along the direction of the rolling shutter of the sCMOS camera and displaying multiple SLM frames per camera acquisition. The sCMOS runs in continuous rolling shutter mode and the SLM keeps the readout-line always inside a dark region presenting different SIM patterns before and after the readout/start-exposure line. Using this approach, we reached a raw frame rate of 714 frames per second (fps) resulting in a two-beam SIM acquisition rate of 79 fps with a region of interest (ROI) of 16.5  ×  16.5 μm2.

  18. Spatially multiplexed interferometric microscopy with partially coherent illumination

    NASA Astrophysics Data System (ADS)

    Picazo-Bueno, José Ángel; Zalevsky, Zeev; García, Javier; Ferreira, Carlos; Micó, Vicente

    2016-10-01

    We have recently reported on a simple, low cost, and highly stable way to convert a standard microscope into a holographic one [Opt. Express 22, 14929 (2014)]. The method, named spatially multiplexed interferometric microscopy (SMIM), proposes an off-axis holographic architecture implemented onto a regular (nonholographic) microscope with minimum modifications: the use of coherent illumination and a properly placed and selected one-dimensional diffraction grating. In this contribution, we report on the implementation of partially (temporally reduced) coherent illumination in SMIM as a way to improve quantitative phase imaging. The use of low coherence sources forces the application of phase shifting algorithm instead of off-axis holographic recording to recover the sample's phase information but improves phase reconstruction due to coherence noise reduction. In addition, a less restrictive field of view limitation (1/2) is implemented in comparison with our previously reported scheme (1/3). The proposed modification is experimentally validated in a regular Olympus BX-60 upright microscope considering a wide range of samples (resolution test, microbeads, swine sperm cells, red blood cells, and prostate cancer cells).

  19. Fast Neuronal Imaging using Objective Coupled Planar Illumination Microscopy

    NASA Astrophysics Data System (ADS)

    Tarantino, Walter

    Complex computations performed by the brain are produced by activities of neuronal populations. There is a large diversity in the functions of each individual neuron, and neuronal activities occur in the time scale of milliseconds. In order to gain a fundamental understanding of the neuronal populations, one has to measure activity of each neuron at high temporal resolution, while investigating enough neurons to encapsulate the neuronal diversity. Traditional neurotechniques such as electrophysiology and optical imaging are constrained by the number of neurons whose activities can be simultaneously measured or the speed of measuring such activities. We have developed a novel light-sheet based technique called Objective Coupled Planar Illumination (OCPI) microscopy which is capable of measuring simultaneous activities of thousands of neurons at high speeds. In this thesis I pursue the following two aims: · Improve OCPI microscopy by enhancing the spatial resolution deeper in tissue. Tissue inhomogeneity and refractive index mismatch at the surface of the tissue lead to optical aberrations. We have compensated for such aberrations by (1) miniaturizing the OCPI illumination optics, so as to enable more vertical imaging of the tissue, (2) correcting for the angular defocus caused by the refraction at the immersion fluid/tissue interface, and (3) applying adaptive optics to correct for higher order optical aberrations. The improvement in the depth at which one can image tissue will enable the measurement of activities of neuronal populations in cortical areas. · Measure the diversity in the expression pattern of VSNs responsive to sulfated steroids. Nodari et al. have identified sulfated steroids as a novel family of ligands which activate vomeronasal sensory neurons (VSNs). Due to the experimental constraints, it has not been possible to obtain a comprehensive understanding of the number, location and functional characteristics of the sulfated steroid responsive VSNs

  20. Analysing intracellular deformation of polymer capsules using structured illumination microscopy

    NASA Astrophysics Data System (ADS)

    Chen, Xi; Cui, Jiwei; Sun, Huanli; Müllner, Markus; Yan, Yan; Noi, Ka Fung; Ping, Yuan; Caruso, Frank

    2016-06-01

    Understanding the behaviour of therapeutic carriers is important in elucidating their mechanism of action and how they are processed inside cells. Herein we examine the intracellular deformation of layer-by-layer assembled polymer capsules using super-resolution structured illumination microscopy (SIM). Spherical- and cylindrical-shaped capsules were studied in three different cell lines, namely HeLa (human epithelial cell line), RAW264.7 (mouse macrophage cell line) and differentiated THP-1 (human monocyte-derived macrophage cell line). We observed that the deformation of capsules was dependent on cell line, but independent of capsule shape. This suggests that the mechanical forces, which induce capsule deformation during cell uptake, vary between cell lines, indicating that the capsules are exposed to higher mechanical forces in HeLa cells, followed by RAW264.7 and then differentiated THP-1 cells. Our study demonstrates the use of super-resolution SIM in analysing intracellular capsule deformation, offering important insights into the cellular processing of drug carriers in cells and providing fundamental knowledge of intracellular mechanobiology. Furthermore, this study may aid in the design of novel drug carriers that are sensitive to deformation for enhanced drug release properties.Understanding the behaviour of therapeutic carriers is important in elucidating their mechanism of action and how they are processed inside cells. Herein we examine the intracellular deformation of layer-by-layer assembled polymer capsules using super-resolution structured illumination microscopy (SIM). Spherical- and cylindrical-shaped capsules were studied in three different cell lines, namely HeLa (human epithelial cell line), RAW264.7 (mouse macrophage cell line) and differentiated THP-1 (human monocyte-derived macrophage cell line). We observed that the deformation of capsules was dependent on cell line, but independent of capsule shape. This suggests that the mechanical forces

  1. Active focus stabilization for upright selective plane illumination microscopy

    PubMed Central

    Hedde, Per Niklas; Gratton, Enrico

    2015-01-01

    Due to its sectioning capability, large field of view, and minimal light exposure, selective plane illumination microscopy has become the preferred choice for 3D time lapse imaging. Single cells in a dish can be conveniently imaged using an upright/inverted configuration. However, for measurements on long time scales (hours to days), mechanical drift is a problem; especially for studies of mammalian cells that typically require heating to 37°C which causes a thermal gradient across the instrument. Since the light sheet diverges towards the edges of the field of view, such a drift leads to a decrease in axial resolution over time. Or, even worse, the specimen could move out of the imaging volume. Here, we present a simple, cost-effective way to stabilize the axial position using the microscope camera to track the sample position. Thereby, sample loss is prevented and an optimal axial resolution is maintained by keeping the sample at the position where the light sheet is at its thinnest. We demonstrate the virtue of our approach by measurements of the light sheet thickness and 3D time lapse imaging of a cell monolayer at physiological conditions. PMID:26072829

  2. Full-color structured illumination optical sectioning microscopy

    PubMed Central

    Qian, Jia; Lei, Ming; Dan, Dan; Yao, Baoli; Zhou, Xing; Yang, Yanlong; Yan, Shaohui; Min, Junwei; Yu, Xianghua

    2015-01-01

    In merits of super-resolved resolution and fast speed of three-dimensional (3D) optical sectioning capability, structured illumination microscopy (SIM) has found variety of applications in biomedical imaging. So far, most SIM systems use monochrome CCD or CMOS cameras to acquire images and discard the natural color information of the specimens. Although multicolor integration scheme are employed, multiple excitation sources and detectors are required and the spectral information is limited to a few of wavelengths. Here, we report a new method for full-color SIM with a color digital camera. A data processing algorithm based on HSV (Hue, Saturation, and Value) color space is proposed, in which the recorded color raw images are processed in the Hue, Saturation, Value color channels, and then reconstructed to a 3D image with full color. We demonstrated some 3D optical sectioning results on samples such as mixed pollen grains, insects, micro-chips and the surface of coins. The presented technique is applicable to some circumstance where color information plays crucial roles, such as in materials science and surface morphology. PMID:26415516

  3. Advancing ovarian folliculometry with selective plane illumination microscopy

    PubMed Central

    Lin, Hsiao-Chun Amy; Dutta, Rahul; Mandal, Subhamoy; Kind, Alexander; Schnieke, Angelika; Razansky, Daniel

    2016-01-01

    Determination of ovarian status and follicle monitoring are common methods of diagnosing female infertility. We evaluated the suitability of selective plane illumination microscopy (SPIM) for the study of ovarian follicles. The large field of view and fast acquisition speed of our SPIM system enables rendering of volumetric image stacks from intact whole porcine ovarian follicles, clearly visualizing follicular features including follicle volume and average diameter (70 μm–2.5 mm), their spherical asymmetry parameters, size of developing cumulus oophorus complexes (40 μm–110 μm), and follicular wall thickness (90 μm–120 μm). Follicles at all developmental stages were identified. A distribution of the theca thickness was measured for each follicle, and a relationship between these distributions and the stages of follicular development was discerned. The ability of the system to non-destructively generate sub-cellular resolution 3D images of developing follicles, with excellent image contrast and high throughput capacity compared to conventional histology, suggests that it can be used to monitor follicular development and identify structural abnormalities indicative of ovarian ailments. Accurate folliculometric measurements provided by SPIM images can immensely help the understanding of ovarian physiology and provide important information for the proper management of ovarian diseases. PMID:27905503

  4. Advancing ovarian folliculometry with selective plane illumination microscopy

    NASA Astrophysics Data System (ADS)

    Lin, Hsiao-Chun Amy; Dutta, Rahul; Mandal, Subhamoy; Kind, Alexander; Schnieke, Angelika; Razansky, Daniel

    2016-12-01

    Determination of ovarian status and follicle monitoring are common methods of diagnosing female infertility. We evaluated the suitability of selective plane illumination microscopy (SPIM) for the study of ovarian follicles. The large field of view and fast acquisition speed of our SPIM system enables rendering of volumetric image stacks from intact whole porcine ovarian follicles, clearly visualizing follicular features including follicle volume and average diameter (70 μm–2.5 mm), their spherical asymmetry parameters, size of developing cumulus oophorus complexes (40 μm–110 μm), and follicular wall thickness (90 μm–120 μm). Follicles at all developmental stages were identified. A distribution of the theca thickness was measured for each follicle, and a relationship between these distributions and the stages of follicular development was discerned. The ability of the system to non-destructively generate sub-cellular resolution 3D images of developing follicles, with excellent image contrast and high throughput capacity compared to conventional histology, suggests that it can be used to monitor follicular development and identify structural abnormalities indicative of ovarian ailments. Accurate folliculometric measurements provided by SPIM images can immensely help the understanding of ovarian physiology and provide important information for the proper management of ovarian diseases.

  5. Full-color structured illumination optical sectioning microscopy

    NASA Astrophysics Data System (ADS)

    Qian, Jia; Lei, Ming; Dan, Dan; Yao, Baoli; Zhou, Xing; Yang, Yanlong; Yan, Shaohui; Min, Junwei; Yu, Xianghua

    2015-09-01

    In merits of super-resolved resolution and fast speed of three-dimensional (3D) optical sectioning capability, structured illumination microscopy (SIM) has found variety of applications in biomedical imaging. So far, most SIM systems use monochrome CCD or CMOS cameras to acquire images and discard the natural color information of the specimens. Although multicolor integration scheme are employed, multiple excitation sources and detectors are required and the spectral information is limited to a few of wavelengths. Here, we report a new method for full-color SIM with a color digital camera. A data processing algorithm based on HSV (Hue, Saturation, and Value) color space is proposed, in which the recorded color raw images are processed in the Hue, Saturation, Value color channels, and then reconstructed to a 3D image with full color. We demonstrated some 3D optical sectioning results on samples such as mixed pollen grains, insects, micro-chips and the surface of coins. The presented technique is applicable to some circumstance where color information plays crucial roles, such as in materials science and surface morphology.

  6. Active focus stabilization for upright selective plane illumination microscopy.

    PubMed

    Hedde, Per Niklas; Gratton, Enrico

    2015-06-01

    Due to its sectioning capability, large field of view, and minimal light exposure, selective plane illumination microscopy has become the preferred choice for 3D time lapse imaging. Single cells in a dish can be conveniently imaged using an upright/inverted configuration. However, for measurements on long time scales (hours to days), mechanical drift is a problem; especially for studies of mammalian cells that typically require heating to 37°C which causes a thermal gradient across the instrument. Since the light sheet diverges towards the edges of the field of view, such a drift leads to a decrease in axial resolution over time. Or, even worse, the specimen could move out of the imaging volume. Here, we present a simple, cost-effective way to stabilize the axial position using the microscope camera to track the sample position. Thereby, sample loss is prevented and an optimal axial resolution is maintained by keeping the sample at the position where the light sheet is at its thinnest. We demonstrate the virtue of our approach by measurements of the light sheet thickness and 3D time lapse imaging of a cell monolayer at physiological conditions.

  7. Dynactin 3D structure: implications for assembly and dynein binding.

    PubMed

    Imai, Hiroshi; Narita, Akihiro; Maéda, Yuichiro; Schroer, Trina A

    2014-09-23

    The multisubunit protein complex, dynactin, is an essential component of the cytoplasmic dynein motor. High-resolution structural work on dynactin and the dynein/dynactin supercomplex has been limited to small subunits and recombinant fragments that do not report fully on either ≈1MDa assembly. In the present study, we used negative-stain electron microscopy and image analysis based on random conical tilt reconstruction to obtain a three-dimensional (3D) structure of native vertebrate dynactin. The 35-nm-long dynactin molecule has a V-shaped shoulder at one end and a flattened tip at the other end, both offset relative to the long axis of the actin-related protein (Arp) backbone. The shoulder projects dramatically away from the Arp filament core in a way that cannot be appreciated in two-dimensional images, which has implications for the mechanism of dynein binding. The 3D structure allows the helical parameters of the entire Arp filament core, which includes the actin capping protein, CP, to be determined for the first time. This structure exhibits near identity to F-actin and can be well fitted into the dynactin envelope. Molecular fitting of modeled CP-Arp polymers into the envelope shows that the filament contains between 7 and 9 Arp protomers and is capped at both ends. In the 7 Arp model, which agrees best with measured Arp stoichiometry and other structural information, actin capping protein (CP) is not present at the distal tip of the structure, unlike what is seen in the other models. The 3D structure suggests a mechanism for dynactin assembly and length specification.

  8. Resolution doubling using confocal microscopy via analogy with structured illumination microscopy

    NASA Astrophysics Data System (ADS)

    Hayashi, Shinichi

    2016-08-01

    Structured illumination microscopy (SIM) is a super-resolution fluorescence microscopy with a 2-fold higher lateral resolution than conventional wide-field fluorescence (WF) microscopy. Confocal fluorescence (CF) microscopy has approximately the same optical cutoff frequency as SIM; however, the maximum theoretical increase in lateral resolution over that of WF is 1.4-fold with an infinitesimal pinhole diameter. Quantitative comparisons based on an analytical imaging formula revealed that modulation transfer functions (MTFs) of SIM reconstructed images before postprocessing are nearly identical to those of CF images recorded with an infinitesimal pinhole diameter. Here, we propose a new method using an adequate pinhole diameter combined with the use of an apodized Fourier inverse filter to increase the lateral resolution of CF images to as much as that SIM images without significant noise degradation in practice. Furthermore, the proposed method does not require a posteriori parameterization and has reproducibility. This approach can be easily applied to conventional laser scanning CF, spinning disk CF, and multiphoton microscopies.

  9. Automated modeling of RNA 3D structure.

    PubMed

    Rother, Kristian; Rother, Magdalena; Skiba, Pawel; Bujnicki, Janusz M

    2014-01-01

    This chapter gives an overview over the current methods for automated modeling of RNA structures, with emphasis on template-based methods. The currently used approaches to RNA modeling are presented with a side view on the protein world, where many similar ideas have been used. Two main programs for automated template-based modeling are presented: ModeRNA assembling structures from fragments and MacroMoleculeBuilder performing a simulation to satisfy spatial restraints. Both approaches have in common that they require an alignment of the target sequence to a known RNA structure that is used as a modeling template. As a way to find promising template structures and to align the target and template sequences, we propose a pipeline combining the ParAlign and Infernal programs on RNA family data from Rfam. We also briefly summarize template-free methods for RNA 3D structure prediction. Typically, RNA structures generated by automated modeling methods require local or global optimization. Thus, we also discuss methods that can be used for local or global refinement of RNA structures.

  10. Adaptive optimisation of illumination beam profiles in fluorescence microscopy

    NASA Astrophysics Data System (ADS)

    Mitchell, T. J.; Saunter, C. D.; O'Nions, W.; Girkin, J. M.; Love, G. D.

    2015-03-01

    Wide-field fluorescence microscope techniques such as single/selective plane illumination microscope (SPIM) are typically configured to image large regions of a sample at once. Here the illumination beam provides uniform excitation of several biological features across the region, `sliced' to a thickness of between 5-10 microns. In this paper we propose a simple alteration to the optical configuration of a SPIM by switching the light-sheet- forming cylindrical lens with a spatial light modulator. This has the potential to adaptively reconfigure the light sheet geometry to improve the optical sectioning of specific biological features, rather than the thicker sectioning of several features at once across a larger observation field-of-view. We present a prototype version of such a system, referred to as an Adaptive-SPIM (A-SPIM) system. We then suggest that the direct recording of illumination beam shapes within the working microscope system can better facilitate the analysis and subsequent re-configuration of the illumination beam to a specific geometry, and summarise the design and operation of a device that we have developed specifically for this purpose. We finally present reconstructed quantitative three dimensional flux maps of illumination beams from three microscope configurations taken using this miniature high-dynamic range beam profiling device, comparing the beam geometry of a regular SPIM system with our prototype A-SPIM system, and suggesting future improvements.

  11. Illumination, wavelength selection, and detection in fluorescence microscopy.

    PubMed

    Spring, K R

    1991-07-01

    The presently available devices for the illumination, changing of wavelengths, and detection of the resultant fluorescence of biological samples viewed in the light microscope have been described and compared. The optimal choice for illumination is a xenon arc lamp with a filter wheel wavelength selector. The optimal choice for an imaging detector is an intensified CCD (charge-coupled-device) camera. These combinations produce the most rapid, stable, and reproducible results when fluorescence measurements are made on living epithelial cells or isolated renal tubules. Techniques for the simultaneous acquisition of fluorescence and differential interference contrast (DIC) images have also been described and compared.

  12. Differences in nuclear DNA organization between lymphocytes, Hodgkin and Reed-Sternberg cells revealed by structured illumination microscopy.

    PubMed

    Righolt, Christiaan H; Guffei, Amanda; Knecht, Hans; Young, Ian T; Stallinga, Sjoerd; van Vliet, Lucas J; Mai, Sabine

    2014-08-01

    Advances in light microscopy have enabled the visualization of DNA in the interphase nucleus with more detail than is visible with conventional light microscopy. The nuclear architecture is assumed to be different in cancer cells compared to normal cells. In this paper we have studied, for the first time, the organization of nuclear DNA and that of DNA-free space in control lymphocytes, Hodgkin cells and Reed-Sternberg cells using 3D structured illumination microscopy (SIM). We have observed detail in these SIM images that was not observed in conventional widefield images. We have measured the size distribution of the DNA structure using granulometry and noted a significant, progressive increase in the amount of sub-micron structures from control lymphocytes to Hodgkin cells to Reed-Sternberg cells. The DNA-free space changes as well; "holes" in the DNA distribution start to appear in the malignant cells. We have studied whether these "holes" are nucleoli by staining for upstream binding factor (UBF), a protein associated with the nucleolus. We have found that the relative UBF content progressively and significantly decreases-or is absent-in the DNA-free space when measured as either the Pearson correlation coefficient with the DNA-free space or as the number of "holes" that contain UBF. Similar differences exist within the population of Reed-Sternberg cells between binucleated and multinucleated cells with four or more subnuclei. To our knowledge, this is the first study that investigates the changes of the nuclear DNA structure in any disease with superresolution light microscopy.

  13. Transillumination spatially modulated illumination microscopy for human chromosome imaging

    NASA Astrophysics Data System (ADS)

    Pitris, Costas; Heracleous, Peter; Patsalis, Philippos

    2005-03-01

    Human chromosome analysis is an essential task in cytogenetics, especially in prenatal screening, genetic syndrome diagnosis, cancer pathology research and mutagen dosimetry. Chromosomal analysis begins with the creation of a karyotype, which is a layout of chromosome images organized by decreasing size in pairs. Both manual and automatic classification of chromosomes are limited by the resolution of the microscope and imaging system used. One way to improve the results of classification and even detect subtleties now remaining undetected, is to enhance the resolution of the images. It is possible to achieve lateral resolution beyond the classical limit, by using spatially modulated illumination (SMI) in a wide-field, non-confocal microscope. In this case, the sample is illuminated with spatially modulated light, which makes normally inaccessible high-resolution information visible in the observed image by shifting higher frequencies within the OTF limits of the microscope. Although, SMI microscopes have been reported in the past, this manuscript reports the development of a transillumination microscope for opaque, non-fluorescent samples. The illumination path consisted of a light source illuminating a ruled grating which was subsequently imaged on the sample. The grating was mounted on a rotating and translating stage so that the magnification and rotation of the pattern could be adjusted. The imaging lens was a 1.25 NA oil immersion objective. Test samples showed resolution improvement, as judged from a comparison of the experimentally obtained FWHM. Further studies using smaller fringe distance or laser interference pattern illumination will be evaluated to further optimize the SMI results.

  14. Computational modeling of RNA 3D structures and interactions.

    PubMed

    Dawson, Wayne K; Bujnicki, Janusz M

    2016-04-01

    RNA molecules have key functions in cellular processes beyond being carriers of protein-coding information. These functions are often dependent on the ability to form complex three-dimensional (3D) structures. However, experimental determination of RNA 3D structures is difficult, which has prompted the development of computational methods for structure prediction from sequence. Recent progress in 3D structure modeling of RNA and emerging approaches for predicting RNA interactions with ions, ligands and proteins have been stimulated by successes in protein 3D structure modeling.

  15. Super-resolution two-photon microscopy via scanning patterned illumination

    NASA Astrophysics Data System (ADS)

    Urban, Ben E.; Yi, Ji; Chen, Siyu; Dong, Biqin; Zhu, Yongling; DeVries, Steven H.; Backman, Vadim; Zhang, Hao F.

    2015-04-01

    We developed two-photon scanning patterned illumination microscopy (2P-SPIM) for super-resolution two-photon imaging. Our approach used a traditional two-photon microscopy setup with temporally modulated excitation to create patterned illumination fields. Combing nine different illuminations and structured illumination reconstruction, super-resolution imaging was achieved in two-photon microscopy. Using 2P-SPIM we achieved a lateral resolution of 141 nm, which represents an improvement by a factor of 1.9 over the corresponding diffraction limit. We further demonstrated super-resolution cellular imaging by 2P-SPIM to image actin cytoskeleton in mammalian cells and three-dimensional imaging in highly scattering retinal tissue.

  16. 3D Structures of Responsive Nanocompartmentalized Microgels.

    PubMed

    Gelissen, Arjan P H; Oppermann, Alex; Caumanns, Tobias; Hebbeker, Pascal; Turnhoff, Sarah K; Tiwari, Rahul; Eisold, Sabine; Simon, Ulrich; Lu, Yan; Mayer, Joachim; Richtering, Walter; Walther, Andreas; Wöll, Dominik

    2016-11-09

    Compartmentalization in soft matter is important for segregating and coordinating chemical reactions, sequestering (re)active components, and integrating multifunctionality. Advances depend crucially on quantitative 3D visualization in situ with high spatiotemporal resolution. Here, we show the direct visualization of different compartments within adaptive microgels using a combination of in situ electron and super-resolved fluorescence microscopy. We unravel new levels of structural details and address the challenge of reconstructing 3D information from 2D projections for nonuniform soft matter as opposed to monodisperse proteins. Moreover, we visualize the thermally induced shrinkage of responsive core-shell microgels live in water. This strategy opens doors for systematic in situ studies of soft matter systems and their application as smart materials.

  17. Second-harmonic illumination to enhance multispectral digital lensless holographic microscopy.

    PubMed

    Mendoza-Yero, Omel; Carbonell-Leal, Miguel; Lancis, Jesús; Garcia-Sucerquia, Jorge

    2016-03-01

    Multispectral digital lensless holographic microscopy (MDLHM) operating with second-harmonic illumination is shown. Added to the improvement of the spatial resolution of the previously reported MDLHM operating with near-infrared illumination, this second-harmonic MDLHM shows promise as a tool to study the behavior of biological samples under a broad spectral illumination. This illumination is generated by focusing a highly spatially coherent ultrashort pulsed radiation into an uncoated Type 1 β-BaB2O4 (BBO) nonlinear crystal. The second-harmonic MDLHM allows achieving multispectral images of biological samples with enhanced micrometer spatial resolution. The illumination wavelength of the second-harmonic MDLHM can be tuned by displacing a focusing optics with respect to a pinhole; spatially resolved information at different wavelengths of the sample can then be retrieved.

  18. A bright and long-pulse illumination for ultrahigh-speed microscopy of living specimens.

    PubMed

    Nakano, Hitoshi; Yokoi, Sayoko; Yoshida, Shigeru; Yamada, Makoto; Takeuchi, Takeshi; Takehara, Kosei; Etoh, T Goji

    2010-01-01

    Ultrahigh-speed microscopy of living specimens requires ultrabright illumination. Moreover, the duration of illumination should be sufficiently long, on the order of at least several tens of milliseconds, in order to investigate the dynamic state of living specimens. However, specimens are exposed to a high risk of damage by the intense illumination. The brightness and pulse duration of illumination have to be continuously controlled for use in the ultrahigh-speed microscopy of living specimens. Commercial or laboratory-made illumination systems do not satisfy the abovementioned requirements. In this paper, the development of a bright and long-pulse illumination system for ultrahigh-speed microscopy of living specimens is presented. A xenon flashlamp with an arc length of 1.5 mm has been used as the light source. The electrical power supply consists of a voltage-regulated circuit, a capacitor bank, and a control circuit including an insulated-gate bipolar transistor as a gating device, which provides a large rectangular current pulse with the duration in the range to the order of several tens of milliseconds. The brightness, pulse duration, and repetition rate can be easily and continuously controlled. The illumination developed in the present study is installed in an inverted fluorescence microscope equipped with a high-speed camera in order to evaluate the performance as an illumination source. A fluorescent image of the living spermatozoa of a mouse obtained at a frame rate of 8 kHz shows good contrast. Such an image cannot be obtained using a commercial illumination system.

  19. A bright and long-pulse illumination for ultrahigh-speed microscopy of living specimens

    NASA Astrophysics Data System (ADS)

    Nakano, Hitoshi; Yokoi, Sayoko; Yoshida, Shigeru; Yamada, Makoto; Takeuchi, Takeshi; Takehara, Kosei; Etoh, T. Goji

    2010-01-01

    Ultrahigh-speed microscopy of living specimens requires ultrabright illumination. Moreover, the duration of illumination should be sufficiently long, on the order of at least several tens of milliseconds, in order to investigate the dynamic state of living specimens. However, specimens are exposed to a high risk of damage by the intense illumination. The brightness and pulse duration of illumination have to be continuously controlled for use in the ultrahigh-speed microscopy of living specimens. Commercial or laboratory-made illumination systems do not satisfy the abovementioned requirements. In this paper, the development of a bright and long-pulse illumination system for ultrahigh-speed microscopy of living specimens is presented. A xenon flashlamp with an arc length of 1.5 mm has been used as the light source. The electrical power supply consists of a voltage-regulated circuit, a capacitor bank, and a control circuit including an insulated-gate bipolar transistor as a gating device, which provides a large rectangular current pulse with the duration in the range to the order of several tens of milliseconds. The brightness, pulse duration, and repetition rate can be easily and continuously controlled. The illumination developed in the present study is installed in an inverted fluorescence microscope equipped with a high-speed camera in order to evaluate the performance as an illumination source. A fluorescent image of the living spermatozoa of a mouse obtained at a frame rate of 8 kHz shows good contrast. Such an image cannot be obtained using a commercial illumination system.

  20. Super-resolution digital holographic microscopy using multi-point light sources illumination

    NASA Astrophysics Data System (ADS)

    Phan, Anh-Hoang; Park, Jae-Hyeung; Kim, Nam; Jeon, Seok-Hee

    2010-02-01

    In this paper, we use multi-point source illumination to enhance the resolution of digital holographic microscopy without shifting the CCD camera. The specimen is illuminated from many directions by using multi-point sources which are easily created by a lens-array. The high frequency information of the specimen can be captured at a fixed position of CCD camera. All information is then synthesized to increase the resolution.

  1. Super-resolution video microscopy of live cells by structured illumination.

    PubMed

    Kner, Peter; Chhun, Bryant B; Griffis, Eric R; Winoto, Lukman; Gustafsson, Mats G L

    2009-05-01

    Structured-illumination microscopy can double the resolution of the widefield fluorescence microscope but has previously been too slow for dynamic live imaging. Here we demonstrate a high-speed structured-illumination microscope that is capable of 100-nm resolution at frame rates up to 11 Hz for several hundred time points. We demonstrate the microscope by video imaging of tubulin and kinesin dynamics in living Drosophila melanogaster S2 cells in the total internal reflection mode.

  2. Three-dimensional super-resolution structured illumination microscopy with maximum a posteriori probability image estimation.

    PubMed

    Lukeš, Tomáš; Křížek, Pavel; Švindrych, Zdeněk; Benda, Jakub; Ovesný, Martin; Fliegel, Karel; Klíma, Miloš; Hagen, Guy M

    2014-12-01

    We introduce and demonstrate a new high performance image reconstruction method for super-resolution structured illumination microscopy based on maximum a posteriori probability estimation (MAP-SIM). Imaging performance is demonstrated on a variety of fluorescent samples of different thickness, labeling density and noise levels. The method provides good suppression of out of focus light, improves spatial resolution, and allows reconstruction of both 2D and 3D images of cells even in the case of weak signals. The method can be used to process both optical sectioning and super-resolution structured illumination microscopy data to create high quality super-resolution images.

  3. Differences in Nuclear DNA Organization Between Lymphocytes, Hodgkin and Reed–Sternberg Cells Revealed by Structured Illumination Microscopy

    PubMed Central

    Righolt, Christiaan H; Guffei, Amanda; Knecht, Hans; Young, Ian T; Stallinga, Sjoerd; van Vliet, Lucas J; Mai, Sabine

    2014-01-01

    Advances in light microscopy have enabled the visualization of DNA in the interphase nucleus with more detail than is visible with conventional light microscopy. The nuclear architecture is assumed to be different in cancer cells compared to normal cells. In this paper we have studied, for the first time, the organization of nuclear DNA and that of DNA-free space in control lymphocytes, Hodgkin cells and Reed–Sternberg cells using 3D structured illumination microscopy (SIM). We have observed detail in these SIM images that was not observed in conventional widefield images. We have measured the size distribution of the DNA structure using granulometry and noted a significant, progressive increase in the amount of sub-micron structures from control lymphocytes to Hodgkin cells to Reed–Sternberg cells. The DNA-free space changes as well; “holes” in the DNA distribution start to appear in the malignant cells. We have studied whether these “holes” are nucleoli by staining for upstream binding factor (UBF), a protein associated with the nucleolus. We have found that the relative UBF content progressively and significantly decreases—or is absent—in the DNA-free space when measured as either the Pearson correlation coefficient with the DNA-free space or as the number of “holes” that contain UBF. Similar differences exist within the population of Reed–Sternberg cells between binucleated and multinucleated cells with four or more subnuclei. To our knowledge, this is the first study that investigates the changes of the nuclear DNA structure in any disease with superresolution light microscopy. J. Cell. Biochem. 115: 1441–1448, 2014. © 2014 The Authors. Journal of Cellular Biochemistry published by Wiley Periodicals, Inc. This is an open access article under the terms of the Creative Commons Attribution-NonCommercial-NoDerivs License, which permits use and distribution in any medium, provided the original work is properly cited, the use is non

  4. Generation of apodized X-ray illumination and its application to scanning and diffraction microscopy.

    PubMed

    Khakurel, Krishna P; Kimura, Takashi; Nakamori, Hiroki; Goto, Takumi; Matsuyama, Satoshi; Sasaki, Tomoya; Takei, Masashi; Kohmura, Yoshiki; Ishikawa, Tetsuya; Yamauchi, Kazuto; Nishino, Yoshinori

    2017-01-01

    X-ray science has greatly benefited from the progress in X-ray optics. Advances in the design and the manufacturing techniques of X-ray optics are key to the success of various microscopic and spectroscopic techniques practiced today. Here the generation of apodized X-ray illumination using a two-stage deformable Kirkpatrick-Baez mirror system is presented. Such apodized illumination is marked by the suppression of the side-lobe intensities of the focused beam. Thus generated apodized illumination was employed to improve the image quality in scanning X-ray fluorescence microscopy. Imaging of a non-isolated object by coherent X-ray diffractive imaging with apodized illumination in a non-scanning mode is also presented.

  5. Complete Tem-Tomography: 3D Structure of Gems Cluster

    NASA Technical Reports Server (NTRS)

    Matsuno, J.; Miyake, A.; Tsuchiyama, A.; Messenger, S.; Nakamura-Messenger, K.

    2015-01-01

    GEMS (glass with embedded metal and sulfide) grains in interplanetary dust particles (IDPs) are considered to be one of the ubiquitous and fundamental building blocks of solids in the Solar System. They have been considered to be interstellar silicate dust that survived various metamorphism or alteration processes in the protoplanetary disk but the elemental and isotopic composition measurements suggest that most of them have been formed in the protoplanetary disk as condensates from high temperature gas. This formation model is also supported by the formation of GEMS-like grains with respect to the size, mineral assemblage, texture and infrared spectrum by condensation experiments from mean GEMS composition materials. Previous GEMS studies were performed only with 2D observation by transmission electron microscopy (TEM) or scanning TEM (STEM). However, the 3D shape and structure of GEMS grains and the spatial distribution of Fe/FeS's has critical information about their formation and origin. Recently, the 3D structure of GEMS grains in ultrathin sections of cluster IDPs was revealed by electron tomography using a TEM/STEM (JEM-2100F, JEOL). However, CT images of thin sections mounted on Cu grids acquired by conventional TEM-tomography are limited to low tilt angles (e. g., less than absolute value of 75 deg. In fact, previous 3D TEM observations of GEMS were affected by some artifacts related to the limited tilt range in the TEM used. Complete tomographic images should be acquired by rotating the sample tilt angle over a range of more than absolute value of 80 deg otherwise the CT images lose their correct structures. In order to constrain the origin and formation process of GEMS grains more clearly, we performed complete electron tomography for GEMS grains. Here we report the sample preparation method we have developed for this study, and the preliminary results.

  6. Three-dimensional fluorescent microscopy via simultaneous illumination and detection at multiple planes

    PubMed Central

    Ma, Qian; Khademhosseinieh, Bahar; Huang, Eric; Qian, Haoliang; Bakowski, Malina A.; Troemel, Emily R.; Liu, Zhaowei

    2016-01-01

    The conventional optical microscope is an inherently two-dimensional (2D) imaging tool. The objective lens, eyepiece and image sensor are all designed to capture light emitted from a 2D ‘object plane’. Existing technologies, such as confocal or light sheet fluorescence microscopy have to utilize mechanical scanning, a time-multiplexing process, to capture a 3D image. In this paper, we present a 3D optical microscopy method based upon simultaneously illuminating and detecting multiple focal planes. This is implemented by adding two diffractive optical elements to modify the illumination and detection optics. We demonstrate that the image quality of this technique is comparable to conventional light sheet fluorescent microscopy with the advantage of the simultaneous imaging of multiple axial planes and reduced number of scans required to image the whole sample volume. PMID:27527813

  7. Digital micromirror device-based laser-illumination Fourier ptychographic microscopy

    PubMed Central

    Kuang, Cuifang; Ma, Ye; Zhou, Renjie; Lee, Justin; Barbastathis, George; Dasari, Ramachandra R.; Yaqoob, Zahid; So, Peter T. C.

    2015-01-01

    We report a novel approach to Fourier ptychographic microscopy (FPM) by using a digital micromirror device (DMD) and a coherent laser source (532 nm) for generating spatially modulated sample illumination. Previously demonstrated FPM systems are all based on partially-coherent illumination, which offers limited throughput due to insufficient brightness. Our FPM employs a high power coherent laser source to enable shot-noise limited high-speed imaging. For the first time, a digital micromirror device (DMD), imaged onto the back focal plane of the illumination objective, is used to generate spatially modulated sample illumination field for ptychography. By coding the on/off states of the micromirrors, the illumination plane wave angle can be varied at speeds more than 4 kHz. A set of intensity images, resulting from different oblique illuminations, are used to numerically reconstruct one high-resolution image without obvious laser speckle. Experiments were conducted using a USAF resolution target and a fiber sample, demonstrating high-resolution imaging capability of our system. We envision that our approach, if combined with a coded-aperture compressive-sensing algorithm, will further improve the imaging speed in DMD-based FPM systems. PMID:26480361

  8. Volumetric structured illumination microscopy enabled by a tunable-focus lens.

    PubMed

    Hinsdale, Taylor; Malik, Bilal H; Olsovsky, Cory; Jo, Javier A; Maitland, Kristen C

    2015-11-01

    We present a mechanical-scan-free method for volumetric imaging of biological tissue. The optical sectioning is provided by structured illumination, and the depth of the imaging plane is varied using an electrically tunable-focus lens. We characterize and evaluate the ability of this axial-scanning mechanism in structured illumination microscopy and demonstrate its ability to perform subcellular resolution imaging in oral mucosa ex vivo. The proposed mechanism can potentially convert any wide-field microscope to a 3D-imaging platform without the need for mechanical scanning of imaging optics and/or sample.

  9. DMD-based LED-illumination super-resolution and optical sectioning microscopy.

    PubMed

    Dan, Dan; Lei, Ming; Yao, Baoli; Wang, Wen; Winterhalder, Martin; Zumbusch, Andreas; Qi, Yujiao; Xia, Liang; Yan, Shaohui; Yang, Yanlong; Gao, Peng; Ye, Tong; Zhao, Wei

    2013-01-01

    Super-resolution three-dimensional (3D) optical microscopy has incomparable advantages over other high-resolution microscopic technologies, such as electron microscopy and atomic force microscopy, in the study of biological molecules, pathways and events in live cells and tissues. We present a novel approach of structured illumination microscopy (SIM) by using a digital micromirror device (DMD) for fringe projection and a low-coherence LED light for illumination. The lateral resolution of 90 nm and the optical sectioning depth of 120 μm were achieved. The maximum acquisition speed for 3D imaging in the optical sectioning mode was 1.6×10(7) pixels/second, which was mainly limited by the sensitivity and speed of the CCD camera. In contrast to other SIM techniques, the DMD-based LED-illumination SIM is cost-effective, ease of multi-wavelength switchable and speckle-noise-free. The 2D super-resolution and 3D optical sectioning modalities can be easily switched and applied to either fluorescent or non-fluorescent specimens.

  10. Nonlinear structured-illumination microscopy: Wide-field fluorescence imaging with theoretically unlimited resolution

    PubMed Central

    Gustafsson, Mats G. L.

    2005-01-01

    Contrary to the well known diffraction limit, the fluorescence microscope is in principle capable of unlimited resolution. The necessary elements are spatially structured illumination light and a nonlinear dependence of the fluorescence emission rate on the illumination intensity. As an example of this concept, this article experimentally demonstrates saturated structured-illumination microscopy, a recently proposed method in which the nonlinearity arises from saturation of the excited state. This method can be used in a simple, wide-field (nonscanning) microscope, uses only a single, inexpensive laser, and requires no unusual photophysical properties of the fluorophore. The practical resolving power is determined by the signal-to-noise ratio, which in turn is limited by photobleaching. Experimental results show that a 2D point resolution of <50 nm is possible on sufficiently bright and photostable samples. PMID:16141335

  11. Fluorescence microscopy gets faster and clearer: roles of photochemistry and selective illumination.

    PubMed

    Wolenski, Joseph S; Julich, Doerthe

    2014-03-01

    Significant advances in fluorescence microscopy tend be a balance between two competing qualities wherein improvements in resolution and low light detection are typically accompanied by losses in acquisition rate and signal-to-noise, respectively. These trade-offs are becoming less of a barrier to biomedical research as recent advances in optoelectronic microscopy and developments in fluorophore chemistry have enabled scientists to see beyond the diffraction barrier, image deeper into live specimens, and acquire images at unprecedented speed. Selective plane illumination microscopy has provided significant gains in the spatial and temporal acquisition of fluorescence specimens several mm in thickness. With commercial systems now available, this method promises to expand on recent advances in 2-photon deep-tissue imaging with improved speed and reduced photobleaching compared to laser scanning confocal microscopy. Superresolution microscopes are also available in several modalities and can be coupled with selective plane illumination techniques. The combination of methods to increase resolution, acquisition speed, and depth of collection are now being married to common microscope systems, enabling scientists to make significant advances in live cell and in situ imaging in real time. We show that light sheet microscopy provides significant advantages for imaging live zebrafish embryos compared to laser scanning confocal microscopy.

  12. RNA Structure: Advances and Assessment of 3D Structure Prediction.

    PubMed

    Miao, Zhichao; Westhof, Eric

    2017-03-30

    Biological functions of RNA molecules are dependent upon sustained specific three-dimensional (3D) structures of RNA, with or without the help of proteins. Understanding of RNA structure is frequently based on 2D structures, which describe only the Watson-Crick (WC) base pairs. Here, we hierarchically review the structural elements of RNA and how they contribute to RNA 3D structure. We focus our analysis on the non-WC base pairs and on RNA modules. Several computer programs have now been designed to predict RNA modules. We describe the RNA-Puzzles initiative, which is a community-wide, blind assessment of RNA 3D structure prediction programs to determine the capabilities and bottlenecks of current predictions. The assessment metrics used in RNA-Puzzles are briefly described. The detection of RNA 3D modules from sequence data and their automatic implementation belong to the current challenges in RNA 3D structure prediction. Expected final online publication date for the Annual Review of Biophysics Volume 46 is May 20, 2017. Please see http://www.annualreviews.org/page/journal/pubdates for revised estimates.

  13. Formal representation of 3D structural geological models

    NASA Astrophysics Data System (ADS)

    Wang, Zhangang; Qu, Honggang; Wu, Zixing; Yang, Hongjun; Du, Qunle

    2016-05-01

    The development and widespread application of geological modeling methods has increased demands for the integration and sharing services of three dimensional (3D) geological data. However, theoretical research in the field of geological information sciences is limited despite the widespread use of Geographic Information Systems (GIS) in geology. In particular, fundamental research on the formal representations and standardized spatial descriptions of 3D structural models is required. This is necessary for accurate understanding and further applications of geological data in 3D space. In this paper, we propose a formal representation method for 3D structural models using the theory of point set topology, which produces a mathematical definition for the major types of geological objects. The spatial relationships between geologic boundaries, structures, and units are explained in detail using the 9-intersection model. Reasonable conditions for describing the topological space of 3D structural models are also provided. The results from this study can be used as potential support for the standardized representation and spatial quality evaluation of 3D structural models, as well as for specific needs related to model-based management, query, and analysis.

  14. Designing 3D Structure by 5-7 Kirigami

    NASA Astrophysics Data System (ADS)

    Gong, Xingting; Cho, Yigil; Castle, Toen; Sussman, Daniel; Kamien, Randall

    2015-03-01

    The purpose of this talk is to explore how one can create 3D structures from 2D materials through the art of kirigami. Kirigami expands upon origami by allowing not only folds, but also cuts, into materials. If we take an incompressible material such as paper and remove a hole from it, the paper will buckle into the third dimension once that hole is sealed in order to relieve strain. Thus, orienting cuts and folds in certain places throughout a sheet of paper can influence its ``pop-up,'' 3D structure. To narrow down the inverse design problem, we confined ourselves to making only one kind of cut (which we call the ``5-7 cut'') on a honeycomb grid, and we show how this single cut can give rise to arbitrarily complex three dimensional structures. A simple set of rules exists: (a) one 5-7 cut divides the material into 2 sections which can choose to pop-up or down independently of each other, (b) rows of uniform cuts must pop up or down in unison, giving (nearly) arbitrary 2D structure, and (c) the 5-7 cuts can be arranged in various ways to create 6 basic pop-up ``modes,'' which can then be arranged to give (nearly) arbitrary 3D structure. These simple rules allow a framework for designing targeted 3D structure from an initial 2D sheet of material. This work was supported by NSF EFRI-ODISSEI Grant EFRI 13-31583.

  15. Coarse-grained modeling of RNA 3D structure.

    PubMed

    Dawson, Wayne K; Maciejczyk, Maciej; Jankowska, Elzbieta J; Bujnicki, Janusz M

    2016-07-01

    Functional RNA molecules depend on three-dimensional (3D) structures to carry out their tasks within the cell. Understanding how these molecules interact to carry out their biological roles requires a detailed knowledge of RNA 3D structure and dynamics as well as thermodynamics, which strongly governs the folding of RNA and RNA-RNA interactions as well as a host of other interactions within the cellular environment. Experimental determination of these properties is difficult, and various computational methods have been developed to model the folding of RNA 3D structures and their interactions with other molecules. However, computational methods also have their limitations, especially when the biological effects demand computation of the dynamics beyond a few hundred nanoseconds. For the researcher confronted with such challenges, a more amenable approach is to resort to coarse-grained modeling to reduce the number of data points and computational demand to a more tractable size, while sacrificing as little critical information as possible. This review presents an introduction to the topic of coarse-grained modeling of RNA 3D structures and dynamics, covering both high- and low-resolution strategies. We discuss how physics-based approaches compare with knowledge based methods that rely on databases of information. In the course of this review, we discuss important aspects in the reasoning process behind building different models and the goals and pitfalls that can result.

  16. In vivo volumetric fluorescence sectioning microscopy with mechanical-scan-free hybrid illumination imaging

    PubMed Central

    Lin, Chen-Yen; Lin, Wei-Hsin; Chien, Ju-Hsuan; Tsai, Jui-Chang; Luo, Yuan

    2016-01-01

    Optical sectioning microscopy in wide-field fashion has been widely used to obtain three-dimensional images of biological samples; however, it requires scanning in depth and considerable time to acquire multiple depth information of a volumetric sample. In this paper, in vivo optical sectioning microscopy with volumetric hybrid illumination, with no mechanical moving parts, is presented. The proposed system is configured such that the optical sectioning is provided by hybrid illumination using a digital micro-mirror device (DMD) for uniform and non-uniform pattern projection, while the depth of imaging planes is varied by using an electrically tunable-focus lens with invariant magnification and resolution. We present and characterize the design, implementation, and experimentally demonstrate the proposed system’s ability through 3D imaging of in vivo Canenorhabditis elegans’ growth cones. PMID:27867708

  17. Imaging multicellular specimens with real-time optimized tiling light-sheet selective plane illumination microscopy

    PubMed Central

    Fu, Qinyi; Martin, Benjamin L.; Matus, David Q.; Gao, Liang

    2016-01-01

    Despite the progress made in selective plane illumination microscopy, high-resolution 3D live imaging of multicellular specimens remains challenging. Tiling light-sheet selective plane illumination microscopy (TLS-SPIM) with real-time light-sheet optimization was developed to respond to the challenge. It improves the 3D imaging ability of SPIM in resolving complex structures and optimizes SPIM live imaging performance by using a real-time adjustable tiling light sheet and creating a flexible compromise between spatial and temporal resolution. We demonstrate the 3D live imaging ability of TLS-SPIM by imaging cellular and subcellular behaviours in live C. elegans and zebrafish embryos, and show how TLS-SPIM can facilitate cell biology research in multicellular specimens by studying left-right symmetry breaking behaviour of C. elegans embryos. PMID:27004937

  18. Measuring the size of biological nanostructures with spatially modulated illumination microscopy.

    PubMed

    Martin, Sonya; Failla, Antonio Virgilio; Spöri, Udo; Cremer, Christoph; Pombo, Ana

    2004-05-01

    Spatially modulated illumination fluorescence microscopy can in theory measure the sizes of objects with a diameter ranging between 10 and 200 nm and has allowed accurate size measurement of subresolution fluorescent beads ( approximately 40-100 nm). Biological structures in this size range have so far been measured by electron microscopy. Here, we have labeled sites containing the active, hyperphosphorylated form of RNA polymerase II in the nucleus of HeLa cells by using the antibody H5. The spatially modulated illumination-microscope was compared with confocal laser scanning and electron microscopes and found to be suitable for measuring the size of cellular nanostructures in a biological setting. The hyperphosphorylated form of polymerase II was found in structures with a diameter of approximately 70 nm, well below the 200-nm resolution limit of standard fluorescence microscopes.

  19. Pipeline for illumination correction of images for high-throughput microscopy.

    PubMed

    Singh, S; Bray, M-A; Jones, T R; Carpenter, A E

    2014-12-01

    The presence of systematic noise in images in high-throughput microscopy experiments can significantly impact the accuracy of downstream results. Among the most common sources of systematic noise is non-homogeneous illumination across the image field. This often adds an unacceptable level of noise, obscures true quantitative differences and precludes biological experiments that rely on accurate fluorescence intensity measurements. In this paper, we seek to quantify the improvement in the quality of high-content screen readouts due to software-based illumination correction. We present a straightforward illumination correction pipeline that has been used by our group across many experiments. We test the pipeline on real-world high-throughput image sets and evaluate the performance of the pipeline at two levels: (a) Z'-factor to evaluate the effect of the image correction on a univariate readout, representative of a typical high-content screen, and (b) classification accuracy on phenotypic signatures derived from the images, representative of an experiment involving more complex data mining. We find that applying the proposed post-hoc correction method improves performance in both experiments, even when illumination correction has already been applied using software associated with the instrument. To facilitate the ready application and future development of illumination correction methods, we have made our complete test data sets as well as open-source image analysis pipelines publicly available. This software-based solution has the potential to improve outcomes for a wide-variety of image-based HTS experiments.

  20. Saturated two-photon excitation fluorescence microscopy with core-ring illumination.

    PubMed

    Oketani, Ryosuke; Doi, Atsushi; Smith, Nicholas I; Nawa, Yasunori; Kawata, Satoshi; Fujita, Katsumasa

    2017-02-01

    We demonstrated resolution improvement in two-photon excitation microscopy by combining saturated excitation (SAX) of fluorescence and pupil manipulation. We theoretically estimated the resolution improvement and the sidelobe effect in the point spread function with various pupil designs and found that the combination of SAX and core-ring illumination can effectively enhance the spatial resolution in 3D and suppress sidelobe artifacts. The experimental demonstration shows that the proposed technique is effective for observation with a depth of 100 μm in a tissue phantom and can be applied to 3D observations of tissue samples with higher spatial resolution than conventional two-photon excitation microscopy.

  1. Three-Dimensional Resolution Doubling in Wide-Field Fluorescence Microscopy by Structured Illumination

    PubMed Central

    Gustafsson, Mats G. L.; Shao, Lin; Carlton, Peter M.; Wang, C. J. Rachel; Golubovskaya, Inna N.; Cande, W. Zacheus; Agard, David A.; Sedat, John W.

    2008-01-01

    Structured illumination microscopy is a method that can increase the spatial resolution of wide-field fluorescence microscopy beyond its classical limit by using spatially structured illumination light. Here we describe how this method can be applied in three dimensions to double the axial as well as the lateral resolution, with true optical sectioning. A grating is used to generate three mutually coherent light beams, which interfere in the specimen to form an illumination pattern that varies both laterally and axially. The spatially structured excitation intensity causes normally unreachable high-resolution information to become encoded into the observed images through spatial frequency mixing. This new information is computationally extracted and used to generate a three-dimensional reconstruction with twice as high resolution, in all three dimensions, as is possible in a conventional wide-field microscope. The method has been demonstrated on both test objects and biological specimens, and has produced the first light microscopy images of the synaptonemal complex in which the lateral elements are clearly resolved. PMID:18326650

  2. Dynamic contrast enhancement in widefield microscopy using projector-generated illumination patterns

    NASA Astrophysics Data System (ADS)

    Carlo Samson, Edward; Mar Blanca, Carlo

    2007-10-01

    We present a simple and cost-effective optical protocol to realize contrast-enhancement imaging (such as dark-field, optical-staining and oblique illumination microscopy) of transparent samples on a conventional widefield microscope using commercial multimedia projectors. The projector functions as both light source and mask generator implemented by creating slideshows of the filters projected along the illumination planes of the microscope. The projected optical masks spatially modulate the distribution of the incident light to selectively enhance structures within the sample according to spatial frequency thereby increasing the image contrast of translucent biological specimens. Any amplitude filter can be customized and dynamically controlled so that switching from one imaging modality to another involves a simple slide transition and can be executed at a keystroke with no physical filters and no moving optical parts. The method yields an image contrast of 89 96% comparable with standard enhancement techniques. The polarization properties of the projector are then utilized to discriminate birefringent and non-birefringent sites on the sample using single-shot, simultaneous polarization and optical-staining microscopy. In addition to dynamic pattern generation and polarization, the projector also provides high illumination power and spectral excitation selectivity through its red-green-blue (RGB) channels. We exploit this last property to explore the feasibility of using video projectors to selectively excite stained samples and perform fluorescence imaging in tandem with reflectance and polarization reflectance microscopy.

  3. A review of RGB-LED based mixed-color illumination system for machine vision and microscopy

    NASA Astrophysics Data System (ADS)

    Hou, Lexin; Wang, Hexin; Xu, Min

    2016-09-01

    The theory and application of RGB-LED based mixed-color illumination system for use in machine vision and optical microscopy systems are presented. For machine vision system, relationship of various color sources and output image sharpness is discussed. From the viewpoint of gray scale images, evaluation and optimization methods of optimal illumination for machine vision are concluded. The image quality under monochromatic and mixed color illumination is compared. For optical microscopy system, demand of light source is introduced and design thoughts of RGB-LED based mixed-color illumination system are concluded. The problems need to be solved in this field are pointed out.

  4. Optically sectioned in vivo imaging with speckle illumination HiLo microscopy

    NASA Astrophysics Data System (ADS)

    Lim, Daryl; Ford, Tim N.; Chu, Kengyeh K.; Mertz, Jerome

    2011-01-01

    We present a simple wide-field imaging technique, called HiLo microscopy, that is capable of producing optically sectioned images in real time, comparable in quality to confocal laser scanning microscopy. The technique is based on the fusion of two raw images, one acquired with speckle illumination and another with standard uniform illumination. The fusion can be numerically adjusted, using a single parameter, to produce optically sectioned images of varying thicknesses with the same raw data. Direct comparison between our HiLo microscope and a commercial confocal laser scanning microscope is made on the basis of sectioning strength and imaging performance. Specifically, we show that HiLo and confocal 3-D imaging of a GFP-labeled mouse brain hippocampus are comparable in quality. Moreover, HiLo microscopy is capable of faster, near video rate imaging over larger fields of view than attainable with standard confocal microscopes. The goal of this paper is to advertise the simplicity, robustness, and versatility of HiLo microscopy, which we highlight with in vivo imaging of common model organisms including planaria, C. elegans, and zebrafish.

  5. RNA and protein 3D structure modeling: similarities and differences.

    PubMed

    Rother, Kristian; Rother, Magdalena; Boniecki, Michał; Puton, Tomasz; Bujnicki, Janusz M

    2011-09-01

    In analogy to proteins, the function of RNA depends on its structure and dynamics, which are encoded in the linear sequence. While there are numerous methods for computational prediction of protein 3D structure from sequence, there have been very few such methods for RNA. This review discusses template-based and template-free approaches for macromolecular structure prediction, with special emphasis on comparison between the already tried-and-tested methods for protein structure modeling and the very recently developed "protein-like" modeling methods for RNA. We highlight analogies between many successful methods for modeling of these two types of biological macromolecules and argue that RNA 3D structure can be modeled using "protein-like" methodology. We also highlight the areas where the differences between RNA and proteins require the development of RNA-specific solutions.

  6. Differential interference contrast microscopy using light-emitting diode illumination in conjunction with dual optical traps.

    PubMed

    Battle, C; Lautscham, L; Schmidt, C F

    2013-05-01

    Differential interference contrast (DIC) microscopy is a common mode of biological light microscopy used to achieve maximal resolution and contrast with label-free, weakly absorbing specimens such as cells. Maintaining the polarization state of the illuminating light is essential for the technique, and this requirement can conflict with optical trapping. We describe how to optimize DIC imaging using a light-emitting diode illumination source in a microscope while integrating a dual optical trap into the set up. Every time a polarized light beam reflects off or transmits through a dichroic mirror in the beam path, its polarization state will change if it is not polarized exactly parallel (p) or perpendicular (s) to the plane of incidence. We observe wavelength-dependent optical rotation and depolarization effects in our illumination light upon reflection from/transmission through dichroic mirrors in the beam path, resulting in significant degradation of image quality. We describe a method to compensate for these effects by introducing quarter-waveplates and a laser clean-up filter into the imaging pathway. We show that this approach achieves a full recovery of image quality.

  7. Automated 3D structure composition for large RNAs

    PubMed Central

    Popenda, Mariusz; Szachniuk, Marta; Antczak, Maciej; Purzycka, Katarzyna J.; Lukasiak, Piotr; Bartol, Natalia; Blazewicz, Jacek; Adamiak, Ryszard W.

    2012-01-01

    Understanding the numerous functions that RNAs play in living cells depends critically on knowledge of their three-dimensional structure. Due to the difficulties in experimentally assessing structures of large RNAs, there is currently great demand for new high-resolution structure prediction methods. We present the novel method for the fully automated prediction of RNA 3D structures from a user-defined secondary structure. The concept is founded on the machine translation system. The translation engine operates on the RNA FRABASE database tailored to the dictionary relating the RNA secondary structure and tertiary structure elements. The translation algorithm is very fast. Initial 3D structure is composed in a range of seconds on a single processor. The method assures the prediction of large RNA 3D structures of high quality. Our approach needs neither structural templates nor RNA sequence alignment, required for comparative methods. This enables the building of unresolved yet native and artificial RNA structures. The method is implemented in a publicly available, user-friendly server RNAComposer. It works in an interactive mode and a batch mode. The batch mode is designed for large-scale modelling and accepts atomic distance restraints. Presently, the server is set to build RNA structures of up to 500 residues. PMID:22539264

  8. Automated 3D structure composition for large RNAs.

    PubMed

    Popenda, Mariusz; Szachniuk, Marta; Antczak, Maciej; Purzycka, Katarzyna J; Lukasiak, Piotr; Bartol, Natalia; Blazewicz, Jacek; Adamiak, Ryszard W

    2012-08-01

    Understanding the numerous functions that RNAs play in living cells depends critically on knowledge of their three-dimensional structure. Due to the difficulties in experimentally assessing structures of large RNAs, there is currently great demand for new high-resolution structure prediction methods. We present the novel method for the fully automated prediction of RNA 3D structures from a user-defined secondary structure. The concept is founded on the machine translation system. The translation engine operates on the RNA FRABASE database tailored to the dictionary relating the RNA secondary structure and tertiary structure elements. The translation algorithm is very fast. Initial 3D structure is composed in a range of seconds on a single processor. The method assures the prediction of large RNA 3D structures of high quality. Our approach needs neither structural templates nor RNA sequence alignment, required for comparative methods. This enables the building of unresolved yet native and artificial RNA structures. The method is implemented in a publicly available, user-friendly server RNAComposer. It works in an interactive mode and a batch mode. The batch mode is designed for large-scale modelling and accepts atomic distance restraints. Presently, the server is set to build RNA structures of up to 500 residues.

  9. Detection of single quantum dots in model organisms with sheet illumination microscopy

    SciTech Connect

    Friedrich, Mike; Nozadze, Revaz; Gan, Qiang; Zelman-Femiak, Monika; Ermolayev, Vladimir; Wagner, Toni U.; Harms, Gregory S.

    2009-12-18

    Single-molecule detection and tracking is important for observing biomolecule interactions in the microenvironment. Here we report selective plane illumination microscopy (SPIM) with single-molecule detection in living organisms, which enables fast imaging and single-molecule tracking and optical penetration beyond 300 {mu}m. We detected single nanocrystals in Drosophila larvae and zebrafish embryo. We also report our first tracking of single quantum dots during zebrafish development, which displays a transition from flow to confined motion prior to the blastula stage. The new SPIM setup represents a new technique, which enables fast single-molecule imaging and tracking in living systems.

  10. Computational illumination for real-time gigapixel phase microscopy (Conference Presentation)

    NASA Astrophysics Data System (ADS)

    Waller, Laura

    2016-03-01

    This talk will describe new source coding methods for rapid acquisition of high-resolution phase images across a very large field of view, using a modified commercial microscope with LED array illumination. Our methods are inspired by a hybrid combination of Fourier Ptychography and differential phase contrast imaging, combined with source multiplexing strategies for fast capture and significantly reduced data requirements. Such computational approaches to optical microscopy add significant new capabilities to commercial microscopes without much cost or hardware modification. We demonstrate our system on live samples in vivo, validating our results for unstained, label-free samples against other popular phase retrieval methods.

  11. Dynamic multimodal full-field optical coherence tomography and fluorescence structured illumination microscopy

    NASA Astrophysics Data System (ADS)

    Thouvenin, Olivier; Fink, Mathias; Boccara, Claude

    2017-02-01

    We report on the development of a configuration of a multimodal full-field optical coherence tomography (FF-OCT) and fluorescence microscope. Our system can simultaneously acquire FF-OCT and structured illumination microscopy images. Dynamic parallel evolution of tissue microstructures and biochemical environments can be visualized. We use high numerical aperture objectives to optimize the combination of the two modalities. We imaged the propagation of mechanical waves initiated by calcium waves in a heart wall to illustrate the interest of simultaneous recording of mechanical and biochemical information.

  12. Perfect optical vortex enhanced surface plasmon excitation for plasmonic structured illumination microscopy imaging

    NASA Astrophysics Data System (ADS)

    Zhang, Chonglei; Min, Changjun; Du, Luping; Yuan, X.-C.

    2016-05-01

    We propose an all-optical technique for plasmonic structured illumination microscopy (PSIM) with perfect optical vortex (POV). POV can improve the efficiency of the excitation of surface plasma and reduce the background noise of the excited fluorescence. The plasmonic standing wave patterns are excited by POV with fractional topological charges for accurate phase shift of {-2π/3, 0, and 2π/3}. The imaging resolution of less than 200 nm was produced. This PSIM technique is expected to be used as a wide field, super resolution imaging technique in dynamic biological imaging.

  13. Quantification of in vitro mesenchymal stem cell invasion into tumor spheroids using selective plane illumination microscopy

    NASA Astrophysics Data System (ADS)

    Rühland, Svenja; Wechselberger, Alexandra; Spitzweg, Christine; Huss, Ralf; Nelson, Peter J.; Harz, Hartmann

    2015-04-01

    Mesenchymal stem cell (MSC) homing and integration into tumors are under evaluation for clinical application. This approach requires the identification of conditions for optimal tumor invasion. We describe a tool for the in vitro comparison of parameters influencing invasion. Human MSC added to experimental tumor spheroids variably migrates toward the center of the structure. To determine MSC distribution inside the three-dimensional specimen, spatial analysis was performed using selective plane illumination microscopy. A standardized method to quantify and compare the invasion potential of variably treated MSC into experimental tumor environments allows efficient screening for optimizing conditions.

  14. Two-photon excitation improves multifocal structured illumination microscopy in thick scattering tissue

    PubMed Central

    Ingaramo, Maria; York, Andrew G.; Wawrzusin, Peter; Milberg, Oleg; Hong, Amy; Weigert, Roberto; Shroff, Hari; Patterson, George H.

    2014-01-01

    Multifocal structured illumination microscopy (MSIM) provides a twofold resolution enhancement beyond the diffraction limit at sample depths up to 50 µm, but scattered and out-of-focus light in thick samples degrades MSIM performance. Here we implement MSIM with a microlens array to enable efficient two-photon excitation. Two-photon MSIM gives resolution-doubled images with better sectioning and contrast in thick scattering samples such as Caenorhabditis elegans embryos, Drosophila melanogaster larval salivary glands, and mouse liver tissue. PMID:24706872

  15. Fast optical sectioning obtained by structured illumination microscopy using a digital mirror device.

    PubMed

    Xu, Dongli; Jiang, Tao; Li, Anan; Hu, Bihe; Feng, Zhao; Gong, Hui; Zeng, Shaoqun; Luo, Qingming

    2013-06-01

    High-throughput optical imaging is critical to obtain large-scale neural connectivity information of brain in neuroscience. Using a digital mirror device and a scientific complementary metal-oxide semiconductor camera, we report a significant speed improvement of structured illumination microscopy (SIM), which produces a maximum SIM net frame rate of 133 Hz. We perform three-dimensional (3-D) imaging of mouse brain slices at diffraction-limited resolution and demonstrate the fast 3-D imaging capability to a large sample with an imaging rate of 6.9×10(7)  pixel/s of our system, an order of magnitude faster than previously reported.

  16. Fast optical sectioning obtained by structured illumination microscopy using a digital mirror device

    NASA Astrophysics Data System (ADS)

    Xu, Dongli; Jiang, Tao; Li, Anan; Hu, Bihe; Feng, Zhao; Gong, Hui; Zeng, Shaoqun; Luo, Qingming

    2013-06-01

    High-throughput optical imaging is critical to obtain large-scale neural connectivity information of brain in neuroscience. Using a digital mirror device and a scientific complementary metal-oxide semiconductor camera, we report a significant speed improvement of structured illumination microscopy (SIM), which produces a maximum SIM net frame rate of 133 Hz. We perform three-dimensional (3-D) imaging of mouse brain slices at diffraction-limited resolution and demonstrate the fast 3-D imaging capability to a large sample with an imaging rate of 6.9 pixel/s of our system, an order of magnitude faster than previously reported.

  17. Partially coherent illumination in full-field interferometric synthetic aperture microscopy

    PubMed Central

    Marks, Daniel L.; Davis, Brynmor J.; Boppart, Stephen A.; Carney, P. Scott

    2010-01-01

    A model is developed for optical coherence tomography and interferometric synthetic aperture microscopy (ISAM) systems employing full-field frequency-scanned illumination with partial spatial coherence. This model is used to derive efficient ISAM inverse scattering algorithms that give diffraction-limited resolution in regions typically regarded as out of focus. Partial spatial coherence of the source is shown to have the advantage of mitigating multiple-scattering effects that can otherwise produce significant artifacts in full-field coherent imaging. PMID:19183692

  18. A Guide to Structured Illumination TIRF Microscopy at High Speed with Multiple Colors

    PubMed Central

    Young, Laurence J.; Ströhl, Florian; Kaminski, Clemens F.

    2016-01-01

    Optical super-resolution imaging with structured illumination microscopy (SIM) is a key technology for the visualization of processes at the molecular level in the chemical and biomedical sciences. Although commercial SIM systems are available, systems that are custom designed in the laboratory can outperform commercial systems, the latter typically designed for ease of use and general purpose applications, both in terms of imaging fidelity and speed. This article presents an in-depth guide to building a SIM system that uses total internal reflection (TIR) illumination and is capable of imaging at up to 10 Hz in three colors at a resolution reaching 100 nm. Due to the combination of SIM and TIRF, the system provides better image contrast than rival technologies. To achieve these specifications, several optical elements are used to enable automated control over the polarization state and spatial structure of the illumination light for all available excitation wavelengths. Full details on hardware implementation and control are given to achieve synchronization between excitation light pattern generation, wavelength, polarization state, and camera control with an emphasis on achieving maximum acquisition frame rate. A step-by-step protocol for system alignment and calibration is presented and the achievable resolution improvement is validated on ideal test samples. The capability for video-rate super-resolution imaging is demonstrated with living cells. PMID:27285848

  19. A Guide to Structured Illumination TIRF Microscopy at High Speed with Multiple Colors.

    PubMed

    Young, Laurence J; Ströhl, Florian; Kaminski, Clemens F

    2016-05-30

    Optical super-resolution imaging with structured illumination microscopy (SIM) is a key technology for the visualization of processes at the molecular level in the chemical and biomedical sciences. Although commercial SIM systems are available, systems that are custom designed in the laboratory can outperform commercial systems, the latter typically designed for ease of use and general purpose applications, both in terms of imaging fidelity and speed. This article presents an in-depth guide to building a SIM system that uses total internal reflection (TIR) illumination and is capable of imaging at up to 10 Hz in three colors at a resolution reaching 100 nm. Due to the combination of SIM and TIRF, the system provides better image contrast than rival technologies. To achieve these specifications, several optical elements are used to enable automated control over the polarization state and spatial structure of the illumination light for all available excitation wavelengths. Full details on hardware implementation and control are given to achieve synchronization between excitation light pattern generation, wavelength, polarization state, and camera control with an emphasis on achieving maximum acquisition frame rate. A step-by-step protocol for system alignment and calibration is presented and the achievable resolution improvement is validated on ideal test samples. The capability for video-rate super-resolution imaging is demonstrated with living cells.

  20. Programmable Colored Illumination Microscopy (PCIM): A practical and flexible optical staining approach for microscopic contrast enhancement

    NASA Astrophysics Data System (ADS)

    Zuo, Chao; Sun, Jiasong; Feng, Shijie; Hu, Yan; Chen, Qian

    2016-03-01

    Programmable colored illumination microscopy (PCIM) has been proposed as a flexible optical staining technique for microscopic contrast enhancement. In this method, we replace the condenser diaphragm of a conventional microscope with a programmable thin film transistor-liquid crystal display (TFT-LCD). By displaying different patterns on the LCD, numerous established imaging modalities can be realized, such as bright field, dark field, phase contrast, oblique illumination, and Rheinberg illuminations, which conventionally rely on intricate alterations in the respective microscope setups. Furthermore, the ease of modulating both the color and the intensity distribution at the aperture of the condenser opens the possibility to combine multiple microscopic techniques, or even realize completely new methods for optical color contrast staining, such as iridescent dark-field and iridescent phase-contrast imaging. The versatility and effectiveness of PCIM is demonstrated by imaging of several transparent colorless specimens, such as unstained lung cancer cells, diatom, textile fibers, and a cryosection of mouse kidney. Finally, the potentialities of PCIM for RGB-splitting imaging with stained samples are also explored by imaging stained red blood cells and a histological section.

  1. High-contrast single-particle tracking by selective focal plane illumination microscopy.

    PubMed

    Ritter, Jörg G; Veith, Roman; Siebrasse, Jan-Peter; Kubitscheck, Ulrich

    2008-05-12

    Wide-field single molecule microscopy is a versatile tool for analyzing dynamics and molecular interactions in biological systems. In extended three-dimensional systems, however, the method suffers from intrinsic out-of-focus fluorescence. We constructed a high-resolution selective plane illumination microscope (SPIM) to efficiently solve this problem. The instrument is an optical sectioning microscope featuring the high speed and high sensitivity of a video microscope. We present theoretical calculations and quantitative measurements of the illumination light sheet thickness yielding 1.7 microm (FWHM) at 543 nm, 2.0 microm at 633 nm, and a FWHM of the axial point spread function of 1.13 microm. A direct comparison of selective plane and epi-illumination of model samples with intrinsic background fluorescence illustrated the clear advantage of SPIM for such samples. Single fluorescent quantum dots in aqueous solution are readily visualized and tracked proving the suitability of our setup for the study of fast and dynamic processes in spatially extended biological specimens.

  2. Selective-plane illumination microscopy for high-content volumetric biological imaging

    NASA Astrophysics Data System (ADS)

    McGorty, Ryan; Huang, Bo

    2016-03-01

    Light-sheet microscopy, also named selective-plane illumination microscopy, enables optical sectioning with minimal light delivered to the sample. Therefore, it allows one to gather volumetric datasets of developing embryos and other light-sensitive samples over extended times. We have configured a light-sheet microscope that, unlike most previous designs, can image samples in formats compatible with high-content imaging. Our microscope can be used with multi-well plates or with microfluidic devices. In designing our optical system to accommodate these types of sample holders we encounter large optical aberrations. We counter these aberrations with both static optical components in the imaging path and with adaptive optics. Potential applications of this microscope include studying the development of a large number of embryos in parallel and over long times with subcellular resolution and doing high-throughput screens on organisms or cells where volumetric data is necessary.

  3. Dynamic structured illumination microscopy: Focused imaging and optical sectioning for moving objects

    NASA Astrophysics Data System (ADS)

    Krzewina, Leo G.; Kim, Myung K.

    2006-02-01

    Structured illumination microscopy (SIM) is a valuable tool for three-dimensional microscopy and has numerous applications in bioscience. Its success has been limited to static objects, though, as three sequential image acquisitions are required per final processed, focused image. To overcome this problem we have developed a multicolored grid which when used in tandem with a color camera is capable of performing SIM with just a single exposure. Images and movies demonstrating optical sectioning of three-dimensional objects are presented, and results of applying color SIM for wide-field focused imaging are compared to those of SIM. From computer modeling and analytical calculations a theoretical estimate of the maximum observable object velocity in both the lateral and axial directions is available, implying that the new method will be capable of imaging a variety of live objects. Sample images of the technique applied to lens paper and a pigeon feather are included to show both advantages and disadvantages of CSIM.

  4. Visualization of podocyte substructure with structured illumination microscopy (SIM): a new approach to nephrotic disease

    PubMed Central

    Pullman, James M.; Nylk, Jonathan; Campbell, Elaine C.; Gunn-Moore, Frank J.; Prystowsky, Michael B.; Dholakia, Kishan

    2016-01-01

    A detailed microscopic analysis of renal podocyte substructure is essential to understand and diagnose nephrotic kidney disease. Currently only time consuming electron microscopy (EM) can resolve this substructure. We used structured illumination microscopy (SIM) to examine frozen sections of renal biopsies stained with an immunofluorescence marker for podocin, a protein localized to the perimeter of the podocyte foot processes and compared them with EM in both normal and nephrotic disease biopsies. SIM images of normal glomeruli revealed curvilinear patterns of podocin densely covering capillary walls similar to podocyte foot processes seen by EM. Podocin staining of all nephrotic disease biopsies were significantly different than normal, corresponding to and better visualizing effaced foot processes seen by EM. The findings support the first potential use of SIM in the diagnosis of nephrotic disease. PMID:26977341

  5. Improved quantitative phase imaging in lensless microscopy by single-shot multi-wavelength illumination using a fast convergence algorithm.

    PubMed

    Sanz, Martín; Picazo-Bueno, José Angel; García, Javier; Micó, Vicente

    2015-08-10

    We report on a novel algorithm for high-resolution quantitative phase imaging in a new concept of lensless holographic microscope based on single-shot multi-wavelength illumination. This new microscope layout, reported by Noom et al. along the past year and named by us as MISHELF (initials incoming from Multi-Illumination Single-Holographic-Exposure Lensless Fresnel) microscopy, rises from the simultaneous illumination and recording of multiple diffraction patterns in the Fresnel domain. In combination with a novel and fast iterative phase retrieval algorithm, MISHELF microscopy is capable of high-resolution (micron range) phase-retrieved (twin image elimination) biological imaging of dynamic events. In this contribution, MISHELF microscopy is demonstrated through qualitative concept description, algorithm implementation, and experimental validation using both a synthetic object (resolution test target) and a biological sample (swine sperm sample) for the case of three (RGB) illumination wavelengths. The proposed method becomes in an alternative instrument improving the capabilities of existing lensless microscopes.

  6. Fast functional imaging of multiple brain regions in intact zebrafish larvae using selective plane illumination microscopy.

    PubMed

    Panier, Thomas; Romano, Sebastián A; Olive, Raphaël; Pietri, Thomas; Sumbre, Germán; Candelier, Raphaël; Debrégeas, Georges

    2013-01-01

    The optical transparency and the small dimensions of zebrafish at the larval stage make it a vertebrate model of choice for brain-wide in-vivo functional imaging. However, current point-scanning imaging techniques, such as two-photon or confocal microscopy, impose a strong limit on acquisition speed which in turn sets the number of neurons that can be simultaneously recorded. At 5 Hz, this number is of the order of one thousand, i.e., approximately 1-2% of the brain. Here we demonstrate that this limitation can be greatly overcome by using Selective-plane Illumination Microscopy (SPIM). Zebrafish larvae expressing the genetically encoded calcium indicator GCaMP3 were illuminated with a scanned laser sheet and imaged with a camera whose optical axis was oriented orthogonally to the illumination plane. This optical sectioning approach was shown to permit functional imaging of a very large fraction of the brain volume of 5-9-day-old larvae with single- or near single-cell resolution. The spontaneous activity of up to 5,000 neurons was recorded at 20 Hz for 20-60 min. By rapidly scanning the specimen in the axial direction, the activity of 25,000 individual neurons from 5 different z-planes (approximately 30% of the entire brain) could be simultaneously monitored at 4 Hz. Compared to point-scanning techniques, this imaging strategy thus yields a ≃20-fold increase in data throughput (number of recorded neurons times acquisition rate) without compromising the signal-to-noise ratio (SNR). The extended field of view offered by the SPIM method allowed us to directly identify large scale ensembles of neurons, spanning several brain regions, that displayed correlated activity and were thus likely to participate in common neural processes. The benefits and limitations of SPIM for functional imaging in zebrafish as well as future developments are briefly discussed.

  7. Structured illumination-based super-resolution optical microscopy for hemato- and cyto-pathology applications.

    PubMed

    Zhang, Tieqiao; Osborn, Samantha; Brandow, Chloe; Dwyre, Denis; Green, Ralph; Lane, Stephen; Wachsmann-Hogiu, Sebastian

    2013-01-01

    Structured illumination fluorescence microscopy utilizes interfering light and the moiré effect to enhance spatial resolution to about a half of that of conventional light microscopy, i.e. approximately 90 nm. In addition to the enhancement in the x and y directions, it also allows enhancement of resolution in the z- direction by the same factor of two (to approximately 220 nm), making it a powerful tool for 3-D morphology studies of fluorescently labeled cells or thin tissue sections. In this report, we applied this technique to several types of blood cells that are commonly seen in hematopathology. Compared with standard brightfield and ordinary fluorescence microscopy images, the 3-D morphology results clearly reveal the morphological features of different types of normal blood cells. We have also used this technique to evaluate morphologies of abnormal erythrocytes and compare them with those recorded on normal cells. The results give a very intuitive presentation of morphological structures of erythrocytes with great details. This research illustrates the potential of this technique to be used in hematology and cyto-pathology studies aimed at identifying nanometer-sized features that cannot be distinguished otherwise with conventional optical microscopy.

  8. Imaging dendritic spines of rat primary hippocampal neurons using structured illumination microscopy.

    PubMed

    Schouten, Marijn; De Luca, Giulia M R; Alatriste González, Diana K; de Jong, Babette E; Timmermans, Wendy; Xiong, Hui; Krugers, Harm; Manders, Erik M M; Fitzsimons, Carlos P

    2014-05-04

    Dendritic spines are protrusions emerging from the dendrite of a neuron and represent the primary postsynaptic targets of excitatory inputs in the brain. Technological advances have identified these structures as key elements in neuron connectivity and synaptic plasticity. The quantitative analysis of spine morphology using light microscopy remains an essential problem due to technical limitations associated with light's intrinsic refraction limit. Dendritic spines can be readily identified by confocal laser-scanning fluorescence microscopy. However, measuring subtle changes in the shape and size of spines is difficult because spine dimensions other than length are usually smaller than conventional optical resolution fixed by light microscopy's theoretical resolution limit of 200 nm. Several recently developed super resolution techniques have been used to image cellular structures smaller than the 200 nm, including dendritic spines. These techniques are based on classical far-field operations and therefore allow the use of existing sample preparation methods and to image beyond the surface of a specimen. Described here is a working protocol to apply super resolution structured illumination microscopy (SIM) to the imaging of dendritic spines in primary hippocampal neuron cultures. Possible applications of SIM overlap with those of confocal microscopy. However, the two techniques present different applicability. SIM offers higher effective lateral resolution, while confocal microscopy, due to the usage of a physical pinhole, achieves resolution improvement at the expense of removal of out of focus light. In this protocol, primary neurons are cultured on glass coverslips using a standard protocol, transfected with DNA plasmids encoding fluorescent proteins and imaged using SIM. The whole protocol described herein takes approximately 2 weeks, because dendritic spines are imaged after 16-17 days in vitro, when dendritic development is optimal. After completion of the

  9. A simple, straightforward correlative live-cell-imaging-structured-illumination-microscopy approach for studying organelle dynamics.

    PubMed

    Sherman, Shachar; Nachmias, Dikla; Elia, Natalie

    2015-09-01

    Most cellular organelles are highly dynamic and continuously undergo membrane fission and fusion to mediate their function. Documenting organelle dynamics under physiological conditions, therefore, requires high temporal resolution of the recording system. Concurrently, these structures are relatively small and determining their substructural organization is often impossible using conventional microscopy. Structured Illumination Microscopy (SIM) is a super resolution technique providing a two-fold increase in resolution. Importantly, SIM is versatile because it allows the use of any fluorescent dye or protein and, hence, is highly applicable for cell biology. However, similar to other SR techniques, the applicability of SIM to high-speed live cell imaging is limited. Here we present an easy, straightforward methodology for coupling of high-speed live cell recordings, using spinning disk (SD) microscopy, with SIM. Using this simple methodology, we are able to track individual mitochondrial membrane fission and fusion events in real time and to determine the network connectivity and substructural organization of the membrane at high resolution. Applying this methodology to other cellular organelles such as, ER, golgi, and cilia will no doubt contribute to our understanding of membrane dynamics in cells.

  10. Comparison of rotational imaging optical coherence tomography and selective plane illumination microscopy for embryonic study

    NASA Astrophysics Data System (ADS)

    Wu, Chen; Ran, Shihao; Le, Henry H.; Singh, Manmohan; Larina, Irina V.; Mayerich, David; Dickinson, Mary E.; Larin, Kirill V.

    2016-03-01

    The mouse is a common model for studying developmental diseases. Different optical techniques have been developed to investigate mouse embryos, but each has its own set of limitations and restrictions. In this study, we imaged the same E9.5 mouse embryo with rotational imaging Optical Coherence Tomography (RI-OCT) and Selective Plane Illumination Microscopy (SPIM), and compared the two techniques. Results demonstrate that both methods can provide images with micrometer-scale spatial resolution. The RI-OCT technique was developed to increase imaging depth of OCT by performing traditional OCT imaging at multiple sides and co-registering the images. In SPIM, optical sectioning is achieved by illuminating the sample with a sheet of light. In this study, the images acquired from both techniques are compared with each other to evaluate the benefits and drawbacks of each technique for embryonic imaging. Since 3D stacks can be obtained by SPIM from different angles by rotating the sample, it might be possible to build a hybrid setup of two imaging modalities to combine the advantages of each technique.

  11. SLM-based off-axis Fourier filtering in microscopy with white light illumination.

    PubMed

    Steiger, Ruth; Bernet, Stefan; Ritsch-Marte, Monika

    2012-07-02

    In various microscopy applications spatial light modulators (SLMs) are used as programmable Fourier filters to realize different optical contrast enhancement methods. It is often advantageous to use the SLM in off-axis configuration, where the filtered image wave is sent into the first diffraction order of a blazed grating superposed to the phase mask on the SLM. Because of dispersion this approach is, however, typically limited to spectrally narrowband illumination. Here we suggest a method involving a grating for pre-compensation, which allows one to use spectrally broadband (even thermal) light in SLM-based Fourier filtering. The proposed approach is demonstrated by multicolor imaging of amplitude and phase objects, such as a resolution target, onion epidermal cells and human epithelial cheek cells.

  12. Video-rate imaging of microcirculation with single-exposure oblique back-illumination microscopy

    PubMed Central

    Mertz, Jerome

    2013-01-01

    Abstract. Oblique back-illumination microscopy (OBM) is a new technique for simultaneous, independent measurements of phase gradients and absorption in thick scattering tissues based on widefield imaging. To date, OBM has been used with sequential camera exposures, which reduces temporal resolution, and can produce motion artifacts in dynamic samples. Here, a variation of OBM that allows single-exposure operation with wavelength multiplexing and image splitting with a Wollaston prism is introduced. Asymmetric anamorphic distortion induced by the prism is characterized and corrected in real time using a graphics-processing unit. To demonstrate the capacity of single-exposure OBM to perform artifact-free imaging of blood flow, video-rate movies of microcirculation in ovo in the chorioallantoic membrane of the developing chick are presented. Imaging is performed with a high-resolution rigid Hopkins lens suitable for endoscopy. PMID:23733023

  13. Video-rate imaging of microcirculation with single-exposure oblique back-illumination microscopy

    NASA Astrophysics Data System (ADS)

    Ford, Tim N.; Mertz, Jerome

    2013-06-01

    Oblique back-illumination microscopy (OBM) is a new technique for simultaneous, independent measurements of phase gradients and absorption in thick scattering tissues based on widefield imaging. To date, OBM has been used with sequential camera exposures, which reduces temporal resolution, and can produce motion artifacts in dynamic samples. Here, a variation of OBM that allows single-exposure operation with wavelength multiplexing and image splitting with a Wollaston prism is introduced. Asymmetric anamorphic distortion induced by the prism is characterized and corrected in real time using a graphics-processing unit. To demonstrate the capacity of single-exposure OBM to perform artifact-free imaging of blood flow, video-rate movies of microcirculation in ovo in the chorioallantoic membrane of the developing chick are presented. Imaging is performed with a high-resolution rigid Hopkins lens suitable for endoscopy.

  14. Dual-view plane illumination microscopy for rapid and spatially isotropic imaging

    PubMed Central

    Kumar, Abhishek; Wu, Yicong; Christensen, Ryan; Chandris, Panagiotis; Gandler, William; McCreedy, Evan; Bokinsky, Alexandra; Colón-Ramos, Daniel A; Bao, Zhirong; McAuliffe, Matthew; Rondeau, Gary; Shroff, Hari

    2015-01-01

    We describe the construction and use of a compact dual-view inverted selective plane illumination microscope (diSPIM) for time-lapse volumetric (4D) imaging of living samples at subcellular resolution. Our protocol enables a biologist with some prior microscopy experience to assemble a diSPIM from commercially available parts, to align optics and test system performance, to prepare samples, and to control hardware and data processing with our software. Unlike existing light sheet microscopy protocols, our method does not require the sample to be embedded in agarose; instead, samples are prepared conventionally on glass coverslips. Tissue culture cells and Caenorhabditis elegans embryos are used as examples in this protocol; successful implementation of the protocol results in isotropic resolution and acquisition speeds up to several volumes per s on these samples. Assembling and verifying diSPIM performance takes ~6 d, sample preparation and data acquisition take up to 5 d and postprocessing takes 3–8 h, depending on the size of the data. PMID:25299154

  15. Super-resolution imaging of plasmodesmata using three-dimensional structured illumination microscopy.

    PubMed

    Fitzgibbon, Jessica; Bell, Karen; King, Emma; Oparka, Karl

    2010-08-01

    We used three-dimensional structured illumination microscopy (3D-SIM) to obtain subdiffraction ("super-resolution") images of plasmodesmata (PD) expressing a green fluorescent protein-tagged viral movement protein (MP) in tobacco (Nicotiana tabacum). In leaf parenchyma cells, we were able to resolve individual components of PD (neck and central cavities) at twice the resolution of a confocal microscope. Within the phloem, MP-green fluorescent protein filaments extended outward from the specialized pore-PD that connect sieve elements (SEs) with their companion cells (CCs) along the tubular sieve element reticulum (SER). The SER was shown to interconnect individual pore-PD at the SE-CC interface. 3D-SIM resolved fine (less than 100 nm) endoplasmic reticulum threads running into individual pore-PD as well as strands that crossed sieve plate pores, structurally linking SEs within a file. Our data reveal that MP entering the SE from the CC may remain associated with the SER. Fluorescence recovery after photobleaching experiments revealed that this MP pool is relatively immobile compared with the membrane probe 3,3'-dihexyloxacarbocyanine iodide, suggesting that MP may become sequestered by the SER once it has entered the SE. The advent of 3D-SIM offers considerable potential in the subdiffraction imaging of plant cells, bridging an important gap between confocal and electron microscopy.

  16. Volumetric structured illumination microscopy enabled by tunable focus lens (Conference Presentation)

    NASA Astrophysics Data System (ADS)

    Hinsdale, Taylor; Malik, Bilal; Olsovsky, Cory; Jo, Javier A.; Maitland, Kristen C.

    2016-03-01

    We present a volumetric imaging method for biological tissue that is free of mechanically scanning components. The optical sectioning in the system is obtained by structured illumination microscopy (SIM) with the depth of focus being varied by the use of an electronic tunable-focus lens (ETL). The performance of the axial scanning mechanism was evaluated and characterized in conjunction with SIM to ensure volumetric images could be recorded and reconstructed without significant losses in optical section thickness and lateral resolution over the full desired scan range. It was demonstrated that sub-cellular image resolutions were obtainable in both microsphere films and in ex vivo oral mucosa, spanning multiple cell layers, without significant losses in image quality. The mechanism proposed here has the ability to be integrated into any wide-field microscopy system to convert it into a three-dimensional imaging platform without the need for axial scanning of the sample or imaging optics. The ability to axially scan independent of mechanical movement also provides the opportunity for the development of endoscopic systems which can create volumetric images of tissue in vivo.

  17. 3D structures of membrane proteins from genomic sequencing

    PubMed Central

    Hopf, Thomas A.; Colwell, Lucy J.; Sheridan, Robert; Rost, Burkhard; Sander, Chris; Marks, Debora S.

    2012-01-01

    Summary We show that amino acid co-variation in proteins, extracted from the evolutionary sequence record, can be used to fold transmembrane proteins. We use this technique to predict previously unknown, 3D structures for 11 transmembrane proteins (with up to 14 helices) from their sequences alone. The prediction method (EVfold_membrane), applies a maximum entropy approach to infer evolutionary co-variation in pairs of sequence positions within a protein family and then generates all-atom models with the derived pairwise distance constraints. We benchmark the approach with blinded, de novo computation of known transmembrane protein structures from 23 families, demonstrating unprecedented accuracy of the method for large transmembrane proteins. We show how the method can predict oligomerization, functional sites, and conformational changes in transmembrane proteins. With the rapid rise in large-scale sequencing, more accurate and more comprehensive information on evolutionary constraints can be decoded from genetic variation, greatly expanding the repertoire of transmembrane proteins amenable to modelling by this method. PMID:22579045

  18. Myosin filament 3D structure in mammalian cardiac muscle☆

    PubMed Central

    AL-Khayat, Hind A.; Morris, Edward P.; Kensler, Robert W.; Squire, John M.

    2008-01-01

    A number of cardiac myopathies (e.g. familial hypertrophic cardiomyopathy and dilated cardiomyopathy) are linked to mutations in cardiac muscle myosin filament proteins, including myosin and myosin binding protein C (MyBP-C). To understand the myopathies it is necessary to know the normal 3D structure of these filaments. We have carried out 3D single particle analysis of electron micrograph images of negatively stained isolated myosin filaments from rabbit cardiac muscle. Single filament images were aligned and divided into segments about 2 × 430 Å long, each of which was treated as an independent ‘particle’. The resulting 40 Å resolution 3D reconstruction showed both axial and azimuthal (no radial) myosin head perturbations within the 430 Å repeat, with successive crown rotations of approximately 60°, 60° and 0°, rather than the regular 40° for an unperturbed helix. However, it is shown that the projecting density peaks appear to start at low radius from origins closer to those expected for an unperturbed helical filament, and that the azimuthal perturbation especially increases with radius. The head arrangements in rabbit cardiac myosin filaments are very similar to those in fish skeletal muscle myosin filaments, suggesting a possible general structural theme for myosin filaments in all vertebrate striated muscles (skeletal and cardiac). PMID:18472277

  19. The 3D structure of Coronal Mass Ejections

    NASA Astrophysics Data System (ADS)

    Patsourakos, Spiros

    2016-07-01

    Coronal Mass Ejections (CMEs) represent one of the most powerful energy release phenomena in the entire solar system and are a major driver of space weather. Prior to 2006, our observational access to CMEs was limited to single viewpoint remote sensing observations in the inner/outer corona, and in-situ observations further away, e.g. at 1 AU. Taking all these factors together, turned out to be a major obstacle in our understanding and characterizing of the 3D structure and evolution of CMEs. The situation improved dramatically with the availability of multi-viewpoint imaging observations of CMEs, all way through from the Sun to 1 AU, from the STEREO mission since 2006, combined with observations from other missions (SOHO, Hinode, SDO, IRIS). With this talk we will discuss several key recent results in CME science resulting from the analysis of multi-viewpoint observations. This includes: (1) shape and structure; (2) kinematics and energetics; (3) trajectories, deflections and rotations; (4) arrival times and velocities at 1 AU; (5) magnetic field structure; (6) relationships with coronal and interplanetary shocks and solar energetic particles. The implications of these results in terms of CME theories and models will be also addressed. We will conclude with a discussion of important open issues in our understanding of CMEs and how these could be addressed with upcoming (Solar Orbiter, Solar Probe Plus) and under-study missions (e.g., L5).

  20. Synchronous multimodal combination of full-field OCT and structured illumination fluorescence microscopy (Conference Presentation)

    NASA Astrophysics Data System (ADS)

    Thouvenin, Olivier; Fink, Mathias; Boccara, Claude

    2016-03-01

    FF-OCT is a full field high transverse resolution version of temporal domain OCT. It acquires En-face images with an isotropic 3D submicronic resolution deep inside a biological tissue. It can access an optical contrast at a given depth, meaning that FF-OCT is sensitive to variations of optical index. FF-OCT can thus probe the microarchitecture of a tissue without label. However, Ff-OCT lacks of specific molecular contrast. On the contrary, Fluorescence microscopy can reveal labelled molecules with a very good specificity. Structured Illumination Microscopy (SIM) is a technique providing optical sectioning to fluorescence widefield microscopy. However, this technique can be complicated to implement in a tissue, and fails at providing environmental information. Therefore, combining FF-OCT and SIM has many advantages and adds a specific molecular contrast to a microarchitecture image of a biological sample. Combining FF-OCT and SIM has already been reported in the literature. Here, we report on the development of different way to combine FF-OCT and SIM. On the contrary to previously described setups, our setup enables the synchronous detection of both modalities. We believe this is important to access to dynamical events that take place in tissues. With such a technique, we are able to detect fast changes happening both in the environment, and in the behavior of a specific molecule. For now, we applied our technique to detect static structural information in the cornea. By the time of the conference, we expect to use our system to detect dynamical changes in a tissue.

  1. Laser direct writing 3D structures for microfluidic channels: flow meter and mixer

    NASA Astrophysics Data System (ADS)

    Lin, Chih-Lang; Liu, Yi-Jui; Lin, Zheng-Da; Wu, Bo-Long; Lee, Yi-Hsiung; Shin, Chow-Shing; Baldeck, Patrice L.

    2015-03-01

    The 3D laser direct-writing technology is aimed at the modeling of arbitrary three-dimensional (3D) complex microstructures by scanning a laser-focusing point along predetermined trajectories. Through the perspective technique, the details of designed 3D structures can be properly fabricated in a microchannel. This study introduces a direct reading flow meter and a 3D passive mixer fabricated by laser direct writing for microfluidic applications. The flow meter consists of two rod-shaped springs, a pillar, an anchor, and a wedge-shaped indicator, installed inside a microfluidic channel. The indicator is deflected by the flowing fluid while restrained by the spring to establish an equilibrium indication according to the flow rate. The measurement is readily carried out by optical microscopy observation. The 3D passive Archimedes-screw-shaped mixer is designed to disturb the laminar flow 3D direction for enhancing the mixing efficiency. The simulation results indicate that the screw provides 3D disturbance of streamlines in the microchannel. The mixing demonstration for fluids flowing in the micrchannel approximately agrees with the simulation result. Thanks to the advantage of the laser direct writing technology, this study performs the ingenious applications of 3D structures for microchannels.

  2. Rapid diagnostic imaging and pathologic evaluation of whole core biopsies at the point-of-care using structured illumination microscopy

    NASA Astrophysics Data System (ADS)

    Wang, Mei; Sholl, Andrew B.; Kimbrell, Hillary; Tulman, David B.; Elfer, Katherine N.; Brown, J. Quincy

    2015-07-01

    Video-rate structured illumination microscopy (VR-SIM) of fluorescently stained prostate biopsies is demonstrated as a potential tool for rapid diagnosis of prostate biopsies at the point of care. Images of entire biopsies at 1.3 micron lateral resolution are rendered in seconds, and pathologist review of the resulting images achieves 90% accuracy as compared to gold standard histopathology.

  3. Automated Microscopy: Macro Language Controlling a Confocal Microscope and its External Illumination: Adaptation for Photosynthetic Organisms.

    PubMed

    Steinbach, Gábor; Kaňa, Radek

    2016-04-01

    Photosynthesis research employs several biophysical methods, including the detection of fluorescence. Even though fluorescence is a key method to detect photosynthetic efficiency, it has not been applied/adapted to single-cell confocal microscopy measurements to examine photosynthetic microorganisms. Experiments with photosynthetic cells may require automation to perform a large number of measurements with different parameters, especially concerning light conditions. However, commercial microscopes support custom protocols (through Time Controller offered by Olympus or Experiment Designer offered by Zeiss) that are often unable to provide special set-ups and connection to external devices (e.g., for irradiation). Our new system combining an Arduino microcontroller with the Cell⊕Finder software was developed for controlling Olympus FV1000 and FV1200 confocal microscopes and the attached hardware modules. Our software/hardware solution offers (1) a text file-based macro language to control the imaging functions of the microscope; (2) programmable control of several external hardware devices (light sources, thermal controllers, actuators) during imaging via the Arduino microcontroller; (3) the Cell⊕Finder software with ergonomic user environment, a fast selection method for the biologically important cells and precise positioning feature that reduces unwanted bleaching of the cells by the scanning laser. Cell⊕Finder can be downloaded from http://www.alga.cz/cellfinder. The system was applied to study changes in fluorescence intensity in Synechocystis sp. PCC6803 cells under long-term illumination. Thus, we were able to describe the kinetics of phycobilisome decoupling. Microscopy data showed that phycobilisome decoupling appears slowly after long-term (>1 h) exposure to high light.

  4. Video-rate structured illumination microscopy (VR-SIM) for rapid assessment of fresh surgical margins

    NASA Astrophysics Data System (ADS)

    Tulman, David B.; Wang, Mei; Kimbrell, Hillary Z.; Sholl, Andrew B.; Elfer, Katherine N.; Schlichenmeyer, Tyler C.; Mandava, Sree; Lee, Benjamin R.; Lacey, Michelle; Brown, J. Quincy

    2015-07-01

    Intra-operative surgical margin assessment by pathology is labor-intensive and time-consuming and is not practically capable of sampling the entire specimen. Positive surgical margins (PSMs), or tumor extending to the surface of the excised specimen, are associated with increased tumor recurrence and are accepted as poor independent prognostic indicators. Considering the PSM rate is high for patients with prostate and kidney cancer, residual tumor following radical prostatectomy and partial nephrectomy remains a significant problem. To address the unmet clinical need for an imaging tool that can provide sub-cellular resolution images of large areas of excised surgical specimens in an intra-operative timeframe, we have developed a video rate structured illumination microscopy (VR-SIM) system. We conducted a clinical trial using VR-SIM to create gigapixel mosaics of entire margin surfaces for each specimen. In the ongoing study, 5 patients undergoing radical prostatectomy and 4 patients undergoing partial nephrectomy participated to have digital images of their surgical specimens reviewed in comparison to the pathology report. The surfaces of the intact, excised specimens were imaged in an appropriate timeframe and showed visualization of histopathologically relevant structures.

  5. Time-lapse contact microscopy of cell cultures based on non-coherent illumination

    NASA Astrophysics Data System (ADS)

    Gabriel, Marion; Balle, Dorothée; Bigault, Stéphanie; Pornin, Cyrille; Gétin, Stéphane; Perraut, François; Block, Marc R.; Chatelain, François; Picollet-D'Hahan, Nathalie; Gidrol, Xavier; Haguet, Vincent

    2015-10-01

    Video microscopy offers outstanding capabilities to investigate the dynamics of biological and pathological mechanisms in optimal culture conditions. Contact imaging is one of the simplest imaging architectures to digitally record images of cells due to the absence of any objective between the sample and the image sensor. However, in the framework of in-line holography, other optical components, e.g., an optical filter or a pinhole, are placed underneath the light source in order to illuminate the cells with a coherent or quasi-coherent incident light. In this study, we demonstrate that contact imaging with an incident light of both limited temporal and spatial coherences can be achieved with sufficiently high quality for most applications in cell biology, including monitoring of cell sedimentation, rolling, adhesion, spreading, proliferation, motility, death and detachment. Patterns of cells were recorded at various distances between 0 and 1000 μm from the pixel array of the image sensors. Cells in suspension, just deposited or at mitosis focalise light into photonic nanojets which can be visualised by contact imaging. Light refraction by cells significantly varies during the adhesion process, the cell cycle and among the cell population in connection with every modification in the tridimensional morphology of a cell.

  6. Time-lapse contact microscopy of cell cultures based on non-coherent illumination

    PubMed Central

    Gabriel, Marion; Balle, Dorothée; Bigault, Stéphanie; Pornin, Cyrille; Gétin, Stéphane; Perraut, François; Block, Marc R.; Chatelain, François; Picollet-D’hahan, Nathalie; Gidrol, Xavier; Haguet, Vincent

    2015-01-01

    Video microscopy offers outstanding capabilities to investigate the dynamics of biological and pathological mechanisms in optimal culture conditions. Contact imaging is one of the simplest imaging architectures to digitally record images of cells due to the absence of any objective between the sample and the image sensor. However, in the framework of in-line holography, other optical components, e.g., an optical filter or a pinhole, are placed underneath the light source in order to illuminate the cells with a coherent or quasi-coherent incident light. In this study, we demonstrate that contact imaging with an incident light of both limited temporal and spatial coherences can be achieved with sufficiently high quality for most applications in cell biology, including monitoring of cell sedimentation, rolling, adhesion, spreading, proliferation, motility, death and detachment. Patterns of cells were recorded at various distances between 0 and 1000 μm from the pixel array of the image sensors. Cells in suspension, just deposited or at mitosis focalise light into photonic nanojets which can be visualised by contact imaging. Light refraction by cells significantly varies during the adhesion process, the cell cycle and among the cell population in connection with every modification in the tridimensional morphology of a cell. PMID:26459014

  7. Protein 3D structure computed from evolutionary sequence variation.

    PubMed

    Marks, Debora S; Colwell, Lucy J; Sheridan, Robert; Hopf, Thomas A; Pagnani, Andrea; Zecchina, Riccardo; Sander, Chris

    2011-01-01

    The evolutionary trajectory of a protein through sequence space is constrained by its function. Collections of sequence homologs record the outcomes of millions of evolutionary experiments in which the protein evolves according to these constraints. Deciphering the evolutionary record held in these sequences and exploiting it for predictive and engineering purposes presents a formidable challenge. The potential benefit of solving this challenge is amplified by the advent of inexpensive high-throughput genomic sequencing.In this paper we ask whether we can infer evolutionary constraints from a set of sequence homologs of a protein. The challenge is to distinguish true co-evolution couplings from the noisy set of observed correlations. We address this challenge using a maximum entropy model of the protein sequence, constrained by the statistics of the multiple sequence alignment, to infer residue pair couplings. Surprisingly, we find that the strength of these inferred couplings is an excellent predictor of residue-residue proximity in folded structures. Indeed, the top-scoring residue couplings are sufficiently accurate and well-distributed to define the 3D protein fold with remarkable accuracy.We quantify this observation by computing, from sequence alone, all-atom 3D structures of fifteen test proteins from different fold classes, ranging in size from 50 to 260 residues, including a G-protein coupled receptor. These blinded inferences are de novo, i.e., they do not use homology modeling or sequence-similar fragments from known structures. The co-evolution signals provide sufficient information to determine accurate 3D protein structure to 2.7-4.8 Å C(α)-RMSD error relative to the observed structure, over at least two-thirds of the protein (method called EVfold, details at http://EVfold.org). This discovery provides insight into essential interactions constraining protein evolution and will facilitate a comprehensive survey of the universe of protein structures

  8. Programmable Illumination and High-Speed, Multi-Wavelength, Confocal Microscopy Using a Digital Micromirror

    PubMed Central

    Martial, Franck P.; Hartell, Nicholas A.

    2012-01-01

    Confocal microscopy is routinely used for high-resolution fluorescence imaging of biological specimens. Most standard confocal systems scan a laser across a specimen and collect emitted light passing through a single pinhole to produce an optical section of the sample. Sequential scanning on a point-by-point basis limits the speed of image acquisition and even the fastest commercial instruments struggle to resolve the temporal dynamics of rapid cellular events such as calcium signals. Various approaches have been introduced that increase the speed of confocal imaging. Nipkov disk microscopes, for example, use arrays of pinholes or slits on a spinning disk to achieve parallel scanning which significantly increases the speed of acquisition. Here we report the development of a microscope module that utilises a digital micromirror device as a spatial light modulator to provide programmable confocal optical sectioning with a single camera, at high spatial and axial resolution at speeds limited by the frame rate of the camera. The digital micromirror acts as a solid state Nipkov disk but with the added ability to change the pinholes size and separation and to control the light intensity on a mirror-by-mirror basis. The use of an arrangement of concave and convex mirrors in the emission pathway instead of lenses overcomes the astigmatism inherent with DMD devices, increases light collection efficiency and ensures image collection is achromatic so that images are perfectly aligned at different wavelengths. Combined with non-laser light sources, this allows low cost, high-speed, multi-wavelength image acquisition without the need for complex wavelength-dependent image alignment. The micromirror can also be used for programmable illumination allowing spatially defined photoactivation of fluorescent proteins. We demonstrate the use of this system for high-speed calcium imaging using both a single wavelength calcium indicator and a genetically encoded, ratiometric, calcium

  9. Live, video-rate super-resolution microscopy using structured illumination and rapid GPU-based parallel processing.

    PubMed

    Lefman, Jonathan; Scott, Keana; Stranick, Stephan

    2011-04-01

    Structured illumination fluorescence microscopy is a powerful super-resolution method that is capable of achieving a resolution below 100 nm. Each super-resolution image is computationally constructed from a set of differentially illuminated images. However, real-time application of structured illumination microscopy (SIM) has generally been limited due to the computational overhead needed to generate super-resolution images. Here, we have developed a real-time SIM system that incorporates graphic processing unit (GPU) based in-line parallel processing of raw/differentially illuminated images. By using GPU processing, the system has achieved a 90-fold increase in processing speed compared to performing equivalent operations on a multiprocessor computer--the total throughput of the system is limited by data acquisition speed, but not by image processing. Overall, more than 350 raw images (16-bit depth, 512 × 512 pixels) can be processed per second, resulting in a maximum frame rate of 39 super-resolution images per second. This ultrafast processing capability is used to provide immediate feedback of super-resolution images for real-time display. These developments are increasing the potential for sophisticated super-resolution imaging applications.

  10. Cross-Sectional Investigations on Epitaxial Silicon Solar Cells by Kelvin and Conducting Probe Atomic Force Microscopy: Effect of Illumination.

    PubMed

    Narchi, Paul; Alvarez, Jose; Chrétien, Pascal; Picardi, Gennaro; Cariou, Romain; Foldyna, Martin; Prod'homme, Patricia; Kleider, Jean-Paul; I Cabarrocas, Pere Roca

    2016-12-01

    Both surface photovoltage and photocurrent enable to assess the effect of visible light illumination on the electrical behavior of a solar cell. We report on photovoltage and photocurrent measurements with nanometer scale resolution performed on the cross section of an epitaxial crystalline silicon solar cell, using respectively Kelvin probe force microscopy and conducting probe atomic force microscopy. Even though two different setups are used, the scans were performed on locations within 100-μm distance in order to compare data from the same area and provide a consistent interpretation. In both measurements, modifications under illumination are observed in accordance with the theory of PIN junctions. Moreover, an unintentional doping during the deposition of the epitaxial silicon intrinsic layer in the solar cell is suggested from the comparison between photovoltage and photocurrent measurements.

  11. Out-of-focus background subtraction for fast structured illumination super-resolution microscopy of optically thick samples.

    PubMed

    Vermeulen, P; Zhan, H; Orieux, F; Olivo-Marin, J-C; Lenkei, Z; Loriette, V; Fragola, A

    2015-09-01

    We propose a structured illumination microscopy method to combine super resolution and optical sectioning in three-dimensional (3D) samples that allows the use of two-dimensional (2D) data processing. Indeed, obtaining super-resolution images of thick samples is a difficult task if low spatial frequencies are present in the in-focus section of the sample, as these frequencies have to be distinguished from the out-of-focus background. A rigorous treatment would require a 3D reconstruction of the whole sample using a 3D point spread function and a 3D stack of structured illumination data. The number of raw images required, 15 per optical section in this case, limits the rate at which high-resolution images can be obtained. We show that by a succession of two different treatments of structured illumination data we can estimate the contrast of the illumination pattern and remove the out-of-focus content from the raw images. After this cleaning step, we can obtain super-resolution images of optical sections in thick samples using a two-beam harmonic illumination pattern and a limited number of raw images. This two-step processing makes it possible to obtain super resolved optical sections in thick samples as fast as if the sample was two-dimensional.

  12. Dual multispectral and 3D structured light laparoscope

    NASA Astrophysics Data System (ADS)

    Clancy, Neil T.; Lin, Jianyu; Arya, Shobhit; Hanna, George B.; Elson, Daniel S.

    2015-03-01

    Intraoperative feedback on tissue function, such as blood volume and oxygenation would be useful to the surgeon in cases where current clinical practice relies on subjective measures, such as identification of ischaemic bowel or tissue viability during anastomosis formation. Also, tissue surface profiling may be used to detect and identify certain pathologies, as well as diagnosing aspects of tissue health such as gut motility. In this paper a dual modality laparoscopic system is presented that combines multispectral reflectance and 3D surface imaging. White light illumination from a xenon source is detected by a laparoscope-mounted fast filter wheel camera to assemble a multispectral image (MSI) cube. Surface shape is then calculated using a spectrally-encoded structured light (SL) pattern detected by the same camera and triangulated using an active stereo technique. Images of porcine small bowel were acquired during open surgery. Tissue reflectance spectra were acquired and blood volume was calculated at each spatial pixel across the bowel wall and mesentery. SL features were segmented and identified using a `normalised cut' algoritm and the colour vector of each spot. Using the 3D geometry defined by the camera coordinate system the multispectral data could be overlaid onto the surface mesh. Dual MSI and SL imaging has the potential to provide augmented views to the surgeon supplying diagnostic information related to blood supply health and organ function. Future work on this system will include filter optimisation to reduce noise in tissue optical property measurement, and minimise spot identification errors in the SL pattern.

  13. Open-source image reconstruction of super-resolution structured illumination microscopy data in ImageJ

    PubMed Central

    Müller, Marcel; Mönkemöller, Viola; Hennig, Simon; Hübner, Wolfgang; Huser, Thomas

    2016-01-01

    Super-resolved structured illumination microscopy (SR-SIM) is an important tool for fluorescence microscopy. SR-SIM microscopes perform multiple image acquisitions with varying illumination patterns, and reconstruct them to a super-resolved image. In its most frequent, linear implementation, SR-SIM doubles the spatial resolution. The reconstruction is performed numerically on the acquired wide-field image data, and thus relies on a software implementation of specific SR-SIM image reconstruction algorithms. We present fairSIM, an easy-to-use plugin that provides SR-SIM reconstructions for a wide range of SR-SIM platforms directly within ImageJ. For research groups developing their own implementations of super-resolution structured illumination microscopy, fairSIM takes away the hurdle of generating yet another implementation of the reconstruction algorithm. For users of commercial microscopes, it offers an additional, in-depth analysis option for their data independent of specific operating systems. As a modular, open-source solution, fairSIM can easily be adapted, automated and extended as the field of SR-SIM progresses. PMID:26996201

  14. Fast high-contrast imaging of animal development with scanned light sheet-based structured illumination microscopy

    PubMed Central

    Keller, Philipp J.; Schmidt, Annette D.; Santella, Anthony; Khairy, Khaled; Bao, Zhirong; Wittbrodt, Joachim; Stelzer, Ernst H.K.

    2015-01-01

    Recording light microscopic images of large, non-transparent specimens, such as developing multi-cellular organisms, is complicated by decreased contrast due to light scattering. Early zebrafish development can be captured by standard light sheet microscopy; however, new imaging strategies are required to obtain high-quality data of late development or of less transparent organisms. We combined Digital Scanned Laser Light Sheet Fluorescence Microscopy (DSLM) with incoherent structured illumination microscopy and created structured illumination patterns with continuously adjustable frequencies (DSLM-SI). Our method discriminates the specimen-related scattered background from signal fluorescence, thereby removing out-of-focus light and optimizing the contrast of in-focus structures. DSLM-SI provides rapid control of the illumination pattern, exceptional imaging quality and high imaging speeds. We performed long-term imaging of zebrafish development for 58 hours and fast multiple-view imaging of early Drosophila development. We reconstructed cell positions over time from the Drosophila DSLM-SI data and created a Fly Digital Embryo. PMID:20601950

  15. Pupil-segmentation-based adaptive optical microscopy with full-pupil illumination.

    PubMed

    Milkie, Daniel E; Betzig, Eric; Ji, Na

    2011-11-01

    Optical aberrations deteriorate the performance of microscopes. Adaptive optics can be used to improve imaging performance via wavefront shaping. Here, we demonstrate a pupil-segmentation based adaptive optical approach with full-pupil illumination. When implemented in a two-photon fluorescence microscope, it recovers diffraction-limited performance and improves imaging signal and resolution.

  16. Immobilization Techniques of Bacteria for Live Super-resolution Imaging Using Structured Illumination Microscopy.

    PubMed

    Bottomley, Amy L; Turnbull, Lynne; Whitchurch, Cynthia B; Harry, Elizabeth J

    2017-01-01

    Advancements in optical microscopy technology have allowed huge progression in the ability to understand protein structure and dynamics in live bacterial cells using fluorescence microscopy. Paramount to high-quality microscopy is good sample preparation to avoid bacterial cell movement that can result in motion blur during image acquisition. Here, we describe two techniques of sample preparation that reduce unwanted cell movement and are suitable for application to a number of bacterial species and imaging methods.

  17. Incoherent structured illumination improves optical sectioning and contrast in multiphoton super-resolution microscopy.

    PubMed

    Winter, Peter W; Chandris, Panagiotis; Fischer, Robert S; Wu, Yicong; Waterman, Clare M; Shroff, Hari

    2015-02-23

    Three-dimensional super-resolution imaging in thick, semi-transparent biological specimens is hindered by light scattering, which increases background and degrades both contrast and optical sectioning. We describe a simple method that mitigates these issues, improving image quality in our recently developed two-photon instant structured illumination microscope without requiring any hardware modifications to the instrument. By exciting the specimen with three laterally-structured, phase-shifted illumination patterns and post-processing the resulting images, we digitally remove both scattered and out-of-focus emissions that would otherwise contaminate our raw data. We demonstrate the improved performance of our approach in biological samples, including pollen grains, primary mouse aortic endothelial cells cultured in a three-dimensional collagen matrix and live tumor-like cell spheroids.

  18. Spatially isotropic four-dimensional imaging with dual-view plane illumination microscopy

    PubMed Central

    Wu, Yicong; Wawrzusin, Peter; Senseney, Justin; Fischer, Robert S; Christensen, Ryan; Santella, Anthony; York, Andrew G; Winter, Peter W; Waterman, Clare M; Bao, Zhirong; Colón-Ramos, Daniel A; McAuliffe, Matthew; Shroff, Hari

    2014-01-01

    Optimal four-dimensional imaging requires high spatial resolution in all dimensions, high speed and minimal photobleaching and damage. We developed a dual-view, plane illumination microscope with improved spatiotemporal resolution by switching illumination and detection between two perpendicular objectives in an alternating duty cycle. Computationally fusing the resulting volumetric views provides an isotropic resolution of 330 nm. As the sample is stationary and only two views are required, we achieve an imaging speed of 200 images/s (i.e., 0.5 s for a 50-plane volume). Unlike spinning-disk confocal or Bessel beam methods, which illuminate the sample outside the focal plane, we maintain high spatiotemporal resolution over hundreds of volumes with negligible photobleaching. To illustrate the ability of our method to study biological systems that require high-speed volumetric visualization and/or low photobleaching, we describe microtubule tracking in live cells, nuclear imaging over 14 h during nematode embryogenesis and imaging of neural wiring during Caenorhabditis elegans brain development over 5 h. PMID:24108093

  19. Low-resolution characterization of the 3D structure of the Euglena gracilis photoreceptor.

    PubMed

    Barsanti, Laura; Coltelli, Primo; Evangelista, Valtere; Passarelli, Vincenzo; Frassanito, Anna Maria; Vesentini, Nicoletta; Gualtieri, Paolo

    2008-10-24

    This paper deals with the first characterization of the structure of the photoreceptive organelle of the unicellular alga Euglena gracilis (Euglenophyta). This organelle has a three-dimensional organization consisting of up to 50 closely stacked membrane lamellae. Ionically induced unstacking of the photoreceptor lamellae revealed ordered arrays well suited to structural analysis by electron microscopy and image analysis, which ultimately yielded a low-resolution picture of the structure. Each lamella is formed by the photoreceptive membrane protein of the cell assembled within the membrane layer in a hexagonal lattice. The first order diffraction spots in the calculated Fourier transform reveals the presence of 6-fold symmetrized topography (better resolution about 90A). The 2D and 3D structural data are very similar with those recently published on proteorodopsin, a membrane protein used by marine bacterio-plankton as light-driven proton pump. In our opinion these similarity indicate that a photoreceptive protein belonging to the same superfamily of proteorodopsin could form the Euglena photoreceptor.

  20. Low-resolution characterization of the 3D structure of the Euglena gracilis photoreceptor

    SciTech Connect

    Barsanti, Laura; Coltelli, Primo; Evangelista, Valtere; Passarelli, Vincenzo; Frassanito, Anna Maria; Vesentini, Nicoletta; Gualtieri, Paolo

    2008-10-24

    This paper deals with the first characterization of the structure of the photoreceptive organelle of the unicellular alga Euglena gracilis (Euglenophyta). This organelle has a three-dimensional organization consisting of up to 50 closely stacked membrane lamellae. Ionically induced unstacking of the photoreceptor lamellae revealed ordered arrays well suited to structural analysis by electron microscopy and image analysis, which ultimately yielded a low-resolution picture of the structure. Each lamella is formed by the photoreceptive membrane protein of the cell assembled within the membrane layer in a hexagonal lattice. The first order diffraction spots in the calculated Fourier transform reveals the presence of 6-fold symmetrized topography (better resolution about 90 A). The 2D and 3D structural data are very similar with those recently published on proteorodopsin, a membrane protein used by marine bacterio-plankton as light-driven proton pump. In our opinion these similarity indicate that a photoreceptive protein belonging to the same superfamily of proteorodopsin could form the Euglena photoreceptor.

  1. Miniaturized selective plane illumination microscopy for high-contrast in vivo fluorescence imaging.

    PubMed

    Engelbrecht, Christoph J; Voigt, Fabian; Helmchen, Fritjof

    2010-05-01

    Light-sheet-based fluorescence imaging techniques rely on simultaneous excitation of a single optical plane and thus permit high-contrast optically sectioned imaging of extended tissue samples. Here, we introduce a miniaturized fiber-optic implementation of a selective plane-illumination microscope (miniSPIM). The excitation light was delivered through a single-mode optical fiber, and a light-sheet was created with a cylindrical gradient-index lens and a right-angle microprism. Fluorescence emission was collected orthogonally to the light-sheet through a gradient-index lens assembly and a coherent fiber bundle. The end face of the fiber bundle was imaged onto a charge-coupled device camera. The spatial resolutions of the miniSPIM were 3.2 microm laterally and 5.1 microm axially. Images of fluorescent beads and neurons in mouse neocortex exhibited superior axial resolution and contrast in the miniSPIM-mode compared to images recorded in epi-illumination mode. The miniSPIM may thus enable novel in vivo imaging approaches.

  2. Apparatus for Direct Optical Fiber Through-Lens Illumination of Microscopy or Observational Objects

    NASA Technical Reports Server (NTRS)

    Kadogawa, Hiroshi (Inventor)

    2001-01-01

    In one embodiment of the invention, a microscope or other observational apparatus, comprises a hollow tube, a lens mounted to the tube, a light source and at least one flexible optical fiber having an input end and an output end. The input end is positioned to receive light from the light source, and the output end is positioned within the tube so as to directly project light along a straight path to the lens to illuminate an object to be viewed. The path of projected light is uninterrupted and free of light deflecting elements. By passing the light through the lens, the light can be diffused or otherwise defocused to provide more uniform illumination across the surface of the object, increasing the quality of the image of the object seen by the viewer. The direct undeflected and uninterrupted projection of light, without change of direction, eliminates the need for light-deflecting elements, such as beam-splitters, mirrors, prisms, or the like, to direct the projected light towards the object.

  3. Phase retrieval using polychromatic illumination for transmission X-ray microscopy

    PubMed Central

    Liu, Yijin; Andrews, Joy C.; Wang, Junyue; Meirer, Florian; Zhu, Peiping; Wu, Ziyu; Pianetta, Piero

    2011-01-01

    An alternative method for quantitative phase retrieval in a transmission X-ray microscope system at sub-50-nm resolution is presented. As an alternative to moving the sample in the beam direction in order to analyze the propagation-introduced phase effect, we have illuminated the TXM using X-rays of different energy without any motor movement in the TXM system. Both theoretical analysis and experimental studies have confirmed the feasibility and the advantage of our method, because energy tuning can be performed with very high energy resolution using a double crystal monochromator at a synchrotron beam line, and there is zero motor error in TXM system in our approach. High-spatial-resolution phase retrieval is accomplished using the proposed method. PMID:21263593

  4. Efficient linear phase contrast in scanning transmission electron microscopy with matched illumination and detector interferometry

    NASA Astrophysics Data System (ADS)

    Ophus, Colin; Ciston, Jim; Pierce, Jordan; Harvey, Tyler R.; Chess, Jordan; McMorran, Benjamin J.; Czarnik, Cory; Rose, Harald H.; Ercius, Peter

    2016-02-01

    The ability to image light elements in soft matter at atomic resolution enables unprecedented insight into the structure and properties of molecular heterostructures and beam-sensitive nanomaterials. In this study, we introduce a scanning transmission electron microscopy technique combining a pre-specimen phase plate designed to produce a probe with structured phase with a high-speed direct electron detector to generate nearly linear contrast images with high efficiency. We demonstrate this method by using both experiment and simulation to simultaneously image the atomic-scale structure of weakly scattering amorphous carbon and strongly scattering gold nanoparticles. Our method demonstrates strong contrast for both materials, making it a promising candidate for structural determination of heterogeneous soft/hard matter samples even at low electron doses comparable to traditional phase-contrast transmission electron microscopy. Simulated images demonstrate the extension of this technique to the challenging problem of structural determination of biological material at the surface of inorganic crystals.

  5. Efficient linear phase contrast in scanning transmission electron microscopy with matched illumination and detector interferometry

    SciTech Connect

    Ophus, Colin; Ciston, Jim; Pierce, Jordan; Harvey, Tyler R.; Chess, Jordan; McMorran, Benjamin J.; Czarnik, Cory; Rose, Harald H.; Ercius, Peter

    2016-02-29

    The ability to image light elements in soft matter at atomic resolution enables unprecedented insight into the structure and properties of molecular heterostructures and beam-sensitive nanomaterials. In this study, we introduce a scanning transmission electron microscopy technique combining a pre-specimen phase plate designed to produce a probe with structured phase with a high-speed direct electron detector to generate nearly linear contrast images with high efficiency. We demonstrate this method by using both experiment and simulation to simultaneously image the atomic-scale structure of weakly scattering amorphous carbon and strongly scattering gold nanoparticles. Our method demonstrates strong contrast for both materials, making it a promising candidate for structural determination of heterogeneous soft/hard matter samples even at low electron doses comparable to traditional phase-contrast transmission electron microscopy. Ultimately, simulated images demonstrate the extension of this technique to the challenging problem of structural determination of biological material at the surface of inorganic crystals.

  6. Efficient linear phase contrast in scanning transmission electron microscopy with matched illumination and detector interferometry.

    PubMed

    Ophus, Colin; Ciston, Jim; Pierce, Jordan; Harvey, Tyler R; Chess, Jordan; McMorran, Benjamin J; Czarnik, Cory; Rose, Harald H; Ercius, Peter

    2016-02-29

    The ability to image light elements in soft matter at atomic resolution enables unprecedented insight into the structure and properties of molecular heterostructures and beam-sensitive nanomaterials. In this study, we introduce a scanning transmission electron microscopy technique combining a pre-specimen phase plate designed to produce a probe with structured phase with a high-speed direct electron detector to generate nearly linear contrast images with high efficiency. We demonstrate this method by using both experiment and simulation to simultaneously image the atomic-scale structure of weakly scattering amorphous carbon and strongly scattering gold nanoparticles. Our method demonstrates strong contrast for both materials, making it a promising candidate for structural determination of heterogeneous soft/hard matter samples even at low electron doses comparable to traditional phase-contrast transmission electron microscopy. Simulated images demonstrate the extension of this technique to the challenging problem of structural determination of biological material at the surface of inorganic crystals.

  7. Achieving magnification smaller than 1 in lensless microscopy by illumination with a convergent wavefront.

    PubMed

    Perraut, François; Doménès, Mickaël; Grateau, Henri; Josso, Quentin

    2016-11-15

    Lensless or lensfree microscopy is now available commercially. With these new microscopes, it is possible to record images in contact mode with a magnification of 1 or in holographic mode with a magnification larger than 1. In this Letter, we present an original setup that allows us to record the image of an object whose surface is larger than that of the image sensor without placing an optical component between the object and the image sensor.

  8. Lensless phase microscopy and diffraction tomography with multi-angle and multi-wavelength illuminations using a LED matrix.

    PubMed

    Zuo, Chao; Sun, Jiasong; Zhang, Jialin; Hu, Yan; Chen, Qian

    2015-06-01

    We demonstrate lensless quantitative phase microscopy and diffraction tomography based on a compact on-chip platform, using only a CMOS image sensor and a programmable color LED matrix. Based on the multi-wavelength phase retrieval and multi-angle illumination diffraction tomography, this platform offers high quality, depth resolved images with a lateral resolution of 3.72μm and an axial resolution of 5μm, across a wide field-of-view of 24mm2. We experimentally demonstrate the success of our method by imaging cheek cells, micro-beads, and fertilized eggs of Parascaris equorum. Such high-throughput and miniaturized imaging device can provide a cost-effective tool for telemedicine applications and point-of-care diagnostics in resource-limited environments.

  9. Fast volumetric imaging with patterned illumination via digital micro-mirror device-based temporal focusing multiphoton microscopy

    PubMed Central

    Chang, Chia-Yuan; Hu, Yvonne Yuling; Lin, Chun-Yu; Lin, Cheng-Han; Chang, Hsin-Yu; Tsai, Sheng-Feng; Lin, Tzu-Wei; Chen, Shean-Jen

    2016-01-01

    Temporal focusing multiphoton microscopy (TFMPM) has the advantage of area excitation in an axial confinement of only a few microns; hence, it can offer fast three-dimensional (3D) multiphoton imaging. Herein, fast volumetric imaging via a developed digital micromirror device (DMD)-based TFMPM has been realized through the synchronization of an electron multiplying charge-coupled device (EMCCD) with a dynamic piezoelectric stage for axial scanning. The volumetric imaging rate can achieve 30 volumes per second according to the EMCCD frame rate of more than 400 frames per second, which allows for the 3D Brownian motion of one-micron fluorescent beads to be spatially observed. Furthermore, it is demonstrated that the dynamic HiLo structural multiphoton microscope can reject background noise by way of the fast volumetric imaging with high-speed DMD patterned illumination. PMID:27231617

  10. 2PE-STED microscopy with a single Ti:sapphire laser for reduced illumination.

    PubMed

    Li, Qifeng; Wang, Yang; Chen, Da; Wu, Sherry S H

    2014-01-01

    We reported a new effective approach to carry out two-photon excitation stimulated emission depletion (2PE-STED) microscopy using a single Ti:sapphire laser system. With an acoustic-optic Bragg cell, the modulated-CW 2PE STED microscope had the benefits of both CW and pulse approaches: lower input power, simple optical scheme and no complicated synchronization. Additionally, it also took advantages of fluorescence yield increasing. The sub-diffraction-limit resolution was demonstrated using ATTO 425-tagged clathrin-coated vesicles.

  11. Efficient linear phase contrast in scanning transmission electron microscopy with matched illumination and detector interferometry

    DOE PAGES

    Ophus, Colin; Ciston, Jim; Pierce, Jordan; ...

    2016-02-29

    The ability to image light elements in soft matter at atomic resolution enables unprecedented insight into the structure and properties of molecular heterostructures and beam-sensitive nanomaterials. In this study, we introduce a scanning transmission electron microscopy technique combining a pre-specimen phase plate designed to produce a probe with structured phase with a high-speed direct electron detector to generate nearly linear contrast images with high efficiency. We demonstrate this method by using both experiment and simulation to simultaneously image the atomic-scale structure of weakly scattering amorphous carbon and strongly scattering gold nanoparticles. Our method demonstrates strong contrast for both materials, makingmore » it a promising candidate for structural determination of heterogeneous soft/hard matter samples even at low electron doses comparable to traditional phase-contrast transmission electron microscopy. Ultimately, simulated images demonstrate the extension of this technique to the challenging problem of structural determination of biological material at the surface of inorganic crystals.« less

  12. 3D structure of eukaryotic flagella/cilia by cryo-electron tomography.

    PubMed

    Ishikawa, Takashi

    2013-01-01

    Flagella/cilia are motile organelles with more than 400 proteins. To understand the mechanism of such complex systems, we need methods to describe molecular arrange-ments and conformations three-dimensionally in vivo. Cryo-electron tomography enabled us such a 3D structural analysis. Our group has been working on 3D structure of flagella/cilia using this method and revealed highly ordered and beautifully organized molecular arrangement. 3D structure gave us insights into the mechanism to gener-ate bending motion with well defined waveforms. In this review, I summarize our recent structural studies on fla-gella/cilia by cryo-electron tomography, mainly focusing on dynein microtubule-based ATPase motor proteins and the radial spoke, a regulatory protein complex.

  13. Cortical Actin Flow in T Cells Quantified by Spatio-temporal Image Correlation Spectroscopy of Structured Illumination Microscopy Data.

    PubMed

    Ashdown, George; Pandžić, Elvis; Cope, Andrew; Wiseman, Paul; Owen, Dylan

    2015-12-17

    Filamentous-actin plays a crucial role in a majority of cell processes including motility and, in immune cells, the formation of a key cell-cell interaction known as the immunological synapse. F-actin is also speculated to play a role in regulating molecular distributions at the membrane of cells including sub-membranous vesicle dynamics and protein clustering. While standard light microscope techniques allow generalized and diffraction-limited observations to be made, many cellular and molecular events including clustering and molecular flow occur in populations at length-scales far below the resolving power of standard light microscopy. By combining total internal reflection fluorescence with the super resolution imaging method structured illumination microscopy, the two-dimensional molecular flow of F-actin at the immune synapse of T cells was recorded. Spatio-temporal image correlation spectroscopy (STICS) was then applied, which generates quantifiable results in the form of velocity histograms and vector maps representing flow directionality and magnitude. This protocol describes the combination of super-resolution imaging and STICS techniques to generate flow vectors at sub-diffraction levels of detail. This technique was used to confirm an actin flow that is symmetrically retrograde and centripetal throughout the periphery of T cells upon synapse formation.

  14. Fluorescent probes and fluorescence (microscopy) techniques--illuminating biological and biomedical research.

    PubMed

    Drummen, Gregor P C

    2012-11-28

    Fluorescence, the absorption and re-emission of photons with longer wavelengths, is one of those amazing phenomena of Nature. Its discovery and utilization had, and still has, a major impact on biological and biomedical research, since it enables researchers not just to visualize normal physiological processes with high temporal and spatial resolution, to detect multiple signals concomitantly, to track single molecules in vivo, to replace radioactive assays when possible, but also to shed light on many pathobiological processes underpinning disease states, which would otherwise not be possible. Compounds that exhibit fluorescence are commonly called fluorochromes or fluorophores and one of these fluorescent molecules in particular has significantly enabled life science research to gain new insights in virtually all its sub-disciplines: Green Fluorescent Protein. Because fluorescent proteins are synthesized in vivo, integration of fluorescent detection methods into the biological system via genetic techniques now became feasible. Currently fluorescent proteins are available that virtually span the whole electromagnetic spectrum. Concomitantly, fluorescence imaging techniques were developed, and often progress in one field fueled innovation in the other. Impressively, the properties of fluorescence were utilized to develop new assays and imaging modalities, ranging from energy transfer to image molecular interactions to imaging beyond the diffraction limit with super-resolution microscopy. Here, an overview is provided of recent developments in both fluorescence imaging and fluorochrome engineering, which together constitute the “fluorescence toolbox” in life science research.

  15. Double-illumination photoacoustic microscopy of intestinal hemodynamics following massive small bowel resection

    NASA Astrophysics Data System (ADS)

    Yao, Junjie; Rowland, Kathryn J.; Wang, Lidai; Maslov, Konstantin I.; Warner, Brad W.; Wang, Lihong V.

    2012-02-01

    Massive small bowel resection (SBR) results in villus angiogenesis and intestinal adaptation. The exact mechanism that causes intestinal villus angiogenesis remains unknown. We hypothesize that hemodynamic changes within the remnant bowel after SBR will trigger intestinal angiogenesis. To validate this, we used photoacoustic microscopy (PAM) to image the microvascular system of the intestine in C57B6 mice and to measure blood flow and oxygen saturation (sO2) of a supplying artery and vein. Baseline measurements were made 6 cm proximal to the ileal-cecal junction (ICJ) prior to resection. A 50% proximal bowel resection was then performed, and measurements were again recorded at the same location immediately, 1, 3 and 7 days following resection. The results show that arterial and venous sO2 were similar prior to SBR. Immediately following SBR, the arterial and venous sO2 decreased by 14.3 +/- 2.7% and 32.7 +/- 6.6%, respectively, while the arterial and venous flow speed decreased by 62.9 +/- 17.3% and 60.0 +/- 20.1%, respectively. Such significant decreases in sO2 and blood flow indicate a hypoxic state after SBR. Within one week after SBR, both sO2 and blood flow speed had gradually recovered. By 7 days after SBR, arterial and venous sO2 had increased to 101.0 +/- 2.9% and 82.7 +/- 7.3% of the baseline values, respectively, while arterial and venous flow speed had increased to 106.0 +/- 21.4% and 150.0 +/- 29.6% of the baseline values, respectively. Such increases in sO2 and blood flow may result from angiogenesis following SBR.

  16. SimRNAweb: a web server for RNA 3D structure modeling with optional restraints.

    PubMed

    Magnus, Marcin; Boniecki, Michał J; Dawson, Wayne; Bujnicki, Janusz M

    2016-07-08

    RNA function in many biological processes depends on the formation of three-dimensional (3D) structures. However, RNA structure is difficult to determine experimentally, which has prompted the development of predictive computational methods. Here, we introduce a user-friendly online interface for modeling RNA 3D structures using SimRNA, a method that uses a coarse-grained representation of RNA molecules, utilizes the Monte Carlo method to sample the conformational space, and relies on a statistical potential to describe the interactions in the folding process. SimRNAweb makes SimRNA accessible to users who do not normally use high performance computational facilities or are unfamiliar with using the command line tools. The simplest input consists of an RNA sequence to fold RNA de novo. Alternatively, a user can provide a 3D structure in the PDB format, for instance a preliminary model built with some other technique, to jump-start the modeling close to the expected final outcome. The user can optionally provide secondary structure and distance restraints, and can freeze a part of the starting 3D structure. SimRNAweb can be used to model single RNA sequences and RNA-RNA complexes (up to 52 chains). The webserver is available at http://genesilico.pl/SimRNAweb.

  17. Predicting 3D Structure, Flexibility, and Stability of RNA Hairpins in Monovalent and Divalent Ion Solutions

    PubMed Central

    Shi, Ya-Zhou; Jin, Lei; Wang, Feng-Hua; Zhu, Xiao-Long; Tan, Zhi-Jie

    2015-01-01

    A full understanding of RNA-mediated biology would require the knowledge of three-dimensional (3D) structures, structural flexibility, and stability of RNAs. To predict RNA 3D structures and stability, we have previously proposed a three-bead coarse-grained predictive model with implicit salt/solvent potentials. In this study, we further develop the model by improving the implicit-salt electrostatic potential and including a sequence-dependent coaxial stacking potential to enable the model to simulate RNA 3D structure folding in divalent/monovalent ion solutions. The model presented here can predict 3D structures of RNA hairpins with bulges/internal loops (<77 nucleotides) from their sequences at the corresponding experimental ion conditions with an overall improved accuracy compared to the experimental data; the model also makes reliable predictions for the flexibility of RNA hairpins with bulge loops of different lengths at several divalent/monovalent ion conditions. In addition, the model successfully predicts the stability of RNA hairpins with various loops/stems in divalent/monovalent ion solutions. PMID:26682822

  18. All-atom 3D structure prediction of transmembrane β-barrel proteins from sequences

    PubMed Central

    Hayat, Sikander; Sander, Chris; Marks, Debora S.

    2015-01-01

    Transmembrane β-barrels (TMBs) carry out major functions in substrate transport and protein biogenesis but experimental determination of their 3D structure is challenging. Encouraged by successful de novo 3D structure prediction of globular and α-helical membrane proteins from sequence alignments alone, we developed an approach to predict the 3D structure of TMBs. The approach combines the maximum-entropy evolutionary coupling method for predicting residue contacts (EVfold) with a machine-learning approach (boctopus2) for predicting β-strands in the barrel. In a blinded test for 19 TMB proteins of known structure that have a sufficient number of diverse homologous sequences available, this combined method (EVfold_bb) predicts hydrogen-bonded residue pairs between adjacent β-strands at an accuracy of ∼70%. This accuracy is sufficient for the generation of all-atom 3D models. In the transmembrane barrel region, the average 3D structure accuracy [template-modeling (TM) score] of top-ranked models is 0.54 (ranging from 0.36 to 0.85), with a higher (44%) number of residue pairs in correct strand–strand registration than in earlier methods (18%). Although the nonbarrel regions are predicted less accurately overall, the evolutionary couplings identify some highly constrained loop residues and, for FecA protein, the barrel including the structure of a plug domain can be accurately modeled (TM score = 0.68). Lower prediction accuracy tends to be associated with insufficient sequence information and we therefore expect increasing numbers of β-barrel families to become accessible to accurate 3D structure prediction as the number of available sequences increases. PMID:25858953

  19. All-atom 3D structure prediction of transmembrane β-barrel proteins from sequences.

    PubMed

    Hayat, Sikander; Sander, Chris; Marks, Debora S; Elofsson, Arne

    2015-04-28

    Transmembrane β-barrels (TMBs) carry out major functions in substrate transport and protein biogenesis but experimental determination of their 3D structure is challenging. Encouraged by successful de novo 3D structure prediction of globular and α-helical membrane proteins from sequence alignments alone, we developed an approach to predict the 3D structure of TMBs. The approach combines the maximum-entropy evolutionary coupling method for predicting residue contacts (EVfold) with a machine-learning approach (boctopus2) for predicting β-strands in the barrel. In a blinded test for 19 TMB proteins of known structure that have a sufficient number of diverse homologous sequences available, this combined method (EVfold_bb) predicts hydrogen-bonded residue pairs between adjacent β-strands at an accuracy of ∼70%. This accuracy is sufficient for the generation of all-atom 3D models. In the transmembrane barrel region, the average 3D structure accuracy [template-modeling (TM) score] of top-ranked models is 0.54 (ranging from 0.36 to 0.85), with a higher (44%) number of residue pairs in correct strand-strand registration than in earlier methods (18%). Although the nonbarrel regions are predicted less accurately overall, the evolutionary couplings identify some highly constrained loop residues and, for FecA protein, the barrel including the structure of a plug domain can be accurately modeled (TM score = 0.68). Lower prediction accuracy tends to be associated with insufficient sequence information and we therefore expect increasing numbers of β-barrel families to become accessible to accurate 3D structure prediction as the number of available sequences increases.

  20. Photoconversion of Dye-Sensitized Solar Cells with a 3D-Structured Photoelectrode Consisting of Both TiO2 Nanofibers and Nanoparticles

    NASA Astrophysics Data System (ADS)

    Hwang, Tae-Hwan; Kim, Wan-Tae; Choi, Won-Youl

    2016-06-01

    In dye-sensitized solar cells, a three-dimensional (3-D)-structured photoelectrode of TiO2 nanofibers and nanoparticles was successfully fabricated by electro-spinning and screen-printing processes. Structures with one-dimensional nanofibers can be expected to improve the charge transport in a photoelectrode. The microstructure and crystalline structure were observed by field-emission scanning electron microscopy and with an x-ray diffractometer, respectively. The particle size of the TiO2 particles and the diameters of the TiO2 nanofiber in the 3-D-structured photoelectrode were ~30 nm and ~500 nm, respectively. The total thickness of the TiO2 layer in the 3-D-structured photoelectrode, which is composed of a nanoparticle layer of ~12 μm and a nanofiber layer of ~8 μm, was ~20 μm. The crystalline, anatase phase was also determined. In these dye-sensitized solar cells with a 3-D-structured layer, a short-circuit current density of 12.36 mA/cm2, an open-circuit voltage of 0.74 V, a fill factor of 0.46, and an energy conversion efficiency of 4.18% were observed. These values are higher than those of dye-sensitized solar cells with a conventional TiO2 nanoparticle layer. The proposed 3-D-structured photoelectrode consisting of TiO2 nanofibers and nanoparticles can help improve the performance of commercial dye-sensitized solar cells.

  1. Multi-illumination Gabor holography recorded in a single camera snap-shot for high-resolution phase retrieval in digital in-line holographic microscopy

    NASA Astrophysics Data System (ADS)

    Sanz, Martin; Picazo-Bueno, Jose A.; Garcia, Javier; Micó, Vicente

    2015-05-01

    In this contribution we introduce MISHELF microscopy, a new concept and design of a lensless holographic microscope based on wavelength multiplexing, single hologram acquisition and digital image processing. The technique which name comes from Multi-Illumination Single-Holographic-Exposure Lensless Fresnel microscopy, is based on the simultaneous illumination and recording of three diffraction patterns in the Fresnel domain. In combination with a novel and fast iterative phase retrieval algorithm, MISHELF microscopy is capable of high-resolution (micron range) phase-retrieved (twin image elimination) biological imaging of dynamic events (video rate recording speed) since it avoids the time multiplexing needed for the in-line hologram sequence recording when using conventional phase-shifting or phase retrieval algorithms. MISHELF microscopy is validated using two different experimental layouts: one using RGB illumination and detection schemes and another using IRRB as illumination while keeping the RGB color camera as detection device. Preliminary experimental results are provided for both experimental layouts using a synthetic object (USAF resolution test target).

  2. Theoretical assessment of optical resolution enhancement and background fluorescence reduction by three-dimensional nonlinear structured illumination microscopy using stimulated emission depletion

    NASA Astrophysics Data System (ADS)

    Dake, Fumihiro

    2016-08-01

    Three-dimensional structured illumination microscopy (SIM) enlarges frequency cutoff laterally and axially by a factor of two, compared with conventional microscopy. However, its optical resolution is still fundamentally limited. It is necessary to introduce nonlinearity to enlarge frequency cutoff further. We propose three-dimensional nonlinear structured illumination microscopy based on stimulated emission depletion (STED) effect, which has a structured excitation pattern and a structured STED pattern, and both three-dimensional illumination patterns have the same lateral pitch and orientation. Theoretical analysis showed that nonlinearity induced by STED effect, which causes harmonics and contributes to enlarging frequency cutoff, depends on the phase difference between two structured illuminations and that the phase difference of π is the most efficient to increase nonlinearity. We also found that undesirable background fluorescence, which degenerates the contrast of structured pattern and limits the ability of SIM, can be reduced by our method. These results revealed that optical resolution improvement and background fluorescence reduction would be compatible. The feasibility study showed that our method will be realized with commercially available laser, having 3.5 times larger frequency cutoff compared with conventional microscopy.

  3. Gigapixel surface imaging of radical prostatectomy specimens for comprehensive detection of cancer-positive surgical margins using structured illumination microscopy

    PubMed Central

    Wang, Mei; Tulman, David B.; Sholl, Andrew B.; Kimbrell, Hillary Z.; Mandava, Sree H.; Elfer, Katherine N.; Luethy, Samuel; Maddox, Michael M.; Lai, Weil; Lee, Benjamin R.; Brown, J. Quincy

    2016-01-01

    Achieving cancer-free surgical margins in oncologic surgery is critical to reduce the need for additional adjuvant treatments and minimize tumor recurrence; however, there is a delicate balance between completeness of tumor removal and preservation of adjacent tissues critical for normal post-operative function. We sought to establish the feasibility of video-rate structured illumination microscopy (VR-SIM) of the intact removed tumor surface as a practical and non-destructive alternative to intra-operative frozen section pathology, using prostate cancer as an initial target. We present the first images of the intact human prostate surface obtained with pathologically-relevant contrast and subcellular detail, obtained in 24 radical prostatectomy specimens immediately after excision. We demonstrate that it is feasible to routinely image the full prostate circumference, generating gigapixel panorama images of the surface that are readily interpreted by pathologists. VR-SIM confirmed detection of positive surgical margins in 3 out of 4 prostates with pathology-confirmed adenocarcinoma at the circumferential surgical margin, and furthermore detected extensive residual cancer at the circumferential margin in a case post-operatively classified by histopathology as having negative surgical margins. Our results suggest that the increased surface coverage of VR-SIM could also provide added value for detection and characterization of positive surgical margins over traditional histopathology. PMID:27257084

  4. Gigapixel surface imaging of radical prostatectomy specimens for comprehensive detection of cancer-positive surgical margins using structured illumination microscopy

    NASA Astrophysics Data System (ADS)

    Wang, Mei; Tulman, David B.; Sholl, Andrew B.; Kimbrell, Hillary Z.; Mandava, Sree H.; Elfer, Katherine N.; Luethy, Samuel; Maddox, Michael M.; Lai, Weil; Lee, Benjamin R.; Brown, J. Quincy

    2016-06-01

    Achieving cancer-free surgical margins in oncologic surgery is critical to reduce the need for additional adjuvant treatments and minimize tumor recurrence; however, there is a delicate balance between completeness of tumor removal and preservation of adjacent tissues critical for normal post-operative function. We sought to establish the feasibility of video-rate structured illumination microscopy (VR-SIM) of the intact removed tumor surface as a practical and non-destructive alternative to intra-operative frozen section pathology, using prostate cancer as an initial target. We present the first images of the intact human prostate surface obtained with pathologically-relevant contrast and subcellular detail, obtained in 24 radical prostatectomy specimens immediately after excision. We demonstrate that it is feasible to routinely image the full prostate circumference, generating gigapixel panorama images of the surface that are readily interpreted by pathologists. VR-SIM confirmed detection of positive surgical margins in 3 out of 4 prostates with pathology-confirmed adenocarcinoma at the circumferential surgical margin, and furthermore detected extensive residual cancer at the circumferential margin in a case post-operatively classified by histopathology as having negative surgical margins. Our results suggest that the increased surface coverage of VR-SIM could also provide added value for detection and characterization of positive surgical margins over traditional histopathology.

  5. Two-photon instant structured illumination microscopy improves the depth penetration of super-resolution imaging in thick scattering samples.

    PubMed

    Winter, Peter W; York, Andrew G; Nogare, Damian Dalle; Ingaramo, Maria; Christensen, Ryan; Chitnis, Ajay; Patterson, George H; Shroff, Hari

    2014-09-20

    Fluorescence imaging methods that achieve spatial resolution beyond the diffraction limit (super-resolution) are of great interest in biology. We describe a super-resolution method that combines two-photon excitation with structured illumination microscopy (SIM), enabling three-dimensional interrogation of live organisms with ~150 nm lateral and ~400 nm axial resolution, at frame rates of ~1 Hz. By performing optical rather than digital processing operations to improve resolution, our microscope permits super-resolution imaging with no additional cost in acquisition time or phototoxicity relative to the point-scanning two-photon microscope upon which it is based. Our method provides better depth penetration and inherent optical sectioning than all previously reported super-resolution SIM implementations, enabling super-resolution imaging at depths exceeding 100 μm from the coverslip surface. The capability of our system for interrogating thick live specimens at high resolution is demonstrated by imaging whole nematode embryos and larvae, and tissues and organs inside zebrafish embryos.

  6. Analysis of the 3D Structure and Velocity of a CME on 2 January 2008

    NASA Astrophysics Data System (ADS)

    López, F. M.; Cremades, H.

    We perform an analysis of the 3D structure and velocity of a CME (coronal mass ejection) ejected on 2 January 2008. The event was imaged by both STEREO A and B spacecraft (mutual separation of ˜44°), providing polarized images of the event from two different points of view. To obtain information on the 3D structure of the CME from polarized images, a polarization technique (Moran & Davila, Science 305, 66, 2003) is applied. Aided by this method, we have constructed topographical maps which show the height of the various event features from the plane of the sky (i.e. toward or away from the observer) and have dinamically analyzed and compared the real and projected on the plane of the sky velocities.

  7. Linear-time protein 3-D structure searching with insertions and deletions

    PubMed Central

    2010-01-01

    Background Two biomolecular 3-D structures are said to be similar if the RMSD (root mean square deviation) between the two molecules' sequences of 3-D coordinates is less than or equal to some given constant bound. Tools for searching for similar structures in biomolecular 3-D structure databases are becoming increasingly important in the structural biology of the post-genomic era. Results We consider an important, fundamental problem of reporting all substructures in a 3-D structure database of chain molecules (such as proteins) which are similar to a given query 3-D structure, with consideration of indels (i.e., insertions and deletions). This problem has been believed to be very difficult but its exact computational complexity has not been known. In this paper, we first prove that the problem in unbounded dimensions is NP-hard. We then propose a new algorithm that dramatically improves the average-case time complexity of the problem in 3-D in case the number of indels k is bounded by a constant. Our algorithm solves the above problem for a query of size m and a database of size N in average-case O(N) time, whereas the time complexity of the previously best algorithm was O(Nmk+1). Conclusions Our results show that although the problem of searching for similar structures in a database based on the RMSD measure with indels is NP-hard in the case of unbounded dimensions, it can be solved in 3-D by a simple average-case linear time algorithm when the number of indels is bounded by a constant. PMID:20047663

  8. R3D-2-MSA: the RNA 3D structure-to-multiple sequence alignment server

    PubMed Central

    Cannone, Jamie J.; Sweeney, Blake A.; Petrov, Anton I.; Gutell, Robin R.; Zirbel, Craig L.; Leontis, Neocles

    2015-01-01

    The RNA 3D Structure-to-Multiple Sequence Alignment Server (R3D-2-MSA) is a new web service that seamlessly links RNA three-dimensional (3D) structures to high-quality RNA multiple sequence alignments (MSAs) from diverse biological sources. In this first release, R3D-2-MSA provides manual and programmatic access to curated, representative ribosomal RNA sequence alignments from bacterial, archaeal, eukaryal and organellar ribosomes, using nucleotide numbers from representative atomic-resolution 3D structures. A web-based front end is available for manual entry and an Application Program Interface for programmatic access. Users can specify up to five ranges of nucleotides and 50 nucleotide positions per range. The R3D-2-MSA server maps these ranges to the appropriate columns of the corresponding MSA and returns the contents of the columns, either for display in a web browser or in JSON format for subsequent programmatic use. The browser output page provides a 3D interactive display of the query, a full list of sequence variants with taxonomic information and a statistical summary of distinct sequence variants found. The output can be filtered and sorted in the browser. Previous user queries can be viewed at any time by resubmitting the output URL, which encodes the search and re-generates the results. The service is freely available with no login requirement at http://rna.bgsu.edu/r3d-2-msa. PMID:26048960

  9. Proteins without unique 3D structures: biotechnological applications of intrinsically unstable/disordered proteins.

    PubMed

    Uversky, Vladimir N

    2015-03-01

    Intrinsically disordered proteins (IDPs) and intrinsically disordered protein regions (IDPRs) are functional proteins or regions that do not have unique 3D structures under functional conditions. Therefore, from the viewpoint of their lack of stable 3D structure, IDPs/IDPRs are inherently unstable. As much as structure and function of normal ordered globular proteins are determined by their amino acid sequences, the lack of unique 3D structure in IDPs/IDPRs and their disorder-based functionality are also encoded in the amino acid sequences. Because of their specific sequence features and distinctive conformational behavior, these intrinsically unstable proteins or regions have several applications in biotechnology. This review introduces some of the most characteristic features of IDPs/IDPRs (such as peculiarities of amino acid sequences of these proteins and regions, their major structural features, and peculiar responses to changes in their environment) and describes how these features can be used in the biotechnology, for example for the proteome-wide analysis of the abundance of extended IDPs, for recombinant protein isolation and purification, as polypeptide nanoparticles for drug delivery, as solubilization tools, and as thermally sensitive carriers of active peptides and proteins.

  10. R3D-2-MSA: the RNA 3D structure-to-multiple sequence alignment server.

    PubMed

    Cannone, Jamie J; Sweeney, Blake A; Petrov, Anton I; Gutell, Robin R; Zirbel, Craig L; Leontis, Neocles

    2015-07-01

    The RNA 3D Structure-to-Multiple Sequence Alignment Server (R3D-2-MSA) is a new web service that seamlessly links RNA three-dimensional (3D) structures to high-quality RNA multiple sequence alignments (MSAs) from diverse biological sources. In this first release, R3D-2-MSA provides manual and programmatic access to curated, representative ribosomal RNA sequence alignments from bacterial, archaeal, eukaryal and organellar ribosomes, using nucleotide numbers from representative atomic-resolution 3D structures. A web-based front end is available for manual entry and an Application Program Interface for programmatic access. Users can specify up to five ranges of nucleotides and 50 nucleotide positions per range. The R3D-2-MSA server maps these ranges to the appropriate columns of the corresponding MSA and returns the contents of the columns, either for display in a web browser or in JSON format for subsequent programmatic use. The browser output page provides a 3D interactive display of the query, a full list of sequence variants with taxonomic information and a statistical summary of distinct sequence variants found. The output can be filtered and sorted in the browser. Previous user queries can be viewed at any time by resubmitting the output URL, which encodes the search and re-generates the results. The service is freely available with no login requirement at http://rna.bgsu.edu/r3d-2-msa.

  11. Rapid diagnostic imaging and pathologic evaluation of surgical tissue using video rate structured illumination microscopy (VR-SIM) (Conference Presentation)

    NASA Astrophysics Data System (ADS)

    Wang, Mei; Tulman, David; Elfer, Kate; Sholl, Andrew; Brown, J. Quincy

    2016-03-01

    Currently available pathology techniques for obtaining a rapid tissue diagnosis, or for determining the adequacy of specimens intended for downstream analysis, are too slow, labor-intensive, and destructive for point-of-care (POC) applications. We previously demonstrated video-rate structured illumination microscopy (VR-SIM) for accurate, high-throughput, non-destructive diagnostic imaging of fluorescently-stained prostate biopsies in seconds per biopsy, with an area under the ROC curve of 0.82-0.88 after pathologist review. In addition, we have demonstrated that it is feasible to use VR-SIM to routinely image very large gross pathology specimens, such as entire prostate resection surfaces, in relatively short timeframes at subcellular resolution. However, our prior work has focused on applications in prostate cancer; the utility in other organ sites has not been explored. Here we extended our technology to varying size kidney, liver, and lung biopsies. We conducted a validation study of VR-SIM against histopathology on a variety of human tissues, including both small biopsies and large slices of tissue. We conducted a blinded study in which the study pathologist accurately identified the organs based on VR-SIM images alone. The results were then used to create a clinical atlas between VR-SIM and H and E images for the different tissues of interest. This clinical atlas will be used to aid in pathologist interpretation in future POC clinical applications of VR-SIM in kidney, liver, and lung. Such applications could include on-site identification of the presence of kidney glomeruli for to ensure successful downstream IHC analysis, or determination of the adequacy of lung cancer biopsies for genomic analysis.

  12. Monte Carlo generators for studies of the 3D structure of the nucleon

    DOE PAGES

    Avakian, Harut; D'Alesio, U.; Murgia, F.

    2015-01-23

    In this study, extraction of transverse momentum and space distributions of partons from measurements of spin and azimuthal asymmetries requires development of a self consistent analysis framework, accounting for evolution effects, and allowing control of systematic uncertainties due to variations of input parameters and models. Development of realistic Monte-Carlo generators, accounting for TMD evolution effects, spin-orbit and quark-gluon correlations will be crucial for future studies of quark-gluon dynamics in general and 3D structure of the nucleon in particular.

  13. Studies of the 3D Structure of the Nucleon at Jlab

    SciTech Connect

    Avakian, Harut

    2016-07-01

    Studies of the 3D structure of the nucleon encoded in Transverse Momentum Dependent distribution and fragmentation functions of partons and Generalized Parton Distributions are among the key objectives of the JLab 12 GeV upgrade and the Electron Ion Collider. Main challenges in extracting 3D partonic distributions from precision measurements of hard scattering processes include clear understanding of leading twist QCD fundamentals, higher twist effects, and also correlations of hadron production in target and current fragmentation regions. In this contribution we discuss some ongoing studies and future measurements of spin-orbit correlations at Jefferson Lab.

  14. Simulation approach of atomic layer deposition in large 3D structures

    NASA Astrophysics Data System (ADS)

    Schwille, Matthias C.; Barth, Jonas; Schössler, Timo; Schön, Florian; Bartha, Johann W.; Oettel, Martin

    2017-04-01

    We present a new simulation method predicting thicknesses of thin films obtained by atomic layer deposition in high aspect ratio 3D geometries as they appear in MEMS manufacturing. The method features a Monte-Carlo computation of film deposition in free molecular flow, as well as in the Knudsen and diffusive gas regime, applicable for large structures. We compare our approach to analytic and simulation results from the literature. The capability of the method is demonstrated by a comparison to experimental film thicknesses in a large 3D structure. Finally, the feasability to extract process parameters, i.e. sticking coefficients is shown.

  15. Observed 3D Structure, Generation, and Dissipation of Oceanic Mesoscale Eddies in the South China Sea

    NASA Astrophysics Data System (ADS)

    Zhang, Zhiwei; Tian, Jiwei; Qiu, Bo; Zhao, Wei; Chang, Ping; Wu, Dexing; Wan, Xiuquan

    2016-04-01

    Oceanic mesoscale eddies with horizontal scales of 50–300 km are the most energetic form of flows in the ocean. They are the oceanic analogues of atmospheric storms and are effective transporters of heat, nutrients, dissolved carbon, and other biochemical materials in the ocean. Although oceanic eddies have been ubiquitously observed in the world oceans since 1960s, our understanding of their three-dimensional (3D) structure, generation, and dissipation remains fragmentary due to lack of systematic full water-depth measurements. To bridge this knowledge gap, we designed and conducted a multi-months field campaign, called the South China Sea Mesoscale Eddy Experiment (S-MEE), in the northern South China Sea in 2013/2014. The S-MEE for the first time captured full-depth 3D structures of an anticyclonic and cyclonic eddy pair, which are characterized by a distinct vertical tilt of their axes. By observing the eddy evolution at an upstream versus downstream location and conducting an eddy energy budget analysis, the authors further proposed that generation of submesoscale motions most likely constitutes the dominant dissipation mechanism for the observed eddies.

  16. Observed 3D Structure, Generation, and Dissipation of Oceanic Mesoscale Eddies in the South China Sea.

    PubMed

    Zhang, Zhiwei; Tian, Jiwei; Qiu, Bo; Zhao, Wei; Chang, Ping; Wu, Dexing; Wan, Xiuquan

    2016-04-14

    Oceanic mesoscale eddies with horizontal scales of 50-300 km are the most energetic form of flows in the ocean. They are the oceanic analogues of atmospheric storms and are effective transporters of heat, nutrients, dissolved carbon, and other biochemical materials in the ocean. Although oceanic eddies have been ubiquitously observed in the world oceans since 1960s, our understanding of their three-dimensional (3D) structure, generation, and dissipation remains fragmentary due to lack of systematic full water-depth measurements. To bridge this knowledge gap, we designed and conducted a multi-months field campaign, called the South China Sea Mesoscale Eddy Experiment (S-MEE), in the northern South China Sea in 2013/2014. The S-MEE for the first time captured full-depth 3D structures of an anticyclonic and cyclonic eddy pair, which are characterized by a distinct vertical tilt of their axes. By observing the eddy evolution at an upstream versus downstream location and conducting an eddy energy budget analysis, the authors further proposed that generation of submesoscale motions most likely constitutes the dominant dissipation mechanism for the observed eddies.

  17. [MOLECULAR EVOLUTION OF ION CHANNELS: AMINO ACID SEQUENCES AND 3D STRUCTURES].

    PubMed

    Korkosh, V S; Zhorov, B S; Tikhonov, D B

    2016-01-01

    An integral part of modern evolutionary biology is comparative analysis of structure and function of macromolecules such as proteins. The first and critical step to understand evolution of homologous proteins is their amino acid sequence alignment. However, standard algorithms fop not provide unambiguous sequence alignments for proteins of poor homology. More reliable results can be obtained by comparing experimental 3D structures obtained at atomic resolution, for instance, with the aid of X-ray structural analysis. If such structures are lacking, homology modeling is used, which may take into account indirect experimental data on functional roles of individual amino-acid residues. An important problem is that the sequence alignment, which reflects genetic modifications, does not necessarily correspond to the functional homology. The latter depends on three-dimensional structures which are critical for natural selection. Since alignment techniques relying only on the analysis of primary structures carry no information on the functional properties of proteins, including 3D structures into consideration is very important. Here we consider several examples involving ion channels and demonstrate that alignment of their three-dimensional structures can significantly improve sequence alignments obtained by traditional methods.

  18. Molecular Phylogeny and Predicted 3D Structure of Plant beta-D-N-Acetylhexosaminidase

    PubMed Central

    Hossain, Md. Anowar

    2014-01-01

    beta-D-N-Acetylhexosaminidase, a family 20 glycosyl hydrolase, catalyzes the removal of β-1,4-linked N-acetylhexosamine residues from oligosaccharides and their conjugates. We constructed phylogenetic tree of β-hexosaminidases to analyze the evolutionary history and predicted functions of plant hexosaminidases. Phylogenetic analysis reveals the complex history of evolution of plant β-hexosaminidase that can be described by gene duplication events. The 3D structure of tomato β-hexosaminidase (β-Hex-Sl) was predicted by homology modeling using 1now as a template. Structural conformity studies of the best fit model showed that more than 98% of the residues lie inside the favoured and allowed regions where only 0.9% lie in the unfavourable region. Predicted 3D structure contains 531 amino acids residues with glycosyl hydrolase20b domain-I and glycosyl hydrolase20 superfamily domain-II including the (β/α)8 barrel in the central part. The α and β contents of the modeled structure were found to be 33.3% and 12.2%, respectively. Eleven amino acids were found to be involved in ligand-binding site; Asp(330) and Glu(331) could play important roles in enzyme-catalyzed reactions. The predicted model provides a structural framework that can act as a guide to develop a hypothesis for β-Hex-Sl mutagenesis experiments for exploring the functions of this class of enzymes in plant kingdom. PMID:25165734

  19. Molecular phylogeny and predicted 3D structure of plant beta-D-N-acetylhexosaminidase.

    PubMed

    Hossain, Md Anowar; Roslan, Hairul Azman

    2014-01-01

    beta-D-N-Acetylhexosaminidase, a family 20 glycosyl hydrolase, catalyzes the removal of β-1,4-linked N-acetylhexosamine residues from oligosaccharides and their conjugates. We constructed phylogenetic tree of β-hexosaminidases to analyze the evolutionary history and predicted functions of plant hexosaminidases. Phylogenetic analysis reveals the complex history of evolution of plant β-hexosaminidase that can be described by gene duplication events. The 3D structure of tomato β-hexosaminidase (β-Hex-Sl) was predicted by homology modeling using 1now as a template. Structural conformity studies of the best fit model showed that more than 98% of the residues lie inside the favoured and allowed regions where only 0.9% lie in the unfavourable region. Predicted 3D structure contains 531 amino acids residues with glycosyl hydrolase20b domain-I and glycosyl hydrolase20 superfamily domain-II including the (β/α)8 barrel in the central part. The α and β contents of the modeled structure were found to be 33.3% and 12.2%, respectively. Eleven amino acids were found to be involved in ligand-binding site; Asp(330) and Glu(331) could play important roles in enzyme-catalyzed reactions. The predicted model provides a structural framework that can act as a guide to develop a hypothesis for β-Hex-Sl mutagenesis experiments for exploring the functions of this class of enzymes in plant kingdom.

  20. Learning the 3-D structure of objects from 2-D views depends on shape, not format

    PubMed Central

    Tian, Moqian; Yamins, Daniel; Grill-Spector, Kalanit

    2016-01-01

    Humans can learn to recognize new objects just from observing example views. However, it is unknown what structural information enables this learning. To address this question, we manipulated the amount of structural information given to subjects during unsupervised learning by varying the format of the trained views. We then tested how format affected participants' ability to discriminate similar objects across views that were rotated 90° apart. We found that, after training, participants' performance increased and generalized to new views in the same format. Surprisingly, the improvement was similar across line drawings, shape from shading, and shape from shading + stereo even though the latter two formats provide richer depth information compared to line drawings. In contrast, participants' improvement was significantly lower when training used silhouettes, suggesting that silhouettes do not have enough information to generate a robust 3-D structure. To test whether the learned object representations were format-specific or format-invariant, we examined if learning novel objects from example views transfers across formats. We found that learning objects from example line drawings transferred to shape from shading and vice versa. These results have important implications for theories of object recognition because they suggest that (a) learning the 3-D structure of objects does not require rich structural cues during training as long as shape information of internal and external features is provided and (b) learning generates shape-based object representations independent of the training format. PMID:27153196

  1. Observed 3D Structure, Generation, and Dissipation of Oceanic Mesoscale Eddies in the South China Sea

    PubMed Central

    Zhang, Zhiwei; Tian, Jiwei; Qiu, Bo; Zhao, Wei; Chang, Ping; Wu, Dexing; Wan, Xiuquan

    2016-01-01

    Oceanic mesoscale eddies with horizontal scales of 50–300 km are the most energetic form of flows in the ocean. They are the oceanic analogues of atmospheric storms and are effective transporters of heat, nutrients, dissolved carbon, and other biochemical materials in the ocean. Although oceanic eddies have been ubiquitously observed in the world oceans since 1960s, our understanding of their three-dimensional (3D) structure, generation, and dissipation remains fragmentary due to lack of systematic full water-depth measurements. To bridge this knowledge gap, we designed and conducted a multi-months field campaign, called the South China Sea Mesoscale Eddy Experiment (S-MEE), in the northern South China Sea in 2013/2014. The S-MEE for the first time captured full-depth 3D structures of an anticyclonic and cyclonic eddy pair, which are characterized by a distinct vertical tilt of their axes. By observing the eddy evolution at an upstream versus downstream location and conducting an eddy energy budget analysis, the authors further proposed that generation of submesoscale motions most likely constitutes the dominant dissipation mechanism for the observed eddies. PMID:27074710

  2. Functional classification of protein 3D structures from predicted local interaction sites.

    PubMed

    Parasuram, Ramya; Lee, Joslynn S; Yin, Pengcheng; Somarowthu, Srinivas; Ondrechen, Mary Jo

    2010-12-01

    A new approach to the functional classification of protein 3D structures is described with application to some examples from structural genomics. This approach is based on functional site prediction with THEMATICS and POOL. THEMATICS employs calculated electrostatic potentials of the query structure. POOL is a machine learning method that utilizes THEMATICS features and has been shown to predict accurate, precise, highly localized interaction sites. Extension to the functional classification of structural genomics proteins is now described. Predicted functionally important residues are structurally aligned with those of proteins with previously characterized biochemical functions. A 3D structure match at the predicted local functional site then serves as a more reliable predictor of biochemical function than an overall structure match. Annotation is confirmed for a structural genomics protein with the ribulose phosphate binding barrel (RPBB) fold. A putative glucoamylase from Bacteroides fragilis (PDB ID 3eu8) is shown to be in fact probably not a glucoamylase. Finally a structural genomics protein from Streptomyces coelicolor annotated as an enoyl-CoA hydratase (PDB ID 3g64) is shown to be misannotated. Its predicted active site does not match the well-characterized enoyl-CoA hydratases of similar structure but rather bears closer resemblance to those of a dehalogenase with similar fold.

  3. Generation of Multilayered 3D Structures of HepG2 Cells Using a Bio-printing Technique

    PubMed Central

    Jeon, Hyeryeon; Kang, Kyojin; Park, Su A; Kim, Wan Doo; Paik, Seung Sam; Lee, Sang-Hun; Jeong, Jaemin; Choi, Dongho

    2017-01-01

    Background/Aims Chronic liver disease is a major widespread cause of death, and whole liver transplantation is the only definitive treatment for patients with end-stage liver diseases. However, many problems, including donor shortage, surgical complications and cost, hinder their usage. Recently, tissue-engineering technology provided a potential breakthrough for solving these problems. Three-dimensional (3D) printing technology has been used to mimic tissues and organs suitable for transplantation, but applications for the liver have been rare. Methods A 3D bioprinting system was used to construct 3D printed hepatic structures using alginate. HepG2 cells were cultured on these 3D structures for 3 weeks and examined by fluorescence microscopy, histology and immunohistochemistry. The expression of liver-specific markers was quantified on days 1, 7, 14, and 21. Results The cells grew well on the alginate scaffold, and liver-specific gene expression increased. The cells grew more extensively in 3D culture than two-dimensional culture and exhibited better structural aspects of the liver, indicating that the 3D bioprinting method recapitulates the liver architecture. Conclusions The 3D bioprinting of hepatic structures appears feasible. This technology may become a major tool and provide a bridge between basic science and the clinical challenges for regenerative medicine of the liver. PMID:27559001

  4. Improved phase sensitivity in spectral domain phase microscopy using line-field illumination and self phase-referencing

    PubMed Central

    Yaqoob, Zahid; Choi, Wonshik; Oh, Seungeun; Lue, Niyom; Park, Yongkeun; Fang-Yen, Christopher; Dasari, Ramachandra R.; Badizadegan, Kamran; Feld, Michael S.

    2010-01-01

    We report a quantitative phase microscope based on spectral domain optical coherence tomography and line-field illumination. The line illumination allows self phase-referencing method to reject common-mode phase noise. The quantitative phase microscope also features a separate reference arm, permitting the use of high numerical aperture (NA > 1) microscope objectives for high resolution phase measurement at multiple points along the line of illumination. We demonstrate that the path-length sensitivity of the instrument can be as good as 41 pm/Hz, which makes it suitable for nanometer scale study of cell motility. We present the detection of natural motions of cell surface and two-dimensional surface profiling of a HeLa cell. PMID:19550464

  5. 3D structure and conductive thermal field of the Upper Rhine Graben

    NASA Astrophysics Data System (ADS)

    Freymark, Jessica; Sippel, Judith; Scheck-Wenderoth, Magdalena; Bär, Kristian; Stiller, Manfred; Fritsche, Johann-Gerhard; Kracht, Matthias

    2016-04-01

    The Upper Rhine Graben (URG) was formed as part of the European Cenozoic Rift System in a complex extensional setting. At present-day, it has a large socioeconomic relevance as it provides a great potential for geothermal energy production in Germany and France. For the utilisation of this energy resource it is crucial to understand the structure and the observed temperature anomalies in the rift basin. In the framework of the EU-funded "IMAGE" project (Integrated Methods for Advanced Geothermal Exploration), we apply a data-driven numerical modelling approach to quantify the processes and properties controlling the spatial distribution of subsurface temperatures. Typically, reservoir-scale numerical models are developed for predictions on the subsurface hydrothermal conditions and for reducing the risk of drilling non-productive geothermal wells. One major problem related to such models is setting appropriate boundary conditions that define, for instance, how much heat enters the reservoir from greater depths. Therefore, we first build a regional lithospheric-scale 3D structural model, which covers not only the entire URG but also adjacent geological features like the Black Forest and the Vosges Mountains. In particular, we use a multidisciplinary dataset (e.g. well data, seismic reflection data, existing structural models, gravity) to construct the geometries of the sediments, the crust and the lithospheric mantle that control the spatial distribution of thermal conductivity and radiogenic heat production and hence temperatures. By applying a data-based and lithology-dependent parameterisation of this lithospheric-scale 3D structural model and a 3D finite element method, we calculate the steady-state conductive thermal field for the entire region. Available measured temperatures (down to depths of up to 5 km) are considered to validate the 3D thermal model. We present major characteristics of the lithospheric-scale 3D structural model and results of the 3D

  6. GARN: Sampling RNA 3D Structure Space with Game Theory and Knowledge-Based Scoring Strategies.

    PubMed

    Boudard, Mélanie; Bernauer, Julie; Barth, Dominique; Cohen, Johanne; Denise, Alain

    2015-01-01

    Cellular processes involve large numbers of RNA molecules. The functions of these RNA molecules and their binding to molecular machines are highly dependent on their 3D structures. One of the key challenges in RNA structure prediction and modeling is predicting the spatial arrangement of the various structural elements of RNA. As RNA folding is generally hierarchical, methods involving coarse-grained models hold great promise for this purpose. We present here a novel coarse-grained method for sampling, based on game theory and knowledge-based potentials. This strategy, GARN (Game Algorithm for RNa sampling), is often much faster than previously described techniques and generates large sets of solutions closely resembling the native structure. GARN is thus a suitable starting point for the molecular modeling of large RNAs, particularly those with experimental constraints. GARN is available from: http://garn.lri.fr/.

  7. Determination and validation of mTOR kinase-domain 3D structure by homology modeling.

    PubMed

    Lakhlili, Wiame; Chevé, Gwénaël; Yasri, Abdelaziz; Ibrahimi, Azeddine

    2015-01-01

    The AKT/mammalian target of rapamycin (mTOR) pathway is considered as one of the commonly activated and deregulated signaling pathways in human cancer. mTOR is associated with other proteins in two molecular complexes: mTOR complex 1/Raptor and the mTOR complex 2/Rictor. Using the crystal structure of the related lipid kinase PI3Kγ, we built a model of the catalytic region of mTOR. The modeling of the three-dimensional (3D) structure of the mTOR was performed by homology modeling program SWISS-MODEL. The quality and validation of the obtained model were performed using PROCHECK and PROVE softwares. The overall stereochemical property of the protein was assessed by the Ramachandran plot. The model validation was also done by docking of known inhibitors. In this paper, we describe and validate a 3D model for the mTOR catalytic site.

  8. 3D structure of eukaryotic flagella in a quiescent state revealed by cryo-electron tomography

    PubMed Central

    Nicastro, Daniela; McIntosh, J. Richard; Baumeister, Wolfgang

    2005-01-01

    We have used cryo-electron tomography to investigate the 3D structure and macromolecular organization of intact, frozen-hydrated sea urchin sperm flagella in a quiescent state. The tomographic reconstructions provide information at a resolution better than 6 nm about the in situ arrangements of macromolecules that are key for flagellar motility. We have visualized the heptameric rings of the motor domains in the outer dynein arm complex and determined that they lie parallel to the plane that contains the axes of neighboring flagellar microtubules. Both the material associated with the central pair of microtubules and the radial spokes display a plane of symmetry that helps to explain the planar beat pattern of these flagella. Cryo-electron tomography has proven to be a powerful technique for helping us understand the relationships between flagellar structure and function and the design of macromolecular machines in situ. PMID:16246999

  9. Optimal Image Stitching for Concrete Bridge Bottom Surfaces Aided by 3d Structure Lines

    NASA Astrophysics Data System (ADS)

    Liu, Yahui; Yao, Jian; Liu, Kang; Lu, Xiaohu; Xia, Menghan

    2016-06-01

    Crack detection for bridge bottom surfaces via remote sensing techniques is undergoing a revolution in the last few years. For such applications, a large amount of images, acquired with high-resolution industrial cameras close to the bottom surfaces with some mobile platform, are required to be stitched into a wide-view single composite image. The conventional idea of stitching a panorama with the affine model or the homographic model always suffers a series of serious problems due to poor texture and out-of-focus blurring introduced by depth of field. In this paper, we present a novel method to seamlessly stitch these images aided by 3D structure lines of bridge bottom surfaces, which are extracted from 3D camera data. First, we propose to initially align each image in geometry based on its rough position and orientation acquired with both a laser range finder (LRF) and a high-precision incremental encoder, and these images are divided into several groups with the rough position and orientation data. Secondly, the 3D structure lines of bridge bottom surfaces are extracted from the 3D cloud points acquired with 3D cameras, which impose additional strong constraints on geometrical alignment of structure lines in adjacent images to perform a position and orientation optimization in each group to increase the local consistency. Thirdly, a homographic refinement between groups is applied to increase the global consistency. Finally, we apply a multi-band blending algorithm to generate a large-view single composite image as seamlessly as possible, which greatly eliminates both the luminance differences and the color deviations between images and further conceals image parallax. Experimental results on a set of representative images acquired from real bridge bottom surfaces illustrate the superiority of our proposed approaches.

  10. Pigment analysis by Raman microscopy and portable X-ray fluorescence (pXRF) of thirteenth to fourteenth century illuminations and cuttings from Bologna

    NASA Astrophysics Data System (ADS)

    Chaplin, Tracey D.; Clark, Robin J. H.; Jones, Richard; Gibbs, Robert

    2016-12-01

    Non-destructive pigment analysis by Raman microscopy (RM) and portable X-ray fluorescence (pXRF) has been carried out on some Bolognese illuminations and cuttings chosen to represent the beginnings, evolution and height of Bolognese illuminated manuscript production. Dating to the thirteenth and fourteenth centuries and held in a private collection, the study provides evidence for the pigments generally used in this period. The results, which are compared with those obtained for other north Italian artwork, show the developments in usage of artistic materials and technique. Also addressed in this study is an examination of the respective roles of RM and pXRF analysis in this area of technical art history. This article is part of the themed issue "Raman spectroscopy in art and archaeology".

  11. A crust-scale 3D structural model of the Beaufort-Mackenzie Basin (Arctic Canada)

    NASA Astrophysics Data System (ADS)

    Sippel, Judith; Scheck-Wenderoth, Magdalena; Lewerenz, Björn; Kroeger, Karsten Friedrich

    2013-04-01

    The Beaufort-Mackenzie Basin was initiated in the Early Jurassic as part of an Arctic rifted passive continental margin which soon after became overprinted by Cordilleran foreland tectonics. Decades of industrial exploration and scientific research in this petroliferous region have produced a wide spectrum of geological and geophysical data as well as geoscientific knowledge. We have integrated available grids of sedimentary horizons, well data, seismic reflection and refraction data, and the observed regional gravity field into the first crust-scale 3D structural model of the Beaufort-Mackenzie Basin. Many characteristics of this model reflect the complex geodynamic and tectonostratigraphic history of the basin. The Mesozoic-Cenozoic sedimentary part of the model comprises seven clastic units (predominantly sandy shales) of which the modelled thickness distributions allow to retrace the well-established history of the basin comprising a gradual north(east)ward shift of the main depocentres as well as diverse phases of localised erosion. As a result of this development, the present-day configuration of the basin reveals that the sedimentary units tend to be younger, more porous, and thus less dense towards the north at a constant depth level. By integrating three refraction seismic profiles and performing combined isostatic and 3D gravity modelling, we have modelled the sub-sedimentary basement of the Beaufort-Mackenzie Basin. The continental basement spans from unstretched domains (as thick as about 42 km) in the south to extremely thinned domains (of less than 5 km thickness) in the north where it probably represents transitional crust attached to the oceanic crust of the Canada Basin. The uppermost parts of the continental crust are less dense (ρ = 2710 kg/m3) and most probably made up by pre-Mesozoic meta-sediments overlying a heavier igneous and metamorphic crust (ρ = 2850 kg/m3). The presented crust-scale 3D structural model shows that the greatest

  12. 3D structural fluctuation of IgG1 antibody revealed by individual particle electron tomography

    DOE PAGES

    Zhang, Xing; Zhang, Lei; Tong, Huimin; ...

    2015-05-05

    Commonly used methods for determining protein structure, including X-ray crystallography and single-particle reconstruction, often provide a single and unique three-dimensional (3D) structure. However, in these methods, the protein dynamics and flexibility/fluctuation remain mostly unknown. Here, we utilized advances in electron tomography (ET) to study the antibody flexibility and fluctuation through structural determination of individual antibody particles rather than averaging multiple antibody particles together. Through individual-particle electron tomography (IPET) 3D reconstruction from negatively-stained ET images, we obtained 120 ab-initio 3D density maps at an intermediate resolution (~1–3 nm) from 120 individual IgG1 antibody particles. Using these maps as a constraint, wemore » derived 120 conformations of the antibody via structural flexible docking of the crystal structure to these maps by targeted molecular dynamics simulations. Statistical analysis of the various conformations disclosed the antibody 3D conformational flexibility through the distribution of its domain distances and orientations. This blueprint approach, if extended to other flexible proteins, may serve as a useful methodology towards understanding protein dynamics and functions.« less

  13. 3D structural fluctuation of IgG1 antibody revealed by individual particle electron tomography

    SciTech Connect

    Zhang, Xing; Zhang, Lei; Tong, Huimin; Peng, Bo; Rames, Matthew J.; Zhang, Shengli; Ren, Gang

    2015-05-05

    Commonly used methods for determining protein structure, including X-ray crystallography and single-particle reconstruction, often provide a single and unique three-dimensional (3D) structure. However, in these methods, the protein dynamics and flexibility/fluctuation remain mostly unknown. Here, we utilized advances in electron tomography (ET) to study the antibody flexibility and fluctuation through structural determination of individual antibody particles rather than averaging multiple antibody particles together. Through individual-particle electron tomography (IPET) 3D reconstruction from negatively-stained ET images, we obtained 120 ab-initio 3D density maps at an intermediate resolution (~1–3 nm) from 120 individual IgG1 antibody particles. Using these maps as a constraint, we derived 120 conformations of the antibody via structural flexible docking of the crystal structure to these maps by targeted molecular dynamics simulations. Statistical analysis of the various conformations disclosed the antibody 3D conformational flexibility through the distribution of its domain distances and orientations. This blueprint approach, if extended to other flexible proteins, may serve as a useful methodology towards understanding protein dynamics and functions.

  14. Efficient global wave propagation adapted to 3-D structural complexity: a pseudospectral/spectral-element approach

    NASA Astrophysics Data System (ADS)

    Leng, Kuangdai; Nissen-Meyer, Tarje; van Driel, Martin

    2016-12-01

    We present a new, computationally efficient numerical method to simulate global seismic wave propagation in realistic 3-D Earth models. We characterize the azimuthal dependence of 3-D wavefields in terms of Fourier series, such that the 3-D equations of motion reduce to an algebraic system of coupled 2-D meridian equations, which is then solved by a 2-D spectral element method (SEM). Computational efficiency of such a hybrid method stems from lateral smoothness of 3-D Earth models and axial singularity of seismic point sources, which jointly confine the Fourier modes of wavefields to a few lower orders. We show novel benchmarks for global wave solutions in 3-D structures between our method and an independent, fully discretized 3-D SEM with remarkable agreement. Performance comparisons are carried out on three state-of-the-art tomography models, with seismic period ranging from 34 s down to 11 s. It turns out that our method has run up to two orders of magnitude faster than the 3-D SEM, featured by a computational advantage expanding with seismic frequency.

  15. Searching protein 3-D structures for optimal structure alignment using intelligent algorithms and data structures.

    PubMed

    Novosád, Tomáš; Snášel, Václav; Abraham, Ajith; Yang, Jack Y

    2010-11-01

    In this paper, we present a novel algorithm for measuring protein similarity based on their 3-D structure (protein tertiary structure). The algorithm used a suffix tree for discovering common parts of main chains of all proteins appearing in the current research collaboratory for structural bioinformatics protein data bank (PDB). By identifying these common parts, we build a vector model and use some classical information retrieval (IR) algorithms based on the vector model to measure the similarity between proteins--all to all protein similarity. For the calculation of protein similarity, we use term frequency × inverse document frequency ( tf × idf ) term weighing schema and cosine similarity measure. The goal of this paper is to introduce new protein similarity metric based on suffix trees and IR methods. Whole current PDB database was used to demonstrate very good time complexity of the algorithm as well as high precision. We have chosen the structural classification of proteins (SCOP) database for verification of the precision of our algorithm because it is maintained primarily by humans. The next success of this paper would be the ability to determine SCOP categories of proteins not included in the latest version of the SCOP database (v. 1.75) with nearly 100% precision.

  16. Function and 3D Structure of the N-Glycans on Glycoproteins

    PubMed Central

    Nagae, Masamichi; Yamaguchi, Yoshiki

    2012-01-01

    Glycosylation is one of the most common post-translational modifications in eukaryotic cells and plays important roles in many biological processes, such as the immune response and protein quality control systems. It has been notoriously difficult to study glycoproteins by X-ray crystallography since the glycan moieties usually have a heterogeneous chemical structure and conformation, and are often mobile. Nonetheless, recent technical advances in glycoprotein crystallography have accelerated the accumulation of 3D structural information. Statistical analysis of “snapshots” of glycoproteins can provide clues to understanding their structural and dynamic aspects. In this review, we provide an overview of crystallographic analyses of glycoproteins, in which electron density of the glycan moiety is clearly observed. These well-defined N-glycan structures are in most cases attributed to carbohydrate-protein and/or carbohydrate-carbohydrate interactions and may function as “molecular glue” to help stabilize inter- and intra-molecular interactions. However, the more mobile N-glycans on cell surface receptors, the electron density of which is usually missing on X-ray crystallography, seem to guide the partner ligand to its binding site and prevent irregular protein aggregation by covering oligomerization sites away from the ligand-binding site. PMID:22942711

  17. CamMedNP: Building the Cameroonian 3D structural natural products database for virtual screening

    PubMed Central

    2013-01-01

    Background Computer-aided drug design (CADD) often involves virtual screening (VS) of large compound datasets and the availability of such is vital for drug discovery protocols. We present CamMedNP - a new database beginning with more than 2,500 compounds of natural origin, along with some of their derivatives which were obtained through hemisynthesis. These are pure compounds which have been previously isolated and characterized using modern spectroscopic methods and published by several research teams spread across Cameroon. Description In the present study, 224 distinct medicinal plant species belonging to 55 plant families from the Cameroonian flora have been considered. About 80 % of these have been previously published and/or referenced in internationally recognized journals. For each compound, the optimized 3D structure, drug-like properties, plant source, collection site and currently known biological activities are given, as well as literature references. We have evaluated the “drug-likeness” of this database using Lipinski’s “Rule of Five”. A diversity analysis has been carried out in comparison with the ChemBridge diverse database. Conclusion CamMedNP could be highly useful for database screening and natural product lead generation programs. PMID:23590173

  18. 3D structure and dynamics of prominences in IRIS-Hinode collaborative observations

    NASA Astrophysics Data System (ADS)

    Okamoto, J.; De Pontieu, B.; Tarbell, T. D.; Title, A. M.

    2013-12-01

    A new solar physics satellite, Interface Region Imaging Spectrograph (IRIS), was launched on June 27, 2013. IRIS obtains UV spectra and images with high spatial resolution (0.33 arcsec) and high time cadence (1 sec / slit) of the chromosphere and transition region of the Sun. The chromosphere is located between the photosphere and the corona. Recently, the Hinode satellite has revealed that the chromosphere is highly active and suggested that it is a very important region in terms of energy deposit and transfer for coronal heating and solar wind acceleration. However, we cannot have further chromospheric information by Hinode because the Hinode Solar Optical Telescope has only a filtergraph for chromospheric observations. Now we have IRIS. IRIS performs spectroscopic observations to get the missing physical quantities of the dynamic chromosphere. Moreover, IRIS and Hinode is the most powerful collaboration to understand chromospheric activities. Hinode observes extremely high-cadence (1.6 sec) and high-spatial (0.2 arcsec) 2-D images, while IRIS measures line-of-sight velocity and Doppler width of fine structures with temperature dependence. This combination provides information about 3-D structures and dynamic phenomena of chromospheric features. Here we focus on prominence observations performed by IRIS and Hinode, and introduce the initial results of prominence dynamics and magnetic structures such as helical configurations, propagating waves and their damping mechanisms, and formation processes.

  19. Graphene originated 3D structures grown on the assembled nickel particles

    NASA Astrophysics Data System (ADS)

    Paronyan, Tereza; Harutyunyan, Avetik; Honda Research Institute USA Inc. Team

    2013-03-01

    Recently, the fabrication of various morphologies of graphene originated structures became very important due to the perspective of wide range of new applications. Particularly, free standing 3D structured graphene foams could be imperative in energy related areas . Here, we present the new approach of the CVD growth of 3D graphene network by using primarily sintered Ni particle's (~40 μm size) assembles as a template-catalyst via decomposition of low rate of CH4 at 1100° C based on synthesis method described earlier. SEM and Raman spectra analysis revealed the formation of graphene structure containing a single up to few layers grown on the sintered metal particles served as a catalyst-template. After etching the metal frame without using any support polymer, 3D free-standing graphene microporous structure was formed demonstrating high BET surface area. Two probe measurements of frame resistance were ~2-8 Ω. Our approach allows controllable tune the pore size and thereby the surface area of 3D graphene network through the variation of the template-catalyst particles size.

  20. Automatic Prediction of Protein 3D Structures by Probabilistic Multi-template Homology Modeling.

    PubMed

    Meier, Armin; Söding, Johannes

    2015-10-01

    Homology modeling predicts the 3D structure of a query protein based on the sequence alignment with one or more template proteins of known structure. Its great importance for biological research is owed to its speed, simplicity, reliability and wide applicability, covering more than half of the residues in protein sequence space. Although multiple templates have been shown to generally increase model quality over single templates, the information from multiple templates has so far been combined using empirically motivated, heuristic approaches. We present here a rigorous statistical framework for multi-template homology modeling. First, we find that the query proteins' atomic distance restraints can be accurately described by two-component Gaussian mixtures. This insight allowed us to apply the standard laws of probability theory to combine restraints from multiple templates. Second, we derive theoretically optimal weights to correct for the redundancy among related templates. Third, a heuristic template selection strategy is proposed. We improve the average GDT-ha model quality score by 11% over single template modeling and by 6.5% over a conventional multi-template approach on a set of 1000 query proteins. Robustness with respect to wrong constraints is likewise improved. We have integrated our multi-template modeling approach with the popular MODELLER homology modeling software in our free HHpred server http://toolkit.tuebingen.mpg.de/hhpred and also offer open source software for running MODELLER with the new restraints at https://bitbucket.org/soedinglab/hh-suite.

  1. 3D structures of individual mammalian genomes studied by single-cell Hi-C.

    PubMed

    Stevens, Tim J; Lando, David; Basu, Srinjan; Atkinson, Liam P; Cao, Yang; Lee, Steven F; Leeb, Martin; Wohlfahrt, Kai J; Boucher, Wayne; O'Shaughnessy-Kirwan, Aoife; Cramard, Julie; Faure, Andre J; Ralser, Meryem; Blanco, Enrique; Morey, Lluis; Sansó, Miriam; Palayret, Matthieu G S; Lehner, Ben; Di Croce, Luciano; Wutz, Anton; Hendrich, Brian; Klenerman, Dave; Laue, Ernest D

    2017-04-06

    The folding of genomic DNA from the beads-on-a-string-like structure of nucleosomes into higher-order assemblies is crucially linked to nuclear processes. Here we calculate 3D structures of entire mammalian genomes using data from a new chromosome conformation capture procedure that allows us to first image and then process single cells. The technique enables genome folding to be examined at a scale of less than 100 kb, and chromosome structures to be validated. The structures of individual topological-associated domains and loops vary substantially from cell to cell. By contrast, A and B compartments, lamina-associated domains and active enhancers and promoters are organized in a consistent way on a genome-wide basis in every cell, suggesting that they could drive chromosome and genome folding. By studying genes regulated by pluripotency factor and nucleosome remodelling deacetylase (NuRD), we illustrate how the determination of single-cell genome structure provides a new approach for investigating biological processes.

  2. Exploring 3D structural influences of aliphatic and aromatic chemicals on α-cyclodextrin binding.

    PubMed

    Linden, Lukas; Goss, Kai-Uwe; Endo, Satoshi

    2016-04-15

    Binding of solutes to macromolecules is often influenced by steric effects caused by the 3D structures of both binding partners. In this study, the 1:1 α-cyclodextrin (αCD) binding constants (Ka1) for 70 organic chemicals were determined to explore the solute-structural effects on the αCD binding. Ka1 was measured using a three-part partitioning system with either a headspace or a passive sampler serving as the reference phase. The Ka1 values ranged from 1.08 to 4.97 log units. The results show that longer linear aliphatic chemicals form more stable complexes than shorter ones, and that the position of the functional group has a strong influence on Ka1, even stronger than the type of the functional group. Comparison of linear and variously branched aliphatic chemicals indicates that having a sterically unhindered alkyl chain is favorable for binding. These results suggest that only one alkyl chain can enter the binding cavity. Relatively small aromatic chemicals such as 1,3-dichlorobenzene bind to αCD well, while larger ones like tetrachlorobenzene and 3-ring aromatic chemicals show only a weak interaction with αCD, which can be explained by cavity exclusion. The findings of this study help interpret cyclodextrin binding data and facilitate the understanding of binding processes to macromolecules.

  3. Computerized modeling techniques predict the 3D structure of H₄R: facts and fiction.

    PubMed

    Zaid, Hilal; Ismael-Shanak, Siba; Michaeli, Amit; Rayan, Anwar

    2012-01-01

    The functional characterization of proteins presents a daily challenge r biochemical, medical and computational sciences, especially when the structures are undetermined empirically, as in the case of the Histamine H4 Receptor (H₄R). H₄R is a member of the GPCR superfamily that plays a vital role in immune and inflammatory responses. To date, the concept of GPCRs modeling is highlighted in textbooks and pharmaceutical pamphlets, and this group of proteins has been the subject of almost 3500 publications in the scientific literature. The dynamic nature of determining the GPCRs structure was elucidated through elegant and creative modeling methodologies, implemented by many groups around the world. H₄R which belongs to the GPCR family was cloned in 2000; understandably, its biological activity was reported only 65 times in pubmed. Here we attempt to cover the fundamental concepts of H₄R structure modeling and its implementation in drug discovery, especially those that have been experimentally tested and to highlight some ideas that are currently being discussed on the dynamic nature of H₄R and GPCRs computerized techniques for 3D structure modeling.

  4. Qualification of a 3D structured light sensor for a reverse engineering application

    NASA Astrophysics Data System (ADS)

    Guarato, Alexandre Z.; Loja, Alexandre C.; Pereira, Leonardo P.; Braga, Sergio L.; Trevilato, Thales R. B.

    2016-11-01

    This paper deals with the qualification of a 3D structured light scanning system for an application of reverse engineering of a mechanical part. As this white light scanner is an electro-optical device and based on the principle of optical triangulation, the measurement accuracy is affected by the measured part geometry and its position within the scanning window. The effects of the scan depth and the projected angle, characterizing the surface normal of the measured surface to the scanning point of view, on the measurement of accuracy are not considered in the standard calibration process of manufacturers and have been identified by experiments in the present work. The digitization errors are analyzed and characterized thanks to a measurement protocol based on quality indicators. Theses quality indicators are evaluated thanks to simple calibrated artifacts. The aim of this work is to redefine the ideal relative distance and relative angle for minimizing the digitizing errors in relation to those stated by the manufacturer for a reverse engineering application.

  5. EDCs DataBank: 3D-Structure database of endocrine disrupting chemicals.

    PubMed

    Montes-Grajales, Diana; Olivero-Verbel, Jesus

    2015-01-02

    Endocrine disrupting chemicals (EDCs) are a group of compounds that affect the endocrine system, frequently found in everyday products and epidemiologically associated with several diseases. The purpose of this work was to develop EDCs DataBank, the only database of EDCs with three-dimensional structures. This database was built on MySQL using the EU list of potential endocrine disruptors and TEDX list. It contains the three-dimensional structures available on PubChem, as well as a wide variety of information from different databases and text mining tools, useful for almost any kind of research regarding EDCs. The web platform was developed employing HTML, CSS and PHP languages, with dynamic contents in a graphic environment, facilitating information analysis. Currently EDCs DataBank has 615 molecules, including pesticides, natural and industrial products, cosmetics, drugs and food additives, among other low molecular weight xenobiotics. Therefore, this database can be used to study the toxicological effects of these molecules, or to develop pharmaceuticals targeting hormone receptors, through docking studies, high-throughput virtual screening and ligand-protein interaction analysis. EDCs DataBank is totally user-friendly and the 3D-structures of the molecules can be downloaded in several formats. This database is freely available at http://edcs.unicartagena.edu.co.

  6. Cloning, Expression and 3D Structure Prediction of Chitinase from Chitinolyticbacter meiyuanensis SYBC-H1

    PubMed Central

    Hao, Zhikui; Wu, Hangui; Yang, Meiling; Chen, Jianjun; Xi, Limin; Zhao, Weijie; Yu, Jialin; Liu, Jiayang; Liao, Xiangru; Huang, Qingguo

    2016-01-01

    Two CHI genes from Chitinolyticbacter meiyuanensis SYBC-H1 encoding chitinases were identified and their protein 3D structures were predicted. According to the amino acid sequence alignment, CHI1 gene encoding 166 aa had a structural domain similar to the GH18 type II chitinase, and CHI2 gene encoding 383 aa had the same catalytic domain as the glycoside hydrolase family 19 chitinase. In this study, CHI2 chitinase were expressed in Escherichia coli BL21 cells, and this protein was purified by ammonium sulfate precipitation, DEAE-cellulose, and Sephadex G-100 chromatography. Optimal activity of CHI2 chitinase occurred at a temperature of 40 °C and a pH of 6.5. The presence of metal ions Fe3+, Fe2+, and Zn2+ inhibited CHI2 chitinase activity, while Na+ and K+ promoted its activity. Furthermore, the presence of EGTA, EDTA, and β-mercaptoethanol significantly increased the stability of CHI2 chitinase. The CHI2 chitinase was active with p-NP-GlcNAc, with the Km and Vm values of 23.0 µmol/L and 9.1 mM/min at a temperature of 37 °C, respectively. Additionally, the CHI2 chitinase was characterized as an N-acetyl glucosaminidase based on the hydrolysate from chitin. Overall, our results demonstrated CHI2 chitinase with remarkable biochemical properties is suitable for bioconversion of chitin waste. PMID:27240345

  7. Integration of 3D structure from disparity into biological motion perception independent of depth awareness.

    PubMed

    Wang, Ying; Jiang, Yi

    2014-01-01

    Images projected onto the retinas of our two eyes come from slightly different directions in the real world, constituting binocular disparity that serves as an important source for depth perception - the ability to see the world in three dimensions. It remains unclear whether the integration of disparity cues into visual perception depends on the conscious representation of stereoscopic depth. Here we report evidence that, even without inducing discernible perceptual representations, the disparity-defined depth information could still modulate the visual processing of 3D objects in depth-irrelevant aspects. Specifically, observers who could not discriminate disparity-defined in-depth facing orientations of biological motions (i.e., approaching vs. receding) due to an excessive perceptual bias nevertheless exhibited a robust perceptual asymmetry in response to the indistinguishable facing orientations, similar to those who could consciously discriminate such 3D information. These results clearly demonstrate that the visual processing of biological motion engages the disparity cues independent of observers' depth awareness. The extraction and utilization of binocular depth signals thus can be dissociable from the conscious representation of 3D structure in high-level visual perception.

  8. ProFunc: a server for predicting protein function from 3D structure.

    PubMed

    Laskowski, Roman A; Watson, James D; Thornton, Janet M

    2005-07-01

    ProFunc (http://www.ebi.ac.uk/thornton-srv/databases/ProFunc) is a web server for predicting the likely function of proteins whose 3D structure is known but whose function is not. Users submit the coordinates of their structure to the server in PDB format. ProFunc makes use of both existing and novel methods to analyse the protein's sequence and structure identifying functional motifs or close relationships to functionally characterized proteins. A summary of the analyses provides an at-a-glance view of what each of the different methods has found. More detailed results are available on separate pages. Often where one method has failed to find anything useful another may be more forthcoming. The server is likely to be of most use in structural genomics where a large proportion of the proteins whose structures are solved are of hypothetical proteins of unknown function. However, it may also find use in a comparative analysis of members of large protein families. It provides a convenient compendium of sequence and structural information that often hold vital functional clues to be followed up experimentally.

  9. Disulfide Connectivity Prediction Based on Modelled Protein 3D Structural Information and Random Forest Regression.

    PubMed

    Yu, Dong-Jun; Li, Yang; Hu, Jun; Yang, Xibei; Yang, Jing-Yu; Shen, Hong-Bin

    2015-01-01

    Disulfide connectivity is an important protein structural characteristic. Accurately predicting disulfide connectivity solely from protein sequence helps to improve the intrinsic understanding of protein structure and function, especially in the post-genome era where large volume of sequenced proteins without being functional annotated is quickly accumulated. In this study, a new feature extracted from the predicted protein 3D structural information is proposed and integrated with traditional features to form discriminative features. Based on the extracted features, a random forest regression model is performed to predict protein disulfide connectivity. We compare the proposed method with popular existing predictors by performing both cross-validation and independent validation tests on benchmark datasets. The experimental results demonstrate the superiority of the proposed method over existing predictors. We believe the superiority of the proposed method benefits from both the good discriminative capability of the newly developed features and the powerful modelling capability of the random forest. The web server implementation, called TargetDisulfide, and the benchmark datasets are freely available at: http://csbio.njust.edu.cn/bioinf/TargetDisulfide for academic use.

  10. LigandBox: A database for 3D structures of chemical compounds.

    PubMed

    Kawabata, Takeshi; Sugihara, Yusuke; Fukunishi, Yoshifumi; Nakamura, Haruki

    2013-01-01

    A database for the 3D structures of available compounds is essential for the virtual screening by molecular docking. We have developed the LigandBox database (http://ligandbox.protein.osaka-u.ac.jp/ligandbox/) containing four million available compounds, collected from the catalogues of 37 commercial suppliers, and approved drugs and biochemical compounds taken from KEGG_DRUG, KEGG_COMPOUND and PDB databases. Each chemical compound in the database has several 3D conformers with hydrogen atoms and atomic charges, which are ready to be docked into receptors using docking programs. The 3D conformations were generated using our molecular simulation program package, myPresto. Various physical properties, such as aqueous solubility (LogS) and carcinogenicity have also been calculated to characterize the ADME-Tox properties of the compounds. The Web database provides two services for compound searches: a property/chemical ID search and a chemical structure search. The chemical structure search is performed by a descriptor search and a maximum common substructure (MCS) search combination, using our program kcombu. By specifying a query chemical structure, users can find similar compounds among the millions of compounds in the database within a few minutes. Our database is expected to assist a wide range of researchers, in the fields of medical science, chemical biology, and biochemistry, who are seeking to discover active chemical compounds by the virtual screening.

  11. Partial order optimum likelihood (POOL): maximum likelihood prediction of protein active site residues using 3D Structure and sequence properties.

    PubMed

    Tong, Wenxu; Wei, Ying; Murga, Leonel F; Ondrechen, Mary Jo; Williams, Ronald J

    2009-01-01

    A new monotonicity-constrained maximum likelihood approach, called Partial Order Optimum Likelihood (POOL), is presented and applied to the problem of functional site prediction in protein 3D structures, an important current challenge in genomics. The input consists of electrostatic and geometric properties derived from the 3D structure of the query protein alone. Sequence-based conservation information, where available, may also be incorporated. Electrostatics features from THEMATICS are combined with multidimensional isotonic regression to form maximum likelihood estimates of probabilities that specific residues belong to an active site. This allows likelihood ranking of all ionizable residues in a given protein based on THEMATICS features. The corresponding ROC curves and statistical significance tests demonstrate that this method outperforms prior THEMATICS-based methods, which in turn have been shown previously to outperform other 3D-structure-based methods for identifying active site residues. Then it is shown that the addition of one simple geometric property, the size rank of the cleft in which a given residue is contained, yields improved performance. Extension of the method to include predictions of non-ionizable residues is achieved through the introduction of environment variables. This extension results in even better performance than THEMATICS alone and constitutes to date the best functional site predictor based on 3D structure only, achieving nearly the same level of performance as methods that use both 3D structure and sequence alignment data. Finally, the method also easily incorporates such sequence alignment data, and when this information is included, the resulting method is shown to outperform the best current methods using any combination of sequence alignments and 3D structures. Included is an analysis demonstrating that when THEMATICS features, cleft size rank, and alignment-based conservation scores are used individually or in combination

  12. i3Drefine software for protein 3D structure refinement and its assessment in CASP10.

    PubMed

    Bhattacharya, Debswapna; Cheng, Jianlin

    2013-01-01

    Protein structure refinement refers to the process of improving the qualities of protein structures during structure modeling processes to bring them closer to their native states. Structure refinement has been drawing increasing attention in the community-wide Critical Assessment of techniques for Protein Structure prediction (CASP) experiments since its addition in 8(th) CASP experiment. During the 9(th) and recently concluded 10(th) CASP experiments, a consistent growth in number of refinement targets and participating groups has been witnessed. Yet, protein structure refinement still remains a largely unsolved problem with majority of participating groups in CASP refinement category failed to consistently improve the quality of structures issued for refinement. In order to alleviate this need, we developed a completely automated and computationally efficient protein 3D structure refinement method, i3Drefine, based on an iterative and highly convergent energy minimization algorithm with a powerful all-atom composite physics and knowledge-based force fields and hydrogen bonding (HB) network optimization technique. In the recent community-wide blind experiment, CASP10, i3Drefine (as 'MULTICOM-CONSTRUCT') was ranked as the best method in the server section as per the official assessment of CASP10 experiment. Here we provide the community with free access to i3Drefine software and systematically analyse the performance of i3Drefine in strict blind mode on the refinement targets issued in CASP10 refinement category and compare with other state-of-the-art refinement methods participating in CASP10. Our analysis demonstrates that i3Drefine is only fully-automated server participating in CASP10 exhibiting consistent improvement over the initial structures in both global and local structural quality metrics. Executable version of i3Drefine is freely available at http://protein.rnet.missouri.edu/i3drefine/.

  13. Determining the 3-D structure and motion of objects using a scanning laser range sensor

    NASA Astrophysics Data System (ADS)

    Nandhakumar, N.; Smith, Philip W.

    1993-12-01

    In order for the EVAHR robot to autonomously track and grasp objects, its vision system must be able to determine the 3-D structure and motion of an object from a sequence of sensory images. This task is accomplished by the use of a laser radar range sensor which provides dense range maps of the scene. Unfortunately, the currently available laser radar range cameras use a sequential scanning approach which complicates image analysis. Although many algorithms have been developed for recognizing objects from range images, none are suited for use with single beam, scanning, time-of-flight sensors because all previous algorithms assume instantaneous acquisition of the entire image. This assumption is invalid since the EVAHR robot is equipped with a sequential scanning laser range sensor. If an object is moving while being imaged by the device, the apparent structure of the object can be significantly distorted due to the significant non-zero delay time between sampling each image pixel. If an estimate of the motion of the object can be determined, this distortion can be eliminated; but, this leads to the motion-structure paradox - most existing algorithms for 3-D motion estimation use the structure of objects to parameterize their motions. The goal of this research is to design a rigid-body motion recovery technique which overcomes this limitation. The method being developed is an iterative, linear, feature-based approach which uses the non-zero image acquisition time constraint to accurately recover the motion parameters from the distorted structure of the 3-D range maps. Once the motion parameters are determined, the structural distortion in the range images is corrected.

  14. How Does the Methodology of 3D Structure Preparation Influence the Quality of pKa Prediction?

    PubMed

    Geidl, Stanislav; Svobodová Vařeková, Radka; Bendová, Veronika; Petrusek, Lukáš; Ionescu, Crina-Maria; Jurka, Zdeněk; Abagyan, Ruben; Koča, Jaroslav

    2015-06-22

    The acid dissociation constant is an important molecular property, and it can be successfully predicted by Quantitative Structure-Property Relationship (QSPR) models, even for in silico designed molecules. We analyzed how the methodology of in silico 3D structure preparation influences the quality of QSPR models. Specifically, we evaluated and compared QSPR models based on six different 3D structure sources (DTP NCI, Pubchem, Balloon, Frog2, OpenBabel, and RDKit) combined with four different types of optimization. These analyses were performed for three classes of molecules (phenols, carboxylic acids, anilines), and the QSPR model descriptors were quantum mechanical (QM) and empirical partial atomic charges. Specifically, we developed 516 QSPR models and afterward systematically analyzed the influence of the 3D structure source and other factors on their quality. Our results confirmed that QSPR models based on partial atomic charges are able to predict pKa with high accuracy. We also confirmed that ab initio and semiempirical QM charges provide very accurate QSPR models and using empirical charges based on electronegativity equalization is also acceptable, as well as advantageous, because their calculation is very fast. On the other hand, Gasteiger-Marsili empirical charges are not applicable for pKa prediction. We later found that QSPR models for some classes of molecules (carboxylic acids) are less accurate. In this context, we compared the influence of different 3D structure sources. We found that an appropriate selection of 3D structure source and optimization method is essential for the successful QSPR modeling of pKa. Specifically, the 3D structures from the DTP NCI and Pubchem databases performed the best, as they provided very accurate QSPR models for all the tested molecular classes and charge calculation approaches, and they do not require optimization. Also, Frog2 performed very well. Other 3D structure sources can also be used but are not so robust, and

  15. Towards Automated Seismic Moment Tensor Inversion in Australia Using 3D Structural Model

    NASA Astrophysics Data System (ADS)

    Hingee, M.; Tkalcic, H.; Fichtner, A.; Sambridge, M.; Kennett, B. L.; Gorbatov, A.

    2009-12-01

    There is significant seismic activity in the region around Australia, largely due to the plate boundaries to the north and to the east of the mainland. This seismicity poses serious seismic and tsunamigenic hazard in a wider region, and risk to coastal areas of Australia, and is monitored by Geoscience Australia (GA) using a network of permanent broadband seismometers within Australia. Earthquake and tsunami warning systems were established by the Australian Government and have been using the waveforms from the GA seismological network. The permanent instruments are augmented by non-GA seismic stations based both within and outside of Australia. In particular, seismic moment tensor (MT) solutions for events around Australia as well as local distances are useful for both warning systems and geophysical studies in general. These monitoring systems, however, currently use only one dimensional, spherically-symmetric models of the Earth for source parameter determination. Recently, a novel 3D model of Australia and the surrounding area has been developed from spectral element simulations [1], taking into account not only velocity heterogeneities, but also radial anisotropy and seismic attenuation. This development, inter alia, introduces the potential of providing significant improvements in MT solution accuracy. Allowing reliable MT solutions with reduced dependence on non-GA stations is a secondary advantage. We studied the feasibility of using 1D versus 3D structural models. The accuracy of the 3D model has been investigated, confirming that these models are in most cases superior to the 1D models. A full MT inversion method using a point source approximation was developed as the first step, keeping in mind that for more complex source time functions, a finite source inversion will be needed. Synthetic experiments have been performed with random noise added to the signal to test the code in the both 1D and 3D setting, using a precomputed library of structural Greens

  16. Combination of photogrammetric and geoelectric methods to assess 3d structures associated to natural hazards

    NASA Astrophysics Data System (ADS)

    Fargier, Yannick; Dore, Ludovic; Antoine, Raphael; Palma Lopes, Sérgio; Fauchard, Cyrille

    2016-04-01

    The extraction of subsurface materials is a key element for the economy of a nation. However, natural degradation of underground quarries is a major issue from an economic and public safety point of view. Consequently, the quarries stakeholders require relevant tools to define hazards associated to these structures. Safety assessment methods of underground quarries are recent and mainly based on rock physical properties. This kind of method leads to a certain homogeneity assumption of pillar internal properties that can cause an underestimation of the risk. Electrical Resistivity Imaging (ERI) is a widely used method that possesses two advantages to overcome this limitation. The first is to provide a qualitative understanding for the detection and monitoring of anomalies in the pillar body (e.g. faults). The second is to provide a quantitative description of the electrical resistivity distribution inside the pillar. This quantitative description can be interpreted with constitutive laws to help decision support (water content decreases the mechanical resistance of a chalk). However, conventional 2D and 3D Imaging techniques are usually applied to flat surface surveys or to surfaces with moderate topography. A 3D inversion of more complex media (case of the pillar) requires a full consideration of the geometry that was never taken into account before. The Photogrammetric technique presents a cost effective solution to obtain an accurate description of the external geometry of a complex media. However, this method has never been fully coupled with a geophysical method to enhance/improve the inversion process. Consequently we developed a complete procedure showing that photogrammetric and ERI tools can be efficiently combined to assess a complex 3D structure. This procedure includes in a first part a photogrammetric survey, a processing stage with an open source software and a post-processing stage finalizing a 3D surface model. The second part necessitates the

  17. SimRNA: a coarse-grained method for RNA folding simulations and 3D structure prediction

    PubMed Central

    Boniecki, Michal J.; Lach, Grzegorz; Dawson, Wayne K.; Tomala, Konrad; Lukasz, Pawel; Soltysinski, Tomasz; Rother, Kristian M.; Bujnicki, Janusz M.

    2016-01-01

    RNA molecules play fundamental roles in cellular processes. Their function and interactions with other biomolecules are dependent on the ability to form complex three-dimensional (3D) structures. However, experimental determination of RNA 3D structures is laborious and challenging, and therefore, the majority of known RNAs remain structurally uncharacterized. Here, we present SimRNA: a new method for computational RNA 3D structure prediction, which uses a coarse-grained representation, relies on the Monte Carlo method for sampling the conformational space, and employs a statistical potential to approximate the energy and identify conformations that correspond to biologically relevant structures. SimRNA can fold RNA molecules using only sequence information, and, on established test sequences, it recapitulates secondary structure with high accuracy, including correct prediction of pseudoknots. For modeling of complex 3D structures, it can use additional restraints, derived from experimental or computational analyses, including information about secondary structure and/or long-range contacts. SimRNA also can be used to analyze conformational landscapes and identify potential alternative structures. PMID:26687716

  18. A coarse-grained model with implicit salt for RNAs: Predicting 3D structure, stability and salt effect

    SciTech Connect

    Shi, Ya-Zhou; Wang, Feng-Hua; Wu, Yuan-Yan; Tan, Zhi-Jie

    2014-09-14

    To bridge the gap between the sequences and 3-dimensional (3D) structures of RNAs, some computational models have been proposed for predicting RNA 3D structures. However, the existed models seldom consider the conditions departing from the room/body temperature and high salt (1M NaCl), and thus generally hardly predict the thermodynamics and salt effect. In this study, we propose a coarse-grained model with implicit salt for RNAs to predict 3D structures, stability, and salt effect. Combined with Monte Carlo simulated annealing algorithm and a coarse-grained force field, the model folds 46 tested RNAs (≤45 nt) including pseudoknots into their native-like structures from their sequences, with an overall mean RMSD of 3.5 Å and an overall minimum RMSD of 1.9 Å from the experimental structures. For 30 RNA hairpins, the present model also gives the reliable predictions for the stability and salt effect with the mean deviation ∼ 1.0 °C of melting temperatures, as compared with the extensive experimental data. In addition, the model could provide the ensemble of possible 3D structures for a short RNA at a given temperature/salt condition.

  19. Measuring the Thickness and Potential Profiles of the Space-Charge Layer at Organic/Organic Interfaces under Illumination and in the Dark by Scanning Kelvin Probe Microscopy.

    PubMed

    Rojas, Geoffrey A; Wu, Yanfei; Haugstad, Greg; Frisbie, C Daniel

    2016-03-09

    Scanning Kelvin probe microscopy was used to measure band-bending at the model donor/acceptor heterojunction poly(3-hexylthiophene) (P3HT)/fullerene (C60). Specifically, we measured the variation in the surface potential of C60 films with increasing thicknesses grown on P3HT to produce a surface potential profile normal to the substrate both in the dark and under illumination. The results confirm a space-charge carrier region with a thickness of 10 nm, consistent with previous observations. We discuss the possibility that the domain size in bulk heterojunction organic solar cells, which is comparable to the space-charge layer thickness, is actually partly responsible for less than expected electron/hole recombination rates.

  20. Selective plane illumination microscopy with a light sheet of uniform thickness formed by an electrically tunable lens.

    PubMed

    Hedde, Per Niklas; Gratton, Enrico

    2016-06-24

    Light sheet microscopy is a powerful technique for rapid, three-dimensional fluorescence imaging of large specimen such as drosophila and zebrafish embryos. Yet, beam divergence results in a loss of axial resolution at the periphery of the light sheet. Here, we demonstrate how an electrically tunable lens can be utilized to maintain the minimal, diffraction-limited thickness of the light sheet over a wide field of view (>600 µm) at high frame rates (40 fps). This mode of operation is necessary for the application of fluorescence fluctuation spectroscopy in images. Microsc. Res. Tech, 2016. © 2016 Wiley Periodicals, Inc.

  1. 3D structure through planting core-shell Si@TiN into an amorphous carbon slag: improved capacity of lithium-ion anodes.

    PubMed

    Tu, Jiguo; Zhao, Zuochao; Hu, Liwen; Jiao, Shuqiang; Hou, Jungang; Zhu, Hongmin

    2013-07-07

    A 3D-structured anode material, planting core-shell Si@TiN into an amorphous carbon slag (3D STC), was synthesized via a facile pyrolyzing process in assistance with the low-temperature reduction route in a liquid Na-NH3 system. The as-prepared samples were characterized by X-ray diffraction, field emission scanning electron microscopy, transmission electron microscopy, cyclic voltammetry and galvanostatic discharge-charge tests. From morphological analysis, TiN nanoparticles were homogeneously dispersed on the surface of Si to form the Si@TiN core-shell structure, subsequently plating into an amorphous C slag to form the 3D STC composite. The electrochemical capacity of the 3D STC anode was measured at a higher rate of 1 C with the cut-off voltages of 0.01 V and 1.5 V. It was found that the initial charge capacity reached up to 1604.6 mA h g(-1). In particular, the reversible charge capacity was as high as 588.7 mA h g(-1) over 100 cycles, with a small capacity loss of about 0.63% per cycle, exhibiting the excellent cycle stability of the 3D STC anode at the higher rate of 1 C. Furthermore, the reversible capacity of the 3D STC anode decreased from 2048.8 mA h g(-1) to 624.0 mA h g(-1) with increasing the current rate from 0.1 C to 2 C, while it was still maintained at 1419.7 mA h g(-1) as the current rate returned to 0.1 C. Consequentially, the 3D structure with a continuous conductive path could provide facile lithium insertion/extraction and fast electron transfer, making for the high rate capacity and good cycle stability.

  2. High resolution three dimensional microscopy of biological microstructures using zone plate lenses with x-ray laser illumination

    SciTech Connect

    Trebes, J.

    1990-12-12

    One of the goals of biomedical research is the development of imaging techniques capable of producing high resolution ({approximately}300{Angstrom}) three dimensional images of structures within live cells. Recent developments in zone plate lenses at LBL and in x-ray lasers at LLNL indicate that flash three dimensional x-ray microscopy of live biological objects can be achieved in the near term. This concept for a microscope utilizes an x-ray laser to backlit immunogold labeled biological objects. These backlit objects are then imaged with low f-number, high resolution zone plate lenses. Backlighting and imaging along several different directions allows a three dimensional image to be obtained using tomographic techniques.

  3. A tetraphenylethylene core-based 3D structure small molecular acceptor enabling efficient non-fullerene organic solar cells.

    PubMed

    Liu, Yuhang; Mu, Cheng; Jiang, Kui; Zhao, Jingbo; Li, Yunke; Zhang, Lu; Li, Zhengke; Lai, Joshua Yuk Lin; Hu, Huawei; Ma, Tingxuan; Hu, Rongrong; Yu, Demei; Huang, Xuhui; Tang, Ben Zhong; Yan, He

    2015-02-01

    A tetraphenylethylene core-based small molecular acceptor with a unique 3D molecular structure is developed. Bulk-heterojunction blend films with a small feature size (≈20 nm) are obtained, which lead to non-fullerene organic solar cells (OSCs) with 5.5% power conversion efficiency. The work provides a new molecular design approach to efficient non-fullerene OSCs based on 3D-structured small-molecule acceptors.

  4. R3D Align web server for global nucleotide to nucleotide alignments of RNA 3D structures

    PubMed Central

    Rahrig, Ryan R.; Petrov, Anton I.; Leontis, Neocles B.; Zirbel, Craig L.

    2013-01-01

    The R3D Align web server provides online access to ‘RNA 3D Align’ (R3D Align), a method for producing accurate nucleotide-level structural alignments of RNA 3D structures. The web server provides a streamlined and intuitive interface, input data validation and output that is more extensive and easier to read and interpret than related servers. The R3D Align web server offers a unique Gallery of Featured Alignments, providing immediate access to pre-computed alignments of large RNA 3D structures, including all ribosomal RNAs, as well as guidance on effective use of the server and interpretation of the output. By accessing the non-redundant lists of RNA 3D structures provided by the Bowling Green State University RNA group, R3D Align connects users to structure files in the same equivalence class and the best-modeled representative structure from each group. The R3D Align web server is freely accessible at http://rna.bgsu.edu/r3dalign/. PMID:23716643

  5. Nanoimprint of a 3D structure on an optical fiber for light wavefront manipulation.

    PubMed

    Calafiore, Giuseppe; Koshelev, Alexander; Allen, Frances I; Dhuey, Scott; Sassolini, Simone; Wong, Edward; Lum, Paul; Munechika, Keiko; Cabrini, Stefano

    2016-09-16

    Integration of complex photonic structures onto optical fiber facets enables powerful platforms with unprecedented optical functionalities. Conventional nanofabrication technologies, however, do not permit viable integration of complex photonic devices onto optical fibers owing to their low throughput and high cost. In this paper we report the fabrication of a three-dimensional structure achieved by direct nanoimprint lithography on the facet of an optical fiber. Nanoimprint processes and tools were specifically developed to enable a high lithographic accuracy and coaxial alignment of the optical device with respect to the fiber core. To demonstrate the capability of this new approach, a 3D beam splitter has been designed, imprinted and optically characterized. Scanning electron microscopy and optical measurements confirmed the good lithographic capabilities of the proposed approach as well as the desired optical performance of the imprinted structure. The inexpensive solution presented here should enable advancements in areas such as integrated optics and sensing, achieving enhanced portability and versatility of fiber optic components.

  6. Multi Length Scale Imaging of Flocculated Estuarine Sediments; Insights into their Complex 3D Structure

    NASA Astrophysics Data System (ADS)

    Wheatland, Jonathan; Bushby, Andy; Droppo, Ian; Carr, Simon; Spencer, Kate

    2015-04-01

    Suspended estuarine sediments form flocs that are compositionally complex, fragile and irregularly shaped. The fate and transport of suspended particulate matter (SPM) is determined by the size, shape, density, porosity and stability of these flocs and prediction of SPM transport requires accurate measurements of these three-dimensional (3D) physical properties. However, the multi-scaled nature of flocs in addition to their fragility makes their characterisation in 3D problematic. Correlative microscopy is a strategy involving the spatial registration of information collected at different scales using several imaging modalities. Previously, conventional optical microscopy (COM) and transmission electron microscopy (TEM) have enabled 2-dimensional (2D) floc characterisation at the gross (> 1 µm) and sub-micron scales respectively. Whilst this has proven insightful there remains a critical spatial and dimensional gap preventing the accurate measurement of geometric properties and an understanding of how structures at different scales are related. Within life sciences volumetric imaging techniques such as 3D micro-computed tomography (3D µCT) and focused ion beam scanning electron microscopy [FIB-SEM (or FIB-tomography)] have been combined to characterise materials at the centimetre to micron scale. Combining these techniques with TEM enables an advanced correlative study, allowing material properties across multiple spatial and dimensional scales to be visualised. The aims of this study are; 1) to formulate an advanced correlative imaging strategy combining 3D µCT, FIB-tomography and TEM; 2) to acquire 3D datasets; 3) to produce a model allowing their co-visualisation; 4) to interpret 3D floc structure. To reduce the chance of structural alterations during analysis samples were first 'fixed' in 2.5% glutaraldehyde/2% formaldehyde before being embedding in Durcupan resin. Intermediate steps were implemented to improve contrast and remove pore water, achieved by the

  7. Nanoimprint of a 3D structure on an optical fiber for light wavefront manipulation

    NASA Astrophysics Data System (ADS)

    Calafiore, Giuseppe; Koshelev, Alexander; Allen, Frances I.; Dhuey, Scott; Sassolini, Simone; Wong, Edward; Lum, Paul; Munechika, Keiko; Cabrini, Stefano

    2016-09-01

    Integration of complex photonic structures onto optical fiber facets enables powerful platforms with unprecedented optical functionalities. Conventional nanofabrication technologies, however, do not permit viable integration of complex photonic devices onto optical fibers owing to their low throughput and high cost. In this paper we report the fabrication of a three-dimensional structure achieved by direct nanoimprint lithography on the facet of an optical fiber. Nanoimprint processes and tools were specifically developed to enable a high lithographic accuracy and coaxial alignment of the optical device with respect to the fiber core. To demonstrate the capability of this new approach, a 3D beam splitter has been designed, imprinted and optically characterized. Scanning electron microscopy and optical measurements confirmed the good lithographic capabilities of the proposed approach as well as the desired optical performance of the imprinted structure. The inexpensive solution presented here should enable advancements in areas such as integrated optics and sensing, achieving enhanced portability and versatility of fiber optic components.

  8. 3-D structures of crack-tip dislocations and their shielding effect revealed by electron tomography.

    PubMed

    Tanaka, Masaki; Honda, Masaki; Sadamatsu, Sunao; Higashida, Kenji

    2010-08-01

    Three-dimensional structures of crack-tip dislocations in silicon crystals have been examined by combining scanning transmission electron microscopy and computed tomography. Cracks were introduced by a Vickers hardness tester at room temperature, and the sample was heated at 823 K for 1 h in order to introduce dislocations around the crack tips. Dislocation segments cut out from loops were observed around the crack tip, the three-dimensional structure of which was characterized by using by electron tomography. Their Burgers vectors including the sings were also determined by oscillating contrasts along dislocations. In order to investigate the effect of the dislocations on fracture behaviours, local stress intensity factor due to one dislocation was calculated, which indicates the dislocations observed were shielding type to increase fracture toughness.

  9. Morphological study of lipid vesicles in presence of amphotericin B via modification of the microfluidic CellASIC platform and LED illumination microscopy

    NASA Astrophysics Data System (ADS)

    Genova, J.; Decheva-Zarkova, M.; Pavlič, J. I.

    2016-02-01

    Giant lipid vesicles (liposomes) are the simplest model of the biological cell and can be easily formed from natural or synthetic lipid species with controlled composition and properties. This is the reason why they are the preferred objects for various scientific investigations. Amphotericin B (AmB) is a membrane active drug, used for treatment of systemic fungal infections. In this work we studied the morphological behavior of giant SOPC vesicles in asymmetrical presence of amphotericin B antibiotic in the vicinity of the lipid membrane. The visualization of the vesicles was carried out via inverted phase contrast microscopy. The illumination source was modified in a way that tungsten light bulb was replaced by 10 W white LED chip. All the experiments were performed using CellASIC ONIX Microfluidic Platform. The setup has been modified thus opening new opportunities for a variety of experimental realizations. The performed morphological studies showed strong and irreversible effect on the vesicle shape at the presence of amphotericin B in concentration 10-5 g/l in the outer for the liposome's membrane solution. At concentration 10-3 g/l AmB the effect was less visible and in 15-20 minutes the vesicles regained its initial spherical shape.

  10. Preparation and photo-induced charge transfer of the composites based on 3D structural CdS nanocrystals and MEH-PPV

    SciTech Connect

    Deng, Dan; Shi, Minmin; Chen, Fei; Chen, Lin; Jiang, Xiaoxia; Chen, Hongzheng

    2010-05-15

    We report the synthesis of 3D structural CdS nanocrystals by a simple biomolecule-assisted hydrothermal process. The CdS nanocrystals are composed of many branched nanorods with the diameter of about 50 nm, and the length of about 250 nm. The phase and crystallographic properties are characterized by scanning electron microscopy (SEM), transmission electron microscopy (TEM), and X-ray diffractometry (XRD). The composites based on CdS nanocrystals and poly[2-methoxy-5-(2-ethylhexyloxy-p-phenylenevinylene)] (MEH-PPV) have been prepared by spin-coating of the mixture in the common solvent. The optical properties of the composites are investigated using ultraviolet-visible (UV-Vis) absorption and photoluminescence (PL) spectroscopies. A significant fluorescence quenching of MEH-PPV in the composites is observed at high CdS nanocrystals/MEH-PPV ratios, indicating that the photo-induced charge transfer occurred due to the energy level offset between the donor MEH-PPV and the acceptor CdS nanocrystals. The obvious photovoltaic behavior of the solar cell made from this composite further demonstrates the mentioned photo-induced charge transfer process. (author)

  11. 3D structure of the Yersinia entomophaga toxin complex and implications for insecticidal activity

    PubMed Central

    Landsberg, Michael J.; Jones, Sandra A.; Rothnagel, Rosalba; Busby, Jason N.; Marshall, Sean D. G.; Simpson, Robert M.; Lott, J. Shaun; Hankamer, Ben; Hurst, Mark R. H.

    2011-01-01

    Toxin complex (Tc) proteins are a class of bacterial protein toxins that form large, multisubunit complexes. Comprising TcA, B, and C components, they are of great interest because many exhibit potent insecticidal activity. Here we report the structure of a novel Tc, Yen-Tc, isolated from the bacterium Yersinia entomophaga MH96, which differs from the majority of bacterially derived Tcs in that it exhibits oral activity toward a broad range of insect pests, including the diamondback moth (Plutella xylostella). We have determined the structure of the Yen-Tc using single particle electron microscopy and studied its mechanism of toxicity by comparative analyses of two variants of the complex exhibiting different toxicity profiles. We show that the A subunits form the basis of a fivefold symmetric assembly that differs substantially in structure and subunit arrangement from its most well characterized homologue, the Xenorhabdus nematophila toxin XptA1. Histopathological and quantitative dose response analyses identify the B and C subunits, which map to a single, surface-accessible region of the structure, as the sole determinants of toxicity. Finally, we show that the assembled Yen-Tc has endochitinase activity and attribute this to putative chitinase subunits that decorate the surface of the TcA scaffold, an observation that may explain the oral toxicity associated with the complex. PMID:22158901

  12. A large dataset of synthetic SEM images of powder materials and their ground truth 3D structures.

    PubMed

    DeCost, Brian L; Holm, Elizabeth A

    2016-12-01

    This data article presents a data set comprised of 2048 synthetic scanning electron microscope (SEM) images of powder materials and descriptions of the corresponding 3D structures that they represent. These images were created using open source rendering software, and the generating scripts are included with the data set. Eight particle size distributions are represented with 256 independent images from each. The particle size distributions are relatively similar to each other, so that the dataset offers a useful benchmark to assess the fidelity of image analysis techniques. The characteristics of the PSDs and the resulting images are described and analyzed in more detail in the research article "Characterizing powder materials using keypoint-based computer vision methods" (B.L. DeCost, E.A. Holm, 2016) [1]. These data are freely available in a Mendeley Data archive "A large dataset of synthetic SEM images of powder materials and their ground truth 3D structures" (B.L. DeCost, E.A. Holm, 2016) located at http://dx.doi.org/10.17632/tj4syyj9mr.1[2] for any academic, educational, or research purposes.

  13. WebRASP: a server for computing energy scores to assess the accuracy and stability of RNA 3D structures

    PubMed Central

    Norambuena, Tomas; Cares, Jorge F.; Capriotti, Emidio; Melo, Francisco

    2013-01-01

    Summary: The understanding of the biological role of RNA molecules has changed. Although it is widely accepted that RNAs play important regulatory roles without necessarily coding for proteins, the functions of many of these non-coding RNAs are unknown. Thus, determining or modeling the 3D structure of RNA molecules as well as assessing their accuracy and stability has become of great importance for characterizing their functional activity. Here, we introduce a new web application, WebRASP, that uses knowledge-based potentials for scoring RNA structures based on distance-dependent pairwise atomic interactions. This web server allows the users to upload a structure in PDB format, select several options to visualize the structure and calculate the energy profile. The server contains online help, tutorials and links to other related resources. We believe this server will be a useful tool for predicting and assessing the quality of RNA 3D structures. Availability and implementation: The web server is available at http://melolab.org/webrasp. It has been tested on the most popular web browsers and requires Java plugin for Jmol visualization. Contact: fmelo@bio.puc.cl PMID:23929030

  14. Modelling a 3D structure for EgDf1 from shape Echinococcus granulosus: putative epitopes, phosphorylation motifs and ligand

    NASA Astrophysics Data System (ADS)

    Paulino, M.; Esteves, A.; Vega, M.; Tabares, G.; Ehrlich, R.; Tapia, O.

    1998-07-01

    EgDf1 is a developmentally regulated protein from the parasite Echinococcus granulosus related to a family of hydrophobic ligand binding proteins. This protein could play a crucial role during the parasite life cycle development since this organism is unable to synthetize most of their own lipids de novo. Furthermore, it has been shown that two related protein from other parasitic platyhelminths (Fh15 from Fasciola hepatica and Sm14 from Schistosoma mansoni) are able to confer protective inmunity against experimental infection in animal models. A three-dimensional structure would help establishing structure/function relationships on a knowledge based manner. 3D structures for EgDf1 protein were modelled by using myelin P2 (mP2) and intestine fatty acid binding protein (I-FABP) as templates. Molecular dynamics techniques were used to validate the models. Template mP2 yielded the best 3D structure for EgDf1. Palmitic and oleic acids were docked inside EgDf1. The present theoretical results suggest definite location in the secondary structure of the epitopic regions, consensus phosphorylation motifs and oleic acid as a good ligand candidate to EgDf1. This protein might well be involved in the process of supplying hydrophobic metabolites for membrane biosynthesis and for signaling pathways.

  15. Representation of protein 3D structures in spherical (ρ, ϕ, θ) coordinates and two of its potential applications.

    PubMed

    Reyes, Vicente M

    2011-09-01

    Three-dimensional objects can be represented using cartesian, spherical or cylindrical coordinate systems, among many others. Currently all protein 3D structures in the PDB are in cartesian coordinates. We wanted to explore the possibility that protein 3D structures, especially the globular type (spheroproteins), when represented in spherical coordinates might find useful novel applications. A Fortran program was written to transform protein 3D structure files in cartesian coordinates (x,y,z) to spherical coordinates (ρ, ϕ, θ), with the centroid of the protein molecule as origin. We present here two applications, namely, (1) separation of the protein outer layer (OL) from the inner core (IC); and (2) identifying protrusions and invaginations on the protein surface. In the first application, ϕ and θ were partitioned into suitable intervals and the point with maximum ρ in each such 'ϕ-θ bin' was determined. A suitable cutoff value for ρ is adopted, and for each ϕ-θ bin, all points with ρ values less than the cutoff are considered part of the IC, and those with ρ values equal to or greater than the cutoff are considered part of the OL. We show that this separation procedure is successful as it gives rise to an OL that is significantly more enriched in hydrophilic amino acid residues, and an IC that is significantly more enriched in hydrophobic amino acid residues, as expected. In the second application, the point with maximum ρ in each ϕ-θ bin are sequestered and their frequency distribution constructed (i.e., maximum ρ's sorted from lowest to highest, collected into 1.50Å-intervals, and the frequency in each interval plotted). We show in such plots that invaginations on the protein surface give rise to subpeaks or shoulders on the lagging side of the main peak, while protrusions give rise to similar subpeaks or shoulders, but on the leading side of the main peak. We used the dataset of Laskowski et al. (1996) to demonstrate both applications.

  16. Nitrogen and sulfur co-doping of partially exfoliated MWCNTs as 3-D structured electrocatalysts for the oxygen reduction reaction

    SciTech Connect

    Wang, Jie; Wu, Zexing; Han, Lili; Lin, Ruoqian; Xiao, Weiping; Xuan, Cuijuan; Xin, Huolin. L.; Wang, Deli

    2016-03-14

    Preventing the stacking of graphene sheets is of vital importance for highly efficient and stable fuel cell electrocatalysts. Here, we report a 3-D structured carbon nanotube intercalated graphene nanoribbon with N/S co-doping. The nanocomposite is obtained by using high temperature heat-treated thiourea with partially unzipped multi-walled carbon nanotubes. This unique structure preserves both the properties of carbon nanotubes and graphene, exhibiting excellent catalytic performance for the ORR with similar onset and half-wave potentials to those of Pt/C electrocatalysts. Furthermore, the stereo structured composite exhibits distinct advantages in long-term stability and methanol poisoning tolerance in comparison to Pt/C.

  17. Building a 3D Virtual Liver: Methods for Simulating Blood Flow and Hepatic Clearance on 3D Structures

    PubMed Central

    Rezania, Vahid; Tuszynski, Jack

    2016-01-01

    In this paper, we develop a spatio-temporal modeling approach to describe blood and drug flow, as well as drug uptake and elimination, on an approximation of the liver. Extending on previously developed computational approaches, we generate an approximation of a liver, which consists of a portal and hepatic vein vasculature structure, embedded in the surrounding liver tissue. The vasculature is generated via constrained constructive optimization, and then converted to a spatial grid of a selected grid size. Estimates for surrounding upscaled lobule tissue properties are then presented appropriate to the same grid size. Simulation of fluid flow and drug metabolism (hepatic clearance) are completed using discretized forms of the relevant convective-diffusive-reactive partial differential equations for these processes. This results in a single stage, uniformly consistent method to simulate equations for blood and drug flow, as well as drug metabolism, on a 3D structure representative of a liver. PMID:27649537

  18. [3D structure of DKK1 indicates its involvement in both canonical and non-canonical Wnt pathways].

    PubMed

    Khalili, S; Rasaee, M J; Bamdad, T

    2017-01-01

    Dikkoppf-1 (DKK1) is an antagonist of the canonical Wnt signaling pathway. The importance of DKK1 as a diagnostic and therapeutic agent in a wide range of diseases along with its significance in a variety of biological processes accentuate the necessity to decipher its 3D structure that would pave the way towards the development of relevant selective inhibitors. A DKK1 structure model predicted by the Robetta server with structural refinements including a 10 ns molecular dynamics run was subjected to functional and docking analyses. We hypothesize that the N-terminal region of the DKK1 molecule could be functionally important for both canonical and noncanonical Wnt pathways. Moreover, it seems that DKK1 could be involved in interactions with the Frizzled receptors, leading to the activation of the Planar Cell Polarity (PCP) pathway (activation of Jun N-terminal kinase (JNK) Pathway) and Wnt/Ca^(2+) pathway (activation of CamKII).

  19. Computational 3D structures of drug-targeting proteins in the 2009-H1N1 influenza A virus

    NASA Astrophysics Data System (ADS)

    Du, Qi-Shi; Wang, Shu-Qing; Huang, Ri-Bo; Chou, Kuo-Chen

    2010-01-01

    The neuraminidase (NA) and M2 proton channel of influenza virus are the drug-targeting proteins, based on which several drugs were developed. However these once powerful drugs encountered drug-resistant problem to the H5N1 and H1N1 flu. To address this problem, the computational 3D structures of NA and M2 proteins of 2009-H1N1 influenza virus were built using the molecular modeling technique and computational chemistry method. Based on the models the structure features of NA and M2 proteins were analyzed, the docking structures of drug-protein complexes were computed, and the residue mutations were annotated. The results may help to solve the drug-resistant problem and stimulate designing more effective drugs against 2009-H1N1 influenza pandemic.

  20. The Relationship between Oxygen A-band Photon Pathlength Distributions and 3D Structures of Heating Rate Profiles

    NASA Astrophysics Data System (ADS)

    Song, L.; Min, Q.

    2012-12-01

    Broadband heating directly drives the global atmospheric and oceanic circulation and its vertical profiles strongly depend upon cloud three-dimensional (3D) structures. Due to the complexity of cloud 3D problems and the difficulties in observations of broadband heating rate profiles (BBHRP), there are still large uncertainties in the relationship of clouds, radiation and climate feedback. Oxygen A-band photon pathlength distributions (PPLD) contain rich information about the 3D structures of clouds and BBHRP and can be observed by both ground based and space based measurements. Therefore, it is meaningful to explore the possibility of connecting A-band PPLD and BBHRP and consequently to describe the internal relationship between them together with the cloud 3D effects on BBHRP. A 3D Monte Carlo radiative transfer model is applied to simulate solar broadband heating rate profiles and oxygen A-band photon pathlength distributions of several ideal cloud fields and two typical cloud fields generated by cloud resolving model (CRM). Principal components (PCs) and the first four moments are selected to represent the vertical structures of BBHRP and PPLD, respectively. In ideal cloud fields, the moments show clear constraint to PCs of BBHRP. The results demonstrate the feasibility to describe the vertical structures of BBHRP by PPLD. The relationship between moments and PCs turns complicated in CRM cloud fields due to the composition of various 3D effects. However, detailed analysis still show that the moments, the PCs and total cloud optical depth are effective factors in defining BBHRP, especially for the vertical structures of relative low clouds. Further, a statistical fitting between the PCs and the moments by a two-layer neural network is applied to provide a quantitative representation of the linkages.

  1. A Micro CT Study in Patients with Breast Microcalcifications Using a Mathematical Algorithm to Assess 3D Structure

    PubMed Central

    Varga, Zsuzsanna; Heuer, Heike; Dedes, Konstantin J.; Berger, Nicole; Filli, Lukas; Boss, Andreas

    2017-01-01

    Purpose The aim of this study was to evaluate the relevance of the three-dimensional (3D) structure of breast microcalcifications (MC) as a predictor of malignancy using highly resolved micro-computed tomography (micro-CT) datasets of biopsy samples. Material and Methods The study included 28 women with suspicious MC in their mammogram undergoing vacuum-assisted biopsy. Directly after the intervention, the specimens were scanned in a micro-CT with an isometric spatial resolution of 9 μm. Datasets were analysed regarding the number, volume and morphology of suspicious non-monomorphic MC (fl—fine linear, fp—fine pleomorphic, ch—coarse heterogeneous) and the structure model index (SMI). Histological evaluation was performed according to the B-classification: normal tissue or benign (group A: B1, B2), unclear malignant potential or suspicious of malignancy (group B: B3, B4) and malignant lesions (group C: B5). Results In all groups, suspicious non-monomorphic MC were found: group A exhibited fp MC in 38.5% of samples, no fl/ch; group B: fl 14.3%, fp 28.6%, ch 14.3%; group C always had at least one type of suspicious non-monomorphic MC (fl (57.1%) or fp (57.1%)) in each sample. The different histologic groups showed a similar mean SMI (benign: 2.97 ± 0.31, malignant: 3.02 ± 0.10, unclear: 2.90 ± 0.28). Between the three groups, no significant differences were found regarding number, volume or SMI value of MC. Conclusion 3D structure based on the SMI of MC analysed with highest spatial resolution is not significantly associated with the B-classification of breast lesions. Thus, magnification views of MC may be omitted in the analysis of MC detected in mammograms. PMID:28107436

  2. Plane wave illumination for correct phase analysis and alternative phase unwrapping in dual-type (transmission and reflection) three-dimensional digital holographic microscopy

    NASA Astrophysics Data System (ADS)

    Kim, Moonseok; Hong, Sukjoon; Shim, Seongbo; Soh, Kwangsup; Shin, Sanghoon; Son, Jung-Young; Lee, Jaesung; Kim, Jaisoon

    2010-05-01

    The digital holographic microscope (DHM) has emerged as a useful tool for verifying the three-dimensional structure of an object. A dual-type inline DHM that can be used with both transmission and reflection imaging in a single device is developed. The proper modes (between transmission and reflection imaging) can be easily changed according to the characteristics of the object in this system. The optimum condition for retrieving the correct phase information is illuminating a plane wave to an object. In contrast to the transmission imaging, it is difficult to illuminate an object using a plane wave without deformations in the reflection imaging. We developed an adequate relay lens module for illumination that can be adapted to any type of microscope objective without significant aberrations in the reflection imaging. The relationship between the illuminating condition and the measured phase based on the wave optics is analyzed. A specially designed and manufactured phase mask is observed in this system, and an alternative method for overcoming the limitation of phase unwrapping is introduced for the inspection of that object.

  3. Low temperature H2S removal with 3-D structural mesoporous molecular sieves supported ZnO from gas stream.

    PubMed

    Li, L; Sun, T H; Shu, C H; Zhang, H B

    2016-07-05

    A series of 3-dimensional (3-D) structural mesoporous silica materials, SBA-16, MCM-48 and KIT-6, was synthesized and supported with different ZnO loadings (10, 20, 30, and 40 wt%) by the incipient wetness method to evaluate the performances on H2S removal at room temperature. These materials were characterized by N2 adsorption, XRD, and TEM to investigate their textural properties. All the ZnO-loaded adsorbents exhibited the H2S removal capacity of bellow 0.1 ppmv. With the best ZnO loading percentage of 30 wt% on MCM-48 and KIT-6, 20 wt% on SBA-16 according to the results of breakthrough test, further increasing ZnO loading caused the decrease of the adsorption capacity due to the agglomeration of ZnO. Besides, the H2S adsorption capacities of the supports materials varied in the order of KIT-6>MCM-48>SBA-16, which was influenced primarily by their pore volume and pore size. With the largest pores in these 3-D arrangement materials, KIT-6 showed the best performance of supported material for ZnO, due to its retained superior physical properties as well as large pore diameter to allow faster gas-solid interaction and huge pore volume to disperse ZnO on the surface of it.

  4. ForceGen 3D structure and conformer generation: from small lead-like molecules to macrocyclic drugs.

    PubMed

    Cleves, Ann E; Jain, Ajay N

    2017-03-13

    We introduce the ForceGen method for 3D structure generation and conformer elaboration of drug-like small molecules. ForceGen is novel, avoiding use of distance geometry, molecular templates, or simulation-oriented stochastic sampling. The method is primarily driven by the molecular force field, implemented using an extension of MMFF94s and a partial charge estimator based on electronegativity-equalization. The force field is coupled to algorithms for direct sampling of realistic physical movements made by small molecules. Results are presented on a standard benchmark from the Cambridge Crystallographic Database of 480 drug-like small molecules, including full structure generation from SMILES strings. Reproduction of protein-bound crystallographic ligand poses is demonstrated on four carefully curated data sets: the ConfGen Set (667 ligands), the PINC cross-docking benchmark (1062 ligands), a large set of macrocyclic ligands (182 total with typical ring sizes of 12-23 atoms), and a commonly used benchmark for evaluating macrocycle conformer generation (30 ligands total). Results compare favorably to alternative methods, and performance on macrocyclic compounds approaches that observed on non-macrocycles while yielding a roughly 100-fold speed improvement over alternative MD-based methods with comparable performance.

  5. The 3D Structure of Eta Carinae's Nebula: A Definitive Picture from High-Dispersion Near-IR Spectra

    NASA Technical Reports Server (NTRS)

    Smith, N.

    2006-01-01

    High resolution long-slit spectra obtained with the Phoenix spectrograph on Gemini South provide our most accurate probe of the 3D structure of the Homunculus Nebula around Eta Carinae. Emission from molecular hydrogen at 2.122 microns traces a very thin outer skin, which contains the vast majority of the more than 10 solar masses of material in the nebula. This emission, in turn, yields our first definitive picture of the exact shape of the nebula, plus the latitude dependence of the mass-loss rate, speed, kinetic energy, shell thickness, and other properties associated with Eta Car's 19th century explosion. This will be critical for testing any models for the outburst mechanism. A preliminary analysis suggests that explosion from a critically rotating star was the dominant mechanism that gave rise to both the bipolar shape of the nebula and the production of its equatorial disk. [Fe II] emission in the near IR traces a geometrically thicker but less massive shell found on the inner surface of the H2 skin --- this is either a reverse shock that decelerates Eta Car's wind or a warm PDR. [Fe Ill emission also clarifies the structure of an inner "Little Homunculus" seen previously in HST/STlS spectra. Comparing these two tracers of cool molecular gas and warm partially-ionized gas resolves some significant confusion about the complex structure noted in previous studies.

  6. In silico 3D structure modeling and inhibitor binding studies of human male germ cell-associated kinase.

    PubMed

    Tanneeru, Karunakar; Balla, Ashok Raja; Guruprasad, Lalitha

    2015-01-01

    Human male germ cell-associated kinase (hMAK) is an androgen-inducible gene in prostate epithelial cells, and it acts as a coactivator of androgen receptor signaling in prostate cancer. The 3D structure of the hMAK kinase was modeled based on the crystal structure of CDK2 kinase using comparative modeling methods, and the ATP-binding site was characterized. We have collected five inhibitors of hMAK from the literature and docked into the ATP-binding site of the kinase domain. Solvated interaction energies (SIE) of inhibitor binding are calculated from the molecular dynamics simulations trajectories of protein-inhibitor complexes. The contribution from each active site residue in hMAK toward inhibitor binding revealed the nature and extent of interactions between inhibitors and individual residues. The main chain atoms of Met79 invariably form hydrogen bonds with all five inhibitors. The amino acids Leu7, Val15, and Leu129 stabilize the inhibitors via CH-pi interactions. The Asp140 in the active site and Glu77 in hinge region show characteristic hydrogen bonding interactions with inhibitors. From SIE, the residue-wise interactions revealed the nature of non-bonding contacts and modifications required to increase the inhibitor activity. Our work provides 3D model structure of hMAK and molecular basis for the mechanisms of hMAK inhibition at atomic level that aid in designing new potent inhibitors.

  7. ForceGen 3D structure and conformer generation: from small lead-like molecules to macrocyclic drugs

    NASA Astrophysics Data System (ADS)

    Cleves, Ann E.; Jain, Ajay N.

    2017-03-01

    We introduce the ForceGen method for 3D structure generation and conformer elaboration of drug-like small molecules. ForceGen is novel, avoiding use of distance geometry, molecular templates, or simulation-oriented stochastic sampling. The method is primarily driven by the molecular force field, implemented using an extension of MMFF94s and a partial charge estimator based on electronegativity-equalization. The force field is coupled to algorithms for direct sampling of realistic physical movements made by small molecules. Results are presented on a standard benchmark from the Cambridge Crystallographic Database of 480 drug-like small molecules, including full structure generation from SMILES strings. Reproduction of protein-bound crystallographic ligand poses is demonstrated on four carefully curated data sets: the ConfGen Set (667 ligands), the PINC cross-docking benchmark (1062 ligands), a large set of macrocyclic ligands (182 total with typical ring sizes of 12-23 atoms), and a commonly used benchmark for evaluating macrocycle conformer generation (30 ligands total). Results compare favorably to alternative methods, and performance on macrocyclic compounds approaches that observed on non-macrocycles while yielding a roughly 100-fold speed improvement over alternative MD-based methods with comparable performance.

  8. An Acinetobacter trimeric autotransporter adhesin reaped from cells exhibits its nonspecific stickiness via a highly stable 3D structure

    PubMed Central

    Yoshimoto, Shogo; Nakatani, Hajime; Iwasaki, Keita; Hori, Katsutoshi

    2016-01-01

    Trimeric autotransporter adhesins (TAAs), cell surface proteins of Gram-negative bacteria, mediate bacterial adhesion to host cells and extracellular matrix proteins. However, AtaA, a TAA in the nonpathogenic Acinetobacter sp. strain Tol 5, shows nonspecific, high adhesiveness to abiotic material surfaces as well as to biotic surfaces. AtaA is a homotrimer of polypeptides comprising 3,630 amino acids and forms long nanofibers; therefore, it is too large and structurally complex to be produced as a recombinant protein. In this study, we isolated AtaA’s passenger domain (AtaA PSD), which is translocated to the cell surface through the C-terminal transmembrane domain and exhibits biological functions, using a new method. We introduced a protease recognition site and reaped AtaA nanofibers 225 nm in length from the cell surface through proteolytic cleavage with a specific protease. Biochemical and biophysical analyses of the purified native AtaA PSD revealed that it has a stable structure under alkaline and acidic conditions. Temperatures above 80 °C, which disrupted AtaA’s higher-order structure but maintained the full-length AtaA polypeptide, inactivated AtaA’s nonspecific adhesiveness, suggesting that the stickiness of AtaA requires its 3D structure. This finding refutes the widespread but vague speculation that large unfolded polypeptides readily stick to various surfaces. PMID:27305955

  9. Clustering of 3D-Structure Similarity Based Network of Secondary Metabolites Reveals Their Relationships with Biological Activities.

    PubMed

    Ohtana, Yuki; Abdullah, Azian Azamimi; Altaf-Ul-Amin, Md; Huang, Ming; Ono, Naoaki; Sato, Tetsuo; Sugiura, Tadao; Horai, Hisayuki; Nakamura, Yukiko; Morita Hirai, Aki; Lange, Klaus W; Kibinge, Nelson K; Katsuragi, Tetsuo; Shirai, Tsuyoshi; Kanaya, Shigehiko

    2014-12-01

    Developing database systems connecting diverse species based on omics is the most important theme in big data biology. To attain this purpose, we have developed KNApSAcK Family Databases, which are utilized in a number of researches in metabolomics. In the present study, we have developed a network-based approach to analyze relationships between 3D structure and biological activity of metabolites consisting of four steps as follows: construction of a network of metabolites based on structural similarity (Step 1), classification of metabolites into structure groups (Step 2), assessment of statistically significant relations between structure groups and biological activities (Step 3), and 2-dimensional clustering of the constructed data matrix based on statistically significant relations between structure groups and biological activities (Step 4). Applying this method to a data set consisting of 2072 secondary metabolites and 140 biological activities reported in KNApSAcK Metabolite Activity DB, we obtained 983 statistically significant structure group-biological activity pairs. As a whole, we systematically analyzed the relationship between 3D-chemical structures of metabolites and biological activities.

  10. 3D Structure Generation, Molecular Dynamics and Docking Studies of IRHOM2 Protein Involved in Cancer & Rheumatoid Arthritis.

    PubMed

    Raj, Utkarsh; Kumar, Himansu; Varadwaj, Pritish Kumar

    2015-01-01

    A short-lived membrane protein IRHOM2 pedals a cascade of events by regulating Epidermal Growth Factor Receptor (EGFR) signalling in parallel with metalloproteases which results their involvement in cancer as well as in rheumatoid arthritis. Therefore, IRHOM2 is a potential therapeutic drug target for these diseases, but its 3D-structure has not been reported yet. In this study, the three-dimensional structure of the IRHOM2 protein was generated using I-TASSER (Iterative Threading Assembly Refinement) server. The modeled structure of IRHOM2 receptor was validated using various Structural Analysis and Verification Server (SAVES) in which 99.7% of amino acid residues are present in the favoured regions of the Ramachandran Plot. Further, the refined modeled structure was subjected to molecular dynamics simulation & docking analysis. Virtual screening studies were carried out using Glide with various selective libraries containing 24552 compounds and the analysis indicated extensive hydrogen bonding network and hydrophobic interactions which play a significant role in its binding. Docking results were analyzed for high ranking compounds using a consensus based docking score to calculate the binding affinity as a measure of protein-ligand interactions. The top ranking molecule against IRHOM2 active site has a glide g-score of -12.565 kcal/mol and glide e-model score of -74.967 with 3 hydrogen bonds and 11 hydrophobic contacts. This compound may act as probable inhibitor against these chronic diseases but further in vitro studies are required.

  11. The case for intrinsically disordered proteins playing contributory roles in molecular recognition without a stable 3D structure

    PubMed Central

    Uversky, Vladimir N.

    2013-01-01

    The classical ‘lock-and-key’ and ‘induced-fit’ mechanisms for binding both originated in attempts to explain features of enzyme catalysis. For both of these mechanisms and for their recent refinements, enzyme catalysis requires exquisite spatial and electronic complementarity between the substrate and the catalyst. Thus, binding models derived from models originally based on catalysis will be highly biased towards mechanisms that utilize structural complementarity. If mere binding without catalysis is the endpoint, then the structural requirements for the interaction become much more relaxed. Recent observations on specific examples suggest that this relaxation can reach an extreme lack of specific 3D structure, leading to molecular recognition with biological consequences that depend not only upon structural and electrostatic complementarity between the binding partners but also upon kinetic, entropic, and generalized electrostatic effects. In addition to this discussion of binding without fixed structure, examples in which unstructured regions carry out important biological functions not involving molecular recognition will also be discussed. Finally, we discuss whether ‘intrinsically disordered protein’ (IDP) represents a useful new concept. PMID:23361308

  12. PONDEROSA-C/S: client-server based software package for automated protein 3D structure determination.

    PubMed

    Lee, Woonghee; Stark, Jaime L; Markley, John L

    2014-11-01

    Peak-picking Of Noe Data Enabled by Restriction Of Shift Assignments-Client Server (PONDEROSA-C/S) builds on the original PONDEROSA software (Lee et al. in Bioinformatics 27:1727-1728. doi: 10.1093/bioinformatics/btr200, 2011) and includes improved features for structure calculation and refinement. PONDEROSA-C/S consists of three programs: Ponderosa Server, Ponderosa Client, and Ponderosa Analyzer. PONDEROSA-C/S takes as input the protein sequence, a list of assigned chemical shifts, and nuclear Overhauser data sets ((13)C- and/or (15)N-NOESY). The output is a set of assigned NOEs and 3D structural models for the protein. Ponderosa Analyzer supports the visualization, validation, and refinement of the results from Ponderosa Server. These tools enable semi-automated NMR-based structure determination of proteins in a rapid and robust fashion. We present examples showing the use of PONDEROSA-C/S in solving structures of four proteins: two that enable comparison with the original PONDEROSA package, and two from the Critical Assessment of automated Structure Determination by NMR (Rosato et al. in Nat Methods 6:625-626. doi: 10.1038/nmeth0909-625 , 2009) competition. The software package can be downloaded freely in binary format from http://pine.nmrfam.wisc.edu/download_packages.html. Registered users of the National Magnetic Resonance Facility at Madison can submit jobs to the PONDEROSA-C/S server at http://ponderosa.nmrfam.wisc.edu, where instructions, tutorials, and instructions can be found. Structures are normally returned within 1-2 days.

  13. Probing Protein 3D Structures and Conformational Changes Using Electrochemistry-Assisted Isotope Labeling Cross-Linking Mass Spectrometry.

    PubMed

    Zheng, Qiuling; Zhang, Hao; Wu, Shiyong; Chen, Hao

    2016-05-01

    This study presents a new chemical cross-linking mass spectrometry (MS) method in combination with electrochemistry and isotope labeling strategy for probing both protein three-dimensional (3D) structures and conformational changes. For the former purpose, the target protein/protein complex is cross-linked with equal mole of premixed light and heavy isotope labeled cross-linkers carrying electrochemically reducible disulfide bonds (i.e., DSP-d0 and DSP-d8 in this study, DSP = dithiobis[succinimidyl propionate]), digested and then electrochemically reduced followed with online MS analysis. Cross-links can be quickly identified because of their reduced intensities upon electrolysis and the presence of doublet isotopic peak characteristics. In addition, electroreduction converts cross-links into linear peptides, facilitating MS/MS analysis to gain increased information about their sequences and modification sites. For the latter purpose of probing protein conformational changes, an altered procedure is adopted, in which the protein in two different conformations is cross-linked using DSP-d0 and DSP-d8 separately, and then the two protein samples are mixed in 1:1 molar ratio. The merged sample is subjected to digestion and electrochemical mass spectrometric analysis. In such a comparative cross-linking experiment, cross-links could still be rapidly recognized based on their responses to electrolysis. More importantly, the ion intensity ratios of light and heavy isotope labeled cross-links reveal the conformational changes of the protein, as exemplified by examining the effect of Ca(2+) on calmodulin conformation alternation. This new cross-linking MS method is fast and would have high value in structural biology. Graphical Abstract ᅟ.

  14. Probing Protein 3D Structures and Conformational Changes Using Electrochemistry-Assisted Isotope Labeling Cross-Linking Mass Spectrometry

    NASA Astrophysics Data System (ADS)

    Zheng, Qiuling; Zhang, Hao; Wu, Shiyong; Chen, Hao

    2016-05-01

    This study presents a new chemical cross-linking mass spectrometry (MS) method in combination with electrochemistry and isotope labeling strategy for probing both protein three-dimensional (3D) structures and conformational changes. For the former purpose, the target protein/protein complex is cross-linked with equal mole of premixed light and heavy isotope labeled cross-linkers carrying electrochemically reducible disulfide bonds (i.e., DSP-d0 and DSP-d8 in this study, DSP = dithiobis[succinimidyl propionate]), digested and then electrochemically reduced followed with online MS analysis. Cross-links can be quickly identified because of their reduced intensities upon electrolysis and the presence of doublet isotopic peak characteristics. In addition, electroreduction converts cross-links into linear peptides, facilitating MS/MS analysis to gain increased information about their sequences and modification sites. For the latter purpose of probing protein conformational changes, an altered procedure is adopted, in which the protein in two different conformations is cross-linked using DSP-d0 and DSP-d8 separately, and then the two protein samples are mixed in 1:1 molar ratio. The merged sample is subjected to digestion and electrochemical mass spectrometric analysis. In such a comparative cross-linking experiment, cross-links could still be rapidly recognized based on their responses to electrolysis. More importantly, the ion intensity ratios of light and heavy isotope labeled cross-links reveal the conformational changes of the protein, as exemplified by examining the effect of Ca2+ on calmodulin conformation alternation. This new cross-linking MS method is fast and would have high value in structural biology.

  15. Helium Ion Microscopy Visualizes Lipid Nanodomains in Mammalian Cells.

    PubMed

    Schürmann, Matthias; Frese, Natalie; Beyer, André; Heimann, Peter; Widera, Darius; Mönkemöller, Viola; Huser, Thomas; Kaltschmidt, Barbara; Kaltschmidt, Christian; Gölzhäuser, Armin

    2015-11-18

    Cell membranes are composed of 2D bilayers of amphipathic lipids, which allow a lateral movement of the respective membrane components. These components are arranged in an inhomogeneous manner as transient micro- and nanodomains, which are believed to be crucially involved in the regulation of signal transduction pathways in mammalian cells. Because of their small size (diameter 10-200 nm), membrane nanodomains cannot be directly imaged using conventional light microscopy. Here, direct visualization of cell membrane nanodomains by helium ion microscopy (HIM) is presented. It is shown that HIM is capable to image biological specimens without any conductive coating and that HIM images clearly allow the identification of nanodomains in the ultrastructure of membranes with 1.5 nm resolution. The shape of these nanodomains is preserved by fixation of the surrounding unsaturated fatty acids while saturated fatty acids inside the nanodomains are selectively removed. Atomic force microscopy, fluorescence microscopy, 3D structured illumination microscopy, and direct stochastic optical reconstruction microscopy provide additional evidence that the structures in the HIM images of cell membranes originate from membrane nanodomains. The nanodomains observed by HIM have an average diameter of 20 nm and are densely arranged with a minimal nearest neighbor distance of ≈ 15 nm.

  16. Molecular Cloning and 3D Structure Modeling of APEX1, DNA Base Excision Repair Enzyme from the Camel, Camelus dromedarius

    PubMed Central

    Ataya, Farid Shokry; Fouad, Dalia; Malik, Ajamaluddin; Saeed, Hesham Mahmoud

    2012-01-01

    The domesticated one-humped camel, Camelus dromedarius, is one of the most important animals in the Arabian Desert. It is exposed most of its life to both intrinsic and extrinsic genotoxic factors that are known to cause gross DNA alterations in many organisms. Ionic radiation and sunlight are known producers of Reactive Oxygen Species (ROS), one of the causes for DNA lesions. The damaged DNA is repaired by many enzymes, among of them Base Excision Repair enzymes, producing the highly mutagenic apurinic/apyrimidinicsites (AP sites). Therefore, recognition of AP sites is fundamental to cell/organism survival. In the present work, the full coding sequence of a putative cAPEX1 gene was amplified for the first time from C. dromedarius by RT-PCR and cloned (NCBI accession number are HM209828 and ADJ96599 for nucleotides and amino acids, respectively). cDNA sequencing was deduced to be 1041 nucleotides, of which 954 nucleotides encode a protein of 318 amino acids, similar to the coding region of the APEX1 gene and the protein from many other species. The calculated molecular weight and isoelectric point of cAPEX1 using Bioinformatics tools was 35.5 kDa and 8.11, respectively. The relative expressions of cAPEX1 in camel kidney, spleen, lung and testis were examined using qPCR and compared with that of the liver using a 18S ribosomal subunit as endogenous control. The highest level of cAPEX1 transcript was found in the testis; 325% higher than the liver, followed by spleen (87%), kidney (20%) and lung (5%), respectively. The cAPEX1 is 94%–97% similar to their mammalian counterparts. Phylogenetic analysis revealed that cAPEX1 is grouped together with that of S. scrofa. The predicted 3D structure of cAPEX1 has similar folds and topology with the human (hAPEX1). The root-mean-square deviation (rmsd) between cAPEX1 and hAPEX1 was 0.582 and the Q-score was 0.939. PMID:22942721

  17. A study of 3D structure of nighttime electron density enhancement in the mid-latitude ionosphere by GPS tomography

    NASA Astrophysics Data System (ADS)

    Chen, C.; Saito, A.

    2011-12-01

    The mid-latitude summer nighttime anomaly (MSNA) is a feature that the nighttime electron density larger than that in the daytime mid-latitude ionosphere. This anomaly was first detected in the southern hemisphere five decades ago and observed in the northern hemisphere recently by ionosondes and satellites. Previous studies presented the electron density structure of MSNA by using COSMIC occultation data and found that MSNA is clearly seen around 300 km altitude during local summer. However, due to lack of observation, the day-to-day variation of MSNA was not investigated. A GPS tomography method by SPEL of Kyoto University using the total electron content (TEC) data measured by the ground-based GPS receiver network is employed in this study. The wide coverage and continuous observation of GPS receivers are suitable for investigating the spatial and day-to-day variations of ionospheric electron densities. The algorithm of the GPS tomography developed by SPEL of Kyoto University use a constraint condition that the gradient of election density tends to be smooth in the horizontal direction and steep in the vicinity of the F2 peak, instead of inputting the initial conditions. Therefore, the algorithm is independent of any ionospheric and plasmaspheric electron density distribution models. The dense ground-based GPS receiver network around European region is used to study the three dimensional (3D) structure of MSNA with GPS tomography. Results show that the MSNA usually appear around the geomagnetic mid-latitude region during local summer nighttime. The feature of MSNA is most obvious at the ionospheric F2-peak altitudes. The result also shows a day-to-day variation in the formation of MSNA, in terms of the occurrence time, intensity, and spatial extent. The tomographic results are compared with the ionosondes, satellites, and radar measurements. A theoretical model simulation, SAMI2, is also used to further discuss the mechanism of MSNA. The comparison with other

  18. The predicted 3D structure of the human D2 dopamine receptor and the binding site and binding affinities for agonists and antagonists

    NASA Astrophysics Data System (ADS)

    Kalani, M. Yashar S.; Vaidehi, Nagarajan; Hall, Spencer E.; Trabanino, Rene J.; Freddolino, Peter L.; Kalani, Maziyar A.; Floriano, Wely B.; Tak Kam, Victor Wai; Goddard, William A., III

    2004-03-01

    Dopamine neurotransmitter and its receptors play a critical role in the cell signaling process responsible for information transfer in neurons functioning in the nervous system. Development of improved therapeutics for such disorders as Parkinson's disease and schizophrenia would be significantly enhanced with the availability of the 3D structure for the dopamine receptors and of the binding site for dopamine and other agonists and antagonists. We report here the 3D structure of the long isoform of the human D2 dopamine receptor, predicted from primary sequence using first-principles theoretical and computational techniques (i.e., we did not use bioinformatic or experimental 3D structural information in predicting structures). The predicted 3D structure is validated by comparison of the predicted binding site and the relative binding affinities of dopamine, three known dopamine agonists (antiparkinsonian), and seven known antagonists (antipsychotic) in the D2 receptor to experimentally determined values. These structures correctly predict the critical residues for binding dopamine and several antagonists, identified by mutation studies, and give relative binding affinities that correlate well with experiments. The predicted binding site for dopamine and agonists is located between transmembrane (TM) helices 3, 4, 5, and 6, whereas the best antagonists bind to a site involving TM helices 2, 3, 4, 6, and 7 with minimal contacts to TM helix 5. We identify characteristic differences between the binding sites of agonists and antagonists.

  19. Airborne & SAR Synergy Reveals the 3D Structure of Air Bubble Entrainment in Internal Waves and Frontal Zones

    NASA Astrophysics Data System (ADS)

    da Silva, J. C. B.; Magalhaes, J. M.; Batista, M.; Gostiaux, L.; Gerkema, T.; New, A. L.

    2013-03-01

    spectral range 8-12 μm. With a nominal ground resolution of approximately 1.5 meters (at an altitude of 500 meters) it is capable to detect fine structure associated to turbulence. The LiDAR system that has been used is the Leica ALS50-II (1064nm) with a hit rate greater than 1 hit per square meter and a vertical resolution of approximately 15 cm. Both systems were available simultaneously, together with the hyperspectral system and the RCD105 39Mpx digital camera, integrated with the LiDAR navigation system. We analyse the airborne data together with a comprehensive dataset of satellite Synthetic Aperture Radar (SAR) that includes ENVISAT and TerraSAR-X images. In addition, in situ observations in the near-shore zone were obtained in a previous experiment (Project SPOTIWAVE-II POCI/MAR/57836/2004 funded by the Portuguese FCT) during the summer period in 2006. These included thermistor chain measurements along the water column that captured the vertical structure of shoaling internal (tidal) waves and ISWs close to the breaking point. The SAR and airborne images were obtained in light wind conditions, in the near-shore zone, and in the presence of ISWs. The LiDAR images revealed sub-surface structures (some 1-2 m below the sea surface) that were co-located with surface films. These film slicks were induced by the convergent fields of internal waves and upwelling fronts. Some of the sub-surface features were located over the front slopes of the internal waves, which coincides with the internal wave slick band visible in the aerial photos and hyperspectral systems. Our flight measurements revealed thermal features similar to “boils” of cold water within the wake of (admittedly breaking) internal waves. These features are consistent with the previous in situ measurements of breaking ISWs. In this paper we will show coincident multi-sensor airborne and satellite SAR observations that reveal the 3D structure of air bubble entrainment in the internal wave field and frontal

  20. Structured Illumination Microscopy and a Quantitative Image Analysis for the Detection of Positive Margins in a Pre-Clinical Genetically Engineered Mouse Model of Sarcoma

    PubMed Central

    Fu, Henry L.; Mueller, Jenna L.; Whitley, Melodi J.; Cardona, Diana M.; Willett, Rebecca M.; Kirsch, David G.; Brown, J. Quincy; Ramanujam, Nimmi

    2016-01-01

    Intraoperative assessment of surgical margins is critical to ensuring residual tumor does not remain in a patient. Previously, we developed a fluorescence structured illumination microscope (SIM) system with a single-shot field of view (FOV) of 2.1×1.6 mm (3.4 mm2) and sub-cellular resolution (4.4 μm). The goal of this study was to test the utility of this technology for the detection of residual disease in a genetically engineered mouse model of sarcoma. Primary soft tissue sarcomas were generated in the hindlimb and after the tumor was surgically removed, the relevant margin was stained with acridine orange (AO), a vital stain that brightly stains cell nuclei and fibrous tissues. The tissues were imaged with the SIM system with the primary goal of visualizing fluorescent features from tumor nuclei. Given the heterogeneity of the background tissue (presence of adipose tissue and muscle), an algorithm known as maximally stable extremal regions (MSER) was optimized and applied to the images to specifically segment nuclear features. A logistic regression model was used to classify a tissue site as positive or negative by calculating area fraction and shape of the segmented features that were present and the resulting receiver operator curve (ROC) was generated by varying the probability threshold. Based on the ROC curves, the model was able to classify tumor and normal tissue with 77% sensitivity and 81% specificity (Youden’s index). For an unbiased measure of the model performance, it was applied to a separate validation dataset that resulted in 73% sensitivity and 80% specificity. When this approach was applied to representative whole margins, for a tumor probability threshold of 50%, only 1.2% of all regions from the negative margin exceeded this threshold, while over 14.8% of all regions from the positive margin exceeded this threshold. PMID:26799613

  1. Oviductosome-Sperm Membrane Interaction in Cargo Delivery: DETECTION OF FUSION AND UNDERLYING MOLECULAR PLAYERS USING THREE-DIMENSIONAL SUPER-RESOLUTION STRUCTURED ILLUMINATION MICROSCOPY (SR-SIM).

    PubMed

    Al-Dossary, Amal A; Bathala, Pradeepthi; Caplan, Jeffrey L; Martin-DeLeon, Patricia A

    2015-07-17

    Oviductosomes ((OVS), exosomes/microvesicles), which deliver the Ca(2+) efflux pump, plasma membrane Ca(2+)ATPase 4 (PMCA4), to sperm are likely to play an important role in sperm fertilizing ability (Al-Dossary, A. A., Strehler, E. E., and Martin-DeLeon, P. A. (2013) PloS one 8, e80181). It is unknown how exosomes/microvesicles deliver transmembrane proteins such as PMCA4 to sperm. Here we define a novel experimental approach for the assessment of the interaction of OVS with sperm at a nanoscale level, using a lipophilic dye (FM4-64FX) and three-dimensional SR/SIM, which has an 8-fold increase in volumetric resolution, compared with conventional confocal microscopy. Coincubation assays detected fusion of prelabeled OVS with sperm, primarily over the head and midpiece. Immunofluorescence revealed oviductosomal delivery of PMCA4a to WT and Pmca4 KO sperm, and also endogenous PMCA4a on the inner acrosomal membrane. Fusion was confirmed by transmission immunoelectron microscopy, showing immunogold particles in OVS, and fusion stalks on sperm membrane. Immunofluorescence colocalized OVS with the αv integrin subunit which, along with CD9, resides primarily on the sperm head and midpiece. In capacitated and acrosome reacted sperm, fusion was significantly (p < 0.001) inhibited by blocking integrin/ligand interactions via antibodies, exogenous ligands (vitronectin and fibronectin), and their RGD recognition motif. Our results provide evidence that receptor/ligand interactions, involving αvβ3 and α5β1integrins on sperm and OVS, facilitate fusion of OVS in the delivery of transmembrane proteins to sperm. The mechanism uncovered is likely to be also involved in cargo delivery of prostasomes, epididymosomes, and uterosomes.

  2. Demand illumination control apparatus

    NASA Technical Reports Server (NTRS)

    Warren, Carl (Inventor); Arline, Jimmie (Inventor); LaPalme, Julius (Inventor)

    1981-01-01

    Solar illuminating compensating apparatus is disclosed whereby the interior of a building is illuminated to a substantially constant, predetermined level of light intensity by a combination of natural illumination from the sun and artificial illumination from electricity wherein the intensity of said artificial illumination is controlled by fully electronic means which increases the level of artificial illumination when the natural illumination is inadequate and vice versa.

  3. Computing the 3-D structure of viruses from unoriented cryo electron microscope images: a fast algorithm for a statistical approach.

    PubMed

    Lee, Junghoon; Zheng, Yili; Doerschuk, Peter C

    2006-01-01

    In a cryo electron microscopy experiment, the data is noisy 2-D projection images of the 3-D electron scattering intensity where the orientation of the projections is not known. In previous work we have developed a solution for this problem based on a maximum likelihood estimator that is computed by an expectation maximization algorithm. In the expectation maximization algorithm the expensive step is the expectation which requires numerical evaluation of 3- or 5-dimensional integrations of a square matrix of dimension equal to the number of Fourier series coefficients used to describe the 3-D reconstruction. By taking advantage of the rotational properties of spherical harmonics, we can reduce the integrations of a matrix to integrations of a scalar. The key property is that a rotated spherical harmonic can be expressed as a linear combination of the other harmonics of the same order and the weights in the linear combination factor so that each of the three factors is a function of only one of the Euler angles describing the orientation of the projection. Numerical example of the reconstructions is provided based on Nudaurelia Omega Capensis virus.

  4. Illumination design for semiconductor backlight inspection and application extensions

    NASA Astrophysics Data System (ADS)

    Zhou, Wei; Rutherford, Todd; Hart, Darcy

    2013-09-01

    High speed strobe based illumination scheme is one of the most critical factors for high throughput semiconductor defect inspection applications. HB LEDs are always the first and best options for such applications due to numerous unique advantages such as excellent spatial and temporal stability, fast responding time, large and linear intensity dynamic range and no heat issue for the extremely low duty cycle applications. For some applications where a large area is required to be illuminated simultaneously, it remains a great challenge to efficiently package a large amount of HB-LEDs in a highly confined 3D space, to generate a seamless illuminated area with high luminance efficiency and spatial uniformity. A novel 3D structured collimation lens is presented in this paper. The non-circular edge shape reduces the intensity drop at the channel boundaries, while the secondary curvatures on the top of the collimator lens efficiently guides the light into desired angular space. The number of the edges and the radius of the top surface curvature are control parameters for the system level performance and the manufacture cost trade-off. The proposed 3D structured LED collimation lens also maintains the benefits of traditional LED collimation lens such as coupling efficiency and mold manufacture capability. The applications can be extended into other non-illumination area like parallelism measurement and solar panel concentrator etc.

  5. Auto-calibrated scanning-angle prism-type total internal reflection microscopy for nanometer-precision axial position determination and optional variable-illumination-depth pseudo total internal reflection microscopy

    DOEpatents

    Fang, Ning; Sun, Wei

    2015-04-21

    A method, apparatus, and system for improved VA-TIRFM microscopy. The method comprises automatically controlled calibration of one or more laser sources by precise control of presentation of each laser relative a sample for small incremental changes of incident angle over a range of critical TIR angles. The calibration then allows precise scanning of the sample for any of those calibrated angles for higher and more accurate resolution, and better reconstruction of the scans for super resolution reconstruction of the sample. Optionally the system can be controlled for incident angles of the excitation laser at sub-critical angles for pseudo TIRFM. Optionally both above-critical angle and sub critical angle measurements can be accomplished with the same system.

  6. Measurement of replication structures at the nanometer scale using super-resolution light microscopy

    PubMed Central

    Baddeley, D.; Chagin, V. O.; Schermelleh, L.; Martin, S.; Pombo, A.; Carlton, P. M.; Gahl, A.; Domaing, P.; Birk, U.; Leonhardt, H.; Cremer, C.; Cardoso, M. C.

    2010-01-01

    DNA replication, similar to other cellular processes, occurs within dynamic macromolecular structures. Any comprehensive understanding ultimately requires quantitative data to establish and test models of genome duplication. We used two different super-resolution light microscopy techniques to directly measure and compare the size and numbers of replication foci in mammalian cells. This analysis showed that replication foci vary in size from 210 nm down to 40 nm. Remarkably, spatially modulated illumination (SMI) and 3D-structured illumination microscopy (3D-SIM) both showed an average size of 125 nm that was conserved throughout S-phase and independent of the labeling method, suggesting a basic unit of genome duplication. Interestingly, the improved optical 3D resolution identified 3- to 5-fold more distinct replication foci than previously reported. These results show that optical nanoscopy techniques enable accurate measurements of cellular structures at a level previously achieved only by electron microscopy and highlight the possibility of high-throughput, multispectral 3D analyses. PMID:19864256

  7. Measurement of replication structures at the nanometer scale using super-resolution light microscopy.

    PubMed

    Baddeley, D; Chagin, V O; Schermelleh, L; Martin, S; Pombo, A; Carlton, P M; Gahl, A; Domaing, P; Birk, U; Leonhardt, H; Cremer, C; Cardoso, M C

    2010-01-01

    DNA replication, similar to other cellular processes, occurs within dynamic macromolecular structures. Any comprehensive understanding ultimately requires quantitative data to establish and test models of genome duplication. We used two different super-resolution light microscopy techniques to directly measure and compare the size and numbers of replication foci in mammalian cells. This analysis showed that replication foci vary in size from 210 nm down to 40 nm. Remarkably, spatially modulated illumination (SMI) and 3D-structured illumination microscopy (3D-SIM) both showed an average size of 125 nm that was conserved throughout S-phase and independent of the labeling method, suggesting a basic unit of genome duplication. Interestingly, the improved optical 3D resolution identified 3- to 5-fold more distinct replication foci than previously reported. These results show that optical nanoscopy techniques enable accurate measurements of cellular structures at a level previously achieved only by electron microscopy and highlight the possibility of high-throughput, multispectral 3D analyses.

  8. Hotsphere illumination

    NASA Astrophysics Data System (ADS)

    Razavi, Bahar S.; Kuzyakov, Yakov

    2016-04-01

    Soils are the most heterogeneous parts of the biosphere, with an extremely high differentiation of properties and processes at all spatial and temporal scales. Importance of the hotspheres such as rhizosphere, detritusphere, porosphere (including drilosphere and biopores), hyphasphere and spermosphere, calls for spatially explicit methods to illuminate distribution of microbial activities in these hotspheres (Kuzyakov and Blagodatskaya, 2015). Zymography technique has previously been adapted to visualize the spatial dynamics of enzyme activities in rhizosphere (Spohn and Kuzyakov, 2014). Here, we further developed soil zymography to obtain a higher resolution of enzyme activities by enabling direct contact of substrate-saturated membranes with soil. For the first time, we aimed at quantitative imaging of enzyme activities in various hotspheres. We calculated and compared percentage of enzymatic hotspots of five hotspheres: spermosphere, rhizosphere, detritusphere, drilosphere and biopores. Spatial distribution of activities of two enzymes: β-glucosidase and leucine amino peptidase were analyzed in the spermosphere, rhizosphere and detritusphere of maize and lentil. Zymography has been done 3 days (spermosphere), 14 days (rhizosphere) after sowing and 21 days after cutting plant (detritusphere). Spatial resolution of fluorescent images was improved by direct application fluorogenically labelled substrates on the soil surface. Such improvement enabled to visualize enzyme distribution of mycorrhiza hypha on the rhizobox surface. Further, to visualize the 2D distribution of the enzyme activities in porosphere, we placed earthworms (Lumbricus terrestris), (drilosphere) and ground beetle species Platynus dorsalis Pont. (Coleoptera; Carabidae), (biopore), in transparent boxes for 2weeks. The developed in situ zymography visualized the heterogeneity of enzyme activities along and across the roots. Spatial patterns of enzyme activities as a function of distance along the

  9. Constructing a 3D structural block diagram of the Central Basin in Marmara Sea by means of bathymetric and seismic data

    NASA Astrophysics Data System (ADS)

    Demirbağ, Emin; Kurt, Hülya; Düşünür, Doğa; Sarıkavak, Kerim; Çetin, Suna

    2007-12-01

    In this study we made a comparative interpretation of multibeam bathymetric and seismic reflection data with different resolutions and penetration properties collected in the Central Basin of the Marmara Sea. Our main objectives were (i) to investigate and compare the active tectonic deformation observed on the sea bottom and within the uppermost sedimentary layers to that of the deep-seated deformation within the limits of resolution and penetration of the available geophysical data and (ii) to build a three-dimensional (3D) block diagram of the active tectonic and buried features by means of a sliced mapping technique. In this approach, we produced slice maps of the active and buried structural features at selected depths and then combined them to form a 3D structural block diagram. Motivation for our work was to produce a 3D structural diagram to derive a more detailed image of the structural features in the Central Basin where there is no available 3D seismic data. The observations from the bathymetry and seismic data and developed 3D diagram support the presence of a through-going strike-slip fault that forms a rotational depression zone against a right-stepping strike-slip faulting causing a pull-apart basin in the Central Depression zone.

  10. 3D structure prediction of human β1-adrenergic receptor via threading-based homology modeling for implications in structure-based drug designing.

    PubMed

    Ul-Haq, Zaheer; Saeed, Maria; Halim, Sobia Ahsan; Khan, Waqasuddin

    2015-01-01

    Dilated cardiomyopathy is a disease of left ventricular dysfunction accompanied by impairment of the β1-adrenergic receptor (β1-AR) signal cascade. The disturbed β1-AR function may be based on an elevated sympathetic tone observed in patients with heart failure. Prolonged adrenergic stimulation may induce metabolic and electrophysiological disturbances in the myocardium, resulting in tachyarrhythmia that leads to the development of heart failure in human and sudden death. Hence, β1-AR is considered as a promising drug target but attempts to develop effective and specific drug against this tempting pharmaceutical target is slowed down due to the lack of 3D structure of Homo sapiens β1-AR (hsβADR1). This study encompasses elucidation of 3D structural and physicochemical properties of hsβADR1 via threading-based homology modeling. Furthermore, the docking performance of several docking programs including Surflex-Dock, FRED, and GOLD were validated by re-docking and cross-docking experiments. GOLD and Surflex-Dock performed best in re-docking and cross docking experiments, respectively. Consequently, Surflex-Dock was used to predict the binding modes of four hsβADR1 agonists. This study provides clear understanding of hsβADR1 structure and its binding mechanism, thus help in providing the remedial solutions of cardiovascular, effective treatment of asthma and other diseases caused by malfunctioning of the target protein.

  11. Identification of the critical residues for the function of vacuolar H⁺-pyrophosphatase by mutational analysis based on the 3D structure.

    PubMed

    Asaoka, Mariko; Segami, Shoji; Maeshima, Masayoshi

    2014-12-01

    H(+)-translocating pyrophosphatase (H(+)-PPase) converts energy from hydrolysis of inorganic pyrophosphate (PPi) to active H(+) translocation across membranes. From the 3D structure resolved by crystallography, 17 amino acid residues in several domains of mung bean (Vigna radiata) enzyme were selected and substituted with alanine individually. The mutant enzymes were expressed in yeast cells to evaluate their biochemical role. The highly conserved residues in the substrate-binding site (T249, D269, D507 and N534) were shown to be essential for PPi hydrolysis and H(+) pump. The amino acid substitution of residues in the H(+) translocation pathway (I545, L555, N738, V746 and L749) resulted in mild decrease in the PPase activity and strong suppression of the H(+) pump. These results suggest that the decoupling of PPi hydrolysis and active H(+) translocation occurred in these five mutants including I545A. The alanine substitution of the C124 and C132, which form an intra-molecular disulfide bond, did not affect the enzyme activity. The modifications of the other residues in the vacuolar lumen loop, and M15 had relatively mild effect on the enzyme function. Functional roles of the 17 residues are discussed with consideration of the 3D structural information.

  12. On the integration of protein-protein interaction networks with gene expression and 3D structural data: What can be gained?

    NASA Astrophysics Data System (ADS)

    Bertolazzi, Paola; Bock, Mary Ellen; Guerra, Concettina; Paci, Paola; Santoni, Daniele

    2014-06-01

    The biological role of proteins has been analyzed from different perspectives, initially by considering proteins as isolated biological entities, then as cooperating entities that perform their function by interacting with other molecules. There are other dimensions that are important for the complete understanding of the biological processes: time and location. However a protein is rarely annotated with temporal and spatial information. Experimental Protein-Proteins Interaction (PPI) data are static; furthermore they generally do not include transient interactions which are a considerable fraction of the interactome of many organisms. One way to incorporate temporal and condition information is to use other sources of information, such as gene expression data and 3D structural data. Here we review work done to understand the insight that can be gained by enriching PPI data with gene expression and 3D structural data. In particular, we address the following questions: Can the dynamics of a single protein or of an interaction be accurately derived from these data? Can the assembly-disassembly of protein complexes be traced over time? What type of topological changes occur in a PPI network architecture over time?

  13. A combined Raman microscopy, XRF and SEM-EDX study of three valuable objects - A large painted leather screen and two illuminated title pages in 17th century books of ordinances of the Worshipful Company of Barbers, London

    NASA Astrophysics Data System (ADS)

    Chaplin, Tracey D.; Clark, Robin J. H.; Martinón-Torres, Marcos

    2010-07-01

    Raman microscopy has been used to identify the pigments decorating three valuable items owned by the Worshipful Company of Barbers (established in 1308 in London), one being a large leather screen dating to before 1712, the other two being illuminated title pages of books of ordinances of the Company dating to 1605 and 1658. Pigments which could not be fully characterised by this technique (particularly the green paints) have also been subject to XRF or SEM-EDX analysis. The combined analytical approach has shown that the pigments identified on all three items are typical of those in use as artists' pigments in the 17th C and include azurite, indigo, vermilion, red lead, pink and yellow lakes, verdigris, lead white, calcite (and chalk), gypsum, carbon-based black, and gold and silver leaf. However in the case of the screen alone, restoration in the 1980s has been carried out with different pigments - haematite, phthalocyanine green, rutile, and a mixture of azurite, malachite and barium sulfate. This work constitutes the first in-depth study of painted leatherwork and demonstrates that the palette used for this purpose is similar to that used on other works of art of the same date. It has also allowed the original colour schemes of the decorations to be determined where pigment degradation has occurred. The combined analysis has also provided a more complete understanding of the materials used for, or on, objects to which access is limited.

  14. RNA 3D Structure Modeling by Combination of Template-Based Method ModeRNA, Template-Free Folding with SimRNA, and Refinement with QRNAS.

    PubMed

    Piatkowski, Pawel; Kasprzak, Joanna M; Kumar, Deepak; Magnus, Marcin; Chojnowski, Grzegorz; Bujnicki, Janusz M

    2016-01-01

    RNA encompasses an essential part of all known forms of life. The functions of many RNA molecules are dependent on their ability to form complex three-dimensional (3D) structures. However, experimental determination of RNA 3D structures is laborious and challenging, and therefore, the majority of known RNAs remain structurally uncharacterized. To address this problem, computational structure prediction methods were developed that either utilize information derived from known structures of other RNA molecules (by way of template-based modeling) or attempt to simulate the physical process of RNA structure formation (by way of template-free modeling). All computational methods suffer from various limitations that make theoretical models less reliable than high-resolution experimentally determined structures. This chapter provides a protocol for computational modeling of RNA 3D structure that overcomes major limitations by combining two complementary approaches: template-based modeling that is capable of predicting global architectures based on similarity to other molecules but often fails to predict local unique features, and template-free modeling that can predict the local folding, but is limited to modeling the structure of relatively small molecules. Here, we combine the use of a template-based method ModeRNA with a template-free method SimRNA. ModeRNA requires a sequence alignment of the target RNA sequence to be modeled with a template of the known structure; it generates a model that predicts the structure of a conserved core and provides a starting point for modeling of variable regions. SimRNA can be used to fold small RNAs (<80 nt) without any additional structural information, and to refold parts of models for larger RNAs that have a correctly modeled core. ModeRNA can be either downloaded, compiled and run locally or run through a web interface at http://genesilico.pl/modernaserver/ . SimRNA is currently available to download for local use as a precompiled

  15. 3D structure prediction of TAS2R38 bitter receptors bound to agonists phenylthiocarbamide (PTC) and 6-n-propylthiouracil (PROP).

    PubMed

    Tan, Jun; Abrol, Ravinder; Trzaskowski, Bartosz; Goddard, William A

    2012-07-23

    The G protein-coupled receptor (GPCR) TAS2R38 is a bitter taste receptor that can respond to bitter compounds such as phenylthiocarbamide (PTC) and 6-n-propylthiouracil (PROP). This receptor was chosen because its four haplotypes (based on three residue site polymorphism) hTAS2R38PAV, hTAS2R38AVI, hTAS2R38AAI, and hTAS2R38PVV are known to have dramatically different responses to PTC and PROP. We aimed to identify the protein-ligand interaction features that determine whether the bitter taste signal from this receptor is sent to the cortex. To do this we predicted the 3D structures of the TAS2R38 bitter taste receptor using our new BiHelix and SuperBiHelix Monte Carlo methods (No experimental determinations of the 3D structure have been reported for any taste receptors.). We find that residue 262 (2nd position in the polymorphism) is involved in the interhelical hydrogen bond network stabilizing the GPCR structure in tasters (hTAS2R38PAV, hTAS2R38AAI, and hTAS2R38PVV), while it is not in the nontaster (hTAS2R38AVI). This suggests that the hydrogen bond interactions between TM3 and TM6 or between TM5 and TM6 may play a role in activating this GPCR. To further validate these structures, we used the DarwinDock method to predict the binding sites and 3D structures for PTC and PROP bound to hTAS2R38PAV, hTAS2R38AVI, hTAS2R38AAI, and hTAS2R38PVV, respectively. Our results show that PTC and PROP can form H-bonds with the backbone of residue 262 in the tasters (hTAS2R38PAV, hTAS2R38AAI, and hTAS2R38PVV) but not in the nontaster (hTAS2R38AVI). Thus it appears that the hydrogen bond interaction between TM3 and TM6 may activate the receptor to pass the ligand binding signal to intracellular processes and that the H-bond between agonists and residue 262 in tasters is involved in the bitter tasting. This is in agreement with experimental observations, providing validation of the predicted ligand-protein complexes and also a potential activation mechanism for the TAS2R38 receptor.

  16. Reflectance, illumination, and appearance in color constancy

    PubMed Central

    McCann, John J.; Parraman, Carinna; Rizzi, Alessandro

    2013-01-01

    We studied color constancy using a pair of identical 3-D Color Mondrian displays. We viewed one 3-D Mondrian in nearly uniform illumination, and the other in directional, nonuniform illumination. We used the three dimensional structures to modulate the light falling on the painted surfaces. The 3-D structures in the displays were a matching set of wooden blocks. Across Mondrian displays, each corresponding facet had the same paint on its surface. We used only 6 chromatic, and 5 achromatic paints applied to 104 block facets. The 3-D blocks add shadows and multiple reflections not found in flat Mondrians. Both 3-D Mondrians were viewed simultaneously, side-by-side. We used two techniques to measure correlation of appearance with surface reflectance. First, observers made magnitude estimates of changes in the appearances of identical reflectances. Second, an author painted a watercolor of the 3-D Mondrians. The watercolor's reflectances quantified the changes in appearances. While constancy generalizations about illumination and reflectance hold for flat Mondrians, they do not for 3-D Mondrians. A constant paint does not exhibit perfect color constancy, but rather shows significant shifts in lightness, hue and chroma in response to the structure in the nonuniform illumination. Color appearance depends on the spatial information in both the illumination and the reflectances of objects. The spatial information of the quanta catch from the array of retinal receptors generates sensations that have variable correlation with surface reflectance. Models of appearance in humans need to calculate the departures from perfect constancy measured here. This article provides a dataset of measurements of color appearances for computational models of sensation. PMID:24478738

  17. 3D Structured Grid Adaptation

    NASA Technical Reports Server (NTRS)

    Banks, D. W.; Hafez, M. M.

    1996-01-01

    Grid adaptation for structured meshes is the art of using information from an existing, but poorly resolved, solution to automatically redistribute the grid points in such a way as to improve the resolution in regions of high error, and thus the quality of the solution. This involves: (1) generate a grid vis some standard algorithm, (2) calculate a solution on this grid, (3) adapt the grid to this solution, (4) recalculate the solution on this adapted grid, and (5) repeat steps 3 and 4 to satisfaction. Steps 3 and 4 can be repeated until some 'optimal' grid is converged to but typically this is not worth the effort and just two or three repeat calculations are necessary. They also may be repeated every 5-10 time steps for unsteady calculations.

  18. Insights from modeling the 3D structure of NAD(P)H-dependent D-xylose reductase of Pichia stipitis and its binding interactions with NAD and NADP.

    PubMed

    Wang, Jing-Fang; Wei, Dong-Qing; Lin, Ying; Wang, Yong-Hua; Du, Hong-Li; Li, Yi-Xve; Chou, Kuo-Chen

    2007-07-27

    NAD(P)H-dependent d-xylose reductase is a homodimeric oxidoreductase that belongs to the aldo-keto reductase superfamily. The enzyme has the special function to catalyze the first step in the assimilation of xylose into yeast metabolic pathways. Performing this function via reducing the open chain xylose to xylitol, the xylose reductase of Pichia stipitis is one of the most important enzymes that can be used to construct recombinant Saccharomyces cerevisiae strain for utilizing xylose and producing alcohol. To investigate into the interaction mechanism of the enzyme with its ligand NAD and NADP, the 3D structure was developed for the NAD(P)H-dependent d-xylose reductase from P. stipitis. With the 3D structure, the molecular docking operations were conducted to find the most stable bindings of the enzyme with NAD and NADP, respectively. Based on these results, the binding pockets of the enzyme for NAD and NADP have been explicitly defined. It has been found that the residues in forming the binding pockets for both NAD and NADP are almost the same and mainly hydrophilic. These findings may be used to guide mutagenesis studies, providing useful clues to modify the enzyme to improve the utilization of xylose for producing alcohol. Also, because human aldose reductases have the function to reduce the open chain form of glucose to sorbitol, a process physiologically significant for diabetic patients at the time that their blood glucose levels are elevated, the information gained through this study may also stimulate the development of new strategies for therapeutic treatment of diabetes.

  19. Phenylalanine-508 mediates a cytoplasmic-membrane domain contact in the CFTR 3D structure crucial to assembly and channel function.

    PubMed

    Serohijos, Adrian W R; Hegedus, Tamás; Aleksandrov, Andrei A; He, Lihua; Cui, Liying; Dokholyan, Nikolay V; Riordan, John R

    2008-03-04

    Deletion of phenylalanine-508 (Phe-508) from the N-terminal nucleotide-binding domain (NBD1) of the cystic fibrosis transmembrane conductance regulator (CFTR), a member of the ATP-binding cassette (ABC) transporter family, disrupts both its folding and function and causes most cystic fibrosis. Most mutant nascent chains do not pass quality control in the ER, and those that do remain thermally unstable, only partially functional, and are rapidly endocytosed and degraded. Although the lack of the Phe-508 peptide backbone diminishes the NBD1 folding yield, the absence of the aromatic side chain is primarily responsible for defective CFTR assembly and channel gating. However, the site of interdomain contact by the side chain is unknown as is the high-resolution 3D structure of the complete protein. Here we present a 3D structure of CFTR, constructed by molecular modeling and supported biochemically, in which Phe-508 mediates a tertiary interaction between the surface of NBD1 and a cytoplasmic loop (CL4) in the C-terminal membrane-spanning domain (MSD2). This crucial cytoplasmic membrane interface, which is dynamically involved in regulation of channel gating, explains the known sensitivity of CFTR assembly to many disease-associated mutations in CL4 as well as NBD1 and provides a sharply focused target for small molecules to treat CF. In addition to identifying a key intramolecular site to be repaired therapeutically, our findings advance understanding of CFTR structure and function and provide a platform for focused biochemical studies of other features of this unique ABC ion channel.

  20. Computational chemistry study of 3D-structure-function relationships for enzymes based on Markov models for protein electrostatic, HINT, and van der Waals potentials.

    PubMed

    Concu, Riccardo; Podda, Gianni; Uriarte, Eugenio; González-Díaz, Humberto

    2009-07-15

    In a significant work, Dobson and Doig (J Mol Biol 2003, 330, 771) illustrated protein prediction as enzymatic or not from spatial structure without resorting to alignments. They used 52 protein features and a nonlinear support vector machine model to classify more than 1000 proteins collected from the PDB with a 77% overall accuracy. The most useful features were: the secondary-structure content, the amino acid frequencies, the number of disulphide bonds, and the largest cleft size. Working on the same dataset used by D&D, in this article we reported a good and simple model, based on the Markov chain models (MCM), to classify protein 3D structures as enzymatic or not, taking into consideration the spatial structure without resorting to alignments. Here we define, for the first time, a general MCM to calculate the electrostatic potential, molecular vibrations, van der Waals (vdw) interactions, and hydrophobic interactions (HINT) and use them in comparative studies of potential fields and/or protein function prediction. The dataset is composed of 1371 proteins divided into 689 enzymes and 682 nonenzymes, all proteins were collected from the PDB. The best model we found was a linear model carried out with the linear discriminant analysis; it was able to classify 74.18% of the proteins using only two electrostatic potentials. In the work described here, we define 3D-HINT potentials (mu(k)) and use them for the first time to derive a classifier for protein enzymes. We analyzed ROC curves, domain of applicability, parametric assumptions, desirability maps, and also tested other nonlinear artificial neural network models which did not improve the linear model. In closing, this MCM allows a fast calculation and comparison of different potentials deriving into accurate protein 3D structure-function relationships, notably simpler than the previous.

  1. Atomic resolution 3D electron diffraction microscopy

    SciTech Connect

    Miao, Jianwei; Ohsuna, Tetsu; Terasaki, Osamu; O'Keefe, Michael A.

    2002-03-01

    Electron lens aberration is the major barrier limiting the resolution of electron microscopy. Here we describe a novel form of electron microscopy to overcome electron lens aberration. By combining coherent electron diffraction with the oversampling phasing method, we show that the 3D structure of a 2 x 2 x 2 unit cell nano-crystal (framework of LTA [Al12Si12O48]8) can be ab initio determined at the resolution of 1 Angstrom from a series of simulated noisy diffraction pattern projections with rotation angles ranging from -70 degrees to +70 degrees in 5 degrees increments along a single rotation axis. This form of microscopy (which we call 3D electron diffraction microscopy) does not require any reference waves, and can image the 3D structure of nanocrystals, as well as non-crystalline biological and materials science samples, with the resolution limited only by the quality of sample diffraction.

  2. Lunar South Pole Illumination

    NASA Video Gallery

    Simulated illumination conditions over the lunar South Pole region, from ~80°S to the pole. The movie runs for 28 days, centered on the LCROSS impact date on October 9th, 2009. The illumination ca...

  3. Lights illuminate surfaces superluminally

    NASA Astrophysics Data System (ADS)

    Nemiroff, Robert J.; Zhong, Qi; Lilleskov, Elias

    2016-07-01

    When a light bulb is turned on, light moves away from it at speed c, by definition. When light from this bulb illuminates a surface, however, this illumination front is not constrained to move at speed c. A simple proof is given that this illumination front always moves faster than c. Generalized, when any compact light source itself varies, this information spreads across all of the surfaces it illuminates at speeds faster than light.

  4. The lithospheric-scale 3D structural configuration of the North Alpine Foreland Basin constrained by gravity modelling and the calculation of the 3D load distribution

    NASA Astrophysics Data System (ADS)

    Przybycin, Anna M.; Scheck-Wenderoth, Magdalena; Schneider, Michael

    2014-05-01

    The North Alpine Foreland Basin is situated in the northern front of the European Alps and extends over parts of France, Switzerland, Germany and Austria. It formed as a wedge shaped depression since the Tertiary in consequence of the Euro - Adriatic continental collision and the Alpine orogeny. The basin is filled with clastic sediments, the Molasse, originating from erosional processes of the Alps and underlain by Mesozoic sedimentary successions and a Paleozoic crystalline crust. For our study we have focused on the German part of the basin. To investigate the deep structure, the isostatic state and the load distribution of this region we have constructed a 3D structural model of the basin and the Alpine area using available depth and thickness maps, regional scale 3D structural models as well as seismic and well data for the sedimentary part. The crust (from the top Paleozoic down to the Moho (Grad et al. 2008)) has been considered as two-parted with a lighter upper crust and a denser lower crust; the partition has been calculated following the approach of isostatic equilibrium of Pratt (1855). By implementing a seismic Lithosphere-Asthenosphere-Boundary (LAB) (Tesauro 2009) the crustal scale model has been extended to the lithospheric-scale. The layer geometry and the assigned bulk densities of this starting model have been constrained by means of 3D gravity modelling (BGI, 2012). Afterwards the 3D load distribution has been calculated using a 3D finite element method. Our results show that the North Alpine Foreland Basin is not isostatically balanced and that the configuration of the crystalline crust strongly controls the gravity field in this area. Furthermore, our results show that the basin area is influenced by varying lateral load differences down to a depth of more than 150 km what allows a first order statement of the required compensating horizontal stress needed to prevent gravitational collapse of the system. BGI (2012). The International

  5. Genome3D: a UK collaborative project to annotate genomic sequences with predicted 3D structures based on SCOP and CATH domains

    PubMed Central

    Lewis, Tony E.; Sillitoe, Ian; Andreeva, Antonina; Blundell, Tom L.; Buchan, Daniel W.A.; Chothia, Cyrus; Cuff, Alison; Dana, Jose M.; Filippis, Ioannis; Gough, Julian; Hunter, Sarah; Jones, David T.; Kelley, Lawrence A.; Kleywegt, Gerard J.; Minneci, Federico; Mitchell, Alex; Murzin, Alexey G.; Ochoa-Montaño, Bernardo; Rackham, Owen J. L.; Smith, James; Sternberg, Michael J. E.; Velankar, Sameer; Yeats, Corin; Orengo, Christine

    2013-01-01

    Genome3D, available at http://www.genome3d.eu, is a new collaborative project that integrates UK-based structural resources to provide a unique perspective on sequence–structure–function relationships. Leading structure prediction resources (DomSerf, FUGUE, Gene3D, pDomTHREADER, Phyre and SUPERFAMILY) provide annotations for UniProt sequences to indicate the locations of structural domains (structural annotations) and their 3D structures (structural models). Structural annotations and 3D model predictions are currently available for three model genomes (Homo sapiens, E. coli and baker’s yeast), and the project will extend to other genomes in the near future. As these resources exploit different strategies for predicting structures, the main aim of Genome3D is to enable comparisons between all the resources so that biologists can see where predictions agree and are therefore more trusted. Furthermore, as these methods differ in whether they build their predictions using CATH or SCOP, Genome3D also contains the first official mapping between these two databases. This has identified pairs of similar superfamilies from the two resources at various degrees of consensus (532 bronze pairs, 527 silver pairs and 370 gold pairs). PMID:23203986

  6. Combining Amine-Reactive Cross-Linkers and Photo-Reactive Amino Acids for 3D-Structure Analysis of Proteins and Protein Complexes.

    PubMed

    Lössl, Philip; Sinz, Andrea

    2016-01-01

    During the last 15 years, the combination of chemical cross-linking and high-resolution mass spectrometry (MS) has matured into an alternative approach for analyzing 3D-structures of proteins and protein complexes. Using the distance constraints imposed by the cross-links, models of the protein or protein complex under investigation can be created. The majority of cross-linking studies are currently conducted with homobifunctional amine-reactive cross-linkers. We extend this "traditional" cross-linking/MS strategy by adding complementary photo-cross-linking data. For this, the diazirine-containing unnatural amino acids photo-leucine and photo-methionine are incorporated into the proteins and cross-link formation is induced by UV-A irradiation. The advantage of the photo-cross-linking strategy is that it is not restricted to lysine residues and that hydrophobic regions in proteins can be targeted, which is advantageous for investigating membrane proteins. We consider the strategy of combining cross-linkers with orthogonal reactivities and distances to be ideally suited for maximizing the amount of structural information that can be gained from a cross-linking experiment.

  7. Multimodal super-resolution optical microscopy visualizes the close connection between membrane and the cytoskeleton in liver sinusoidal endothelial cell fenestrations

    NASA Astrophysics Data System (ADS)

    Mönkemöller, Viola; Øie, Cristina; Hübner, Wolfgang; Huser, Thomas; McCourt, Peter

    2015-11-01

    Liver sinusoidal endothelial cells (LSECs) act as a filter between blood and the hepatocytes. LSECs are highly fenestrated cells; they contain transcellular pores with diameters between 50 to 200 nm. The small sizes of the fenestrae have so far prohibited any functional analysis with standard and advanced light microscopy techniques. Only the advent of super-resolution optical fluorescence microscopy now permits the recording of such small cellular structures. Here, we demonstrate the complementary use of two different super-resolution optical microscopy modalities, 3D structured illumination microscopy (3D-SIM) and single molecule localization microscopy in a common optical platform to obtain new insights into the association between the cytoskeleton and the plasma membrane that supports the formation of fenestrations. We applied 3D-SIM to multi-color stained LSECs to acquire highly resolved overviews of large sample areas. We then further increased the spatial resolution for imaging fenestrations by single molecule localization microscopy applied to select small locations of interest in the same sample on the same microscope setup. We optimized the use of fluorescent membrane stains for these imaging conditions. The combination of these techniques offers a unique opportunity to significantly improve studies of subcellular ultrastructures such as LSEC fenestrations.

  8. Multimodal super-resolution optical microscopy visualizes the close connection between membrane and the cytoskeleton in liver sinusoidal endothelial cell fenestrations.

    PubMed

    Mönkemöller, Viola; Øie, Cristina; Hübner, Wolfgang; Huser, Thomas; McCourt, Peter

    2015-11-09

    Liver sinusoidal endothelial cells (LSECs) act as a filter between blood and the hepatocytes. LSECs are highly fenestrated cells; they contain transcellular pores with diameters between 50 to 200 nm. The small sizes of the fenestrae have so far prohibited any functional analysis with standard and advanced light microscopy techniques. Only the advent of super-resolution optical fluorescence microscopy now permits the recording of such small cellular structures. Here, we demonstrate the complementary use of two different super-resolution optical microscopy modalities, 3D structured illumination microscopy (3D-SIM) and single molecule localization microscopy in a common optical platform to obtain new insights into the association between the cytoskeleton and the plasma membrane that supports the formation of fenestrations. We applied 3D-SIM to multi-color stained LSECs to acquire highly resolved overviews of large sample areas. We then further increased the spatial resolution for imaging fenestrations by single molecule localization microscopy applied to select small locations of interest in the same sample on the same microscope setup. We optimized the use of fluorescent membrane stains for these imaging conditions. The combination of these techniques offers a unique opportunity to significantly improve studies of subcellular ultrastructures such as LSEC fenestrations.

  9. Fabrication of nano-Fe3O4 3D structure on carbon fibers as a microwave absorber and EMI shielding composite by modified EPD method

    NASA Astrophysics Data System (ADS)

    Gholampoor, Mahdi; Movassagh-Alanagh, Farid; Salimkhani, Hamed

    2017-02-01

    Recently, electromagnetic interference (EMI) shielding materials have absorbed a lot of attention due to a growing need for application in the area of electronic and wireless devices. In this study, a carbon-based EMI shielding composite was fabricated by electrophoretic deposition of Fe3O4 nano-particles on carbon fibers (CFs) as a 3D structure incorporated with an epoxy resin. Co-precipitation method was employed to synthesize Fe3O4 nano-particles. This as-synthesized Fe3O4 nano-powder was then successfully deposited on CFs using a modified multi-step electrophoretic deposition (EPD) method. The results of structural studies showed that the Fe3O4 nano-particles (25 nm) were successfully and uniformly deposited on CFs. The measured magnetic properties of as-synthesized Fe3O4 nano-powder and nano-Fe3O4/CFs composite showed that the saturation magnetization of bare Fe3O4 was decreased from Ms = 72.3 emu/g to Ms = 33.1 emu/g for nano-Fe3O4/CFs composite and also corecivity of Fe3O4 was increased from Hc = 4.9 Oe to Hc = 168 Oe for composite. The results of microwave absorption tests revealed that the reflection loss (RL) of an epoxy-based nano-Fe3O4/CFs composite are significantly influenced by layer thickness. The maximum RL value of -10.21 dB at 10.12 GHz with an effective absorption bandwidth about 2 GHz was obtained for the sample with the thickness of 2 mm. It also exhibited an EMI shielding performance of -23 dB for whole the frequency range of 8.2-12.4 GHz.

  10. Molecular cloning, expression pattern, and 3D structural prediction of the cold inducible RNA-binding protein (CIRP) in Japanese flounder ( Paralichthys olivaceus)

    NASA Astrophysics Data System (ADS)

    Yang, Xiao; Gao, Jinning; Ma, Liman; Li, Zan; Wang, Wenji; Wang, Zhongkai; Yu, Haiyang; Qi, Jie; Wang, Xubo; Wang, Zhigang; Zhang, Quanqi

    2015-02-01

    Cold-inducible RNA-binding protein (CIRP) is a kind of RNA binding proteins that plays important roles in many physiological processes. The CIRP has been widely studied in mammals and amphibians since it was first cloned from mammals. On the contrary, there are little reports in teleosts. In this study, the Po CIRP gene of the Japanese flounder was cloned and sequenced. The genomic sequence consists of seven exons and six introns. The putative PoCIRP protein of flounder was 198 amino acid residues long containing the RNA recognition motif (RRM). Phylogenetic analysis showed that the flounder PoCIRP is highly conserved with other teleost CIRPs. The 5' flanking sequence was cloned by genome walking and many transcription factor binding sites were identified. There is a CpGs region located in promoter and exon I region and the methylation state is low. Quantitative real-time PCR analysis uncovered that Po CIRP gene was widely expressed in adult tissues with the highest expression level in the ovary. The mRNA of the Po CIRP was maternally deposited and the expression level of the gene was regulated up during the gastrula and neurula stages. In order to gain the information how the protein interacts with mRNA, we performed the modeling of the 3D structure of the flounder PoCIRP. The results showed a cleft existing the surface of the molecular. Taken together, the results indicate that the CIRP is a multifunctional molecular in teleosts and the findings about the structure provide valuable information for understanding the basis of this protein's function.

  11. Illumination Under Trees

    SciTech Connect

    Max, N

    2002-08-19

    This paper is a survey of the author's work on illumination and shadows under trees, including the effects of sky illumination, sun penumbras, scattering in a misty atmosphere below the trees, and multiple scattering and transmission between leaves. It also describes a hierarchical image-based rendering method for trees.

  12. Grouping Illumination Frameworks

    ERIC Educational Resources Information Center

    Zdravkovic, Suncica; Economou, Elias; Gilchrist, Alan

    2012-01-01

    According to Koffka (1935), the lightness of a target surface is determined by the relationship between the target and the illumination frame of reference to which it belongs. However, each scene contains numerous illumination frames, and judging each one separately would lead to an enormous amount of computing. Grouping those frames that are in…

  13. Jacaric acid and its octadecatrienoic acid geoisomers induce apoptosis selectively in cancerous human prostate cells: a mechanistic and 3-D structure-activity study.

    PubMed

    Gasmi, Jihane; Thomas Sanderson, J

    2013-06-15

    analysis of structural similarity enabled us to rank geoisomeric fatty acids according to cytotoxic potency, whereas a 2-D positional assessment of cis/trans structure did not. Our findings provide mechanistic evidence that nutrition-derived non-essential fatty acids have chemopreventive biological activities and Exhibit 3-D structure-activity relationships that could be exploited to develop new strategies for the prevention or treatment of prostate cancer regardless of hormone dependency.

  14. An Evaluation of the Observational Capabilities of A Scanning 95-GHz Radar in Studying the 3D Structures of Marine Stratocumulus Clouds

    NASA Astrophysics Data System (ADS)

    Bowley, Kevin

    Marine stratocumulus clouds play a critical role in Earth's radiative balance primarily due to the role of their high albedo reflecting incoming solar radiation, causing a cooling effect, while weakly reflecting outgoing infrared radiation. Characterization of the 3-Dimensional (3D) structure of these cloud systems over scales of 20-40 km is required to accurately account for the role of cloud inhomogeneity and structure on their shortwave forcing and lifetime, which has important applications for Global Climate Models. For first time, such 3D measurements in clouds were made available from a scanning cloud radar during the U.S. Department of Energy (DOE) Atmospheric Radiation Measurement (ARM) program's Clouds, Aerosol, and Precipitation in the Marine Boundary Layer (CAP-MBL) field campaign in the Azores Islands. The scanning radar observations were complemented by a suite of zenith-pointing active and passive remote sensors that were deployed to provide a detailed description of marine stratus over a long-term observation period in the ideal marine environment commonly found at the Azores. The scanning cloud radar observations present a shift from a multi-instrument, vertically pointing 'soda-straw' observation technique to a radar-only, 'radar-centric' observation technique. The scanning radar observations were gridded using a nearest-neighbor type scheme devised to take the natural variability of the observed field into account. The ability of the scheme to capture primary cloud properties (cloud fraction, cloud boundaries, drizzle detection) was assessed using measurements from the vertically pointing sensors. Despite the great sensitivity of the scanning cloud radar (-42.5 dBZ at 1 km range), the drop in sensitivity with range resulted in an artificial thinning of clouds with range from the radar. Drizzle-free cloud structures were undetectable beyond 5 km from the radar. Cloud fields containing drizzle were generally detectable to ranges exceeding 10 km from

  15. Characterization of Polymer Blends: Optical Microscopy (*Polarized, Interference and Phase Contrast Microscopy*) and Confocal Microscopy

    SciTech Connect

    Ramanathan, Nathan Muruganathan; Darling, Seth B.

    2015-01-01

    Chapter 15 surveys the characterization of macro, micro and meso morphologies of polymer blends by optical microscopy. Confocal Microscopy offers the ability to view the three dimensional morphology of polymer blends, popular in characterization of biological systems. Confocal microscopy uses point illumination and a spatial pinhole to eliminate out-of focus light in samples that are thicker than the focal plane.

  16. Shackleton Crater Illumination

    NASA Video Gallery

    Simulated illumination conditions near the lunar South Pole. The 30km x 30km region highlights the Shackleton crater. The movie runs for 28 days, centered on the LCROSS impact date on October 9th, ...

  17. Wood's lamp illumination (image)

    MedlinePlus

    A Wood's lamp emits ultraviolet light and can be a diagnostic aid in determining if someone has a fungal ... is an infection on the area where the Wood's lamp is illuminating, the area will fluoresce. Normally ...

  18. Illuminating black holes

    NASA Astrophysics Data System (ADS)

    Barr, Ian A.; Bull, Anne; O'Brien, Eileen; Drillsma-Milgrom, Katy A.; Milgrom, Lionel R.

    2016-07-01

    Two-dimensional shadows formed by illuminating vortices are shown to be visually analogous to the gravitational action of black holes on light and surrounding matter. They could be useful teaching aids demonstrating some of the consequences of general relativity.

  19. Heterogeneous architecture of vertebrate kinetochores revealed by three-dimensional superresolution fluorescence microscopy

    PubMed Central

    Wynne, David J.; Funabiki, Hironori

    2016-01-01

    The kinetochore is often depicted as having a disk-like architecture in which the outer layer of proteins, which engage microtubules and control checkpoint signaling, are built on a static inner layer directly linked to CENP-A chromatin. Here, applying three-dimensional (3D) structural illumination microscopy (SIM) and stochastic optical reconstruction microscopy (STORM) to Xenopus egg extracts and tissue culture cells, we report various distribution patterns of inner and outer kinetochore proteins. In egg extracts, a configuration in which outer kinetochore proteins surround the periphery of CENP-A chromatin is common, forming an ∼200-nm ring-like organization that may engage a bundle of microtubule ends. Similar rings are observed in Xenopus tissue culture cells at a lower frequency but are enriched in conditions in which the spindle is disorganized. Although rings are rare in human cells, the distribution of both inner and outer kinetochore proteins elongates in the absence of microtubule attachment in a manner dependent on Aurora B. We propose a model in which the 3D organization of both the outer and inner kinetochore regions respond to the progression from lateral to end-on microtubule attachments by coalescing into a tight disk from less uniform distributions early in prometaphase. PMID:27170176

  20. Live-cell superresolution microscopy reveals the organization of RNA polymerase in the bacterial nucleoid

    PubMed Central

    Stracy, Mathew; Lesterlin, Christian; Garza de Leon, Federico; Uphoff, Stephan; Zawadzki, Pawel; Kapanidis, Achillefs N.

    2015-01-01

    Despite the fundamental importance of transcription, a comprehensive analysis of RNA polymerase (RNAP) behavior and its role in the nucleoid organization in vivo is lacking. Here, we used superresolution microscopy to study the localization and dynamics of the transcription machinery and DNA in live bacterial cells, at both the single-molecule and the population level. We used photoactivated single-molecule tracking to discriminate between mobile RNAPs and RNAPs specifically bound to DNA, either on promoters or transcribed genes. Mobile RNAPs can explore the whole nucleoid while searching for promoters, and spend 85% of their search time in nonspecific interactions with DNA. On the other hand, the distribution of specifically bound RNAPs shows that low levels of transcription can occur throughout the nucleoid. Further, clustering analysis and 3D structured illumination microscopy (SIM) show that dense clusters of transcribing RNAPs form almost exclusively at the nucleoid periphery. Treatment with rifampicin shows that active transcription is necessary for maintaining this spatial organization. In faster growth conditions, the fraction of transcribing RNAPs increases, as well as their clustering. Under these conditions, we observed dramatic phase separation between the densest clusters of RNAPs and the densest regions of the nucleoid. These findings show that transcription can cause spatial reorganization of the nucleoid, with movement of gene loci out of the bulk of DNA as levels of transcription increase. This work provides a global view of the organization of RNA polymerase and transcription in living cells. PMID:26224838

  1. Illumination in diverse codimensions

    NASA Technical Reports Server (NTRS)

    Banks, David C.

    1994-01-01

    This paper derives a model of diffuse and specular illumination in arbitrarily large dimensions, based on a few characteristics of material and light in three-space. It then describes how to adjust for the anomaly of excess brightness in large codimensions. If a surface is grooved or furry, it can be illuminated with a hybrid model that incorporates both the one dimensional geometry (the grooves or fur) and the two dimensional geometry (the surface).

  2. Axial Plane Optical Microscopy

    PubMed Central

    Li, Tongcang; Ota, Sadao; Kim, Jeongmin; Wong, Zi Jing; Wang, Yuan; Yin, Xiaobo; Zhang, Xiang

    2014-01-01

    We present axial plane optical microscopy (APOM) that can, in contrast to conventional microscopy, directly image a sample's cross-section parallel to the optical axis of an objective lens without scanning. APOM combined with conventional microscopy simultaneously provides two orthogonal images of a 3D sample. More importantly, APOM uses only a single lens near the sample to achieve selective-plane illumination microscopy, as we demonstrated by three-dimensional (3D) imaging of fluorescent pollens and brain slices. This technique allows fast, high-contrast, and convenient 3D imaging of structures that are hundreds of microns beneath the surfaces of large biological tissues. PMID:25434770

  3. Light sheet microscopy.

    PubMed

    Weber, Michael; Mickoleit, Michaela; Huisken, Jan

    2014-01-01

    This chapter introduces the concept of light sheet microscopy along with practical advice on how to design and build such an instrument. Selective plane illumination microscopy is presented as an alternative to confocal microscopy due to several superior features such as high-speed full-frame acquisition, minimal phototoxicity, and multiview sample rotation. Based on our experience over the last 10 years, we summarize the key concepts in light sheet microscopy, typical implementations, and successful applications. In particular, sample mounting for long time-lapse imaging and the resulting challenges in data processing are discussed in detail.

  4. Natural light illumination system.

    PubMed

    Whang, Allen Jong-Woei; Chen, Yi-Yung; Yang, Shu-Hua; Pan, Po-Hsuan; Chou, Kao-Hsu; Lee, Yu-Chi; Lee, Zong-Yi; Chen, Chi-An; Chen, Cheng-Nan

    2010-12-10

    In recent years, green energy has undergone a lot of development and has been the subject of many applications. Many research studies have focused on illumination with sunlight as a means of saving energy and creating healthy lighting. Natural light illumination systems have collecting, transmitting, and lighting elements. Today, most daylight collectors use dynamic concentrators; these include Sun tracking systems. However, this design is too expensive to be cost effective. To create a low-cost collector that can be easily installed on a large building, we have designed a static concentrator, which is prismatic and cascadable, to collect sunlight for indoor illumination. The transmission component uses a large number of optical fibers. Because optical fibers are expensive, this means that most of the cost for the system will be related to transmission. In this paper, we also use a prismatic structure to design an optical coupler for coupling n to 1. With the n-to-1 coupler, the number of optical fibers necessary can be greatly reduced. Although this new natural light illumination system can effectively guide collected sunlight and send it to the basement or to other indoor places for healthy lighting, previously there has been no way to manage the collected sunlight when lighting was not desired. To solve this problem, we have designed an optical switch and a beam splitter to control and separate the transmitted light. When replacing traditional sources, the lighting should have similar characteristics, such as intensity distribution and geometric parameters, to those of traditional artificial sources. We have designed, simulated, and optimized an illumination lightpipe with a dot pattern to redistribute the collected sunlight from the natural light illumination system such that it equals the qualities of a traditional lighting system. We also provide an active lighting module that provides lighting from the natural light illumination system or LED auxiliary

  5. Bright field illumination system

    NASA Technical Reports Server (NTRS)

    Huber, Edward D. (Inventor)

    1998-01-01

    A Bright Field Illumination system for inspecting a range of characteristically different kinds of defects, depressions, and ridges in a selected material surface. The system has an illumination source placed near a first focus of an elliptical reflector. In addition, a camera facing the inspected area is placed near the illumination source and the first focus. The second focus of the elliptical reflector is located at a distance approximately twice the elliptical reflector's distance above the inspected surface. The elliptical reflector directs the light from the source onto the inspected surface. Due to the shape of the elliptical reflector, light that is specularly reflected from the inspected surface is directed into the camera is which located at the position of the reflected second focus of the ellipse. This system creates a brightly lighted background field against which damage sites appear as high contrast dark objects which can be easily detected by a person or an automated inspection system. In addition, the Bright Field Illumination system and method can be used in combination with a vision inspection system providing for multiplexed illumination and data handling of multiple kinds of surface characteristics including abrupt and gradual surface variations and differences between measured characteristics of different kinds and prior instruments.

  6. A novel 3D structure composed of strings of hierarchical TiO2 spheres formed on TiO2 nanobelts with high photocatalytic properties

    NASA Astrophysics Data System (ADS)

    Jiang, Yongjian; Li, Meicheng; Song, Dandan; Li, Xiaodan; Yu, Yue

    2014-03-01

    A novel hierarchical titanium dioxide (TiO2) composite nanostructure with strings of anatase TiO2 hierarchical micro-spheres and rutile nanobelts framework (TiO2 HSN) is successfully synthesized via a one-step hydrothermal method. Particularly, the strings of hierarchical spheres are assembled by very thin TiO2 nanosheets, which are composed of highly crystallized anatase nanocrystals. Meanwhile, the HSN has a large surface area of 191 m2/g, which is about 3 times larger than Degussa P25. More importantly, the photocatalytic activity of HSN and P25 were evaluated by the photocatalytic oxidation decomposition of methyl orange (MO) under UV light illumination, and the TiO2 HSN shows enhanced photocatalytic activity compared with Degussa P25, as result of its continuous hierarchical structures, special conductive channel and large specific surface area. With these features, the hierarchical TiO2 may have more potential applications in the fields of dye-sensitized solar cells and lithium ion batteries.

  7. HERITAGE: the concept of a giant flux neutron reflectometer for the exploration of 3-d structure of free-liquid and solid interfaces in thin films

    NASA Astrophysics Data System (ADS)

    Mattauch, S.; Ioffe, A.; Lott, D.; Bottyán, L.; Daillant, J.; Markó, M.; Menelle, A.; Sajti, S.; Veres, T.

    2017-01-01

    The instrumental concept of HERITAGE - a reflectometer with a horizontal sample geometry - well fitted to the long pulse structure of a neutron source is presented. It is constitutes a new class of reflectometers achieving the unprecedentedly high flux for classical specular reflectometry combined with off-specular reflectometry and grazing incidence small-angle scattering (GISANS), thus resulting in a complete 3-d exploration of lateral and in depth structures in thin films. This is achieved by specially designed neutron guides. In the horizontal direction (perpendicular to the scattering plane) the guide's elliptic shape focusses the neutrons onto the sample. In the vertical direction a multichannel geometry provides a smooth divergence distribution at the sample position while accepting the entire beam from a compact high-brilliance flat moderator. The modular collimation setup of HERITAGE provides extremely high flexibility in respect to sample geometries and environments, including the possibility to study virtually all types of solid and liquid interfaces, statically or kinetically. The use of multiple beam illumination allows for reflectivity and GISANS measurements at liquid interfaces both from above and below without a need to move the sample. This concept assures the delivery of the maximum possible and usable flux to the sample in both reflectivity and GISANS measurement regimes. The presented design outperforms the flux of all present-day and already for the ESS planned reflectometers and GISANS setups in flux and in measuring time for standard samples.

  8. Open LED Illuminator: A Simple and Inexpensive LED Illuminator for Fast Multicolor Particle Tracking in Neurons

    PubMed Central

    Bosse, Jens B.; Tanneti, Nikhila S.; Hogue, Ian B.; Enquist, Lynn W.

    2015-01-01

    Dual-color live cell fluorescence microscopy of fast intracellular trafficking processes, such as axonal transport, requires rapid switching of illumination channels. Typical broad-spectrum sources necessitate the use of mechanical filter switching, which introduces delays between acquisition of different fluorescence channels, impeding the interpretation and quantification of highly dynamic processes. Light Emitting Diodes (LEDs), however, allow modulation of excitation light in microseconds. Here we provide a step-by-step protocol to enable any scientist to build a research-grade LED illuminator for live cell microscopy, even without prior experience with electronics or optics. We quantify and compare components, discuss our design considerations, and demonstrate the performance of our LED illuminator by imaging axonal transport of herpes virus particles with high temporal resolution. PMID:26600461

  9. OLED area illumination source

    DOEpatents

    Foust, Donald Franklin; Duggal, Anil Raj; Shiang, Joseph John; Nealon, William Francis; Bortscheller, Jacob Charles

    2008-03-25

    The present invention relates to an area illumination light source comprising a plurality of individual OLED panels. The individual OLED panels are configured in a physically modular fashion. Each OLED panel comprising a plurality of OLED devices. Each OLED panel comprises a first electrode and a second electrode such that the power being supplied to each individual OLED panel may be varied independently. A power supply unit capable of delivering varying levels of voltage simultaneously to the first and second electrodes of each of the individual OLED panels is also provided. The area illumination light source also comprises a mount within which the OLED panels are arrayed.

  10. Nonimaging optical illumination system

    DOEpatents

    Winston, R.; Ries, H.

    1996-12-17

    A nonimaging illumination optical device for producing a selected far field illuminance over an angular range. The optical device includes a light source, a light reflecting surface, and a family of light edge rays defined along a reference line with the reflecting surface defined in terms of the reference line as a parametric function R(t) where t is a scalar parameter position and R(t)=k(t)+Du(t) where k(t) is a parameterization of the reference line, and D is a distance from a point on the reference line to the reflection surface along the desired edge ray through the point. 35 figs.

  11. Nonimaging optical illumination system

    DOEpatents

    Winston, R.; Ries, H.

    1998-10-06

    A nonimaging illumination optical device for producing a selected far field illuminance over an angular range. The optical device includes a light source a light reflecting surface, and a family of light edge rays defined along a reference line with the reflecting surface defined in terms of the reference lines a parametric function R(t) where t is a scalar parameter position and R(t)=k(t)+Du(t) where k(t) is a parameterization of the reference line, and D is a distance from a point on the reference line to the reflection surface along the desired edge ray through the point. 35 figs.

  12. A novel 3D structure composed of strings of hierarchical TiO{sub 2} spheres formed on TiO{sub 2} nanobelts with high photocatalytic properties

    SciTech Connect

    Jiang, Yongjian; Li, Meicheng; Song, Dandan; Li, Xiaodan; Yu, Yue

    2014-03-15

    A novel hierarchical titanium dioxide (TiO{sub 2}) composite nanostructure with strings of anatase TiO{sub 2} hierarchical micro-spheres and rutile nanobelts framework (TiO{sub 2} HSN) is successfully synthesized via a one-step hydrothermal method. Particularly, the strings of hierarchical spheres are assembled by very thin TiO{sub 2} nanosheets, which are composed of highly crystallized anatase nanocrystals. Meanwhile, the HSN has a large surface area of 191 m{sup 2}/g, which is about 3 times larger than Degussa P25. More importantly, the photocatalytic activity of HSN and P25 were evaluated by the photocatalytic oxidation decomposition of methyl orange (MO) under UV light illumination, and the TiO{sub 2} HSN shows enhanced photocatalytic activity compared with Degussa P25, as result of its continuous hierarchical structures, special conductive channel and large specific surface area. With these features, the hierarchical TiO{sub 2} may have more potential applications in the fields of dye-sensitized solar cells and lithium ion batteries. -- Graphical abstract: Novel TiO{sub 2} with anatase micro-spheres and rutile nanobelts is synthesized. Enhanced photocatalysis is attributed to hierarchical structures (3D spheres), conductive channel (1D nanobelts) and large specific surface area (2D nanosheet). Highlights: • The novel TiO{sub 2} nanostructure (HSN) is fabricated for the first time. • HSN is composed of strings of anatase hierarchical spheres and rutile nanobelt. • HSN presents a larger S{sub BET} of 191 m{sup 2}/g, 3 times larger than the Degussa P25 (59 m{sup 2}/g). • HSN owns three kinds of dimensional TiO{sub 2} (1D, 2D and 3D) simultaneously. • HSN exhibits better photocatalytic performance compared with Degussa P25.

  13. Nonimaging Optical Illumination System

    DOEpatents

    Winston, Roland

    1994-02-22

    A nonimaging illumination or concentration optical device. An optical device is provided having a light source, a light reflecting surface with an opening and positioned partially around the light source which is opposite the opening of the light reflecting surface. The light reflecting surface is disposed to produce a substantially uniform intensity output with the reflecting surface defined in terms of a radius vector R.sub.i in conjunction with an angle .phi..sub.i between R.sub.i, a direction from the source and an angle .theta..sub.i between direct forward illumination and the light ray reflected once from the reflecting surface. R.sub.i varies as the exponential of tan (.phi..sub.i -.theta..sub.i)/2 integrated over .phi..sub.i.

  14. Nonimaging optical illumination system

    DOEpatents

    Winston, Roland; Ries, Harald

    1996-01-01

    A nonimaging illumination optical device for producing a selected far field illuminance over an angular range. The optical device includes a light source 102, a light reflecting surface 108, and a family of light edge rays defined along a reference line 104 with the reflecting surface 108 defined in terms of the reference line 104 as a parametric function R(t) where t is a scalar parameter position and R(t)=k(t)+Du(t) where k(t) is a parameterization of the reference line 104, and D is a distance from a point on the reference line 104 to the reflection surface 108 along the desired edge ray through the point.

  15. Nonimaging optical illumination system

    DOEpatents

    Winston, Roland; Ries, Harald

    1998-01-01

    A nonimaging illumination optical device for producing a selected far field illuminance over an angular range. The optical device includes a light source 102, a light reflecting surface 108, and a family of light edge rays defined along a reference line 104 with the reflecting surface 108 defined in terms of the reference line 104 as a parametric function R(t) where t is a scalar parameter position and R(t)=k(t)+Du(t) where k(t) is a parameterization of the reference line 104, and D is a distance from a point on the reference line 104 to the reflection surface 108 along the desired edge ray through the point.

  16. Nonimaging optical illumination system

    DOEpatents

    Winston, Roland; Ries, Harald

    2000-01-01

    A nonimaging illumination optical device for producing a selected far field illuminance over an angular range. The optical device includes a light source 102, a light reflecting surface 108, and a family of light edge rays defined along a reference line 104 with the reflecting surface 108 defined in terms of the reference line 104 as a parametric function R(t) where t is a scalar parameter position and R(t)=k(t)+Du(t) where k(t) is a parameterization of the reference line 104, and D is a distance from a point on the reference line 104 to the reflection surface 108 along the desired edge ray through the point.

  17. Visualization of neuritic plaques in Alzheimer’s disease by polarization-sensitive optical coherence microscopy

    NASA Astrophysics Data System (ADS)

    Baumann, Bernhard; Woehrer, Adelheid; Ricken, Gerda; Augustin, Marco; Mitter, Christian; Pircher, Michael; Kovacs, Gabor G.; Hitzenberger, Christoph K.

    2017-03-01

    One major hallmark of Alzheimer’s disease (AD) and cerebral amyloid angiopathy (CAA) is the deposition of extracellular senile plaques and vessel wall deposits composed of amyloid-beta (Aβ). In AD, degeneration of neurons is preceded by the formation of Aβ plaques, which show different morphological forms. Most of them are birefringent owing to the parallel arrangement of amyloid fibrils. Here, we present polarization sensitive optical coherence microscopy (PS-OCM) for imaging mature neuritic Aβ plaques based on their birefringent properties. Formalin-fixed, post-mortem brain samples of advanced stage AD patients were investigated. In several cortical brain regions, neuritic Aβ plaques were successfully visualized in tomographic and three-dimensional (3D) images. Cortical grey matter appeared polarization preserving, whereas neuritic plaques caused increased phase retardation. Consistent with the results from PS-OCM imaging, the 3D structure of senile Aβ plaques was computationally modelled for different illumination settings and plaque sizes. Furthermore, the birefringent properties of cortical and meningeal vessel walls in CAA were investigated in selected samples. Significantly increased birefringence was found in smaller vessels. Overall, these results provide evidence that PS-OCM is able to assess amyloidosis based on intrinsic birefringent properties.

  18. Visualization of neuritic plaques in Alzheimer’s disease by polarization-sensitive optical coherence microscopy

    PubMed Central

    Baumann, Bernhard; Woehrer, Adelheid; Ricken, Gerda; Augustin, Marco; Mitter, Christian; Pircher, Michael; Kovacs, Gabor G.; Hitzenberger, Christoph K.

    2017-01-01

    One major hallmark of Alzheimer’s disease (AD) and cerebral amyloid angiopathy (CAA) is the deposition of extracellular senile plaques and vessel wall deposits composed of amyloid-beta (Aβ). In AD, degeneration of neurons is preceded by the formation of Aβ plaques, which show different morphological forms. Most of them are birefringent owing to the parallel arrangement of amyloid fibrils. Here, we present polarization sensitive optical coherence microscopy (PS-OCM) for imaging mature neuritic Aβ plaques based on their birefringent properties. Formalin-fixed, post-mortem brain samples of advanced stage AD patients were investigated. In several cortical brain regions, neuritic Aβ plaques were successfully visualized in tomographic and three-dimensional (3D) images. Cortical grey matter appeared polarization preserving, whereas neuritic plaques caused increased phase retardation. Consistent with the results from PS-OCM imaging, the 3D structure of senile Aβ plaques was computationally modelled for different illumination settings and plaque sizes. Furthermore, the birefringent properties of cortical and meningeal vessel walls in CAA were investigated in selected samples. Significantly increased birefringence was found in smaller vessels. Overall, these results provide evidence that PS-OCM is able to assess amyloidosis based on intrinsic birefringent properties. PMID:28262719

  19. Optical photon reassignment with increased axial resolution by structured illumination

    NASA Astrophysics Data System (ADS)

    Roth, Stephan; Heintzmann, Rainer

    2016-12-01

    Fluorescent microscopy methods linked to the reassignment principle as image scanning microscopy (ISM), re-scan confocal (RSC), optical photon reassignment (OPRA) and instant structured illumination microscopy (iSIM) have the potential to replace confocal microscopy as the standard microscopy technique. Photon reassignment methods are known to link the most important properties in biological imaging as resolution, sensitivity, imaging speed and combinability with fluorophores in an elegant way. On the example of OPRA, we show how this method could be easily extended to the third dimension. If OPRA is used in combination with a structured illumination pattern the sectioning ability can be improved while maintaining the very high signal intensity. We present a detailed analysis about the imaging properties of OPRA in three dimensions and show experimental results on biological samples.

  20. Nonimaging Optical Illumination System

    DOEpatents

    Winston, Roland

    1994-08-02

    A nonimaging illumination optical device for producing selected intensity output over an angular range. The device includes a light reflecting surface (24, 26) around a light source (22) which is disposed opposite the aperture opening of the light reflecting surface (24, 26). The light source (22) has a characteristic dimension which is small relative to one or more of the distance from the light source (22) to the light reflecting surface (24, 26) or the angle subtended by the light source (22) at the light reflecting surface (24, 26).

  1. Ellipticus CW Illumination System

    DTIC Science & Technology

    2012-08-07

    conceived the idea of making the antenna out of coaxial cable instead of wire and loading the outer conductor with resis- tively damped ferrite ...radiat- ing gap and match the impedance of the antenna as shown in Figure 4. The ferrite bead design, as well as that of the balun, was quite a...Illuminator at Patuxent River NAS. 3.2.1 Wormhole feed concept. The method chosen for driving the radiat- ing gap at the top of the antenna was what Dr

  2. 3D image reconstruction algorithms for cryo-electron-microscopy images of virus particles

    NASA Astrophysics Data System (ADS)

    Doerschuk, Peter C.; Johnson, John E.

    2000-11-01

    A statistical model for the object and the complete image formation process in cryo electron microscopy of viruses is presented. Using this model, maximum likelihood reconstructions of the 3D structure of viruses are computed using the expectation maximization algorithm and an example based on Cowpea mosaic virus is provided.

  3. Parallel hierarchical global illumination

    SciTech Connect

    Snell, Quinn O.

    1997-10-08

    Solving the global illumination problem is equivalent to determining the intensity of every wavelength of light in all directions at every point in a given scene. The complexity of the problem has led researchers to use approximation methods for solving the problem on serial computers. Rather than using an approximation method, such as backward ray tracing or radiosity, the authors have chosen to solve the Rendering Equation by direct simulation of light transport from the light sources. This paper presents an algorithm that solves the Rendering Equation to any desired accuracy, and can be run in parallel on distributed memory or shared memory computer systems with excellent scaling properties. It appears superior in both speed and physical correctness to recent published methods involving bidirectional ray tracing or hybrid treatments of diffuse and specular surfaces. Like progressive radiosity methods, it dynamically refines the geometry decomposition where required, but does so without the excessive storage requirements for ray histories. The algorithm, called Photon, produces a scene which converges to the global illumination solution. This amounts to a huge task for a 1997-vintage serial computer, but using the power of a parallel supercomputer significantly reduces the time required to generate a solution. Currently, Photon can be run on most parallel environments from a shared memory multiprocessor to a parallel supercomputer, as well as on clusters of heterogeneous workstations.

  4. Chip-scale microscopy imaging.

    PubMed

    Zheng, Guoan

    2012-08-01

    Chip-scale microscopy imaging platforms are pivotal for improving the efficiency of modern biomedical and bioscience experiments. Their integration with other lab-on-a-chip techniques would allow rapid, reliable and high-throughput sample analysis for applications in diverse disciplines. In typical chip-scale microscopy imaging platforms, the light path can be generalized to the following steps: photons leave the light source, interact with the sample and finally are detected by the sensor. Based on the light path of these platforms, the current review aims to provide some insights on design strategies for chip-scale microscopy. Specifically, we analyze current chip-scale microscopy approaches from three aspects: illumination design, sample manipulation and substrate/imager modification. We also discuss some opportunities for future developments of chip-scale microscopy, such as time multiplexed structured illumination and hydrodynamic focusing for high throughput sample manipulation.

  5. 3D Structure of Tillage Soils

    NASA Astrophysics Data System (ADS)

    González-Torre, Iván; Losada, Juan Carlos; Falconer, Ruth; Hapca, Simona; Tarquis, Ana M.

    2015-04-01

    Soil structure may be defined as the spatial arrangement of soil particles, aggregates and pores. The geometry of each one of these elements, as well as their spatial arrangement, has a great influence on the transport of fluids and solutes through the soil. Fractal/Multifractal methods have been increasingly applied to quantify soil structure thanks to the advances in computer technology (Tarquis et al., 2003). There is no doubt that computed tomography (CT) has provided an alternative for observing intact soil structure. These CT techniques reduce the physical impact to sampling, providing three-dimensional (3D) information and allowing rapid scanning to study sample dynamics in near real-time (Houston et al., 2013a). However, several authors have dedicated attention to the appropriate pore-solid CT threshold (Elliot and Heck, 2007; Houston et al., 2013b) and the better method to estimate the multifractal parameters (Grau et al., 2006; Tarquis et al., 2009). The aim of the present study is to evaluate the effect of the algorithm applied in the multifractal method (box counting and box gliding) and the cube size on the calculation of generalized fractal dimensions (Dq) in grey images without applying any threshold. To this end, soil samples were extracted from different areas plowed with three tools (moldboard, chissel and plow). Soil samples for each of the tillage treatment were packed into polypropylene cylinders of 8 cm diameter and 10 cm high. These were imaged using an mSIMCT at 155keV and 25 mA. An aluminium filter (0.25 mm) was applied to reduce beam hardening and later several corrections where applied during reconstruction. References Elliot, T.R. and Heck, R.J. 2007. A comparison of 2D and 3D thresholding of CT imagery. Can. J. Soil Sci., 87(4), 405-412. Grau, J, Médez, V.; Tarquis, A.M., Saa, A. and Díaz, M.C.. 2006. Comparison of gliding box and box-counting methods in soil image analysis. Geoderma, 134, 349-359. González-Torres, Iván. Theory and application of multifractal analysis methods in images for the study of soil structure. Master thesis, UPM, 2014. Houston, A.N.; S. Schmidt, A.M. Tarquis, W. Otten, P.C. Baveye, S.M. Hapca. Effect of scanning and image reconstruction settings in X-ray computed tomography on soil image quality and segmentation performance. Geoderma, 207-208, 154-165, 2013a. Houston, A, Otten, W., Baveye, Ph., Hapca, S. Adaptive-Window Indicator Kriging: A Thresholding Method for Computed Tomography, Computers & Geosciences, 54, 239-248, 2013b. Tarquis, A.M., R.J. Heck, D. Andina, A. Alvarez and J.M. Antón. Multifractal analysis and thresholding of 3D soil images. Ecological Complexity, 6, 230-239, 2009. Tarquis, A.M.; D. Giménez, A. Saa, M.C. Díaz. and J.M. Gascó. Scaling and Multiscaling of Soil Pore Systems Determined by Image Analysis. Scaling Methods in Soil Systems. Pachepsky, Radcliffe and Selim Eds., 19-33, 2003. CRC Press, Boca Ratón, Florida. Acknowledgements First author acknowledges the financial support obtained from Soil Imaging Laboratory (University of Gueplh, Canada) in 2014.

  6. Microfabricating 3D Structures by Laser Origami

    DTIC Science & Technology

    2011-11-09

    technique generates 3D microstructures by controlled out-of- plane folding of 2D patterns through a variety of laser-based digital fabrication...processes. Digital microfabrication techniques such as laser direct-write (LDW) offer a viable alternative for generating 3D self-folding designs. These...folding at the microscale where manual or mechanized actuation of the smaller struc- tures is not practical. LDW techniques allow micromachining and

  7. The 3D Structure of the Proton

    NASA Astrophysics Data System (ADS)

    Kaiser, Ralf

    2012-09-01

    When Rutherford, Geiger and Marsden discovered the atomic nucleus in 1909 in Manchester, they at the same time also laid the foundations for the most successful method to study the structure of nuclei and nucleons. They found a point-like scattering centre inside the atom and identified it with the atomic nucleus and the theoretical description of this process has been known as Rutherford scattering ever since. The deviation between the theoretical description for a point-like scattering centre and experimental data has since been used to reveal information about the structure of the nucleus as well as the nucleon. There has been a continuous development from Hofstadters experiments in the 1950s, over the SLAC experiments in the 60s and 70s to the the HERA experiments at DESY and the experimental programme at Jeffersonlab. In this paper I am presenting the most recent results in Deeply Virtual Compton Scattering from the Hermes experiment at DESY, taken with a high density unpolarised target and a recoil detector in 2006/7.

  8. Chemical synthesis and 1H-NMR 3D structure determination of AgTx2-MTX chimera, a new potential blocker for Kv1.2 channel, derived from MTX and AgTx2 scorpion toxins

    PubMed Central

    Pimentel, Cyril; M'Barek, Sarrah; Visan, Violetta; Grissmer, Stephan; Sampieri, François; Sabatier, Jean-Marc; Darbon, Hervé; Fajloun, Ziad

    2008-01-01

    Agitoxin 2 (AgTx2) is a 38-residue scorpion toxin, cross-linked by three disulfide bridges, which acts on voltage-gated K+ (Kv) channels. Maurotoxin (MTX) is a 34-residue scorpion toxin with an uncommon four-disulfide bridge reticulation, acting on both Ca2+-activated and Kv channels. A 39-mer chimeric peptide, named AgTx2-MTX, was designed from the sequence of the two toxins and chemically synthesized. It encompasses residues 1–5 of AgTx2, followed by the complete sequence of MTX. As established by enzyme cleavage, the new AgTx2-MTX molecule displays half-cystine pairings of the type C1–C5, C2–C6, C3–C7, and C4–C8, which is different from that of MTX. The 3D structure of AgTx2-MTX solved by 1H-NMR, revealed both α-helical and β-sheet structures, consistent with a common α/β scaffold of scorpion toxins. Pharmacological assays of AgTx2-MTX revealed that this new molecule is more potent than both original toxins in blocking rat Kv1.2 channel. Docking simulations, performed with the 3D structure of AgTx2-MTX, confirmed this result and demonstrated the participation of the N-terminal domain of AgTx2 in its increased affinity for Kv1.2 through additional molecular contacts. Altogether, the data indicated that replacement of the N-terminal domain of MTX by the one of AgTx2 in the AgTx2-MTX chimera results in a reorganization of the disulfide bridge arrangement and an increase of affinity to the Kv1.2 channel. PMID:18042681

  9. Chemical synthesis and 1H-NMR 3D structure determination of AgTx2-MTX chimera, a new potential blocker for Kv1.2 channel, derived from MTX and AgTx2 scorpion toxins.

    PubMed

    Pimentel, Cyril; M'Barek, Sarrah; Visan, Violetta; Grissmer, Stephan; Sampieri, François; Sabatier, Jean-Marc; Darbon, Hervé; Fajloun, Ziad

    2008-01-01

    Agitoxin 2 (AgTx2) is a 38-residue scorpion toxin, cross-linked by three disulfide bridges, which acts on voltage-gated K(+) (Kv) channels. Maurotoxin (MTX) is a 34-residue scorpion toxin with an uncommon four-disulfide bridge reticulation, acting on both Ca(2+)-activated and Kv channels. A 39-mer chimeric peptide, named AgTx2-MTX, was designed from the sequence of the two toxins and chemically synthesized. It encompasses residues 1-5 of AgTx2, followed by the complete sequence of MTX. As established by enzyme cleavage, the new AgTx2-MTX molecule displays half-cystine pairings of the type C1-C5, C2-C6, C3-C7, and C4-C8, which is different from that of MTX. The 3D structure of AgTx2-MTX solved by (1)H-NMR, revealed both alpha-helical and beta-sheet structures, consistent with a common alpha/beta scaffold of scorpion toxins. Pharmacological assays of AgTx2-MTX revealed that this new molecule is more potent than both original toxins in blocking rat Kv1.2 channel. Docking simulations, performed with the 3D structure of AgTx2-MTX, confirmed this result and demonstrated the participation of the N-terminal domain of AgTx2 in its increased affinity for Kv1.2 through additional molecular contacts. Altogether, the data indicated that replacement of the N-terminal domain of MTX by the one of AgTx2 in the AgTx2-MTX chimera results in a reorganization of the disulfide bridge arrangement and an increase of affinity to the Kv1.2 channel.

  10. Split-illumination electron holography

    SciTech Connect

    Tanigaki, Toshiaki; Aizawa, Shinji; Suzuki, Takahiro; Park, Hyun Soon; Inada, Yoshikatsu; Matsuda, Tsuyoshi; Taniyama, Akira; Shindo, Daisuke; Tonomura, Akira

    2012-07-23

    We developed a split-illumination electron holography that uses an electron biprism in the illuminating system and two biprisms (applicable to one biprism) in the imaging system, enabling holographic interference micrographs of regions far from the sample edge to be obtained. Using a condenser biprism, we split an electron wave into two coherent electron waves: one wave is to illuminate an observation area far from the sample edge in the sample plane and the other wave to pass through a vacuum space outside the sample. The split-illumination holography has the potential to greatly expand the breadth of applications of electron holography.

  11. Illuminated push-button switch

    NASA Astrophysics Data System (ADS)

    Iwagiri, T.

    1983-05-01

    An illuminated push-button switch is described. It is characterized by the fact that is consists of a switch group, an operator button opening and closing the switch group, and a light-emitting element which illuminates the face of the operator button.

  12. Optical tomography with structured illumination.

    PubMed

    Lukic, Vladimir; Markel, Vadim A; Schotland, John C

    2009-04-01

    We consider the image reconstruction problem for optical tomography with structured illumination. A fast image reconstruction algorithm is proposed that reduces the required number of measurements of the optical field compared to methods that utilize point-source illumination. The results are illustrated with numerical simulations.

  13. Do humans discount the illuminant?

    NASA Astrophysics Data System (ADS)

    McCann, John J.

    2005-03-01

    In constancy experiments, humans report very small changes in appearance with substantial illumination changes. Hermann von Helmholtz introduced the term "discounting the illuminant" to describe 19th century thinking about underlying mechanisms of constancy. It uses an indirect approach. Since observers see objects as constant, observers "must" be able to detect the spatial and spectral changes in illumination and automatically compensate by altering the signals from the quanta catches of retinal receptors. Instead of solving the problem directly by calculating an object"s reflectance from the array of scene radiances, Helmholtz chose to solve the problem of identifying the illumination. Twentieth century experiments by Hubel and Wiesel, Campbell, Land, and Gibson demonstrate the power of mechanisms using spatial comparisons. This paper analyses a series of different experiments looking for unequivocal evidence that either supports "discounting the illuminant" or supports spatial comparisons as the underlying mechanism of constancy.

  14. Construction of an instant structured illumination microscope

    PubMed Central

    Curd, Alistair; Cleasby, Alexa; Makowska, Katarzyna; York, Andrew; Shroff, Hari; Peckham, Michelle

    2015-01-01

    A challenge in biological imaging is to capture high-resolution images at fast frame rates in live cells. The “instant structured illumination microscope” (iSIM) is a system designed for this purpose. Similarly to standard structured illumination microscopy (SIM), an iSIM provides a twofold improvement over widefield microscopy, in x, y and z, but also allows much faster image acquisition, with real-time display of super-resolution images. The assembly of an iSIM is reasonably complex, involving the combination and alignment of many optical components, including three micro-optics arrays (two lenslet arrays and an array of pinholes, all with a pitch of 222 μm) and a double-sided scanning mirror. In addition, a number of electronic components must be correctly controlled. Construction of the system is therefore not trivial, but is highly desirable, particularly for live-cell imaging. We report, and provide instructions for, the construction of an iSIM, including minor modifications to a previous design in both hardware and software. The final instrument allows us to rapidly acquire fluorescence images at rates faster than 100 fps, with approximately twofold improvement in resolution in both x–y and z; sub-diffractive biological features have an apparent size (full width at half maximum) of 145 nm (lateral) and 320 nm (axial), using a 1.49 NA objective and 488 nm excitation. PMID:26210400

  15. Construction of an instant structured illumination microscope.

    PubMed

    Curd, Alistair; Cleasby, Alexa; Makowska, Katarzyna; York, Andrew; Shroff, Hari; Peckham, Michelle

    2015-10-15

    A challenge in biological imaging is to capture high-resolution images at fast frame rates in live cells. The "instant structured illumination microscope" (iSIM) is a system designed for this purpose. Similarly to standard structured illumination microscopy (SIM), an iSIM provides a twofold improvement over widefield microscopy, in x, y and z, but also allows much faster image acquisition, with real-time display of super-resolution images. The assembly of an iSIM is reasonably complex, involving the combination and alignment of many optical components, including three micro-optics arrays (two lenslet arrays and an array of pinholes, all with a pitch of 222 μm) and a double-sided scanning mirror. In addition, a number of electronic components must be correctly controlled. Construction of the system is therefore not trivial, but is highly desirable, particularly for live-cell imaging. We report, and provide instructions for, the construction of an iSIM, including minor modifications to a previous design in both hardware and software. The final instrument allows us to rapidly acquire fluorescence images at rates faster than 100 fps, with approximately twofold improvement in resolution in both x-y and z; sub-diffractive biological features have an apparent size (full width at half maximum) of 145 nm (lateral) and 320 nm (axial), using a 1.49 NA objective and 488 nm excitation.

  16. Confocal microscopy in transmitted light

    NASA Astrophysics Data System (ADS)

    Dodt, Hans-Ulrich; Becker, Klaus

    2003-10-01

    We developed a confocal microscope for transmitted light to visualize fine details in phase objects like unstained biological specimens. The main difficulty of confocal microscopy in transmission is the alignment of illumination and detector pinholes. This alignment was achieved by using "electronic pinholes" on the detector side. As a first step, we were able to image cells in onion skin at greater depths and with higher resolution than by using conventional microscopy.

  17. Remodelling of cortical actin where lytic granules dock at natural killer cell immune synapses revealed by super-resolution microscopy.

    PubMed

    Brown, Alice C N; Oddos, Stephane; Dobbie, Ian M; Alakoskela, Juha-Matti; Parton, Richard M; Eissmann, Philipp; Neil, Mark A A; Dunsby, Christopher; French, Paul M W; Davis, Ilan; Davis, Daniel M

    2011-09-01

    Natural Killer (NK) cells are innate immune cells that secrete lytic granules to directly kill virus-infected or transformed cells across an immune synapse. However, a major gap in understanding this process is in establishing how lytic granules pass through the mesh of cortical actin known to underlie the NK cell membrane. Research has been hampered by the resolution of conventional light microscopy, which is too low to resolve cortical actin during lytic granule secretion. Here we use two high-resolution imaging techniques to probe the synaptic organisation of NK cell receptors and filamentous (F)-actin. A combination of optical tweezers and live cell confocal microscopy reveals that microclusters of NKG2D assemble into a ring-shaped structure at the centre of intercellular synapses, where Vav1 and Grb2 also accumulate. Within this ring-shaped organisation of NK cell proteins, lytic granules accumulate for secretion. Using 3D-structured illumination microscopy (3D-SIM) to gain super-resolution of ~100 nm, cortical actin was detected in a central region of the NK cell synapse irrespective of whether activating or inhibitory signals dominate. Strikingly, the periodicity of the cortical actin mesh increased in specific domains at the synapse when the NK cell was activated. Two-colour super-resolution imaging revealed that lytic granules docked precisely in these domains which were also proximal to where the microtubule-organising centre (MTOC) polarised. Together, these data demonstrate that remodelling of the cortical actin mesh occurs at the central region of the cytolytic NK cell immune synapse. This is likely to occur for other types of cell secretion and also emphasises the importance of emerging super-resolution imaging technology for revealing new biology.

  18. Matching illumination of solid objects.

    PubMed

    Pont, Sylvia C; Koenderink, Jan J

    2007-04-01

    The appearance of objects is determined by their surface reflectance and roughness and by the light field. Conversely, human observers might derive properties of the light field from the appearance of objects. The inverse problem has no unique solution, so perceptual interactions between reflectance, roughness, and lightfield are to be expected. In two separate experiments, we tested whether observers are able to match the illumination of spheres under collimated illumination only (matching of illumination direction) and under more or less diffuse illumination (matching of illumination direction and directedness of the beam). We found that observers are quite able to match collimated illumination directions of two rendered Lambertian spheres. Matching of the collimated beam directions of a Lambertian sphere and that of a real object with arbitrary reflectance and roughness properties resulted in similar results for the azimuthal angle, but in higher variance for the polar angle. Translucent objects and a tennis ball were found to be systematic outliers. If the directedness of the beam was also varied, the direction settings showed larger variance for more diffuse illumination. The directedness settings showed an overall quite large variance and, interestingly, interacted with the polar angle settings. We discuss possible photometrical mechanisms behind these effects.

  19. Three-Dimensional Structural Analysis of MgO-Supported Osmium Clusters by Electron Microscopy with Single-Atom Sensitivity

    SciTech Connect

    Aydin, C.; Kulkarni, Apoorva; Chi, Miaofang; Browning, Nigel D.; Gates, Bruce C.

    2013-05-10

    Size, shape, nuclearity: Aberration-corrected scanning transmission electron microscopy was used to determine the 3D structures of MgO-supported Os3, Os4, Os5, and Os10 clusters, which have structures nearly matching those of osmium carbonyl compounds with known crystal structures. The samples are among the best-defined supported catalysts.

  20. Infrared Beam Induced Contrast With Double Illumination

    NASA Astrophysics Data System (ADS)

    Castaldini, A.; Cavallini, A.

    1988-07-01

    Over the last few years scanning optical microscopy (SOM) has been largely developed as a tool to explore the physical properties of materials. In particular the optical beam induced current (OBIC) mode of the SOM has been used to map the electronic properties of semiconducting devices. A new type of scanning microscopy method, as well as some results obtained by it, will be reported in this paper. Though similar, to a certain extent, to the standard scanning optical microscopy, this new investigation technique, from now on refered to as infrared beam induced contrast (IRBIC), differs from it in substance. The chopped light from a quartz halogen lamp is focused by a conventional microscope rearranged on the specimen surface, and a pin-hole is positioned so as to reduce the probe size (not the resolving power) to 1.5um. The resulting beam power density is of the order of 1mW*cm-2. Such a low power density presents some disadvantages in comparison with the traditional laser sources, but, on the other hand, it allows a very high sensitivity in the investigation of the defect electrical activity. With this experimental set-up the specimen front surface is probed with band-gap radiation. Its back surface is illuminated by continuous light in the infrared, coming through a monochromator from a glow-bar. The radiation wavelength can be selected continuously so as the photon energy ranges over the whole valence-to-conduction energy gap. When the specimen is probed, the photoinduced carriers are separated by the built-in field due to the depletion zone of a p-n junction or a Schottky barrier, and the photocurrent is amplified by the lock-in technique. The application of a back-surface radiation of less than the band-gap energy modifies, in some way, the photoconductive response to the band-gap probe since the secondary illumination changes the occupancy of the traps in the forbidden gap active in the photoconductive process. This phenomenon, known as "quenching" of

  1. The role of amino acid electron-donor/acceptor atoms in host-cell binding peptides is associated with their 3D structure and HLA-binding capacity in sterile malarial immunity induction

    SciTech Connect

    Patarroyo, Manuel E.; Almonacid, Hannia; Moreno-Vranich, Armando

    2012-01-20

    Highlights: Black-Right-Pointing-Pointer Fundamental residues located in some HABPs are associated with their 3D structure. Black-Right-Pointing-Pointer Electron-donor atoms present in {beta}-turn, random, distorted {alpha}-helix structures. Black-Right-Pointing-Pointer Electron-donor atoms bound to HLA-DR53. Black-Right-Pointing-Pointer Electron-acceptor atoms present in regular {alpha}-helix structure bound to HLA-DR52. -- Abstract: Plasmodium falciparum malaria continues being one of the parasitic diseases causing the highest worldwide mortality due to the parasite's multiple evasion mechanisms, such as immunological silence. Membrane and organelle proteins are used during invasion for interactions mediated by high binding ability peptides (HABPs); these have amino acids which establish hydrogen bonds between them in some of their critical binding residues. Immunisation assays in the Aotus model using HABPs whose critical residues had been modified have revealed a conformational change thereby enabling a protection-inducing response. This has improved fitting within HLA-DR{beta}1{sup Asterisk-Operator} molecules where amino acid electron-donor atoms present in {beta}-turn, random or distorted {alpha}-helix structures preferentially bound to HLA-DR53 molecules, whilst HABPs having amino acid electron-acceptor atoms present in regular {alpha}-helix structure bound to HLA-DR52. This data has great implications for vaccine development.

  2. Computational modeling and validation studies of 3-D structure of neuraminidase protein of H1N1 influenza A virus and subsequent in silico elucidation of piceid analogues as its potent inhibitors.

    PubMed

    Gupta, Chhedi Lal; Akhtar, Salman; Bajpaib, Preeti; Kandpal, K N; Desai, G S; Tiwari, Ashok K

    2013-01-01

    Emergence of the drug resistant variants of the Influenza A virus in the recent years has aroused a great need for the development of novel neuraminidase inhibitors for controlling the pandemic. The neuraminidase (NA) protein of the influenza virus has been the most potential target for the anti-influenza. However, in the absence of any experimental structure of the drug targeting NA protein of H1N1 influenza A virus as zanamivir and oseltamivir, the comprehensive study of the interaction of the drug molecules with the target protein has been missing. Hence in this study a computational 3-D structure of neuraminidase of H1N1 influenza A virus has been developed using homology modeling technique, and the same was validated for its reliability by ProSA web server in term of energy profile & Z scores and PROCHECK program followed by Ramachandran plot. Further, the developed 3-D model had been employed for docking studies with the class of compounds as Piceid and its analogs. In this context, two novel compounds (ChemBank ID 2110359 and 3075417) were found to be more potent inhibitors of neuraminidase than control drugs as zanamivir and oseltamivir in terms of their robust binding energies, strong inhibition constant (Ki) and better hydrogen bond interactions between the protein-ligand complex. The interaction of these compounds with NA protein has been significantly studied at the molecular level.

  3. Hourly Illumination of Shackleton Crater

    NASA Video Gallery

    Illumination of Shackleton crater, a 21-km-diameter (12.5 mile-diameter) structure situated adjacent to the Moon’s south pole. The resolution is 30 meters (approximately 100 feet) per pixel. Fra...

  4. Laser sources for object illumination

    SciTech Connect

    Albrecht, G.F.

    1994-11-15

    The considerations which formulate the specifications for a laser illuminator are explained, using the example of an underwater object. Depending on the parameters which define the scenario, widely varying laser requirements result.

  5. Scanned Laser Illuminator/Receiver

    DTIC Science & Technology

    1976-11-01

    illustrate parallel development of the PIN diode /CCD sensor hybrid and the 100W laser . Al- though a detailed cost analysis for procurement of this large...pmww^^W .m^n.m .,** ■ —ssa^ AFAL-TR-76-184 \\ SCANNED LASER ILLUMINATOR/RECEIVER ^ R. A. Honzik and F. B. Warren ^•Martin Marietta...NUMBER 4. TITLE (and Sublille) SCANNED LASER ILLUMINATOR/RECEIVER 5, TYPE OF REPORT & PERIOD COVERED Final Technical Report Dec 75

  6. Image plane sweep volume illumination.

    PubMed

    Sundén, Erik; Ynnerman, Anders; Ropinski, Timo

    2011-12-01

    In recent years, many volumetric illumination models have been proposed, which have the potential to simulate advanced lighting effects and thus support improved image comprehension. Although volume ray-casting is widely accepted as the volume rendering technique which achieves the highest image quality, so far no volumetric illumination algorithm has been designed to be directly incorporated into the ray-casting process. In this paper we propose image plane sweep volume illumination (IPSVI), which allows the integration of advanced illumination effects into a GPU-based volume ray-caster by exploiting the plane sweep paradigm. Thus, we are able to reduce the problem complexity and achieve interactive frame rates, while supporting scattering as well as shadowing. Since all illumination computations are performed directly within a single rendering pass, IPSVI does not require any preprocessing nor does it need to store intermediate results within an illumination volume. It therefore has a significantly lower memory footprint than other techniques. This makes IPSVI directly applicable to large data sets. Furthermore, the integration into a GPU-based ray-caster allows for high image quality as well as improved rendering performance by exploiting early ray termination. This paper discusses the theory behind IPSVI, describes its implementation, demonstrates its visual results and provides performance measurements.

  7. Holographic microscopy in low coherence

    NASA Astrophysics Data System (ADS)

    Chmelík, Radim; Petráček, Jiří; Slabá, Michala; Kollárová, Věra; Slabý, Tomáš; Čolláková, Jana; Komrska, Jiří; Dostál, Zbyněk.; Veselý, Pavel

    2016-03-01

    Low coherence of the illumination substantially improves the quality of holographic and quantitative phase imaging (QPI) by elimination of the coherence noise and various artefacts and by improving the lateral resolution compared to the coherent holographic microscopy. Attributes of coherence-controlled holographic microscope (CCHM) designed and built as an off-axis holographic system allowing QPI within the range from complete coherent to incoherent illumination confirmed these expected advantages. Low coherence illumination also furnishes the coherence gating which constraints imaging of some spatial frequencies of an object axially thus forming an optical section in the wide sense. In this way the depth discrimination capability of the microscope is introduced at the price of restricting the axial interval of possible numerical refocusing. We describe theoretically these effects for the whole range of illumination coherence. We also show that the axial refocusing constraints can be overcome using advanced mode of imaging based on mutual lateral shift of reference and object image fields in CCHM. Lowering the spatial coherence of illumination means increasing its numerical aperture. We study how this change of the illumination geometry influences 3D objects QPI and especially the interpretation of live cells QPI in terms of the dry mass density measurement. In this way a strong dependence of the imaging process on the light coherence is demonstrated. The theoretical calculations and numerical simulations are supported by experimental data including a chance of time-lapse watching of live cells even in optically turbid milieu.

  8. Fluorescence Microscopy

    PubMed Central

    Sanderson, Michael J.; Smith, Ian; Parker, Ian; Bootman, Martin D.

    2016-01-01

    Fluorescence microscopy is a major tool with which to monitor cell physiology. Although the concepts of fluorescence and its optical separation using filters remain similar, microscope design varies with the aim of increasing image contrast and spatial resolution. The basics of wide-field microscopy are outlined to emphasize the selection, advantages, and correct use of laser scanning confocal microscopy, two-photon microscopy, scanning disk confocal microscopy, total internal reflection, and super-resolution microscopy. In addition, the principles of how these microscopes form images are reviewed to appreciate their capabilities, limitations, and constraints for operation. PMID:25275114

  9. Specimen illumination apparatus with optical cavity for dark field illumination

    DOEpatents

    Pinkel, Daniel; Sudar, Damir; Albertson, Donna

    1999-01-01

    An illumination apparatus with a specimen slide holder, an illumination source, an optical cavity producing multiple reflection of illumination light to a specimen comprising a first and a second reflective surface arranged to achieve multiple reflections of light to a specimen is provided. The apparatus can further include additional reflective surfaces to achieve the optical cavity, a slide for mounting the specimen, a coverslip which is a reflective component of the optical cavity, one or more prisms for directing light within the optical cavity, antifading solutions for improving the viewing properties of the specimen, an array of materials for analysis, fluorescent components, curved reflective surfaces as components of the optical cavity, specimen detection apparatus, optical detection equipment, computers for analysis of optical images, a plane polarizer, fiberoptics, light transmission apertures, microscopic components, lenses for viewing the specimen, and upper and lower mirrors above and below the specimen slide as components of the optical cavity. Methods of using the apparatus are also provided.

  10. Mixed messages from benthic microbial communities exposed to nanoparticulate and ionic silver: 3D structure picks up nano-specific effects, while EPS and traditional endpoints indicate a concentration-dependent impact of silver ions.

    PubMed

    Kroll, Alexandra; Matzke, Marianne; Rybicki, Marcus; Obert-Rauser, Patrick; Burkart, Corinna; Jurkschat, Kerstin; Verweij, Rudo; Sgier, Linn; Jungmann, Dirk; Backhaus, Thomas; Svendsen, Claus

    2016-03-01

    Silver nanoparticles (AgNP) are currently defined as emerging pollutants in surface water ecosystems. Whether the toxic effects of AgNP towards freshwater organisms are fully explainable by the release of ionic silver (Ag(+)) has not been conclusively elucidated. Long-term effects to benthic microbial communities (periphyton) that provide essential functions in stream ecosystems are unknown. The effects of exposure of periphyton to 2 and 20 μg/L Ag(+) (AgNO3) and AgNP (polyvinylpyrrolidone stabilised) were investigated in artificial indoor streams. The extracellular polymeric substances (EPS) and 3D biofilm structure, biomass, algae species, Ag concentrations in the water phase and bioassociated Ag were analysed. A strong decrease in total Ag was observed within 4 days. Bioassociated Ag was proportional to dissolved Ag indicating a rate limitation by diffusion across the diffusive boundary layer. Two micrograms per liter of AgNO3 or AgNP did not induce significant effects despite detectable bioassociation of Ag. The 20-μg/L AgNO3 affected green algae and diatom communities, biomass and the ratio of polysaccharides to proteins in EPS. The 20-μg/L AgNO3 and AgNP decreased biofilm volume to about 50 %, while the decrease of biomass was lower in 20 μg/L AgNP samples than the 20-μg/L AgNO3 indicating a compaction of the NP-exposed biofilms. Roughness coefficients were lower in 20 μg/L AgNP-treated samples. The more traditional endpoints (biomass and diversity) indicated silver ion concentration-dependent effects, while the newly introduced parameters (3D structure and EPS) indicated both silver ion concentration-dependent effects and effects related to the silver species applied.

  11. Illumination for Worm Tracking and Behavioral Imaging

    PubMed Central

    Yemini, Eviatar; Kerr, Rex A.; Schafer, William R.

    2016-01-01

    Neurobiological research in genetically tractable organisms relies heavily on robust assays for behavioral phenotypes. The simple body plan of the nematode Caenorhabditis elegans makes it particularly amenable to the use of automated microscopy and image analysis to describe behavioral patterns quantitatively. This protocol provides an approach for obtaining uniform illumination during worm tracking. Good lighting can be more of an art than a science. Once the system is set up, it will be necessary to play with it, testing the results after each adjustment to ensure that the analysis software is able to clearly identify the worm and its boundaries. Although the protocol was developed for use in a single-worm tracker, it addresses factors important for the generation of reproducible, standardized images in all systems. PMID:22135668

  12. Multi-contrast Photoacoustic Microscopy

    NASA Astrophysics Data System (ADS)

    Yao, Junjie

    Photoacoustic microscopy is a hybrid imaging modality with high spatial resolution, moderate imaging depth, excellent imaging contrast and functional imaging capability. Taking full advantage of this powerful weapon, we have investigated different anatomical, functional, flow dynamic and metabolic parameter measurements using photoacoustic microscopy. Specifically, Evans-blue dye was used to enhance photoacoustic microscopy of capillaries; label-free transverse and axial blood flow was measured based on bandwidth broadening and time shift of the photoacoustic signals; metabolic rate of oxygen was quantified in vivo from all the five parameters measured by photoacoustic microcopy; whole cross-sectional imaging of small intestine was achieved on a double-illumination photoacoustic microscopy with extended depth of focus and imaging depth; hemodynamic imaging was performed on a MEMS-mirror enhanced photoacoustic microscopy with a cross-sectional imaging rate of 400 Hz. As a maturing imaging technique, PAM is expected to find new applications in both fundamental life science and clinical practice.

  13. Limiting factors in atomic resolution cryo electron microscopy: No simple tricks

    PubMed Central

    Zhang, Xing; Zhou, Z. Hong

    2013-01-01

    To bring cryo electron microscopy (cryoEM) of large biological complexes to atomic resolution, several factors – in both cryoEM image acquisition and 3D reconstruction – that may be neglected at low resolution become significantly limiting. Here we present thorough analyses of four limiting factors: (a) electron-beam tilt, (b) inaccurate determination of defocus values, (c) focus gradient through particles, and (d) particularly for large particles, dynamic (multiple) scattering of electrons. We also propose strategies to cope with these factors: (a) the divergence and direction tilt components of electron-beam tilt could be reduced by maintaining parallel illumination and by using a coma-free alignment procedure, respectively. Moreover, the effect of all beam tilt components, including spiral tilt, could be eliminated by use of a spherical aberration corrector. (b) More accurate measurement of defocus value could be obtained by imaging areas adjacent to the target area at high electron dose and by measuring the image shift induced by tilting the electron beam. (c) Each known Fourier coefficient in the Fourier transform of a cryoEM image is the sum of two Fourier coefficients of the 3D structure, one on each of two curved ‘characteristic surfaces’ in 3D Fourier space. We describe a simple model-based iterative method that could recover these two Fourier coefficients on the two characteristic surfaces. (d) The effect of dynamic scattering could be corrected by deconvolution of a transfer function. These analyses and our proposed strategies offer useful guidance for future experimental designs targeting atomic resolution cryoEM reconstruction. PMID:21627992

  14. Confocal multiview light-sheet microscopy

    PubMed Central

    Medeiros, Gustavo de; Norlin, Nils; Gunther, Stefan; Albert, Marvin; Panavaite, Laura; Fiuza, Ulla-Maj; Peri, Francesca; Hiiragi, Takashi; Krzic, Uros; Hufnagel, Lars

    2015-01-01

    Selective-plane illumination microscopy has proven to be a powerful imaging technique due to its unsurpassed acquisition speed and gentle optical sectioning. However, even in the case of multiview imaging techniques that illuminate and image the sample from multiple directions, light scattering inside tissues often severely impairs image contrast. Here we combine multiview light-sheet imaging with electronic confocal slit detection implemented on modern camera sensors. In addition to improved imaging quality, the electronic confocal slit detection doubles the acquisition speed in multiview setups with two opposing illumination directions allowing simultaneous dual-sided illumination. Confocal multiview light-sheet microscopy eliminates the need for specimen-specific data fusion algorithms, streamlines image post-processing, easing data handling and storage. PMID:26602977

  15. Structured white-light illumination for diagnostic investigations

    NASA Astrophysics Data System (ADS)

    Schau, P.; Brandes, A.; Frenner, K.; Kienle, A.; Osten, W.

    2013-04-01

    The optical coherence tomography (OCT) is an important technology for non-invasive, in vivo medical diagnostics. It enables the high-resolution recording of two-dimensional tomograms or three-dimensional volumes of biological tissue. Two mechanisms help separating the signal from the scattering background. First, reflected or backscattered light from outside the focal spot is suppressed by confocal discrimination. Additionally, the signal modulation is enhanced due to identical optical path lengths of both branches of the white light interferometry setup. Since the OCT relies on the interference between reference light and scattered light, this method cannot be readily extended for fluorescence measurements. An alternative approach is the confocal fluorescence microscopy, which uses confocal microscopy to suppress the fluorescent light from outside the focal spot. Hence, only the fluorescent light in the focal plane, which is 3 to 4 magnitudes lower in intensity than the excitation light, is detected. However, the surrounding area is illuminated with full intensity, which might cause photo-bleaching. There are also other promising approaches such as the two-photon excitation microscopy or fluorescence lifetime microscopy, which we will not cover in more detail. For fluorescence measurements of strongly-scattering samples such as biological tissue but also for technical surfaces, we propose a structured white-light illumination. We present two different approaches for the sample illumination utilizing a white light laser or a white light LED, respectively. We show first simulations of the individual illumination setups and their impact on the scattering within the sample. Furthermore, we investigated the distribution of the fluorescent light that reaches the detection part of the device when excited within a scattering medium, for this purpose we implemented a novel fast-converging algorithm for conditional fluence rate in our Monte Carlo algorithm.

  16. Multiple-illumination photoacoustic tomography

    NASA Astrophysics Data System (ADS)

    Barber, Quinn M.; Zemp, Roger J.

    2016-03-01

    Previously we described the potential for multiple illumination photoacoustic tomography to provide quantitative reconstructions, however this work used only simulated data. We have developed a custom photoacoustic-ultrasound tomography system capable of multiple illuminations and parallel acquisition from a 256 element 5 MHz transducer ring array with 8-cm diameter. The multiple illumination scheme uses a free-space light delivery geometry where a rotational stage scans a pulsed laser beam onto different incident locations around the sample. For each illumination location a photoacoustic image is reconstructed using a modified backprojection algorithm. Images from different source locations have the potential to be combined to form an improved deep-tissue image using our previously developed iterative algorithms. We complement the photoacoustic imaging data with unique ultrasound imaging data. Most previous ultrasound tomography methods have used migration algorithms, iterative ray-based analysis, wave-equation modeling, or frequency-based algorithms that all demand large amounts of data and computational power. We propose a new UST method that offers isotropic resolution, provides scattering contrast, as well as the potential for measuring ultrasound scattering anisotropy and decoupling density and compressibility contributions. The imaging system is driven by a Verasonics scan engine and programmed for both ultrasound and photoacoustic imaging modes. Resolution has been measured to be 150 μm for ultrasound and 200 μm for photoacoustic images. Imaging capabilities are demonstrated on phantoms with custom-tailored ultrasound scattering and optical properties, as well as in murine models.

  17. Analytical Microscopy

    SciTech Connect

    Not Available

    2006-06-01

    In the Analytical Microscopy group, within the National Center for Photovoltaic's Measurements and Characterization Division, we combine two complementary areas of analytical microscopy--electron microscopy and proximal-probe techniques--and use a variety of state-of-the-art imaging and analytical tools. We also design and build custom instrumentation and develop novel techniques that provide unique capabilities for studying materials and devices. In our work, we collaborate with you to solve materials- and device-related R&D problems. This sheet summarizes the uses and features of four major tools: transmission electron microscopy, scanning electron microscopy, the dual-beam focused-ion-beam workstation, and scanning probe microscopy.

  18. En face coherence microscopy [Invited

    PubMed Central

    Thouvenin, Olivier; Grieve, Kate; Xiao, Peng; Apelian, Clement; Boccara, A. Claude

    2017-01-01

    En face coherence microscopy or flying spot or full field optical coherence tomography or microscopy (FF-OCT/FF-OCM) belongs to the OCT family because the sectioning ability is mostly linked to the source coherence length. In this article we will focus our attention on the advantages and the drawbacks of the following approaches: en face versus B scan tomography in terms of resolution, coherent versus incoherent illumination and influence of aberrations, and scanning versus full field imaging. We then show some examples to illustrate the diverse applications of en face coherent microscopy and show that endogenous or exogenous contrasts can add valuable information to the standard morphological image. To conclude we discuss a few domains that appear promising for future development of en face coherence microscopy. PMID:28270972

  19. Laser illuminated flat panel display

    SciTech Connect

    Veligdan, J.T.

    1995-12-31

    A 10 inch laser illuminated flat panel Planar Optic Display (POD) screen has been constructed and tested. This POD screen technology is an entirely new concept in display technology. Although the initial display is flat and made of glass, this technology lends itself to applications where a plastic display might be wrapped around the viewer. The display screen is comprised of hundreds of planar optical waveguides where each glass waveguide represents a vertical line of resolution. A black cladding layer, having a lower index of refraction, is placed between each waveguide layer. Since the cladding makes the screen surface black, the contrast is high. The prototype display is 9 inches wide by 5 inches high and approximately I inch thick. A 3 milliwatt HeNe laser is used as the illumination source and a vector scanning technique is employed.

  20. DARK-FIELD ILLUMINATION SYSTEM

    DOEpatents

    Norgren, D.U.

    1962-07-24

    A means was developed for viewing objects against a dark background from a viewing point close to the light which illuminates the objects and under conditions where the back scattering of light by the objects is minimal. A broad light retro-directing member on the opposite side of the objects from the light returns direct light back towards the source while directing other light away from the viewing point. The viewing point is offset from the light and thus receives only light which is forwardly scattered by an object while returning towards the source. The object is seen, at its true location, against a dark background. The invention is particularly adapted for illuminating and viewing nuclear particle tracks in a liquid hydrogen bubble chamber through a single chamber window. (AEC)

  1. Lunar-illuminated outdoor hologram

    NASA Astrophysics Data System (ADS)

    Jepsen, Mary Lou; Dawson, Paula H.

    1992-05-01

    The first step in the construction of a very deep, large source size, white light illuminated hologram is discussed. We outline the steps taken thus far in the creation of our computer- generated master hologram slit. Our goal is to computer generate a 2 meter master slit for optical transfer via Benton rainbow holography technique. The transfer hologram will ultimately be re-illuminated by the moon and fill a space of approximately 8000 cubic meters. Discussion of the relative merits of synthetic stereographic methods and CGH methods is presented as well as several novel hybrid techniques. The CGH test prints thus far created are evaluated for several types of aberrations and methods of pre-distortion and distortion correction are proposed.

  2. Illumination box and camera system

    DOEpatents

    Haas, Jeffrey S.; Kelly, Fredrick R.; Bushman, John F.; Wiefel, Michael H.; Jensen, Wayne A.; Klunder, Gregory L.

    2002-01-01

    A hand portable, field-deployable thin-layer chromatography (TLC) unit and a hand portable, battery-operated unit for development, illumination, and data acquisition of the TLC plates contain many miniaturized features that permit a large number of samples to be processed efficiently. The TLC unit includes a solvent tank, a holder for TLC plates, and a variety of tool chambers for storing TLC plates, solvent, and pipettes. After processing in the TLC unit, a TLC plate is positioned in a collapsible illumination box, where the box and a CCD camera are optically aligned for optimal pixel resolution of the CCD images of the TLC plate. The TLC system includes an improved development chamber for chemical development of TLC plates that prevents solvent overflow.

  3. Illumination influences working memory: an EEG study.

    PubMed

    Park, Jin Young; Min, Byoung-Kyong; Jung, Young-Chul; Pak, Hyensou; Jeong, Yeon-Hong; Kim, Eosu

    2013-09-05

    Illumination conditions appear to influence working efficacy in everyday life. In the present study, we obtained electroencephalogram (EEG) correlates of working-memory load, and investigated how these waveforms are modulated by illumination conditions. We hypothesized that illumination conditions may affect cognitive performance. We designed an EEG study to monitor and record participants' EEG during the Sternberg working memory task under four different illumination conditions. Illumination conditions were generated with a factorial design of two color-temperatures (3000 and 7100 K) by two illuminance levels (150 and 700 lx). During a working memory task, we observed that high illuminance led to significantly lower frontal EEG theta activity than did low illuminance. These differences persisted despite no significant difference in task performance between illumination conditions. We found that the latency of an early event-related potential component, such as N1, was significantly modulated by the illumination condition. The fact that the illumination condition affects brain activity but not behavioral performance suggests that the lighting conditions used in the present study did not influence the performance stage of behavioral processing. Nevertheless, our findings provide objective evidence that illumination conditions modulate brain activity. Further studies are necessary to refine the optimal lighting parameters for facilitating working memory.

  4. Correlative Microscopy

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Microscopy and Imaging offers many opportunities to collaborate and cooperate with scientists in many different fields nationally and internationally. Images have proven to be very important components in basic research, product development and understanding structure/function relationships in addit...

  5. Correlative microscopy.

    PubMed

    Loussert Fonta, Céline; Humbel, Bruno M

    2015-09-01

    In recent years correlative microscopy, combining the power and advantages of different imaging system, e.g., light, electrons, X-ray, NMR, etc., has become an important tool for biomedical research. Among all the possible combinations of techniques, light and electron microscopy, have made an especially big step forward and are being implemented in more and more research labs. Electron microscopy profits from the high spatial resolution, the direct recognition of the cellular ultrastructure and identification of the organelles. It, however, has two severe limitations: the restricted field of view and the fact that no live imaging can be done. On the other hand light microscopy has the advantage of live imaging, following a fluorescently tagged molecule in real time and at lower magnifications the large field of view facilitates the identification and location of sparse individual cells in a large context, e.g., tissue. The combination of these two imaging techniques appears to be a valuable approach to dissect biological events at a submicrometer level. Light microscopy can be used to follow a labelled protein of interest, or a visible organelle such as mitochondria, in time, then the sample is fixed and the exactly same region is investigated by electron microscopy. The time resolution is dependent on the speed of penetration and fixation when chemical fixatives are used and on the reaction time of the operator for cryo-fixation. Light microscopy can also be used to identify cells of interest, e.g., a special cell type in tissue or cells that have been modified by either transfections or RNAi, in a large population of non-modified cells. A further application is to find fluorescence labels in cells on a large section to reduce searching time in the electron microscope. Multiple fluorescence labelling of a series of sections can be correlated with the ultrastructure of the individual sections to get 3D information of the distribution of the marked proteins: array

  6. Applications of TM polarized illumination

    NASA Astrophysics Data System (ADS)

    Smith, Bruce; Zhou, Jianming; Xie, Peng

    2008-03-01

    The use of transverse electric (TE) polarization has dominated illumination schemes as selective polarization is used for high-NA patterning. The benefits of TE polarization are clear - the interference of diffracted beams remains absolute at oblique angles. Transverse magnetic (TM) polarization is usually considered less desirable as imaging modulation from interference at large angle falls off rapidly as the 1/cosθ. Significant potential remains, however, for the use of TM polarization at large angles when its reflection component is utilized. By controlling the resist/substrate interface reflectivity, high modulation for TM polarization can be maintained for angles up to 90° in the resist. This can potentially impact the design of illumination away from most recent TE-only schemes for oblique imaging angles (high NA). We demonstrate several cases of TM illumination combined with tuned substrate reflectivity for 0.93NA, 1.20NA, and 1.35NA and compare results to TE and unpolarized cases. The goal is to achieve a flat response through polarization at large imaging angles. An additional application of TM illumination is its potential use for double patterning. As double patterning and double exposure approaches are sought in order to meet the needs of 32nm device generations and beyond, materials and process engineering challenges become prohibitive. We have devised a method for frequency doubling in a single exposure using an unconventional means of polarization selection and by making use of the reflective component produced at the photoresist/substrate interface. In doing so, patterns can be deposited into a photoresist film with double density. As an example, using a projection system numerical aperture of 1.20, with water as an immersion fluid, and a conventional polyacrylate 193nm photoresist, pattern resolution at 20nm half-pitch are obtainable (which is 0.125lambda/NA). The process to transfer this geometry into a hardmask layer uses conventional materials

  7. Synchrotron-based EUV lithography illuminator simulator

    DOEpatents

    Naulleau, Patrick P.

    2004-07-27

    A lithographic illuminator to illuminate a reticle to be imaged with a range of angles is provided. The illumination can be employed to generate a pattern in the pupil of the imaging system, where spatial coordinates in the pupil plane correspond to illumination angles in the reticle plane. In particular, a coherent synchrotron beamline is used along with a potentially decoherentizing holographic optical element (HOE), as an experimental EUV illuminator simulation station. The pupil fill is completely defined by a single HOE, thus the system can be easily modified to model a variety of illuminator fill patterns. The HOE can be designed to generate any desired angular spectrum and such a device can serve as the basis for an illuminator simulator.

  8. Atmospheric effects on active illumination

    NASA Astrophysics Data System (ADS)

    Shaw, Scot E. J.; Kansky, Jan E.

    2005-08-01

    For some beam-control applications, we can rely on the cooperation of the target when gathering information about the target location and the state of the atmosphere between the target and the beam-control system. The typical example is a cooperative point-source beacon on the target. Light from such a beacon allows the beam-control system to track the target accurately, and, if higher-order adaptive optics is to be employed, to make wave-front measurements and apply appropriate corrections with a deformable mirror. In many applications, including directed-energy weapons, the target is not cooperative. In the absence of a cooperative beacon, we must find other ways to collect the relevant information. This can be accomplished with an active-illumination system. Typically, this means shining one or more lasers at the target and observing the reflected light. In this paper, we qualitatively explore a number of difficulties inherent to active illumination, and suggest some possible mitigation techniques.

  9. Expansion Microscopy

    PubMed Central

    Chen, Fei; Tillberg, Paul W.; Boyden, Edward S.

    2014-01-01

    In optical microscopy, fine structural details are resolved by using refraction to magnify images of a specimen. Here we report the discovery that, by synthesizing a swellable polymer network within a specimen, it can be physically expanded, resulting in physical magnification. By covalently anchoring specific labels located within the specimen directly to the polymer network, labels spaced closer than the optical diffraction limit can be isotropically separated and optically resolved, a process we call expansion microscopy (ExM). Thus, this process can be used to perform scalable super-resolution microscopy with diffraction-limited microscopes. We demonstrate ExM with effective ~70 nm lateral resolution in both cultured cells and brain tissue, performing three-color super-resolution imaging of ~107 μm3 of the mouse hippocampus with a conventional confocal microscope. PMID:25592419

  10. Segmentation and estimation of spatially varying illumination.

    PubMed

    Lin Gu; Huynh, Cong Phuoc; Robles-Kelly, Antonio

    2014-08-01

    In this paper, we present an unsupervised method for segmenting the illuminant regions and estimating the illumination power spectrum from a single image of a scene lit by multiple light sources. Here, illuminant region segmentation is cast as a probabilistic clustering problem in the image spectral radiance space. We formulate the problem in an optimization setting, which aims to maximize the likelihood of the image radiance with respect to a mixture model while enforcing a spatial smoothness constraint on the illuminant spectrum. We initialize the sample pixel set under each illuminant via a projection of the image radiance spectra onto a low-dimensional subspace spanned by a randomly chosen subset of spectra. Subsequently, we optimize the objective function in a coordinate-ascent manner by updating the weights of the mixture components, sample pixel set under each illuminant, and illuminant posterior probabilities. We then estimate the illuminant power spectrum per pixel making use of these posterior probabilities. We compare our method with a number of alternatives for the tasks of illumination region segmentation, illumination color estimation, and color correction. Our experiments show the effectiveness of our method as applied to one hyperspectral and three trichromatic image data sets.

  11. Background illumination and automated perimetry.

    PubMed

    Klewin, K M; Radius, R L

    1986-03-01

    Visual field function in the right and left eyes of 31 normal volunteers was evaluated with an automated projection perimeter (OCTOPUS). Serial visual field evaluations were repeated in these same eyes with neutral filters of increasing optical density. We compared the results of threshold determinations with the different neutral filters in place before the examined eye. Significant reduction in threshold sensitivity at several test spots throughout the central 30 degrees visual field was seen with neutral density filters of 0.5 log units or greater. The low level of background illumination of the OCTOPUS perimeter (4.0 apostilbs) may allow relatively minor reduction in light transmission by the ocular media to produce significant changes in the recorded level of threshold sensitivity during visual field evaluation.

  12. Fully depleted back illuminated CCD

    DOEpatents

    Holland, Stephen Edward

    2001-01-01

    A backside illuminated charge coupled device (CCD) is formed of a relatively thick high resistivity photon sensitive silicon substrate, with frontside electronic circuitry, and an optically transparent backside ohmic contact for applying a backside voltage which is at least sufficient to substantially fully deplete the substrate. A greater bias voltage which overdepletes the substrate may also be applied. One way of applying the bias voltage to the substrate is by physically connecting the voltage source to the ohmic contact. An alternate way of applying the bias voltage to the substrate is to physically connect the voltage source to the frontside of the substrate, at a point outside the depletion region. Thus both frontside and backside contacts can be used for backside biasing to fully deplete the substrate. Also, high resistivity gaps around the CCD channels and electrically floating channel stop regions can be provided in the CCD array around the CCD channels. The CCD array forms an imaging sensor useful in astronomy.

  13. Free-form illumination optics

    NASA Astrophysics Data System (ADS)

    Mohedano, Rubén; Chaves, Julio; Hernández, Maikel

    2016-04-01

    In many illumination problems, the beam pattern needed and/or some geometrical constraints lead to very asymmetric design conditions. These asymmetries have been solved in the past by means of arrangements of rotationally symmetric or linear lamps aimed in different directions whose patterns overlap to provide the asymmetric prescriptions or by splitting one single lamp into several sections, each one providing a part of the pattern. The development of new design methods yielding smooth continuous free-form optical surfaces to solve these challenging design problems, combined with the proper CAD modeling tools plus the development of multiple axes diamond turn machines, give birth to a new generation of optics. These are able to offer the performance and other advanced features, such as efficiency, compactness, or aesthetical advantages, and can be manufactured at low cost by injection molding. This paper presents two examples of devices with free-form optical surfaces, a camera flash, and a car headlamp.

  14. Off-axis illumination of lithography tool

    NASA Astrophysics Data System (ADS)

    Xing, Han; Lin, Li; Bin, Ma

    2013-12-01

    Lithography tool is a necessary part for LSI and VLSI. The illumination system design is an important part in the lithography optical system design. Off-axis illumination technology is an effective way to reducing resolution of lithography. The paper introduction the basic components of lithography tool, the principle of off-axis illumination reducing the resolution of lithography and focus on the two implementations of OAI technology, finally point out advantages and disadvantage of the two implementations.

  15. Simultaneous differential spinning disk fluorescence optical sectioning microscopy and nanomechanical mapping atomic force microscopy

    SciTech Connect

    Miranda, Adelaide; De Beule, Pieter A. A.

    2015-09-15

    Combined microscopy techniques offer the life science research community a powerful tool to investigate complex biological systems and their interactions. Here, we present a new combined microscopy platform based on fluorescence optical sectioning microscopy through aperture correlation microscopy with a Differential Spinning Disk (DSD) and nanomechanical mapping with an Atomic Force Microscope (AFM). The illumination scheme of the DSD microscope unit, contrary to standard single or multi-point confocal microscopes, provides a time-independent illumination of the AFM cantilever. This enables a distortion-free simultaneous operation of fluorescence optical sectioning microscopy and atomic force microscopy with standard probes. In this context, we discuss sample heating due to AFM cantilever illumination with fluorescence excitation light. Integration of a DSD fluorescence optical sectioning unit with an AFM platform requires mitigation of mechanical noise transfer of the spinning disk. We identify and present two solutions to almost annul this noise in the AFM measurement process. The new combined microscopy platform is applied to the characterization of a DOPC/DOPS (4:1) lipid structures labelled with a lipophilic cationic indocarbocyanine dye deposited on a mica substrate.

  16. Polarimetric target detection under uneven illumination.

    PubMed

    Huang, Bingjing; Liu, Tiegen; Han, Jiahui; Hu, Haofeng

    2015-09-07

    In polarimetric imaging, the uneven illumination could cause the significant spatial intensity fluctuations in the scene, and thus hampers the target detection. In this paper, we propose a method of illumination compensation and contrast optimization for Stokes polarimetric imaging, which allows significantly increasing the performance of target detection under uneven illumination. We show with numerical simulation and real-world experiment that, based on the intensity information contained in the polarization information, the contrast can be effectively enhanced by proper approach, which is of particular importance in practical applications with spatial illumination fluctuations, such as remote sensing.

  17. Illumination system characterization for hyperspectral imaging

    NASA Astrophysics Data System (ADS)

    Katrašnik, Jaka; Pernuš, Franjo; Likar, Boštjan

    2011-03-01

    Near-infrared hyperspectral imaging is becoming a popular tool in the biomedical field, especially for detection and analysis of different types of cancers, analysis of skin burns and bruises, imaging of blood vessels and for many other applications. As in all imaging systems, proper illumination is crucial to attain optimal image quality that is needed for best performance of image analysis algorithms. In hyperspectral imaging based on filters (AOTF, LCTF and filter wheel) the acquired spectral signature has to be representative in all parts of the imaged object. Therefore, the whole object must be equally well illuminated - without shadows and specular reflections. As there are no restrictions imposed on the material and geometry of the object, the desired object illumination can only be achieved with completely diffuse illumination. In order to minimize shadows and specular reflections in diffuse illumination the light illuminating the object must be spatially, angularly and spectrally uniform. We present and test two diffuse illumination system designs that try to achieve optimal uniformity of the above mentioned properties. The illumination uniformity properties were measured with an AOTF based hyperspectral imaging system utilizing a standard white diffuse reflectance target and a specially designed calibration target for estimating the spatial and angular illumination uniformity.

  18. Illumination discrimination in real and simulated scenes

    PubMed Central

    Radonjić, Ana; Pearce, Bradley; Aston, Stacey; Krieger, Avery; Dubin, Hilary; Cottaris, Nicolas P.; Brainard, David H.; Hurlbert, Anya C.

    2016-01-01

    Characterizing humans' ability to discriminate changes in illumination provides information about the visual system's representation of the distal stimulus. We have previously shown that humans are able to discriminate illumination changes and that sensitivity to such changes depends on their chromatic direction. Probing illumination discrimination further would be facilitated by the use of computer-graphics simulations, which would, in practice, enable a wider range of stimulus manipulations. There is no a priori guarantee, however, that results obtained with simulated scenes generalize to real illuminated scenes. To investigate this question, we measured illumination discrimination in real and simulated scenes that were well-matched in mean chromaticity and scene geometry. Illumination discrimination thresholds were essentially identical for the two stimulus types. As in our previous work, these thresholds varied with illumination change direction. We exploited the flexibility offered by the use of graphics simulations to investigate whether the differences across direction are preserved when the surfaces in the scene are varied. We show that varying the scene's surface ensemble in a manner that also changes mean scene chromaticity modulates the relative sensitivity to illumination changes along different chromatic directions. Thus, any characterization of sensitivity to changes in illumination must be defined relative to the set of surfaces in the scene.

  19. Dynamic Mechanical Responses of Arabidopsis Thylakoid Membranes during PSII-Specific Illumination

    PubMed Central

    Clausen, Casper H.; Brooks, Matthew D.; Li, Tai-De; Grob, Patricia; Kemalyan, Gigi; Nogales, Eva; Niyogi, Krishna K.; Fletcher, Daniel A.

    2014-01-01

    Remodeling of thylakoid membranes in response to illumination is an important process for the regulation of photosynthesis. We investigated the thylakoid network from Arabidopsis thaliana using atomic force microscopy to capture dynamic changes in height, elasticity, and viscosity of isolated thylakoid membranes caused by changes in illumination. We also correlated the mechanical response of the thylakoid network with membrane ultrastructure using electron microscopy. We find that the elasticity of the thylakoid membranes increases immediately upon PSII-specific illumination, followed by a delayed height change. Direct visualization by electron microscopy confirms that there is a significant change in the packing repeat distance of the membrane stacks in response to illumination. Although experiments with Gramicidin show that the change in elasticity depends primarily on the transmembrane pH gradient, the height change requires both the pH gradient and STN7-kinase-dependent phosphorylation of LHCII. Our studies indicate that lumen expansion in response to illumination is not simply a result of the influx of water, and we propose a dynamic model in which protein interactions within the lumen drive these changes. PMID:24806918

  20. Positron microscopy

    SciTech Connect

    Hulett, L.D. Jr.; Xu, J.

    1995-02-01

    The negative work function property that some materials have for positrons make possible the development of positron reemission microscopy (PRM). Because of the low energies with which the positrons are emitted, some unique applications, such as the imaging of defects, can be made. The history of the concept of PRM, and its present state of development will be reviewed. The potential of positron microprobe techniques will be discussed also.

  1. Use of astronomy filters in fluorescence microscopy.

    PubMed

    Piper, Jörg

    2012-02-01

    Monochrome astronomy filters are well suited for use as excitation or suppression filters in fluorescence microscopy. Because of their particular optical design, such filters can be combined with standard halogen light sources for excitation in many fluorescent probes. In this "low energy excitation," photobleaching (fading) or other irritations of native specimens are avoided. Photomicrographs can be taken from living motile fluorescent specimens also with a flash so that fluorescence images can be created free from indistinctness caused by movement. Special filter cubes or dichroic mirrors are not needed for our method. By use of suitable astronomy filters, fluorescence microscopy can be carried out with standard laboratory microscopes equipped with condensers for bright-field (BF) and dark-field (DF) illumination in transmitted light. In BF excitation, the background brightness can be modulated in tiny steps up to dark or black. Moreover, standard industry microscopes fitted with a vertical illuminator for examinations of opaque probes in DF or BF illumination based on incident light (wafer inspections, for instance) can also be used for excitation in epi-illumination when adequate astronomy filters are inserted as excitatory and suppression filters in the illuminating and imaging light path. In all variants, transmission bands can be modulated by transmission shift.

  2. Near-infrared hyperspectral reflective confocal microscopy

    NASA Astrophysics Data System (ADS)

    Huang, Wei; Zhang, Yunhai; Miao, Xin; Xue, Xiaojun; Xiao, Yun

    2016-10-01

    A Near-Infrared HyperSpectral Reflective Confocal Microscopy (NIHS-RCM) is proposed in order to get high resolution images of deep biological tissues such as skin. The microscopy system uses a super-continuum laser for illumination, an acousto-optic tunable filter (AOTF) for rapid selection of near-infrared spectrum, a resonant galvanometer scanner for high speed imaging (15f/s) and near-infrared avalanche diode as detector. Porcine skin and other experiments show that the microscopy system could get deep tissue images (180 μm), and show the different ingredients of tissue with different wavelength of illumination. The system has the ability of selectively imaging of multiple ingredients at deep tissue which can be used in skin diseases diagnosis and other fields.

  3. A near-field optical microscopy nanoarray

    SciTech Connect

    Semin, D.J.; Ambrose, W.P.; Goodwin, P.M.; Kwller, A.; Wendt, J.R.

    1996-12-31

    Multiplexing near-field scanning optical microscopy (NSOM) by the use of a nanoarray with parallel imaging is studied. The fabrication, characterization, and utilization of nanoarrays with {approximately} 100 nm diameter apertures spaced 500 nm center-to- center is presented. Extremely uniform nanoarrays with {approximately} 10{sup 8} apertures were fabricated by electron beam lithography and reactive ion etching. The nanoarrays were characterized by atomic force microscopy (AFM) and scanning electron microscopy (SEM). In this paper we utilize these nanoarrays in a laser-illuminated microscope with parallel detection on a charge- coupled device (CCD). Detection of B-phycoerythrin (B-PE) molecules using near-field illumination is presented. In principle, our system can be used to obtain high lateral resolution NSOM images over a wide-field of view (e.g. 50-100 {mu}m) within seconds.

  4. Köhler integrators embedded into illumination optics add functionality

    NASA Astrophysics Data System (ADS)

    Dross, O.; Mohedano, R.; Hernández, M.; Cvetkovic, A.; Miñano, J. C.; Benítez, P.

    2008-09-01

    The Köhler illumination concept was originally invented to achieve uniform illumination in microscopy1. Köhler integrators can also be formed by arrays of lenticulations that can be any combination of reflective and/or refractive surfaces, organized in corresponding pairs. Arrays of integrating facets can be arranged not only on flat surfaces but on rotationally symmetric and even freeform surfaces6. Currently flat lenslet arrays are widely applied as homogenizing optics2 for lithography, machine vision illumination, and projection. Adding Köhler facets onto already designed surfaces can improve the optical system performance, while respecting its original function. In general, the optics output can be made somewhat independent of the source characteristics, although at the expense of a slight ètendue dilution or efficiency losses. This work revises the Köhler concept and its application to different kind of optics, ranging from photovoltaic concentrators to automotive LED headlights. In the former, irradiance peaks on the solar cell can be avoided, while preserving high aiming tolerance (acceptance) of the solar concentrator. In the latter, LEDs drawbacks like large source image sizes, source misalignments, ill defined source edges, and low source radiance can be compensated.

  5. Illumination preference, illumination constancy and colour discrimination by bumblebees in an environment with patchy light.

    PubMed

    Arnold, Sarah E J; Chittka, Lars

    2012-07-01

    Patchy illumination presents foraging animals with a challenge, as the targets being sought may appear to vary in colour depending on the illumination, compromising target identification. We sought to explore how the bumblebee Bombus terrestris copes with tasks involving flower colour discrimination under patchy illumination. Light patches varied between unobscured daylight and leaf-shade, as a bee might encounter in and around woodland. Using a flight arena and coloured filters, as well as one or two different colours of artificial flower, we quantified how bees chose to forage when presented with foraging tasks under patchy illumination. Bees were better at discriminating a pair of similar colours under simulated unobscured daylight illumination than when foraging under leaf-shade illumination. Accordingly, we found that bees with prior experience of simulated daylight but not leaf-shade illumination initially preferred to forage in simulated daylight when all artificial flowers contained rewards as well as when only one colour was rewarding, whereas bees with prior experience of both illuminants did not exhibit this preference. Bees also switched between illuminants less than expected by chance. This means that bees prefer illumination conditions with which they are familiar, and in which rewarding flower colours are easily distinguishable from unrewarding ones. Under patchy illumination, colour discrimination performance was substantially poorer than in homogenous light. The bees' abilities at coping with patchy light may therefore impact on foraging behaviour in the wild, particularly in woodlands, where illumination can change over short spatial scales.

  6. Secure communication via quantum illumination

    NASA Astrophysics Data System (ADS)

    Shapiro, Jeffrey H.; Zhang, Zheshen; Wong, Franco N. C.

    2014-10-01

    In the quantum illumination protocol for secure communication, Alice prepares entangled signal and idler beams via spontaneous parametric downconversion. She sends the signal beam to Bob, while retaining the idler. Bob imposes message modulation on the beam he receives from Alice, amplifies it, and sends it back to her. Alice then decodes Bob's information by making a joint quantum measurement on the light she has retained and the light she has received from him. The basic performance analysis for this protocol—which demonstrates its immunity to passive eavesdropping, in which Eve can only listen to Alice and Bob's transmissions—is reviewed, along with the results of its first proof-of-principle experiment. Further analysis is then presented, showing that secure data rates in excess of 1 Gbps may be possible over 20-km-long fiber links with technology that is available or under development. Finally, an initial scheme for thwarting active eavesdropping, in which Eve injects her own light into Bob's terminal, is proposed and analyzed.

  7. Extinction-Optimized Volume Illumination.

    PubMed

    Ament, Marco; Zirr, Tobias; Dachsbacher, Carsten

    2016-05-16

    We present a novel method to optimize the attenuation of light for the single scattering model in direct volume rendering. A common problem of single scattering is the high dynamic range between lit and shadowed regions due to the exponential attenuation of light along a ray. Moreover, light is often attenuated too strong between a sample point and the camera, hampering the visibility of important features. Our algorithm employs an importance function to selectively illuminate important structures and make them visible from the camera. With the importance function, more light can be transmitted to the features of interest, while contextual structures cast shadows which provide visual cues for perception of depth. At the same time, more scattered light is transmitted from the sample point to the camera to improve the primary visibility of important features. We formulate a minimization problem that automatically determines the extinction along a view or shadow ray to obtain a good balance between sufficient transmittance and attenuation. In contrast to previous approaches, we do not require a computationally expensive solution of a global optimization, but instead provide a closed-form solution for each sampled extinction value along a view or shadow ray and thus achieve interactive performance.

  8. Super-resolved spatial light interference microscopy.

    PubMed

    Chu, Kaiqin; Smith, Zachary J; Wachsmann-Hogiu, Sebastian; Lane, Stephen

    2012-03-01

    We report a scheme to achieve resolution beyond the diffraction limit in spatial light interference microscopy (SLIM). By adding a grating to the optical path, the structured illumination technique can be used to improve the resolution by a factor of 2. We show that a direct application of the structured illumination technique, however, has proved to be unsuccessful. Through two crucial modifications, namely, one to the pupil plane of the objective and the other to the demodulation procedure, faithful phase information of the object is recovered and the resolution is improved by a factor of 2.

  9. 29 CFR 1926.56 - Illumination.

    Code of Federal Regulations, 2012 CFR

    2012-07-01

    ... shall be lighted to not less than the minimum illumination intensities listed in Table D-3 while any work is in progress: Table D-3—Minimum Illumination Intensities in Foot-Candles Foot-candles Area or... drilling, mucking, and scaling. Bureau of Mines approved cap lights shall be acceptable for use in...

  10. 29 CFR 1926.56 - Illumination.

    Code of Federal Regulations, 2013 CFR

    2013-07-01

    ... shall be lighted to not less than the minimum illumination intensities listed in Table D-3 while any work is in progress: Table D-3—Minimum Illumination Intensities in Foot-Candles Foot-candles Area or... drilling, mucking, and scaling. Bureau of Mines approved cap lights shall be acceptable for use in...

  11. Translating Rimbaud's "Illuminations": Games with Words

    ERIC Educational Resources Information Center

    Slote, Daniel

    1978-01-01

    Rimbaud's "Illuminations," one vast word-game, is used as an example of one of the most interesting challenges in translation--the rendering of plays on words. The process is discussed and illustrated by the analysis of numerous segments from "Illuminations." It is concluded that a satisfactory translation is almost impossible.…

  12. Reflectance and Illumination Recovery in the Wild.

    PubMed

    Lombardi, Stephen; Nishino, Ko

    2016-01-01

    The appearance of an object in an image encodes invaluable information about that object and the surrounding scene. Inferring object reflectance and scene illumination from an image would help us decode this information: reflectance can reveal important properties about the materials composing an object; the illumination can tell us, for instance, whether the scene is indoors or outdoors. Recovering reflectance and illumination from a single image in the real world, however, is a difficult task. Real scenes illuminate objects from every visible direction and real objects vary greatly in reflectance behavior. In addition, the image formation process introduces ambiguities, like color constancy, that make reversing the process ill-posed. To address this problem, we propose a Bayesian framework for joint reflectance and illumination inference in the real world. We develop a reflectance model and priors that precisely capture the space of real-world object reflectance and a flexible illumination model that can represent real-world illumination with priors that combat the deleterious effects of image formation. We analyze the performance of our approach on a set of synthetic data and demonstrate results on real-world scenes. These contributions enable reliable reflectance and illumination inference in the real world.

  13. The 2015 super-resolution microscopy roadmap

    NASA Astrophysics Data System (ADS)

    Hell, Stefan W.; Sahl, Steffen J.; Bates, Mark; Zhuang, Xiaowei; Heintzmann, Rainer; Booth, Martin J.; Bewersdorf, Joerg; Shtengel, Gleb; Hess, Harald; Tinnefeld, Philip; Honigmann, Alf; Jakobs, Stefan; Testa, Ilaria; Cognet, Laurent; Lounis, Brahim; Ewers, Helge; Davis, Simon J.; Eggeling, Christian; Klenerman, David; Willig, Katrin I.; Vicidomini, Giuseppe; Castello, Marco; Diaspro, Alberto; Cordes, Thorben

    2015-11-01

    Far-field optical microscopy using focused light is an important tool in a number of scientific disciplines including chemical, (bio)physical and biomedical research, particularly with respect to the study of living cells and organisms. Unfortunately, the applicability of the optical microscope is limited, since the diffraction of light imposes limitations on the spatial resolution of the image. Consequently the details of, for example, cellular protein distributions, can be visualized only to a certain extent. Fortunately, recent years have witnessed the development of ‘super-resolution’ far-field optical microscopy (nanoscopy) techniques such as stimulated emission depletion (STED), ground state depletion (GSD), reversible saturated optical (fluorescence) transitions (RESOLFT), photoactivation localization microscopy (PALM), stochastic optical reconstruction microscopy (STORM), structured illumination microscopy (SIM) or saturated structured illumination microscopy (SSIM), all in one way or another addressing the problem of the limited spatial resolution of far-field optical microscopy. While SIM achieves a two-fold improvement in spatial resolution compared to conventional optical microscopy, STED, RESOLFT, PALM/STORM, or SSIM have all gone beyond, pushing the limits of optical image resolution to the nanometer scale. Consequently, all super-resolution techniques open new avenues of biomedical research. Because the field is so young, the potential capabilities of different super-resolution microscopy approaches have yet to be fully explored, and uncertainties remain when considering the best choice of methodology. Thus, even for experts, the road to the future is sometimes shrouded in mist. The super-resolution optical microscopy roadmap of Journal of Physics D: Applied Physics addresses this need for clarity. It provides guidance to the outstanding questions through a collection of short review articles from experts in the field, giving a thorough

  14. Illuminant spectrum estimation at a pixel.

    PubMed

    Ratnasingam, Sivalogeswaran; Hernández-Andrés, Javier

    2011-04-01

    In this paper, an algorithm is proposed to estimate the spectral power distribution of a light source at a pixel. The first step of the algorithm is forming a two-dimensional illuminant invariant chromaticity space. In estimating the illuminant spectrum, generalized inverse estimation and Wiener estimation methods were applied. The chromaticity space was divided into small grids and a weight matrix was used to estimate the illuminant spectrum illuminating the pixels that fall within a grid. The algorithm was tested using a different number of sensor responses to determine the optimum number of sensors for accurate colorimetric and spectral reproduction. To investigate the performance of the algorithm realistically, the responses were multiplied with Gaussian noise and then quantized to 10 bits. The algorithm was tested with standard and measured data. Based on the results presented, the algorithm can be used with six sensors to obtain a colorimetrically good estimate of the illuminant spectrum at a pixel.

  15. Chromatic illumination discrimination ability reveals that human colour constancy is optimised for blue daylight illuminations.

    PubMed

    Pearce, Bradley; Crichton, Stuart; Mackiewicz, Michal; Finlayson, Graham D; Hurlbert, Anya

    2014-01-01

    The phenomenon of colour constancy in human visual perception keeps surface colours constant, despite changes in their reflected light due to changing illumination. Although colour constancy has evolved under a constrained subset of illuminations, it is unknown whether its underlying mechanisms, thought to involve multiple components from retina to cortex, are optimised for particular environmental variations. Here we demonstrate a new method for investigating colour constancy using illumination matching in real scenes which, unlike previous methods using surface matching and simulated scenes, allows testing of multiple, real illuminations. We use real scenes consisting of solid familiar or unfamiliar objects against uniform or variegated backgrounds and compare discrimination performance for typical illuminations from the daylight chromaticity locus (approximately blue-yellow) and atypical spectra from an orthogonal locus (approximately red-green, at correlated colour temperature 6700 K), all produced in real time by a 10-channel LED illuminator. We find that discrimination of illumination changes is poorer along the daylight locus than the atypical locus, and is poorest particularly for bluer illumination changes, demonstrating conversely that surface colour constancy is best for blue daylight illuminations. Illumination discrimination is also enhanced, and therefore colour constancy diminished, for uniform backgrounds, irrespective of the object type. These results are not explained by statistical properties of the scene signal changes at the retinal level. We conclude that high-level mechanisms of colour constancy are biased for the blue daylight illuminations and variegated backgrounds to which the human visual system has typically been exposed.

  16. Chromatic Illumination Discrimination Ability Reveals that Human Colour Constancy Is Optimised for Blue Daylight Illuminations

    PubMed Central

    Pearce, Bradley; Crichton, Stuart; Mackiewicz, Michal; Finlayson, Graham D.; Hurlbert, Anya

    2014-01-01

    The phenomenon of colour constancy in human visual perception keeps surface colours constant, despite changes in their reflected light due to changing illumination. Although colour constancy has evolved under a constrained subset of illuminations, it is unknown whether its underlying mechanisms, thought to involve multiple components from retina to cortex, are optimised for particular environmental variations. Here we demonstrate a new method for investigating colour constancy using illumination matching in real scenes which, unlike previous methods using surface matching and simulated scenes, allows testing of multiple, real illuminations. We use real scenes consisting of solid familiar or unfamiliar objects against uniform or variegated backgrounds and compare discrimination performance for typical illuminations from the daylight chromaticity locus (approximately blue-yellow) and atypical spectra from an orthogonal locus (approximately red-green, at correlated colour temperature 6700 K), all produced in real time by a 10-channel LED illuminator. We find that discrimination of illumination changes is poorer along the daylight locus than the atypical locus, and is poorest particularly for bluer illumination changes, demonstrating conversely that surface colour constancy is best for blue daylight illuminations. Illumination discrimination is also enhanced, and therefore colour constancy diminished, for uniform backgrounds, irrespective of the object type. These results are not explained by statistical properties of the scene signal changes at the retinal level. We conclude that high-level mechanisms of colour constancy are biased for the blue daylight illuminations and variegated backgrounds to which the human visual system has typically been exposed. PMID:24586299

  17. Long-range laser-illuminated imaging

    NASA Astrophysics Data System (ADS)

    Dayton, David C.; Browne, Stephen L.; Sandven, Steven C.; Gonglewski, John D.; Gallegos, Joe; Shilko, Michael L., Sr.

    2000-11-01

    We demonstrate the utility of laser illuminated imaging for clandestine night time surveillance from a simulated airborne platform at standoff ranges in excess 20 km. In order to reduce the necessary laser per pulse energy required for illumination at such long ranges, and to mitigate atmospheric turbulence effects on image resolution, we have investigated a unique multi-frame post-processing technique. It is shown that in the presence of atmospheric turbulence and coherent speckle effects, this approach can produce superior results to conventional scene flood illumination.

  18. RGB digital lensless holographic microscopy

    NASA Astrophysics Data System (ADS)

    Garcia-Sucerquia, Jorge

    2013-11-01

    The recent introduction of color digital lensless holographic microscopy (CDLHM) has shown the possibility of imaging microscopic specimens at full color without the need of lenses. Owing to the simplicity, robustness, and compactness of the digital lensless holographic microscopes (DLHM), they have been presented as the ideal candidates to being developed into portable holographic microscopes. However, in the case of CDLHM the utilization of three independent lasers hinders the portability option for this microscope. In this contribution an alternative to reduce the complexity of CDLHM aimed to recover the portability of this microscopy technology is presented. A super-bright white-light light-emitting diode (LED) is spectrally and spatially filtered to produce the needed illumination by CDLHM to work. CDLHM with LED illumination is used to image at full color a section of the head of a drosophila melanogaster fly (fruit fly). The LED-CDLHM method shows the capability of imaging objects of 2μm size in comparison with the micrometer resolution reported for LASER-CDLHM.

  19. Illumination control apparatus for compensating solar light

    NASA Technical Reports Server (NTRS)

    Owens, L. J. (Inventor)

    1978-01-01

    An illumination control apparatus is presented for supplementing light from solar radiation with light from an artificial light source to compensate for periods of insufficient levels of solar light. The apparatus maintains a desired illumination level within an interior space comprising an artificial light source connected to an electrical power source with a switch means for selectively energizing said light source. An actuator means for controlling the on-off operation of the switch means is connected to a light sensor which responses to the illumination level of the interior space. A limit switch carried adjacent to the actuator limits the movement of the actuator within a predetermined range so as to prevent further movement thereof during detection of erroneous illumination conditions.

  20. The Freeform Reflector for Uniform Rectangular Illumination

    NASA Astrophysics Data System (ADS)

    Ding, Yi; Gu, Pei-Fu; Zheng, Zhen-Rong

    2007-12-01

    Energy from the source was rearranged through reflection by a freeform reflector, in order to get uniform rectangular illumination. The numerical results of partial differential equation sets were investigated to obtain the freeform reflector and these equations were obtained upon the determination of the characters of source and the desired illumination. As an example, a light emitting diode (LED) with a Lambertian light-emitting surface of 1 × 1 mm2 and a viewing angle of 120° was applied as the source, and the target plane was a 4:3 rectangle with uniform illumination. The projective length of the reflector on x-axis is about 23 mm, and on y-axis is about 21 mm. Thus the illumination system is very compact.

  1. An illuminated flute needle for vitreoretinal surgery.

    PubMed

    Davison, C N; Rosen, P H

    1994-06-01

    We have developed a simple self-illuminated flute needle for internal drainage of subretinal fluid during three-port vitrectomy. This instrument facilitates visualization and drainage through peripheral retinal breaks.

  2. 49 CFR 230.86 - Required illumination.

    Code of Federal Regulations, 2013 CFR

    2013-10-01

    ... provides illumination sufficient for a steam locomotive engineer in the cab to see, in a clear atmosphere... other than to pick up a detached portion of its train or to make terminal movements, it shall also...

  3. 49 CFR 230.86 - Required illumination.

    Code of Federal Regulations, 2014 CFR

    2014-10-01

    ... provides illumination sufficient for a steam locomotive engineer in the cab to see, in a clear atmosphere... other than to pick up a detached portion of its train or to make terminal movements, it shall also...

  4. 49 CFR 230.86 - Required illumination.

    Code of Federal Regulations, 2010 CFR

    2010-10-01

    ... provides illumination sufficient for a steam locomotive engineer in the cab to see, in a clear atmosphere... other than to pick up a detached portion of its train or to make terminal movements, it shall also...

  5. Coded multi-angular illumination for Fourier ptychography based on Hadamard codes

    NASA Astrophysics Data System (ADS)

    Sun, Jiasong; Zhang, Yuzhen; Zuo, Chao; Chen, Qian; Feng, Shijie; Hu, Yan; Zhang, Jialin

    2015-07-01

    Fourier ptychographic microscopy (FPM) is a newly developed super-resolution technique, which employs angularly varying illumination and a phase retrieval algorithm to surpass the diffraction limit of the objective lens. Specifically, FP captures a set of low-resolution (LR) images, under angularly varying illuminations, and stitches them together in the Fourier domain. However, because the requisite large number of incident illumination angles, the long capturing process becomes an obvious limiting factor. Furthermore, in order to acquire high-dynamic range images, the time can be increased several times over. In this work, utilizing the Hadamard code principle, we propose a highly efficient method, which applies coded multi-angular illumination for FPM, to shorten the exposure time of each raw image. High acquisition efficiency is achieved by employing an optimal multi-angular illumination scheme by using two set of Hadamard coded multiplexing patterns. Both simulation and experimental results indicate that the proposed multi-angular illumination process could shorten the acquisition time of conventional FPM.

  6. Insulator Surface Flashover Due to UV Illumination

    DTIC Science & Technology

    2009-06-01

    The surface of an insulator under vacuum and under electrical charge will flashover when illuminated by a critical dose of ultra-violet (UV...fluence (energy per unit area) required to induce surface flashover of vacuum insulators for some candid insulator materials: High Density... Insulator Surface Flashover Due to UV Illumination1 J. B. Javedani, T.L. Houck, D.A. Lahowe, G.E. Vogtlin and D.A. Goerz Lawrence Livermore

  7. Optical microscopy using a single-molecule light source

    PubMed

    Michaelis; Hettich; Mlynek; Sandoghdar

    2000-05-18

    Rapid progress in science on nanoscopic scales has promoted increasing interest in techniques of ultrahigh-resolution optical microscopy. The diffraction limit can be surpassed by illuminating an object in the near field through a sub-wavelength aperture at the end of a sharp metallic probe. Proposed modifications of this technique involve replacing the physical aperture by a nanoscopic active light source. Advances in the spatial and spectral detection of individual fluorescent molecules, using near-field and far-field methods, suggest the possibility of using a single molecule as the illumination source. Here we present optical images taken with a single molecule as a point-like source of illumination, by combining fluorescence excitation spectroscopy with shear-force microscopy. Our single-molecule probe has potential for achieving molecular resolution in optical microscopy; it should also facilitate controlled studies of nanometre-scale phenomena (such as resonant energy transfer) with improved lateral and axial spatial resolution.

  8. Circumventing photodamage in live-cell microscopy

    PubMed Central

    Magidson, Valentin; Khodjakov, Alexey

    2013-01-01

    Fluorescence microscopy has become an essential tool in cell biology. This technique allows researchers to visualize the dynamics of tissue, cells, individual organelles and macromolecular assemblies inside the cell. Unfortunately, fluorescence microscopy is not completely ‘non-invasive’ as the high-intensity excitation light required for excitation of fluorophores is inherently toxic for live cells. Physiological changes induced by excessive illumination can lead to artifacts and abnormal responses. In this chapter we review major factors that contribute to phototoxicity and discuss practical solutions for circumventing photodamage. These solutions include the proper choice of image acquisition parameters, optimization of filter sets, hardware synchronization, and the use of intelligent illumination to avoid unnecessary light exposure. PMID:23931522

  9. The Effect of Subbandgap Illumination on the Bulk Resistivity of CdZnTe

    SciTech Connect

    Wright, Jonathan S.; Washington II, Aaron L.; Duff, Martine C.; Burger, Arnold; Groza, Michael; Matei, Liviu; Buliga, Vladimir

    2013-08-24

    The variation in bulk resistivity during infrared (IR) illumination above 950 nm of state-of-the-art CdZnTe (CZT) crystals grown using the traveling heating method or the modified Bridgman method is documented. The change in steady-state current with and without illumination is also evaluated. The influence of secondary phases (SP) on current-voltage (I-V) characteristics is discussed using IR transmission microscopy to determine the defect concentration within the crystal bulk. SP present within the CZT are connected to the existence of deep, IR-excitable traps within the bandgap.

  10. Surface color perception under two illuminants: the second illuminant reduces color constancy

    NASA Technical Reports Server (NTRS)

    Yang, Joong Nam; Shevell, Steven K.

    2003-01-01

    This study investigates color perception in a scene with two different illuminants. The two illuminants, in opposite corners, simultaneously shine on a (simulated) scene with an opaque dividing wall, which controls how much of the scene is illuminated by each source. In the first experiment, the height of the dividing wall was varied. This changed the amount of each illuminant reaching objects on the opposite side of the wall. Results showed that the degree of color constancy decreased when a region on one side of the wall had cues to both illuminants, suggesting that cues from the second illuminant are detrimental to color constancy. In a later experiment, color constancy was found to improve when the specular highlight cues from the second illuminant were altered to be consistent with the first illuminant. This corroborates the influence of specular highlights in surface color perception, and suggests that the reduced color constancy in the first experiment is due to the inconsistent, though physically correct, cues from the two illuminants.

  11. Quantitative analysis of platelets aggregates in 3D by digital holographic microscopy

    PubMed Central

    Boudejltia, Karim Zouaoui; Ribeiro de Sousa, Daniel; Uzureau, Pierrick; Yourassowsky, Catherine; Perez-Morga, David; Courbebaisse, Guy; Chopard, Bastien; Dubois, Frank

    2015-01-01

    Platelet spreading and retraction play a pivotal role in the platelet plugging and the thrombus formation. In routine laboratory, platelet function tests include exhaustive information about the role of the different receptors present at the platelet surface without information on the 3D structure of platelet aggregates. In this work, we develop, a method in Digital Holographic Microscopy (DHM) to characterize the platelet and aggregate 3D shapes using the quantitative phase contrast imaging. This novel method is suited to the study of platelets physiology in clinical practice as well as the development of new drugs. PMID:26417523

  12. Scanning Probe Microscopy of Organic Solar Cells

    NASA Astrophysics Data System (ADS)

    Reid, Obadiah G.

    EFM, and of greater utility in identifying local changes in steady-state charge density that can be associated with charge trapping. In the second case, we have developed a new understanding of charge transport between a sharp AFM tip and planar substrates applicable to conductive and photoconductive atomic force microscopy, and shown that hole-only transport characteristics can be easily obtained including quantitative values of the charge carrier mobility. Finally, we have shown that intensity-dependent photoconductive atomic force microscopy measurements can be used to infer the 3D structure of organic photovoltaic materials, and gained new insight into the influence vertical composition of the these devices can have on their open-circuit voltage and its intensity dependence.

  13. Photobleaching imprinting microscopy: seeing clearer and deeper.

    PubMed

    Gao, Liang; Garcia-Uribe, Alejandro; Liu, Yan; Li, Chiye; Wang, Lihong V

    2014-01-15

    We present a generic sub-diffraction-limited imaging method - photobleaching imprinting microscopy (PIM) - for biological fluorescence imaging. A lateral resolution of 110 nm was measured, more than a twofold improvement over the optical diffraction limit. Unlike other super-resolution imaging techniques, PIM does not require complicated illumination modules or specific fluorescent dyes. PIM is expected to facilitate the conversion of super-resolution imaging into a routine lab tool, making it accessible to a much broader biological research community. Moreover, we show that PIM can increase the image contrast of biological tissue, effectively extending the fundamental depth limit of multi-photon fluorescence microscopy.

  14. Scanning Tunneling Optical Resonance Microscopy

    NASA Technical Reports Server (NTRS)

    Bailey, Sheila; Wilt, Dave; Raffaelle, Ryne; Gennett, Tom; Tin, Padetha; Lau, Janice; Castro, Stephanie; Jenkins, Philip; Scheiman, Dave

    2003-01-01

    Scanning tunneling optical resonance microscopy (STORM) is a method, now undergoing development, for measuring optoelectronic properties of materials and devices on the nanoscale by means of a combination of (1) traditional scanning tunneling microscopy (STM) with (2) tunable laser spectroscopy. In STORM, an STM tip probing a semiconductor is illuminated with modulated light at a wavelength in the visible-to-near-infrared range and the resulting photoenhancement of the tunneling current is measured as a function of the illuminating wavelength. The photoenhancement of tunneling current occurs when the laser photon energy is sufficient to excite charge carriers into the conduction band of the semiconductor. Figure 1 schematically depicts a proposed STORM apparatus. The light for illuminating the semiconductor specimen at the STM would be generated by a ring laser that would be tunable across the wavelength range of interest. The laser beam would be chopped by an achromatic liquid-crystal modulator. A polarization-maintaining optical fiber would couple the light to the tip/sample junction of a commercial STM. An STM can be operated in one of two modes: constant height or constant current. A STORM apparatus would be operated in the constant-current mode, in which the height of the tip relative to the specimen would be varied in order to keep the tunneling current constant. In this mode, a feedback control circuit adjusts the voltage applied to a piezoelectric actuator in the STM that adjusts the height of the STM tip to keep the tunneling current constant. The exponential relationship between the tunneling current and tip-to-sample distance makes it relatively easy to implement this mode of operation. The choice of method by which the photoenhanced portion of the tunneling current would be measured depends on choice of the frequency at which the input illumination would be modulated (chopped). If the frequency of modulation were low enough (typically < 10 Hz) that the

  15. Deterministic phase retrieval employing spherical illumination

    NASA Astrophysics Data System (ADS)

    Martínez-Carranza, J.; Falaggis, K.; Kozacki, T.

    2015-05-01

    Deterministic Phase Retrieval techniques (DPRTs) employ a series of paraxial beam intensities in order to recover the phase of a complex field. These paraxial intensities are usually generated in systems that employ plane-wave illumination. This type of illumination allows a direct processing of the captured intensities with DPRTs for recovering the phase. Furthermore, it has been shown that intensities for DPRTs can be acquired from systems that use spherical illumination as well. However, this type of illumination presents a major setback for DPRTs: the captured intensities change their size for each position of the detector on the propagation axis. In order to apply the DPRTs, reescalation of the captured intensities has to be applied. This condition can increase the error sensitivity of the final phase result if it is not carried out properly. In this work, we introduce a novel system based on a Phase Light Modulator (PLM) for capturing the intensities when employing spherical illumination. The proposed optical system enables us to capture the diffraction pattern of under, in, and over-focus intensities. The employment of the PLM allows capturing the corresponding intensities without displacing the detector. Moreover, with the proposed optical system we can control accurately the magnification of the captured intensities. Thus, the stack of captured intensities can be used in DPRTs, overcoming the problems related with the resizing of the images. In order to prove our claims, the corresponding numerical experiments will be carried out. These simulations will show that the retrieved phases with spherical illumination are accurate and can be compared with those that employ plane wave illumination. We demonstrate that with the employment of the PLM, the proposed optical system has several advantages as: the optical system is compact, the beam size on the detector plane is controlled accurately, and the errors coming from mechanical motion can be suppressed easily.

  16. Optical microscopy versus scanning electron microscopy in urolithiasis.

    PubMed

    Marickar, Y M Fazil; Lekshmi, P R; Varma, Luxmi; Koshy, Peter

    2009-10-01

    Stone analysis is incompletely done in many clinical centers. Identification of the stone component is essential for deciding future prophylaxis. X-ray diffraction, Fourier transform infrared spectroscopy, and scanning electron microscopy (SEM) still remains a distant dream for routine hospital work. It is in this context that optical microscopy is suggested as an alternate procedure. The objective of this article was to assess the utility of an optical microscope which gives magnification of up to 40x and gives clear picture of the surface of the stones. In order to authenticate the morphological analysis of urinary stones, SEM and elemental distribution analysis were performed. A total of 250 urinary stones of different compositions were collected from stone clinic, photographed, observed under an optical microscope, and optical photographs were taken at different angles. Twenty-five representative samples among these were gold sputtered to make them conductive and were fed into the SEM machine. Photographs of the samples were taken at different angles at magnifications up to 4,000. Elemental distribution analysis (EDAX) was done to confirm the composition. The observations of the two studies were compared. The different appearances of the stones under optical illuminated microscopy were mostly standardized appearances, namely bosselations of pure whewellite, spiculations of weddellite, bright yellow colored appearance of uric acid, and dirty white amorphous appearance of phosphates. SEM and EDAX gave clearer pictures and gave added confirmation of the stone composition. From the references thus obtained, it was possible to confirm the composition by studying the optical microscopic pictures. Higher magnification capacity of the SEM and the EDAX patterns are useful to give reference support for performing optical microscopy work. After standardization, routine analysis can be performed with optical microscopy. The advantage of the optical microscope is that, it

  17. The 3D Structure of the Galactic Bulge

    NASA Astrophysics Data System (ADS)

    Zoccali, Manuela; Valenti, Elena

    2016-06-01

    We review the observational evidences concerning the three-dimensional structure of the Galactic bulge. Although the inner few kpc of our Galaxy are normally referred to as the bulge, all the observations demonstrate that this region is dominated by a bar, i.e., the bulge is a bar. The bar has a boxy/peanut (X-shaped) structure in its outer regions, while it seems to become less and less elongated in its innermost region. A thinner and longer structure departing from the main bar has also been found, although the observational evidences that support the scenario of two separate structures has been recently challenged. Metal-poor stars ([Fe/H] ≲ -0.5 dex) trace a different structure, and also have different kinematics.

  18. The 3D structures of VDAC represent a native conformation

    PubMed Central

    Hiller, Sebastian; Abramson, Jeff; Mannella, Carmen; Wagner, Gerhard; Zeth, Kornelius

    2010-01-01

    The most abundant protein of the mitochondrial outer membrane is the voltage-dependent anion channel (VDAC), which facilitates the exchange of ions and molecules between mitochondria and cytosol and is regulated by interactions with other proteins and small molecules. VDAC has been extensively studied for more than three decades, and last year three independent investigations revealed a structure of VDAC-1 exhibiting 19 transmembrane β-strands, constituting a unique structural class of β-barrel membrane proteins. Here, we provide a historical perspective on VDAC research and give an overview of the experimental design used to obtain these structures. Furthermore, we validate the protein refolding approach and summarize biochemical and biophysical evidence that links the 19-stranded structure to the native form of VDAC. PMID:20708406

  19. 3D structure of nearby groups of galaxies

    NASA Astrophysics Data System (ADS)

    Makarova, L.; Makarov, D.; Klypin, A.; Gottlöber, S.

    2016-10-01

    Using high accuracy distance estimates, we study the three-dimensional distribution of galaxies in five galaxy groups at a distance less than 5 Mpc from the Milky Way. Due to proximity of these groups our sample of galaxies is nearly complete down to extremely small dwarf galaxies with absolute magnitudes M B = -12. We find that the average number-density profile of the groups shows a steep power-law decline dn/dV ˜ R-3 at distances R=(100-500) kpc consistent with predictions of the standard cosmological model. We also find that there is no indication of a truncation or a cutoff in the density at the expected virial radius: the density profile extends at least to 1.5 Mpc. Vast majority of galaxies within 1.5 Mpc radius around group centres are gas-rich star-forming galaxies. Early-type galaxies are found only in the central ˜ 300 kpc region. Lack of dwarf spheroidal and dwarf elliptical galaxies in the field and in the outskirts of large groups is a clear indication that these galaxies experienced morphological transformation when they came close to the central region of forming galaxy group.

  20. Small-angle scattering and 3D structure interpretation.

    PubMed

    Trewhella, Jill

    2016-10-01

    This review focuses on advances in the application of solution small-angle scattering (SAS) in structural analysis of biomolecules and the complexes they form. Examples highlighted illustrate the unique contribution of SAS, using both X-rays and neutrons, in hybrid or integrative modelling methods. The increased information content when neutron scattering with contrast variation is used is a particular focus. Finally, progress toward an agreed reporting framework, the development of open data and model archives, and the importance of these initiatives is covered.

  1. 3-D Structure of Sunspots using Imaging Spectroscopy

    NASA Technical Reports Server (NTRS)

    Balasubramaniam, K. S.; Gary, G. Allen; Reardon, K.

    2006-01-01

    We use the Interferometric BIdimensional Spectrometer (IBIS) of the INAF/Arcetri Astrophysical Observatory and installed at the National Solar Observatory (NSO) Dunn Solar Telescope, to understand the structure of sunspots. Using the spectral lines FeI 6301.5 A, FeII 7224.4 A, and CaII 8542.6 A, we examine the spectroscopic variation of sunspot penumbral and umbral structures at the heights of formation of these lines. These high resolution observations were acquired on 2004 July 30-31, of active region NOAA 10654, using the high order NSO adaptive optics system. We map the spatio-temporal variation of Doppler signatures in these spectral lines, from the photosphere to the chromosphere. From a 70-minute temporal average of individual 32-second cadence Doppler observations we find that the averaged velocities decrease with height, about 3.5 times larger in the deeper photosphere (FeII 7224.4 A; height-of-formation approx. 50 km) than in the upper photosphere FeI 6301.5 A; height-of-formation approx. 350 km), There is a remarkable coherence of Doppler signals over the height difference of 300 km. From a high-speed animation of the Doppler sequence we find evidence for what appears to be ejection of high speed gas concentrations from edges of penumbral filaments into the surrounding granular photosphere. The Evershed flow persists a few arcseconds beyond the traditionally demarcated penumbra-granulation boundary. We present these and other results and discuss the implications of these measurements for sunspot models.

  2. Advancements in 3D Structural Analysis of Geothermal Systems

    SciTech Connect

    Siler, Drew L; Faulds, James E; Mayhew, Brett; McNamara, David

    2013-06-23

    Robust geothermal activity in the Great Basin, USA is a product of both anomalously high regional heat flow and active fault-controlled extension. Elevated permeability associated with some fault systems provides pathways for circulation of geothermal fluids. Constraining the local-scale 3D geometry of these structures and their roles as fluid flow conduits is crucial in order to mitigate both the costs and risks of geothermal exploration and to identify blind (no surface expression) geothermal resources. Ongoing studies have indicated that much of the robust geothermal activity in the Great Basin is associated with high density faulting at structurally complex fault intersection/interaction areas, such as accommodation/transfer zones between discrete fault systems, step-overs or relay ramps in fault systems, intersection zones between faults with different strikes or different senses of slip, and horse-tailing fault terminations. These conceptualized models are crucial for locating and characterizing geothermal systems in a regional context. At the local scale, however, pinpointing drilling targets and characterizing resource potential within known or probable geothermal areas requires precise 3D characterization of the system. Employing a variety of surface and subsurface data sets, we have conducted detailed 3D geologic analyses of two Great Basin geothermal systems. Using EarthVision (Dynamic Graphics Inc., Alameda, CA) we constructed 3D geologic models of both the actively producing Brady’s geothermal system and a ‘greenfield’ geothermal prospect at Astor Pass, NV. These 3D models allow spatial comparison of disparate data sets in 3D and are the basis for quantitative structural analyses that can aid geothermal resource assessment and be used to pinpoint discrete drilling targets. The relatively abundant data set at Brady’s, ~80 km NE of Reno, NV, includes 24 wells with lithologies interpreted from careful analysis of cuttings and core, a 1:24,000 scale detailed geologic map and cross-sections, 2D seismic reflection profiles and other geophysical data, and downhole temperature data. The 3D geologic model based on these data consists of 61 fault planes, 25 distinct stratigraphic units, and 2 intrusive bodies. Geothermal fluids are produced from a left step-over/relay ramp within the Brady’s Fault Zone (BFZ). Under local stress conditions, fault segments that strike NNE-to-NE are most likely to slip and/or dilate, and therefore transmit geothermal fluids. The 3D model defines the locations of discrete fault intersections within the BFZ and indicates that the densest zones of structurally controlled fracture permeability are ~10-to-10s of meters in diameter and plunge ~55° NW-NNW beneath the heart of the BFZ step over. The locations of high intersection density, high fault slip and dilation tendency, high subsurface temperature, and lithologies known to support high fracture permeability are combined to produce 3D ‘fairway’ maps useful in both assessments of geothermal resource potential and for defining drilling targets. Astor Pass is located on the Pyramid Lake Paiute Reservation, ~80 km north of Reno, NV. It is a prospective ‘greenfield’ geothermal area, and thus subsurface data are relatively sparse. Available data include: two relatively deep wells (~1400 m) and one shallower well (~500 m) with lithologies interpreted from drill cuttings, several 2D seismic reflection profiles, a 1:24,000 scale geologic map and cross-section, a shallow temperature survey, and downhole temperature data. 3D modeling based on these data has defined 19 distinct fault planes and 16 stratigraphic units. Based on the stress field calculated from borehole breakouts, drilling induced tensile cracks and petal-centerline cracks in the two relatively deep wells, 3D slip and dilation tendency analysis indicates that northerly striking fault segments are most likely to slip and/or dilate, and therefore transmit geothermal fluids. Analysis of fault intersection density indicates that the highest density of structurally controlled permeability within the field lies in a narrow (10-to-10s of m) zone plunging moderately (~35°) to the NNW beneath Pleistocene tufa deposits. This zone of increased fracture density, which we interpret as the primary upflow zone, is controlled by the intersection of N-to-NNW striking normal faults and a WNW striking dextral fault zone and represents the most promising target for future drilling. Construction of a 3D geologic model involves integration of a variety of data into an internally consistent framework. A robust model allows for spatial comparison between the various types of data (structural, stratigraphic, geophysical, temperature, etc.) that are commonly used independently to site geothermal wells. Furthermore, highly detailed 3D geologic models provide the basis for additional quantitative analysis, including 3D fault slip and dilation tendency analysis and the precise location of structurally controlled permeability pathways. These analyses provide detailed information relating to the internal dynamics of geothermal systems and can mitigate the costs and risks of geothermal exploration and development by contributing to better well targeting and more accurate evaluations of resource potential.

  3. MUFOLD: A new solution for protein 3D structure prediction.

    PubMed

    Zhang, Jingfen; Wang, Qingguo; Barz, Bogdan; He, Zhiquan; Kosztin, Ioan; Shang, Yi; Xu, Dong

    2010-04-01

    There have been steady improvements in protein structure prediction during the past 2 decades. However, current methods are still far from consistently predicting structural models accurately with computing power accessible to common users. Toward achieving more accurate and efficient structure prediction, we developed a number of novel methods and integrated them into a software package, MUFOLD. First, a systematic protocol was developed to identify useful templates and fragments from Protein Data Bank for a given target protein. Then, an efficient process was applied for iterative coarse-grain model generation and evaluation at the Calpha or backbone level. In this process, we construct models using interresidue spatial restraints derived from alignments by multidimensional scaling, evaluate and select models through clustering and static scoring functions, and iteratively improve the selected models by integrating spatial restraints and previous models. Finally, the full-atom models were evaluated using molecular dynamics simulations based on structural changes under simulated heating. We have continuously improved the performance of MUFOLD by using a benchmark of 200 proteins from the Astral database, where no template with >25% sequence identity to any target protein is included. The average root-mean-square deviation of the best models from the native structures is 4.28 A, which shows significant and systematic improvement over our previous methods. The computing time of MUFOLD is much shorter than many other tools, such as Rosetta. MUFOLD demonstrated some success in the 2008 community-wide experiment for protein structure prediction CASP8.

  4. ProSAT+: visualizing sequence annotations on 3D structure.

    PubMed

    Stank, Antonia; Richter, Stefan; Wade, Rebecca C

    2016-08-01

    PRO: tein S: tructure A: nnotation T: ool-plus (ProSAT(+)) is a new web server for mapping protein sequence annotations onto a protein structure and visualizing them simultaneously with the structure. ProSAT(+) incorporates many of the features of the preceding ProSAT and ProSAT2 tools but also provides new options for the visualization and sharing of protein annotations. Data are extracted from the UniProt KnowledgeBase, the RCSB PDB and the PDBe SIFTS resource, and visualization is performed using JSmol. User-defined sequence annotations can be added directly to the URL, thus enabling visualization and easy data sharing. ProSAT(+) is available at http://prosat.h-its.org.

  5. Lunar Polar Illumination for Power Analysis

    NASA Technical Reports Server (NTRS)

    Fincannon, James

    2008-01-01

    This paper presents illumination analyses using the latest Earth-based radar digital elevation model (DEM) of the lunar south pole and an independently developed analytical tool. These results enable the optimum sizing of solar/energy storage lunar surface power systems since they quantify the timing and durations of illuminated and shadowed periods. Filtering and manual editing of the DEM based on comparisons with independent imagery were performed and a reduced resolution version of the DEM was produced to reduce the analysis time. A comparison of the DEM with lunar limb imagery was performed in order to validate the absolute heights over the polar latitude range, the accuracy of which affects the impact of long range, shadow-casting terrain. Average illumination and energy storage duration maps of the south pole region are provided for the worst and best case lunar day using the reduced resolution DEM. Average illumination fractions and energy storage durations are presented for candidate low energy storage duration south pole sites. The best site identified using the reduced resolution DEM required a 62 hr energy storage duration using a fast recharge power system. Solar and horizon terrain elevations as well as illumination fraction profiles are presented for the best identified site and the data for both the reduced resolution and high resolution DEMs compared. High resolution maps for three low energy storage duration areas are presented showing energy storage duration for the worst case lunar day, surface height, and maximum absolute surface slope.

  6. Strong confinement of two-photon excitation field by photonic nanojet with radial polarization illumination

    NASA Astrophysics Data System (ADS)

    Liu, Y.; Kuang, C. F.; Ding, Z. H.

    2011-09-01

    The fluorescence excitation field by photonic nanojet has been investigated, where a SiO 2 microsphere is illuminated by the radial polarized beam. We show that radial polarization illumination can achieve very strong three-dimensional confinement of photonic nanojet below the diffraction limit, especially the longitudinal light field. It is also noted that the location of maximum excitation field intensity spot is not almost change by wavelength, but the volume of excitation field is depended on wavelength. Compared to single-photon excitation, two-photon excitation based on photonic nanojet can obtain the least illumination volume, with lateral FWHM ≈ λ/4 and axial FWHM < λ/10. This offers a broad range of application in single-molecule detection, ultra-resolution microscopy and nanopatterning.

  7. 30 CFR 57.17001 - Illumination of surface working areas.

    Code of Federal Regulations, 2011 CFR

    2011-07-01

    ... 30 Mineral Resources 1 2011-07-01 2011-07-01 false Illumination of surface working areas. 57.17001... Illumination § 57.17001 Illumination of surface working areas. Illumination sufficient to provide safe working conditions shall be provided in and on all surface structures, paths, walkways, stairways, switch...

  8. 10 CFR 431.202 - Definitions concerning illuminated exit signs.

    Code of Federal Regulations, 2010 CFR

    2010-01-01

    ... 10 Energy 3 2010-01-01 2010-01-01 false Definitions concerning illuminated exit signs. 431.202... COMMERCIAL AND INDUSTRIAL EQUIPMENT Illuminated Exit Signs § 431.202 Definitions concerning illuminated exit signs. Basic model means, with respect to illuminated exit signs, all units of a given type...

  9. 10 CFR 431.202 - Definitions concerning illuminated exit signs.

    Code of Federal Regulations, 2011 CFR

    2011-01-01

    ... 10 Energy 3 2011-01-01 2011-01-01 false Definitions concerning illuminated exit signs. 431.202... COMMERCIAL AND INDUSTRIAL EQUIPMENT Illuminated Exit Signs § 431.202 Definitions concerning illuminated exit signs. Basic model means, with respect to illuminated exit signs, all units of a given type...

  10. 30 CFR 57.17001 - Illumination of surface working areas.

    Code of Federal Regulations, 2010 CFR

    2010-07-01

    ... 30 Mineral Resources 1 2010-07-01 2010-07-01 false Illumination of surface working areas. 57.17001... Illumination § 57.17001 Illumination of surface working areas. Illumination sufficient to provide safe working conditions shall be provided in and on all surface structures, paths, walkways, stairways, switch...

  11. 30 CFR 56.17001 - Illumination of surface working areas.

    Code of Federal Regulations, 2010 CFR

    2010-07-01

    ... 30 Mineral Resources 1 2010-07-01 2010-07-01 false Illumination of surface working areas. 56.17001... NONMETAL MINE SAFETY AND HEALTH SAFETY AND HEALTH STANDARDS-SURFACE METAL AND NONMETAL MINES Illumination § 56.17001 Illumination of surface working areas. Illumination sufficient to provide safe...

  12. Content adaptive illumination for Fourier ptychography.

    PubMed

    Bian, Liheng; Suo, Jinli; Situ, Guohai; Zheng, Guoan; Chen, Feng; Dai, Qionghai

    2014-12-01

    Fourier ptychography (FP) is a recently reported technique, for large field-of-view and high-resolution imaging. Specifically, FP captures a set of low-resolution images, under angularly varying illuminations, and stitches them together in the Fourier domain. One of FP's main disadvantages is its long capturing process, due to the requisite large number of incident illumination angles. In this Letter, utilizing the sparsity of natural images in the Fourier domain, we propose a highly efficient method, termed adaptive Fourier ptychography (AFP), which applies content adaptive illumination for FP, to capture the most informative parts of the scene's spatial spectrum. We validate the effectiveness and efficiency of the reported framework, with both simulated and real experiments. Results show that the proposed AFP could shorten the acquisition time of conventional FP, by around 30%-60%.

  13. Contrast edge colors under different natural illuminations.

    PubMed

    Nieves, Juan Luis; Nascimento, Sérgio M C; Romero, Javier

    2012-02-01

    Essential to sensory processing in the human visual system is natural illumination, which can vary considerably not only across space but also along the day depending on the atmospheric conditions and the sun's position in the sky. In this work, edges derived from the three postreceptoral Luminance, Red-Green, and Blue-Yellow signals were computed from hyperspectral images of natural scenes rendered with daylights of Correlated Color Temperatures (CCTs) from 2735 to 25,889 K; for low CCT, the same analysis was performed using Planckian illuminants up to 800 K. It was found that average luminance and chromatic edge contrasts were maximal for low correlated color temperatures and almost constants above 10,000 K. The magnitude of these contrast changes was, however, only about 2% across the tested daylights. Results suggest that the postreceptoral opponent and nonopponent color vision mechanisms produce almost constant responses for color edge detection under natural illumination.

  14. Quantum Estimation Methods for Quantum Illumination

    NASA Astrophysics Data System (ADS)

    Sanz, M.; Las Heras, U.; García-Ripoll, J. J.; Solano, E.; Di Candia, R.

    2017-02-01

    Quantum illumination consists in shining quantum light on a target region immersed in a bright thermal bath with the aim of detecting the presence of a possible low-reflective object. If the signal is entangled with the receiver, then a suitable choice of the measurement offers a gain with respect to the optimal classical protocol employing coherent states. Here, we tackle this detection problem by using quantum estimation techniques to measure the reflectivity parameter of the object, showing an enhancement in the signal-to-noise ratio up to 3 dB with respect to the classical case when implementing only local measurements. Our approach employs the quantum Fisher information to provide an upper bound for the error probability, supplies the concrete estimator saturating the bound, and extends the quantum illumination protocol to non-Gaussian states. As an example, we show how Schrödinger's cat states may be used for quantum illumination.

  15. Quantum Estimation Methods for Quantum Illumination.

    PubMed

    Sanz, M; Las Heras, U; García-Ripoll, J J; Solano, E; Di Candia, R

    2017-02-17

    Quantum illumination consists in shining quantum light on a target region immersed in a bright thermal bath with the aim of detecting the presence of a possible low-reflective object. If the signal is entangled with the receiver, then a suitable choice of the measurement offers a gain with respect to the optimal classical protocol employing coherent states. Here, we tackle this detection problem by using quantum estimation techniques to measure the reflectivity parameter of the object, showing an enhancement in the signal-to-noise ratio up to 3 dB with respect to the classical case when implementing only local measurements. Our approach employs the quantum Fisher information to provide an upper bound for the error probability, supplies the concrete estimator saturating the bound, and extends the quantum illumination protocol to non-Gaussian states. As an example, we show how Schrödinger's cat states may be used for quantum illumination.

  16. Condenser-free contrast methods for transmitted-light microscopy

    PubMed Central

    WEBB, K F

    2015-01-01

    Phase contrast microscopy allows the study of highly transparent yet detail-rich specimens by producing intensity contrast from phase objects within the sample. Presented here is a generalized phase contrast illumination schema in which condenser optics are entirely abrogated, yielding a condenser-free yet highly effective method of obtaining phase contrast in transmitted-light microscopy. A ring of light emitting diodes (LEDs) is positioned within the light-path such that observation of the objective back focal plane places the illuminating ring in appropriate conjunction with the phase ring. It is demonstrated that true Zernike phase contrast is obtained, whose geometry can be flexibly manipulated to provide an arbitrary working distance between illuminator and sample. Condenser-free phase contrast is demonstrated across a range of magnifications (4–100×), numerical apertures (0.13–1.65NA) and conventional phase positions. Also demonstrated is condenser-free darkfield microscopy as well as combinatorial contrast including Rheinberg illumination and simultaneous, colour-contrasted, brightfield, darkfield and Zernike phase contrast. By providing enhanced and arbitrary working space above the preparation, a range of concurrent imaging and electrophysiological techniques will be technically facilitated. Condenser-free phase contrast is demonstrated in conjunction with scanning ion conductance microscopy (SICM), using a notched ring to admit the scanned probe. The compact, versatile LED illumination schema will further lend itself to novel next-generation transmitted-light microscopy designs. The condenser-free illumination method, using rings of independent or radially-scanned emitters, may be exploited in future in other electromagnetic wavebands, including X-rays or the infrared. PMID:25226859

  17. Illumination-compensated non-contact imaging photoplethysmography via dual-mode temporally coded illumination

    NASA Astrophysics Data System (ADS)

    Amelard, Robert; Scharfenberger, Christian; Wong, Alexander; Clausi, David A.

    2015-03-01

    Non-contact camera-based imaging photoplethysmography (iPPG) is useful for measuring heart rate in conditions where contact devices are problematic due to issues such as mobility, comfort, and sanitation. Existing iPPG methods analyse the light-tissue interaction of either active or passive (ambient) illumination. Many active iPPG methods assume the incident ambient light is negligible to the active illumination, resulting in high power requirements, while many passive iPPG methods assume near-constant ambient conditions. These assumptions can only be achieved in environments with controlled illumination and thus constrain the use of such devices. To increase the number of possible applications of iPPG devices, we propose a dual-mode active iPPG system that is robust to changes in ambient illumination variations. Our system uses a temporally-coded illumination sequence that is synchronized with the camera to measure both active and ambient illumination interaction for determining heart rate. By subtracting the ambient contribution, the remaining illumination data can be attributed to the controlled illuminant. Our device comprises a camera and an LED illuminant controlled by a microcontroller. The microcontroller drives the temporal code via synchronizing the frame captures and illumination time at the hardware level. By simulating changes in ambient light conditions, experimental results show our device is able to assess heart rate accurately in challenging lighting conditions. By varying the temporal code, we demonstrate the trade-off between camera frame rate and ambient light compensation for optimal blood pulse detection.

  18. Super-resolution spinning-disk confocal microscopy using optical photon reassignment.

    PubMed

    Azuma, Takuya; Kei, Takayuki

    2015-06-01

    Spinning-disk confocal microscopy is a proven technology for investigating 3D structures of biological specimens. Here we report a super-resolution method based on spinning-disk confocal microscopy that optically improves lateral resolution by a factor of 1.37 with a single exposure. Moreover, deconvolution yields twofold improvement over the diffraction limit. With the help of newly modified Nipkow disk which comprises pinholes and micro-lenses on the front and back respectively, emitted photons from specimen can be optically reassigned to the most probable locations they originate from. Consequently, the improvement in resolution is achieved preserving inherent sectioning capabilities of confocal microscopy. This extremely simple implementation will enable reliable observations at super high resolution in biomedical routine research.

  19. Concentrated and piped sunlight for indoor illumination.

    PubMed

    Fraas, L M; Pyle, W R; Ryason, P R

    1983-02-15

    A concept for indoor illumination of buildings using sunlight is described. For this system, a tracking concentrator on the building roof follows the sun and focuses sunlight into a lightguide. A system of transparent lightguides distributes the sunlight to interior rooms. Recent advances in the transparency of acrylic plastic optical fibers suggest that acrylic lightguides could be successfully used for piping sunlight. The proposed system displaces electricity currently used for indoor lighting. It is argued that using sunlight directly for indoor illumination would be about twenty-five times more cost-effective than using sunlight to generate electricity with solar cells for powering electric lamps for indoor lighting.

  20. Organic light emitting devices for illumination

    DOEpatents

    Hack, Michael [Lambertville, NJ; Lu, Min-Hao Michael [Lawrenceville, NJ; Weaver, Michael S [Princeton, NJ

    2012-01-24

    An organic light emitting device an a method of obtaining illumination from such a device is provided. The device has a plurality of regions, each region having an organic emissive layer adapted to emit a different spectrum of light. The regions in combination emit light suitable for illumination purposes. The area of each region may be selected such that the device is more efficient than an otherwise equivalent device having regions of equal size. The regions may have an aspect ratio of at least about four. All parts of any given region may be driven at the same current.

  1. Resolution enhancement using simultaneous couple illumination

    NASA Astrophysics Data System (ADS)

    Hussain, Anwar; Martínez Fuentes, José Luis

    2016-10-01

    A super-resolution technique based on structured illumination created by a liquid crystal on silicon spatial light modulator (LCOS-SLM) is presented. Single and simultaneous pairs of tilted beams are generated to illuminate a target object. Resolution enhancement of an optical 4f system is demonstrated by using numerical simulations. The resulting intensity images are recorded at a charged couple device (CCD) and stored in the computer memory for further processing. One dimension enhancement can be performed with only 15 images. Two dimensional complete improvement requires 153 different images. The resolution of the optical system is extended three times compared to the band limited system.

  2. Fiber-type dosimeter with improved illuminator

    DOEpatents

    Fox, Richard J.

    1987-01-01

    A single-piece, molded plastic, Cassigrainian-type condenser arrangement is incorporated in a tubular-shaped personal pocket dosimeter of the type which combines an ionization chamber with an optically-read fiber electrometer to provide improved illumination of the electrometer fiber. The condenser routes incoming light from one end of the dosimeter tubular housing around a central axis charging pin assembly and focuses the light at low angles to the axis so that it falls within the acceptance angle of the electrometer fiber objective lens viewed through an eyepiece lens disposed in the opposite end of the dosimeter. This results in improved fiber illumination and fiber image contrast.

  3. Fiber-type dosimeter with improved illuminator

    DOEpatents

    Fox, R.J.

    1985-12-23

    A single-piece, molded plastic, Cassigrainian-type condenser arrangement is incorporated in a tubular-shaped personal pocket dosimeter of the type which combines an ionization chamber with an optically-read fiber electrometer to provide improved illumination of the electrometer fiber. The condenser routes incoming light from one end of the dosimeter tubular housing around a central axis charging pin assembly and focuses the light at low angles to the axis so that it falls within the acceptance angle of the electrometer fiber objective lens viewed through an eyepiece lens disposed in the opposite end of the dosimeter. This results in improved fiber illumination and fiber image contrast.

  4. Extending Whole Slide Imaging: Color Darkfield Internal Reflection Illumination (DIRI) for Biological Applications

    PubMed Central

    Namiki, Kana; Miyawaki, Atsushi; Ishikawa, Takuji

    2017-01-01

    Whole slide imaging (WSI) is a useful tool for multi-modal imaging, and in our work, we have often combined WSI with darkfield microscopy. However, traditional darkfield microscopy cannot use a single condenser to support high- and low-numerical-aperture objectives, which limits the modality of WSI. To overcome this limitation, we previously developed a darkfield internal reflection illumination (DIRI) microscope using white light-emitting diodes (LEDs). Although the developed DIRI is useful for biological applications, substantial problems remain to be resolved. In this study, we propose a novel illumination technique called color DIRI. The use of three-color LEDs dramatically improves the capability of the system, such that color DIRI (1) enables optimization of the illumination color; (2) can be combined with an oil objective lens; (3) can produce fluorescence excitation illumination; (4) can adjust the wavelength of light to avoid cell damage or reactions; and (5) can be used as a photostimulator. These results clearly illustrate that the proposed color DIRI can significantly extend WSI modalities for biological applications. PMID:28085892

  5. Extending Whole Slide Imaging: Color Darkfield Internal Reflection Illumination (DIRI) for Biological Applications.

    PubMed

    Kawano, Yoshihiro; Namiki, Kana; Miyawaki, Atsushi; Ishikawa, Takuji

    2017-01-01

    Whole slide imaging (WSI) is a useful tool for multi-modal imaging, and in our work, we have often combined WSI with darkfield microscopy. However, traditional darkfield microscopy cannot use a single condenser to support high- and low-numerical-aperture objectives, which limits the modality of WSI. To overcome this limitation, we previously developed a darkfield internal reflection illumination (DIRI) microscope using white light-emitting diodes (LEDs). Although the developed DIRI is useful for biological applications, substantial problems remain to be resolved. In this study, we propose a novel illumination technique called color DIRI. The use of three-color LEDs dramatically improves the capability of the system, such that color DIRI (1) enables optimization of the illumination color; (2) can be combined with an oil objective lens; (3) can produce fluorescence excitation illumination; (4) can adjust the wavelength of light to avoid cell damage or reactions; and (5) can be used as a photostimulator. These results clearly illustrate that the proposed color DIRI can significantly extend WSI modalities for biological applications.

  6. Quantum-enhanced microscopy with binary-outcome photon counting

    NASA Astrophysics Data System (ADS)

    Jin, G. R.; Yang, W.; Sun, C. P.

    2017-01-01

    Polarized light microscopy using path-entangled N -photon states (i.e., the N00N states) has been demonstrated to surpass the shot-noise limit at very low light illumination. However, the microscopy images suffer from divergence of phase sensitivity, which inevitably reduces the image quality. Here we show that due to experimental imperfections, such a singularity also takes place in the microscopy that uses twin-Fock states of light for illumination. We propose two schemes to completely eliminate this singularity: (i) locking the phase shift sensed by the beams at the optimal working point using a spatially dependent offset phase; (ii) a combination of two binary-outcome photon counting measurements, one with a fixed offset phase and the other without any offset phase. Our observations remain valid for any kind of binary-outcome measurement and may open the way for quantum-enhanced microscopy with high N photon states.

  7. Nonlinear Structured Illumination Using a Fluorescent Protein Activating at the Readout Wavelength

    PubMed Central

    Hou, Wenya; Kielhorn, Martin; Arai, Yoshiyuki; Nagai, Takeharu; Kessels, Michael M.; Qualmann, Britta; Heintzmann, Rainer

    2016-01-01

    Structured illumination microscopy (SIM) is a wide-field technique in fluorescence microscopy that provides fast data acquisition and two-fold resolution improvement beyond the Abbe limit. We observed a further resolution improvement using the nonlinear emission response of a fluorescent protein. We demonstrated a two-beam nonlinear structured illumination microscope by introducing only a minor change into the system used for linear SIM (LSIM). To achieve the required nonlinear dependence in nonlinear SIM (NL-SIM) we exploited the photoswitching of the recently introduced fluorophore Kohinoor. It is particularly suitable due to its positive contrast photoswitching characteristics. Contrary to other reversibly photoswitchable fluorescent proteins which only have high photostability in living cells, Kohinoor additionally showed little degradation in fixed cells over many switching cycles. PMID:27783656

  8. Extended-resolution structured illumination imaging of endocytic and cytoskeletal dynamics

    PubMed Central

    Li, Dong; Shao, Lin; Chen, Bi-Chang; Zhang, Xi; Zhang, Mingshu; Moses, Brian; Milkie, Daniel E.; Beach, Jordan R.; Hammer, John A.; Pasham, Mithun; Kirchhausen, Tomas; Baird, Michelle A.; Davidson, Michael W.; Xu, Pingyong; Betzig, Eric

    2015-01-01

    Super-resolution fluorescence microscopy is distinct among nanoscale imaging tools in its ability to image protein dynamics in living cells. Structured illumination microscopy (SIM) stands out in this regard because of its high speed and low illumination intensities, but typically offers only a twofold resolution gain. We extended the resolution of live-cell SIM through two approaches: ultrahigh numerical aperture SIM at 84-nanometer lateral resolution for more than 100 multicolor frames, and nonlinear SIM with patterned activation at 45- to 62-nanometer resolution for approximately 20 to 40 frames. We applied these approaches to image dynamics near the plasma membrane of spatially resolved assemblies of clathrin and caveolin, Rab5a in early endosomes, and a-actinin, often in relationship to cortical actin. In addition, we examined mitochondria, actin, and the Golgi apparatus dynamics in three dimensions. PMID:26315442

  9. Phase Aberrations in Diffraction Microscopy

    SciTech Connect

    Marchesini, S; Chapman, H N; Barty, A; Howells, M R; Spence, J H; Cui, C; Weierstall, U; Minor, A M

    2005-09-29

    In coherent X-ray diffraction microscopy the diffraction pattern generated by a sample illuminated with coherent x-rays is recorded, and a computer algorithm recovers the unmeasured phases to synthesize an image. By avoiding the use of a lens the resolution is limited, in principle, only by the largest scattering angles recorded. However, the imaging task is shifted from the experiment to the computer, and the algorithm's ability to recover meaningful images in the presence of noise and limited prior knowledge may produce aberrations in the reconstructed image. We analyze the low order aberrations produced by our phase retrieval algorithms. We present two methods to improve the accuracy and stability of reconstructions.

  10. Fused off-axis object illumination direct-to-digital holography with a plurality of illumination sources

    DOEpatents

    Price, Jeffery R.; Bingham, Philip R.

    2005-11-08

    Systems and methods are described for rapid acquisition of fused off-axis illumination direct-to-digital holography. A method of recording a plurality of off-axis object illuminated spatially heterodyne holograms, each of the off-axis object illuminated spatially heterodyne holograms including spatially heterodyne fringes for Fourier analysis, includes digitally recording, with a first illumination source of an interferometer, a first off-axis object illuminated spatially heterodyne hologram including spatially heterodyne fringes for Fourier analysis; and digitally recording, with a second illumination source of the interferometer, a second off-axis object illuminated spatially heterodyne hologram including spatially heterodyne fringes for Fourier analysis.

  11. 29 CFR 1918.92 - Illumination.

    Code of Federal Regulations, 2010 CFR

    2010-07-01

    ... operations, illumination for cargo transfer operations shall be of a minimum light intensity of five foot-candles (54 lux). Where work tasks require more light to be performed safely, supplemental lighting shall be used. (b) Intensity measurement. The lighting intensity shall be measured at the...

  12. 29 CFR 1918.92 - Illumination.

    Code of Federal Regulations, 2012 CFR

    2012-07-01

    ... operations, illumination for cargo transfer operations shall be of a minimum light intensity of five foot-candles (54 lux). Where work tasks require more light to be performed safely, supplemental lighting shall be used. (b) Intensity measurement. The lighting intensity shall be measured at the...

  13. 29 CFR 1918.92 - Illumination.

    Code of Federal Regulations, 2014 CFR

    2014-07-01

    ... operations, illumination for cargo transfer operations shall be of a minimum light intensity of five foot-candles (54 lux). Where work tasks require more light to be performed safely, supplemental lighting shall be used. (b) Intensity measurement. The lighting intensity shall be measured at the...

  14. 29 CFR 1918.92 - Illumination.

    Code of Federal Regulations, 2011 CFR

    2011-07-01

    ... operations, illumination for cargo transfer operations shall be of a minimum light intensity of five foot-candles (54 lux). Where work tasks require more light to be performed safely, supplemental lighting shall be used. (b) Intensity measurement. The lighting intensity shall be measured at the...

  15. Adaptive Illumination Patterns for Radar Applications

    DTIC Science & Technology

    2006-03-01

    Losses . . . . . . . . . . . . . . . . . . . . . . . . . 44 Ri Range to the i th range ring . . . . . . . . . . . . . . . . . 44 R Signal-Dependent...Interval . . . . . . . . . . . . . . . . . . . 23 R /H Range-to-Height Ratio . . . . . . . . . . . . . . . . . . . . 51 RMB Reed Mallet and Brennan Rule...as follows: 16 AIP-TIPD Planar Array Radar Model Extensions I - I l r rr r l t i Space Time Illumination Patterns (STIP) i Ill i ti tt r ( I

  16. Lighting design for globally illuminated volume rendering.

    PubMed

    Zhang, Yubo; Ma, Kwan-Liu

    2013-12-01

    With the evolution of graphics hardware, high quality global illumination becomes available for real-time volume rendering. Compared to local illumination, global illumination can produce realistic shading effects which are closer to real world scenes, and has proven useful for enhancing volume data visualization to enable better depth and shape perception. However, setting up optimal lighting could be a nontrivial task for average users. There were lighting design works for volume visualization but they did not consider global light transportation. In this paper, we present a lighting design method for volume visualization employing global illumination. The resulting system takes into account view and transfer-function dependent content of the volume data to automatically generate an optimized three-point lighting environment. Our method fully exploits the back light which is not used by previous volume visualization systems. By also including global shadow and multiple scattering, our lighting system can effectively enhance the depth and shape perception of volumetric features of interest. In addition, we propose an automatic tone mapping operator which recovers visual details from overexposed areas while maintaining sufficient contrast in the dark areas. We show that our method is effective for visualizing volume datasets with complex structures. The structural information is more clearly and correctly presented under the automatically generated light sources.

  17. Diffuse-Illumination Systems for Growing Plants

    NASA Technical Reports Server (NTRS)

    May, George; Ryan, Robert

    2010-01-01

    Agriculture in both terrestrial and space-controlled environments relies heavily on artificial illumination for efficient photosynthesis. Plant-growth illumination systems require high photon flux in the spectral range corresponding with plant photosynthetic active radiation (PAR) (400 700 nm), high spatial uniformity to promote uniform growth, and high energy efficiency to minimize electricity usage. The proposed plant-growth system takes advantage of the highly diffuse reflective surfaces on the interior of a sphere, hemisphere, or other nearly enclosed structure that is coated with highly reflective materials. This type of surface and structure uniformly mixes discrete light sources to produce highly uniform illumination. Multiple reflections from within the domelike structures are exploited to obtain diffuse illumination, which promotes the efficient reuse of photons that have not yet been absorbed by plants. The highly reflective surfaces encourage only the plant tissue (placed inside the sphere or enclosure) to absorb the light. Discrete light sources, such as light emitting diodes (LEDs), are typically used because of their high efficiency, wavelength selection, and electronically dimmable properties. The light sources are arranged to minimize shadowing and to improve uniformity. Different wavelengths of LEDs (typically blue, green, and red) are used for photosynthesis. Wavelengths outside the PAR range can be added for plant diagnostics or for growth regulation

  18. 29 CFR 1926.56 - Illumination.

    Code of Federal Regulations, 2010 CFR

    2010-07-01

    ... Illumination. (a) General. Construction areas, ramps, runways, corridors, offices, shops, and storage areas... tunnel heading.) 10 General construction plant and shops (e.g., batch plants, screening plants, mechanical and electrical equipment rooms, carpenter shops, rigging lofts and active storerooms, barracks...

  19. Freeform LED lens for rectangularly prescribed illumination

    NASA Astrophysics Data System (ADS)

    Wang, Kai; Liu, Sheng; Chen, Fei; Qin, Zong; Liu, Zongyuan; Luo, Xiaobing

    2009-10-01

    Freeform lenses are playing a more and more important role in LED secondary optics design. In this study, based on the new light energy mapping relationship, edge ray principle, Snell's law and error control of surface construction, a modified discontinuous freeform lens design method was presented for rectangularly prescribed illumination, with the advantages of a flexible energy mapping relationship, accurate light irradiation control and easier to manufacture. A polymethyl methacrylate (PMMA) discontinuous freeform lens was designed as an example for LED tunnel illumination according to this method. The numerical simulation results demonstrated that the light pattern of the lens was in good agreement with the expected illumination performance when using a point source. Tolerance analyses were also conducted. An extended light source had little effect on the light output efficiency (LOE) of the lens but significantly decreased the effective illumination area. Installation errors had more effect on the uniformity and shape of the light pattern than the LOE of the lens. The tolerances of vertical, horizontal and rotational deviation of this lens were 0.4 mm, 0.4 mm and 2°, respectively.

  20. Multi-Perspective Illumination: A Case Study.

    ERIC Educational Resources Information Center

    Melton, R. F.; Zimmer, R. S.

    1987-01-01

    This description of an illuminative evaluation process as a qualitative research method focuses on a study of concerns about financial constraints in the British Open University. Interviews and open-ended discussions were used to encourage participation and input by a wide variety of individuals with differing perspectives on the problem. (LRW)

  1. High Resolution Quantitative Angle-Scanning Widefield Surface Plasmon Microscopy

    PubMed Central

    Tan, Han-Min; Pechprasarn, Suejit; Zhang, Jing; Pitter, Mark C.; Somekh, Michael G.

    2016-01-01

    We describe the construction of a prismless widefield surface plasmon microscope; this has been applied to imaging of the interactions of protein and antibodies in aqueous media. The illumination angle of spatially incoherent diffuse laser illumination was controlled with an amplitude spatial light modulator placed in a conjugate back focal plane to allow dynamic control of the illumination angle. Quantitative surface plasmon microscopy images with high spatial resolution were acquired by post-processing a series of images obtained as a function of illumination angle. Experimental results are presented showing spatially and temporally resolved binding of a protein to a ligand. We also show theoretical results calculated by vector diffraction theory that accurately predict the response of the microscope on a spatially varying sample thus allowing proper quantification and interpretation of the experimental results. PMID:26830146

  2. Superresolution imaging with optical fluctuation using speckle patterns illumination

    PubMed Central

    Kim, MinKwan; Park, ChungHyun; Rodriguez, Christophe; Park, YongKeun; Cho, Yong-Hoon

    2015-01-01

    Superresolution fluorescence microscopy possesses an important role for the study of processes in biological cells with subdiffraction resolution. Recently, superresolution methods employing the emission properties of fluorophores have rapidly evolved due to their technical simplicity and direct applicability to existing microscopes. However, the application of these methods has been limited to samples labeled with fluorophores that can exhibit intrinsic emission properties at a restricted timescale, especially stochastic blinking. Here, we present a superresolution method that can be performed using general fluorophores, regardless of this intrinsic property. Utilizing speckle patterns illumination, temporal emission fluctuation of fluorophores is induced and controlled, from which a superresolution image can be obtained exploiting its statistical property. Using this method, we demonstrate, theoretically and experimentally, the capability to produce subdiffraction resolution images. A spatial resolution of 500 nm, 300 nm and 140 nm with 0.4, 0.5 and 1.4 NA objective lenses respectively was achieved in various samples with an enhancement factor of 1.6 compared to conventional fluorescence microscopy. PMID:26572283

  3. Image simulation for biological microscopy: microlith

    PubMed Central

    Mehta, Shalin B.; Oldenbourg, Rudolf

    2014-01-01

    Image simulation remains under-exploited for the most widely used biological phase microscopy methods, because of difficulties in simulating partially coherent illumination. We describe an open-source toolbox, microlith (https://code.google.com/p/microlith), which accurately predicts three-dimensional images of a thin specimen observed with any partially coherent imaging system, as well as images of coherently illuminated and self-luminous incoherent specimens. Its accuracy is demonstrated by comparing simulated and experimental bright-field and dark-field images of well-characterized amplitude and phase targets, respectively. The comparison provides new insights about the sensitivity of the dark-field microscope to mass distributions in isolated or periodic specimens at the length-scale of 10nm. Based on predictions using microlith, we propose a novel approach for detecting nanoscale structural changes in a beating axoneme using a dark-field microscope. PMID:24940543

  4. LED illuminator for a microdisplay projector

    NASA Astrophysics Data System (ADS)

    Magarill, Simon

    2012-10-01

    An illumination system for a microdisplay projector with a two-step imaging system is described here. In the first step, an imaging condenser creates an image of the LED at the color combiner entrance window. In the second step, we relay the image of the integrator exit window onto the micro-display. The illuminator demonstrates high collection efficiency, small footprint, and efficient mixing of light from RGB LEDs that provides required uniformity. A variety of approaches to collecting light emitted from LEDs of various types are compared, leading to the two-step design. A design example using a 0.55" diagonal DLP-based optical engine is presented with the following characteristics: Footprint: 3.9"x3.3"x2.0" (25.7 cubic inches) Light output: 338 white lumens Efficiency: 4.7 lm/watt

  5. Pulsed laser illumination of photovoltaic cells

    NASA Technical Reports Server (NTRS)

    Yater, Jane A.; Lowe, Roland A.; Jenkins, Phillip P.; Landis, Geoffrey A.

    1994-01-01

    In future space missions, free electron lasers (FEL) may be used to illuminate photovoltaic array receivers to provide remote power. Both the radio-frequency (RF) and induction FEL provide FEL produce pulsed rather than continuous output. In this work we investigate cell response to pulsed laser light which simulates the RF FEL format. The results indicate that if the pulse repetition is high, cell efficiencies are only slightly reduced compared to constant illumination at the same wavelength. The frequency response of the cells is weak, with both voltage and current outputs essentially dc in nature. Comparison with previous experiments indicates that the RF FEL pulse format yields more efficient photovoltaic conversion than does an induction FEL pulse format.

  6. Pulsed laser illumination of photovoltaic cells

    NASA Technical Reports Server (NTRS)

    Yater, Jane A.; Lowe, Roland A.; Jenkins, Phillip P.; Landis, Geoffrey A.

    1995-01-01

    In future space missions, free electron lasers (FEL) may be used to illuminate photovoltaic receivers to provide remote power. Both the radio-frequency (RF) and induction FEL produce pulsed rather than continuous output. In this work we investigate cell response to pulsed laser light which simulates the RF FEL format. The results indicate that if the pulse repetition is high, cell efficiencies are only slightly reduced compared to constant illumination at the same wavelength. The frequency response of the cells is weak, with both voltage and current outputs essentially dc in nature. Comparison with previous experiments indicates that the RF FEL pulse format yields more efficient photovoltaic conversion than does an induction FEL format.

  7. Microscope illumination systems for 157 nm

    NASA Astrophysics Data System (ADS)

    Pesch, Alexander; Uhlendorf, Kristina; Deparnay, Arnaud; Erdmann, Lars; Kuschnerus, Peter; Engel, Thomas; Brunner, Robert

    2003-05-01

    The image quality of an inspection microscope depends strongly on the performance of the illumination system. Especially in the case of laser-based illumination it is necessary to transform the original beam profile into a homogeneous light spot with a flat top field distribution. Simultaneously, speckles caused by the coherence of the laser have to be reduced. Here we discuss different ways to homogenize the multi mode beam profile of a pulsed compact 157 nm excimer laser. A variety of setups, combining dynamic acting diffusers, microlens arrays and primary lenses were realized and characterized in several geometrical arrangements. The homogenizers were evaluated and characterized especially with respect to the statistical behavior on the integrated pulse number.

  8. Insulator Surface Flashover Due to UV Illumination

    SciTech Connect

    Javedani, J B; Houck, T L; Lahowe, D A; Vogtlin, G E; Goerz, D A

    2009-07-27

    The surface of an insulator under vacuum and under electrical charge will flashover when illuminated by a critical dose of ultra-violet (UV) radiation - depending on the insulator size and material, insulator cone angle, the applied voltage and insulator shot-history. A testbed comprised of an excimer laser (KrF, 248 nm, {approx}16 MW, 30 ns FWHM,), a vacuum chamber, and a negative polarity dc high voltage power supply ({le} -60 kV) were assembled to test 1.0 cm thick angled insulators for surface-flashover. Several candidate insulator materials, e.g. High Density Polyethylene (HDPE), Rexolite{reg_sign} 1400, Macor{trademark} and Mycalex, of varying cone angles were tested against UV illumination. Commercial energy meters were used to measure the UV fluence of the pulsed laser beam. In-house designed and fabricated capacitive probes (D-dots, >12 GHz bandwidth) were embedded in the anode electrode underneath the insulator to determine the time of UV arrival and time of flashover. Of the tested insulators, the +45 degree Rexolite insulator showed more resistance to UV for surface flashover; at UV fluence level of less than 13 mJ/cm{sup 2}, it was not possible to induce a flashover for up to -60 kV of DC potential across the insulator's surface. The probes also permitted the electrical charge on the insulator before and after flashover to be inferred. Photon to electron conversion efficiency for the surface of Rexolite insulator was determined from charge-balance equation. In order to understand the physical mechanism leading to flashover, we further experimented with the +45 degree Rexolite insulator by masking portions of the UV beam to illuminate only a section of the insulator surface; (1) the half nearest the cathode and subsequently, (2) the half nearest the anode. The critical UV fluence and time to flashover were measured and the results in each case were then compared with the base case of full-beam illumination. It was discovered that the time for the

  9. Compact laser illumination system for endoscopic interventions.

    PubMed

    Blase, Bastian

    2015-08-01

    External cold light sources as well as LEDs are commonly used for abdominal illumination in minimally invasive surgery. Still, both feature certain disadvantages. A new illumination system for endoscopes based on laser diodes is placed in the handle. No external light cables are needed. High conversion and coupling efficiencies and small package size allow for several diodes to be integrated, enabling color mixing and the adjustment of color temperatures. An optical module to collimate and combine the light is described. The heat to be dissipated is stored in a passive latent heat storage based on phase change materials surrounding the optical module. Thereby, operation time is considerably extended, as the handle's temperature is stabilized. To reduce the negative effect of coherent light on optical rough surfaces leading to patterns of spots, several devices for speckle reduction are developed and tested. By combining these components, an assembly of a powerful RGB laser light module for the integration in standard sized endoscopes is formed.

  10. Improving the opto-microwave performance of SiGe/Si phototransistor through edge-illuminated structure

    NASA Astrophysics Data System (ADS)

    Tegegne, Z. G.; Viana, C.; Polleux, J. L.; Grzeskowiak, M.; Richalot, E.

    2016-03-01

    This paper demonstrates the experimental study of edge and top illuminated SiGe phototransistors (HPT) implemented using the existing industrial SiGe2RF Telefunken GmbH BiCMOS technology for opto-microwave (OM) applications using 850nm Multi-Mode Fibers (MMF). Its technology and structure are described. Two different optical window size HPTs with top illumination (5x5μm2, 10x10μm2) and an edge illuminated HPTs having 5μm x5μm size are presented and compared. A two-step post fabrication process was used to create an optical access on the edge of the HPT for lateral illumination with a lensed MMF through simple polishing and dicing techniques. We perform Opto-microwave Scanning Near-field Optical Microscopy (OM-SNOM) analysis on edge and top illuminated HPTs in order to observe the fastest and the highest sensitive regions of the HPTs. This analysis also allows understanding the parasitic effect from the substrate, and thus draws a conclusion on the design aspect of SiGe/Si HPT. A low frequency OM responsivity of 0.45A/W and a cutoff frequency, f-3dB, of 890MHz were measured for edge illuminated HPT. Compared to the top illuminated HPT of the same size, the edge illuminated HPT improves the f-3dB by a factor of more than two and also improves the low frequency responsivity by a factor of more than four. These results demonstrate that a simple etched HPT is still enough to achieve performance improvements compared to the top illuminated HPT without requiring a complex coupling structure. Indeed, it also proves the potential of edge coupled SiGe HPT in the ultra-low-cost silicon based optoelectronics circuits with a new approach of the optical packaging and system integration to 850nm MMF.

  11. Alternative Packaging for Back-Illuminated Imagers

    NASA Technical Reports Server (NTRS)

    Pain, Bedabrata

    2009-01-01

    An alternative scheme has been conceived for packaging of silicon-based back-illuminated, back-side-thinned complementary metal oxide/semiconductor (CMOS) and charge-coupled-device image-detector integrated circuits, including an associated fabrication process. This scheme and process are complementary to those described in "Making a Back-Illuminated Imager With Back-Side Connections" (NPO-42839), NASA Tech Briefs, Vol. 32, No. 7 (July 2008), page 38. To avoid misunderstanding, it should be noted that in the terminology of imaging integrated circuits, "front side" or "back side" does not necessarily refer to the side that, during operation, faces toward or away from a source of light or other object to be imaged. Instead, "front side" signifies that side of a semiconductor substrate upon which the pixel pattern and the associated semiconductor devices and metal conductor lines are initially formed during fabrication, and "back side" signifies the opposite side. If the imager is of the type called "back-illuminated," then the back side is the one that faces an object to be imaged. Initially, a back-illuminated, back-side-thinned image-detector is fabricated with its back side bonded to a silicon handle wafer. At a subsequent stage of fabrication, the front side is bonded to a glass wafer (for mechanical support) and the silicon handle wafer is etched away to expose the back side. The frontside integrated circuitry includes metal input/output contact pads, which are rendered inaccessible by the bonding of the front side to the glass wafer. Hence, one of the main problems is to make the input/output contact pads accessible from the back side, which is ultimately to be the side accessible to the external world. The present combination of an alternative packaging scheme and associated fabrication process constitute a solution of the problem.

  12. Reinterpretation and improvement of Talbot array illuminators.

    PubMed

    Arrizón, V; Rojo-Velázquez, G

    2000-09-10

    We show that the transmittance of a finite Talbot array illuminator (TAI) can be expressed by the phase distribution of a pixelated lens, modulated by a discrete phase grating (G). Thus the TAI reconstruction field is given by the convolution of the grating's Fourier transform, with the point-spread function of the pixelated lens. On the basis of this approach we propose a method to improve the performance of a finite TAI by modifying the basic cell of the grating factor G.

  13. Moonbase night power by laser illumination

    NASA Technical Reports Server (NTRS)

    Landis, Geoffrey A.

    1992-01-01

    Moonbase solar-power concepts must somehow address the energy storage problem posed by the 354-hour lunar night. Attention is presently given to the feasibility of laser-array illumination of a lunar base, using technology that is projected to be available in the near term. Beam-spreading due to atmospheric distortions could be reduced through the use of adaptive optics to compensate for atmospheric turbulence.

  14. Shape, Illumination, and Reflectance from Shading

    DTIC Science & Technology

    2013-05-29

    Shape, Illumination, and Reflectance from Shading Jonathan Barron Jitendra Malik Electrical Engineering and Computer Sciences University of...S) AND ADDRESS(ES) University of California at Berkeley, Electrical Engineering and Computer Sciences,Berkeley,CA,94720 8. PERFORMING ORGANIZATION...images [2], [3]. We will construct priors similar to those used � J.T. Barron and J. Malik are with the Department of Electrical Engi- neering and

  15. Illumination analysis of LAPAN's IR micro bolometer

    NASA Astrophysics Data System (ADS)

    Bustanul, A.; Irwan, P.; Andi M., T.

    2016-10-01

    We have since 2 years ago been doing a research in term of an IR Micrometer Bolometer which aims to fulfill our office, LAPAN, desire to put it as one of payloads into LAPAN's next micro satellite project, either at LAPAN A4 or at LAPAN A5. Due to the lack of experience on the subject, everything had been initiated by spectral radiance analysis adjusted by catastrophes sources in Indonesia, mainly wild fire (forest fire) and active volcano. Based on the result of the appropriate spectral radiance wavelength, 3.8 - 4 μm, and field of view (FOV), we, then, went through the further analysis, optical analysis. Focusing in illumination matter, the process was done by using Zemax software. Optical pass Interference and Stray light were two things that become our concern throughout the work. They could also be an evaluation of the performance optimization of illumination analysis of our optical design. The results, graphs, show that our design performance is close diffraction limited and the image blur of the geometrical produced by Lapan's IR Micro Bolometer lenses is in the pixel area range. Therefore, our optical design performance is relatively good and will produce image with high quality. In this paper, the Illumination analysis and process of LAPAN's Infra Red (IR) Micro Bolometer is presented.

  16. Optical mapping at increased illumination intensities

    NASA Astrophysics Data System (ADS)

    Kanaporis, Giedrius; Martišienė, Irma; Jurevičius, Jonas; Vosyliūtė, Rūta; Navalinskas, Antanas; Treinys, Rimantas; Matiukas, Arvydas; Pertsov, Arkady M.

    2012-09-01

    Voltage-sensitive fluorescent dyes have become a major tool in cardiac and neuro-electrophysiology. Achieving high signal-to-noise ratios requires increased illumination intensities, which may cause photobleaching and phototoxicity. The optimal range of illumination intensities varies for different dyes and must be evaluated individually. We evaluate two dyes: di-4-ANBDQBS (excitation 660 nm) and di-4-ANEPPS (excitation 532 nm) in the guinea pig heart. The light intensity varies from 0.1 to 5 mW/mm2, with the upper limit at 5 to 10 times above values reported in the literature. The duration of illumination was 60 s, which in guinea pigs corresponds to 300 beats at a normal heart rate. Within the identified duration and intensity range, neither dye shows significant photobleaching or detectable phototoxic effects. However, light absorption at higher intensities causes noticeable tissue heating, which affects the electrophysiological parameters. The most pronounced effect is a shortening of the action potential duration, which, in the case of 532-nm excitation, can reach ˜30%. At 660-nm excitation, the effect is ˜10%. These findings may have important implications for the design of optical mapping protocols in biomedical applications.

  17. Pulsed Laser Illumination of Photovoltaic Cells

    NASA Technical Reports Server (NTRS)

    Yater, Jane A.; Lowe, Roland; Jenkins, Philip; Landis, Geoffrey A.

    1994-01-01

    In future space missions, free electron lasers (FEL) may be used to illuminate photovoltaic array receivers to provide remote power. The induction FEL and the radio-frequency (RF) FEL both produce pulsed rather than continuous output. In this work, we investigate cell response to pulsed laser light which simulates the RF FEL format, producing 50 ps pulses at a frequency of 78 MHz. A variety of Si, GaAs, CaSb and CdInSe2 (CIS) solar cells are tested at average incident powers between 4 mW/sq cm and 425 mW/sq cm. The results indicate that if the pulse repetition is high, cell efficiencies are only slightly reduced by using a pulsed laser source compared to constant illumination at the same wavelength. Because the pulse separation is less than or approximately equal to the minority carrier lifetime, the illumination conditions are effectively those of a continuous wave laser. The time dependence of the voltage and current response of the cells are also measured using a sampling oscilloscope equipped with a high frequency voltage probe and current transformer. The frequency response of the cells is weak, with both voltage and current outputs essentially dc in nature. Comparison with previous experiments shows that the RF FEL pulse format yields much more efficient photovoltaic conversion of light than does an induction FEL pulse format.

  18. 47 CFR 80.969 - Illumination of operating controls.

    Code of Federal Regulations, 2011 CFR

    2011-10-01

    ... illuminate the operating controls at the principal operating position. (b) Instead of dial lights, a light from an electric lamp may be provided to illuminate the operating controls of the radiotelephone at...

  19. 47 CFR 80.969 - Illumination of operating controls.

    Code of Federal Regulations, 2010 CFR

    2010-10-01

    ... illuminate the operating controls at the principal operating position. (b) Instead of dial lights, a light from an electric lamp may be provided to illuminate the operating controls of the radiotelephone at...

  20. Super-Resolution Laser Scanning Microscopy through Spatiotemporal Modulation

    PubMed Central

    Lu, Ju; Min, Wei; Conchello, José-Angel; Xie, Xiaoliang Sunney; Lichtman, Jeff W.

    2009-01-01

    Super-resolution optical microscopy has attracted great interest among researchers in many fields, especially in biology where the scale of physical structures and molecular processes fall below the diffraction limit of resolution for light. As one of the emerging techniques, structured illumination microscopy can double the resolution by shifting unresolvable spatial frequencies into the pass-band of the microscope through spatial frequency mixing with a wide-field structured illumination pattern. However, such a wide-field scheme typically can only image optically thin samples and is incompatible with multiphoton processes such as two-photon fluorescence, which require point scanning with a focused laser beam. Here, we propose two new super-resolution schemes for laser scanning microscopy by generalizing the concept of a spatially nonuniform imaging system. One scheme, scanning patterned illumination (SPIN) microscopy, employs modulation of the excitation combined with temporally cumulative imaging by a nondescanned array detector. The other scheme, scanning patterned detection (SPADE) microscopy, utilizes detection modulation together with spatially cumulative imaging, in this case by a nondescanned single-element detector. When combined with multiphoton excitation, both schemes can image thick samples with three-dimensional optical sectioning and much improved resolution. PMID:19743870

  1. Cryo-electron microscopy and cryo-electron tomography of nanoparticles.

    PubMed

    Stewart, Phoebe L

    2017-03-01

    Cryo-transmission electron microscopy (cryo-TEM or cryo-EM) and cryo-electron tomography (cryo-ET) offer robust and powerful ways to visualize nanoparticles. These techniques involve imaging of the sample in a frozen-hydrated state, allowing visualization of nanoparticles essentially as they exist in solution. Cryo-TEM grid preparation can be performed with the sample in aqueous solvents or in various organic and ionic solvents. Two-dimensional (2D) cryo-TEM provides a direct way to visualize the polydispersity within a nanoparticle preparation. Fourier transforms of cryo-TEM images can confirm the structural periodicity within a sample. While measurement of specimen parameters can be performed with 2D TEM images, determination of a three-dimensional (3D) structure often facilitates more spatially accurate quantization. 3D structures can be determined in one of two ways. If the nanoparticle has a homogeneous structure, then 2D projection images of different particles can be averaged using a computational process referred to as single particle reconstruction. Alternatively, if the nanoparticle has a heterogeneous structure, then a structure can be generated by cryo-ET. This involves collecting a tilt-series of 2D projection images for a defined region of the grid, which can be used to generate a 3D tomogram. Occasionally it is advantageous to calculate both a single particle reconstruction, to reveal the regular portions of a nanoparticle structure, and a cryo-electron tomogram, to reveal the irregular features. A sampling of 2D cryo-TEM images and 3D structures are presented for protein based, DNA based, lipid based, and polymer based nanoparticles. WIREs Nanomed Nanobiotechnol 2017, 9:e1417. doi: 10.1002/wnan.1417 For further resources related to this article, please visit the WIREs website.

  2. Cellulose Nanocrystals as Chiral Inducers: Enantioselective Catalysis and Transmission Electron Microscopy 3D Characterization.

    PubMed

    Kaushik, Madhu; Basu, Kaustuv; Benoit, Charles; Cirtiu, Ciprian M; Vali, Hojatollah; Moores, Audrey

    2015-05-20

    Cellulose nanocrystals (CNCs), derived from cellulose, provide us with an opportunity to devise more sustainable solutions to current technological challenges. Enantioselective catalysis, especially heterogeneous, is the preferred method for the synthesis of pure chiral molecules in the fine chemical industries. Cellulose has been long sought as a chiral inducer in enantioselective catalysis. We report herein an unprecedentedly high enantiomeric excess (ee) for Pd patches deposited onto CNCs used as catalysts for the hydrogenation of prochiral ketones in water at room temperature and 4 bar H2. Our system, where CNCs acted as support and sole chiral source, achieved an ee of 65% with 100% conversions. Cryo-electron microscopy, high-resolution transmission electron microscopy, and tomography were used for the first time to study the 3D structure of a metal functionalized CNC hybrid. It established the presence of sub-nanometer-thick Pd patches at the surface of CNCs and provided insight into the chiral induction mechanism.

  3. 21 CFR 886.1160 - Color vision plate illuminator.

    Code of Federal Regulations, 2012 CFR

    2012-04-01

    ... 21 Food and Drugs 8 2012-04-01 2012-04-01 false Color vision plate illuminator. 886.1160 Section... (CONTINUED) MEDICAL DEVICES OPHTHALMIC DEVICES Diagnostic Devices § 886.1160 Color vision plate illuminator. (a) Identification. A color vision plate illuminator is an AC-powered device that is a lamp...

  4. 21 CFR 886.1160 - Color vision plate illuminator.

    Code of Federal Regulations, 2011 CFR

    2011-04-01

    ... 21 Food and Drugs 8 2011-04-01 2011-04-01 false Color vision plate illuminator. 886.1160 Section... (CONTINUED) MEDICAL DEVICES OPHTHALMIC DEVICES Diagnostic Devices § 886.1160 Color vision plate illuminator. (a) Identification. A color vision plate illuminator is an AC-powered device that is a lamp...

  5. 21 CFR 886.1160 - Color vision plate illuminator.

    Code of Federal Regulations, 2013 CFR

    2013-04-01

    ... 21 Food and Drugs 8 2013-04-01 2013-04-01 false Color vision plate illuminator. 886.1160 Section... (CONTINUED) MEDICAL DEVICES OPHTHALMIC DEVICES Diagnostic Devices § 886.1160 Color vision plate illuminator. (a) Identification. A color vision plate illuminator is an AC-powered device that is a lamp...

  6. 21 CFR 886.1160 - Color vision plate illuminator.

    Code of Federal Regulations, 2014 CFR

    2014-04-01

    ... 21 Food and Drugs 8 2014-04-01 2014-04-01 false Color vision plate illuminator. 886.1160 Section... (CONTINUED) MEDICAL DEVICES OPHTHALMIC DEVICES Diagnostic Devices § 886.1160 Color vision plate illuminator. (a) Identification. A color vision plate illuminator is an AC-powered device that is a lamp...

  7. 21 CFR 886.1160 - Color vision plate illuminator.

    Code of Federal Regulations, 2010 CFR

    2010-04-01

    ... 21 Food and Drugs 8 2010-04-01 2010-04-01 false Color vision plate illuminator. 886.1160 Section... (CONTINUED) MEDICAL DEVICES OPHTHALMIC DEVICES Diagnostic Devices § 886.1160 Color vision plate illuminator. (a) Identification. A color vision plate illuminator is an AC-powered device that is a lamp...

  8. 30 CFR 56.17001 - Illumination of surface working areas.

    Code of Federal Regulations, 2011 CFR

    2011-07-01

    ... 30 Mineral Resources 1 2011-07-01 2011-07-01 false Illumination of surface working areas. 56.17001... § 56.17001 Illumination of surface working areas. Illumination sufficient to provide safe working conditions shall be provided in and on all surface structures, paths, walkways, stairways, switch...

  9. 32 CFR 707.10 - Wake illumination light.

    Code of Federal Regulations, 2013 CFR

    2013-07-01

    ... 32 National Defense 5 2013-07-01 2013-07-01 false Wake illumination light. 707.10 Section 707.10... RESPECT TO ADDITIONAL STATION AND SIGNAL LIGHTS § 707.10 Wake illumination light. Naval vessels may display a white spot light located near the stern to illuminate the wake....

  10. 32 CFR 707.10 - Wake illumination light.

    Code of Federal Regulations, 2011 CFR

    2011-07-01

    ... 32 National Defense 5 2011-07-01 2011-07-01 false Wake illumination light. 707.10 Section 707.10... RESPECT TO ADDITIONAL STATION AND SIGNAL LIGHTS § 707.10 Wake illumination light. Naval vessels may display a white spot light located near the stern to illuminate the wake....

  11. 32 CFR 707.10 - Wake illumination light.

    Code of Federal Regulations, 2012 CFR

    2012-07-01

    ... 32 National Defense 5 2012-07-01 2012-07-01 false Wake illumination light. 707.10 Section 707.10... RESPECT TO ADDITIONAL STATION AND SIGNAL LIGHTS § 707.10 Wake illumination light. Naval vessels may display a white spot light located near the stern to illuminate the wake....

  12. 30 CFR 75.1719-1 - Illumination in working places.

    Code of Federal Regulations, 2011 CFR

    2011-07-01

    ... working place, the areas within a miner's normal field of vision which shall be illuminated in the working... are operated, the areas which shall be illuminated shall be as follows: (i) The face, and (ii) The... equipment is operated to load material, the areas which shall be illuminated shall be as follows: (i)...

  13. 30 CFR 75.1719-1 - Illumination in working places.

    Code of Federal Regulations, 2010 CFR

    2010-07-01

    ... working place, the areas within a miner's normal field of vision which shall be illuminated in the working... are operated, the areas which shall be illuminated shall be as follows: (i) The face, and (ii) The... equipment is operated to load material, the areas which shall be illuminated shall be as follows: (i)...

  14. 32 CFR 707.10 - Wake illumination light.

    Code of Federal Regulations, 2010 CFR

    2010-07-01

    ... 32 National Defense 5 2010-07-01 2010-07-01 false Wake illumination light. 707.10 Section 707.10... RESPECT TO ADDITIONAL STATION AND SIGNAL LIGHTS § 707.10 Wake illumination light. Naval vessels may display a white spot light located near the stern to illuminate the wake....

  15. 21 CFR 892.1890 - Radiographic film illuminator.

    Code of Federal Regulations, 2014 CFR

    2014-04-01

    ... 21 Food and Drugs 8 2014-04-01 2014-04-01 false Radiographic film illuminator. 892.1890 Section... (CONTINUED) MEDICAL DEVICES RADIOLOGY DEVICES Diagnostic Devices § 892.1890 Radiographic film illuminator. (a) Identification. A radiographic film illuminator is a device containing a visible light source covered with...

  16. 21 CFR 892.1890 - Radiographic film illuminator.

    Code of Federal Regulations, 2012 CFR

    2012-04-01

    ... 21 Food and Drugs 8 2012-04-01 2012-04-01 false Radiographic film illuminator. 892.1890 Section... (CONTINUED) MEDICAL DEVICES RADIOLOGY DEVICES Diagnostic Devices § 892.1890 Radiographic film illuminator. (a) Identification. A radiographic film illuminator is a device containing a visible light source covered with...

  17. 21 CFR 892.1890 - Radiographic film illuminator.

    Code of Federal Regulations, 2013 CFR

    2013-04-01

    ... 21 Food and Drugs 8 2013-04-01 2013-04-01 false Radiographic film illuminator. 892.1890 Section... (CONTINUED) MEDICAL DEVICES RADIOLOGY DEVICES Diagnostic Devices § 892.1890 Radiographic film illuminator. (a) Identification. A radiographic film illuminator is a device containing a visible light source covered with...

  18. 21 CFR 892.1890 - Radiographic film illuminator.

    Code of Federal Regulations, 2011 CFR

    2011-04-01

    ... 21 Food and Drugs 8 2011-04-01 2011-04-01 false Radiographic film illuminator. 892.1890 Section... (CONTINUED) MEDICAL DEVICES RADIOLOGY DEVICES Diagnostic Devices § 892.1890 Radiographic film illuminator. (a) Identification. A radiographic film illuminator is a device containing a visible light source covered with...

  19. 21 CFR 892.1890 - Radiographic film illuminator.

    Code of Federal Regulations, 2010 CFR

    2010-04-01

    ... 21 Food and Drugs 8 2010-04-01 2010-04-01 false Radiographic film illuminator. 892.1890 Section... (CONTINUED) MEDICAL DEVICES RADIOLOGY DEVICES Diagnostic Devices § 892.1890 Radiographic film illuminator. (a) Identification. A radiographic film illuminator is a device containing a visible light source covered with...

  20. 32 CFR 707.10 - Wake illumination light.

    Code of Federal Regulations, 2014 CFR

    2014-07-01

    ... 32 National Defense 5 2014-07-01 2014-07-01 false Wake illumination light. 707.10 Section 707.10... RESPECT TO ADDITIONAL STATION AND SIGNAL LIGHTS § 707.10 Wake illumination light. Naval vessels may display a white spot light located near the stern to illuminate the wake....