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Sample records for cryogenic light microscopy

  1. High-aperture cryogenic light microscopy

    Science.gov (United States)

    LE GROS, M.A.; McDERMOTT, G.; UCHIDA, M.; KNOECHEL, C.G.; LARABELL, C.A.

    2012-01-01

    Summary We report here the development of instruments and protocols for carrying out high numerical aperture immersion light microscopy on cryogenic specimens. Imaging by this modality greatly increases the lifetimes of fluorescence probes, including those commonly used for protein localization studies, while retaining the ability to image the specimen with high fidelity and spatial resolution. The novel use of a cryogenic immersion fluid also minimizes the refractive index mismatch between the sample and lens, leading to a more efficient coupling of the light from the sample to the image forming system. This enhancement is applicable to both fluorescence and transmitted light microscopy techniques. The design concepts used for the cryogenic microscope can be applied to virtually any existing light-based microscopy technique. This prospect is particularly exciting in the context of ‘super-resolution’ techniques, where enhanced fluorescence lifetime probes are especially useful. Thus, using this new modality it is now possible to observe dynamic events in a live cell, and then rapidly vitrify the specimen at a specific time point prior to carrying out high-resolution imaging. The techniques described can be used in conjunction with other imaging modalities in correlated studies. We have also developed instrumentation to perform cryo-light imaging together with soft X-ray tomography on the same cryo-fixed specimen as a means of carrying out high content, quantifiable correlated imaging analyses. These methods are equally applicable to correlated light and electron microscopy of frozen biological objects. PMID:19566622

  2. Soft X-ray Tomography and Cryogenic Light Microscopy: The Cool Combination in Cellular Imaging

    Science.gov (United States)

    McDermott, Gerry; Le Gros, Mark A.; Knoechel, Christian G.; Uchida, Maho; Larabell, Carolyn A.

    2012-01-01

    Soft x-ray tomography (SXT) is ideally suited to imaging sub-cellular architecture and organization, particularly in eukaryotic cells. SXT is similar in concept to the well-established medical diagnostic technique computed axial tomography (CAT), except SXT is capable of imaging with a spatial resolution of 50 nm, or better. In soft x-ray tomography (SXT) cells are imaged using photons from a region of the spectrum known as the ‘water window’. This results in quantitative, high-contrast images of intact, fully hydrated cells without the need to use contrast-enhancing agents. Cells are therefore visualized very close to their native, fully functional state. The utility of SXT has recently been enhanced by the development of high numerical aperture cryogenic light microscopy for correlated imaging. Taking this multi-modal approach now allows labeled molecules to be localized in the context of a high-resolution 3-dimensional tomographic reconstruction of the cell. PMID:19818625

  3. Laboratory-Based Cryogenic Soft X-ray Tomography with Correlative Cryo-Light and Electron Microscopy

    Energy Technology Data Exchange (ETDEWEB)

    Carlson, David B.; Gelb, Jeff; Palshin, Vadim; Evans, James E.

    2013-02-01

    Here we present a novel laboratory-based cryogenic soft X-ray microscope for whole cell tomography of frozen hydrated samples. We demonstrate the capabilities of this compact cryogenic microscope by visualizing internal sub-cellular structures of Saccharomyces cerevisiae cells. The microscope is shown to achieve better than 50 nm spatial resolution with a Siemens star test sample. For whole biological cells, the microscope can image specimens up to 5 micrometers thick. Structures as small as 90 nm can be detected in tomographic reconstructions at roughly 70 nm spatial resolution following a low cumulative radiation dose of only 7.2 MGy. Furthermore, the design of the specimen chamber utilizes a standard sample support that permits multimodal correlative imaging of the exact same unstained yeast cell via cryo-fluorescence light microscopy, cryo-soft x-ray microscopy and cryo-transmission electron microscopy. This completely laboratory-based cryogenic soft x-ray microscope will therefore enable greater access to three-dimensional ultrastructure determination of biological whole cells without chemical fixation or physical sectioning.

  4. High-numerical-aperture cryogenic light microscopy for increased precision of superresolution reconstructions.

    Science.gov (United States)

    Nahmani, Marc; Lanahan, Conor; DeRosier, David; Turrigiano, Gina G

    2017-04-11

    Superresolution microscopy has fundamentally altered our ability to resolve subcellular proteins, but improving on these techniques to study dense structures composed of single-molecule-sized elements has been a challenge. One possible approach to enhance superresolution precision is to use cryogenic fluorescent imaging, reported to reduce fluorescent protein bleaching rates, thereby increasing the precision of superresolution imaging. Here, we describe an approach to cryogenic photoactivated localization microscopy (cPALM) that permits the use of a room-temperature high-numerical-aperture objective lens to image frozen samples in their native state. We find that cPALM increases photon yields and show that this approach can be used to enhance the effective resolution of two photoactivatable/switchable fluorophore-labeled structures in the same frozen sample. This higher resolution, two-color extension of the cPALM technique will expand the accessibility of this approach to a range of laboratories interested in more precise reconstructions of complex subcellular targets.

  5. Cryogenic coherent X-ray diffraction imaging of biological samples at SACLA: a correlative approach with cryo-electron and light microscopy.

    Science.gov (United States)

    Takayama, Yuki; Yonekura, Koji

    2016-03-01

    Coherent X-ray diffraction imaging at cryogenic temperature (cryo-CXDI) allows the analysis of internal structures of unstained, non-crystalline, whole biological samples in micrometre to sub-micrometre dimensions. Targets include cells and cell organelles. This approach involves preparing frozen-hydrated samples under controlled humidity, transferring the samples to a cryo-stage inside a vacuum chamber of a diffractometer, and then exposing the samples to coherent X-rays. Since 2012, cryo-coherent diffraction imaging (CDI) experiments have been carried out with the X-ray free-electron laser (XFEL) at the SPring-8 Ångstrom Compact free-electron LAser (SACLA) facility in Japan. Complementary use of cryo-electron microscopy and/or light microscopy is highly beneficial for both pre-checking samples and studying the integrity or nature of the sample. This article reports the authors' experience in cryo-XFEL-CDI of biological cells and organelles at SACLA, and describes an attempt towards reliable and higher-resolution reconstructions, including signal enhancement with strong scatterers and Patterson-search phasing.

  6. Coherent light microscopy

    CERN Document Server

    Ferraro, Pietro; Zalevsky, Zeev

    2011-01-01

    This book deals with the latest achievements in the field of optical coherent microscopy. While many other books exist on microscopy and imaging, this book provides a unique resource dedicated solely to this subject. Similarly, many books describe applications of holography, interferometry and speckle to metrology but do not focus on their use for microscopy. The coherent light microscopy reference provided here does not focus on the experimental mechanics of such techniques but instead is meant to provide a users manual to illustrate the strengths and capabilities of developing techniques. Th

  7. Polarized Light Microscopy

    Science.gov (United States)

    Frandsen, Athela F.

    2016-01-01

    Polarized light microscopy (PLM) is a technique which employs the use of polarizing filters to obtain substantial optical property information about the material which is being observed. This information can be combined with other microscopy techniques to confirm or elucidate the identity of an unknown material, determine whether a particular contaminant is present (as with asbestos analysis), or to provide important information that can be used to refine a manufacturing or chemical process. PLM was the major microscopy technique in use for identification of materials for nearly a century since its introduction in 1834 by William Fox Talbot, as other techniques such as SEM (Scanning Electron Microscopy), FTIR (Fourier Transform Infrared spectroscopy), XPD (X-ray Powder Diffraction), and TEM (Transmission Electron Microscopy) had not yet been developed. Today, it is still the only technique approved by the Environmental Protection Agency (EPA) for asbestos analysis, and is often the technique first applied for identification of unknown materials. PLM uses different configurations in order to determine different material properties. With each configuration additional clues can be gathered, leading to a conclusion of material identity. With no polarizing filter, the microscope can be used just as a stereo optical microscope, and view qualities such as morphology, size, and number of phases. With a single polarizing filter (single polars), additional properties can be established, such as pleochroism, individual refractive indices, and dispersion staining. With two polarizing filters (crossed polars), even more can be deduced: isotropy vs. anisotropy, extinction angle, birefringence/degree of birefringence, sign of elongation, and anomalous polarization colors, among others. With the use of PLM many of these properties can be determined in a matter of seconds, even for those who are not highly trained. McCrone, a leader in the field of polarized light microscopy, often

  8. Correlated Light Microscopy and Electron Microscopy

    NARCIS (Netherlands)

    Sjollema, Klaas A.; Schnell, Ulrike; Kuipers, Jeroen; Kalicharan, Ruby; Giepmans, Ben N. G.; MullerReichert, T; Verkade, P

    2012-01-01

    Understanding where, when, and how biomolecules (inter)act is crucial to uncover fundamental mechanisms in cell biology. Recent developments in fluorescence light microscopy (FLM) allow protein imaging in living cells and at the near molecular level. However, fluorescence microscopy only reveals

  9. Correlated Light Microscopy and Electron Microscopy

    NARCIS (Netherlands)

    Sjollema, Klaas A.; Schnell, Ulrike; Kuipers, Jeroen; Kalicharan, Ruby; Giepmans, Ben N. G.; MullerReichert, T; Verkade, P

    2012-01-01

    Understanding where, when, and how biomolecules (inter)act is crucial to uncover fundamental mechanisms in cell biology. Recent developments in fluorescence light microscopy (FLM) allow protein imaging in living cells and at the near molecular level. However, fluorescence microscopy only reveals sel

  10. SINGLE: single photon sensitive cryogenic light detectors

    Science.gov (United States)

    Biassoni, Matteo; SINGLE Collaboration

    2017-09-01

    Thermal detectors operated at few mK as calorimeters are a powerful tool for the study of rare particle physics processes. In order to implement particle identification, light detection can be effectively performed by means of other thermal detectors operated as light sensors. This configuration can be used also in large scale, thousand-channels setups, but the light sensors must be sensitive enough to detect few, possibly a single, photons. The SINGLE project described here aims at producing silicon based, large area devices that can be operated as thermal detectors with single-photon sensitivity, and demonstrate the reliability of the performance, scalability of the production process and integrability with present and next generation cryogenic experiments for the search for rare events.

  11. Spatial light interference microscopy (SLIM).

    Science.gov (United States)

    Wang, Zhuo; Millet, Larry; Mir, Mustafa; Ding, Huafeng; Unarunotai, Sakulsuk; Rogers, John; Gillette, Martha U; Popescu, Gabriel

    2011-01-17

    We present spatial light interference microscopy (SLIM) as a new optical microscopy technique, capable of measuring nanoscale structures and dynamics in live cells via interferometry. SLIM combines two classic ideas in light imaging: Zernike's phase contrast microscopy, which renders high contrast intensity images of transparent specimens, and Gabor's holography, where the phase information from the object is recorded. Thus, SLIM reveals the intrinsic contrast of cell structures and, in addition, renders quantitative optical path-length maps across the sample. The resulting topographic accuracy is comparable to that of atomic force microscopy, while the acquisition speed is 1,000 times higher. We illustrate the novel insight into cell dynamics via SLIM by experiments on primary cell cultures from the rat brain. SLIM is implemented as an add-on module to an existing phase contrast microscope, which may prove instrumental in impacting the light microscopy field at a large scale.

  12. Advances in cryogenic transmission electron microscopy for the characterization of dynamic self-assembling nanostructures.

    Science.gov (United States)

    Newcomb, Christina J; Moyer, Tyson J; Lee, Sungsoo S; Stupp, Samuel I

    2012-12-01

    Elucidating the structural information of nanoscale materials in their solvent-exposed state is crucial, as a result, cryogenic transmission electron microscopy (cryo-TEM) has become an increasingly popular technique in the materials science, chemistry, and biology communities. Cryo-TEM provides a method to directly visualize the specimen structure in a solution-state through a thin film of vitrified solvent. This technique complements X-ray, neutron, and light scattering methods that probe the statistical average of all species present; furthermore, cryo-TEM can be used to observe changes in structure over time. In the area of self-assembly, this tool has been particularly powerful for the characterization of natural and synthetic small molecule assemblies, as well as hybrid organic-inorganic composites. In this review, we discuss recent advances in cryogenic TEM in the context of self-assembling systems with emphasis on characterization of transitions observed in response to external stimuli.

  13. Contrast enhancement in light microscopy.

    Science.gov (United States)

    Ernst Keller, H; Watkins, Simon

    2013-01-01

    The optical microscope is a fundamental component of an image cytometry system. This unit covers the basic concepts of light microscopy, including Köhler illumination, resolution, contrast, and numerical aperture, and reviews the many types of instruments and techniques for contrast enhancement.

  14. Light microscopy - Methods and protocols

    Directory of Open Access Journals (Sweden)

    CarloAlberto Redi

    2011-11-01

    Full Text Available The first part of the book (six chapters is devoted to some selected applications of bright-field microscopy while the second part (eight chapters to some fluorescence microscopy studies. Both animal and plant biology investigations are presented covering multiple fields like immunology, cell signaling, cancer biology and, surprisingly to me, ecology. This chapter is titled: Light microscopy in aquatic ecology: Methods for plankton communities studies and it is due to Maria Carolina S. Soares and colleagues from the Laboratory of Aquatic Ecology, Dept. of Biology, Federal University of Juiz de Fora (Brazil. Here they present methods to quantify the different component of planktonic communities in a step-by-step manner so that virus, bacteria, algae and animals pertaining to different taxa can be recognized and the contribution they made to the plankton composition evaluated. It descends that even how the plankton composition is changing due to environmental variations can be accurately determined....

  15. Cryogenic transmission electron microscopy nanostructural study of shed microparticles.

    Directory of Open Access Journals (Sweden)

    Liron Issman

    Full Text Available Microparticles (MPs are sub-micron membrane vesicles (100-1000 nm shed from normal and pathologic cells due to stimulation or apoptosis. MPs can be found in the peripheral blood circulation of healthy individuals, whereas elevated concentrations are found in pregnancy and in a variety of diseases. Also, MPs participate in physiological processes, e.g., coagulation, inflammation, and angiogenesis. Since their clinical properties are important, we have developed a new methodology based on nano-imaging that provides significant new data on MPs nanostructure, their composition and function. We are among the first to characterize by direct-imaging cryogenic transmitting electron microscopy (cryo-TEM the near-to-native nanostructure of MP systems isolated from different cell types and stimulation procedures. We found that there are no major differences between the MP systems we have studied, as most particles were spherical, with diameters from 200 to 400 nm. However, each MP population is very heterogeneous, showing diverse morphologies. We investigated by cryo-TEM the effects of standard techniques used to isolate and store MPs, and found that either high-g centrifugation of MPs for isolation purposes, or slow freezing to -80 °C for storage introduce morphological artifacts, which can influence MP nanostructure, and thus affect the efficiency of these particles as future diagnostic tools.

  16. Image scanning microscopy with radially polarized light

    Science.gov (United States)

    Xiao, Yun; Zhang, Yunhai; Wei, Tongda; Huang, Wei; Shi, Yaqin

    2017-03-01

    In order to improve the resolution of image scanning microscopy, we present a method based on image scanning microscopy and radially polarized light. According to the theory of image scanning microscopy, we get the effective point spread function of image scanning microscopy with the longitudinal component of radially polarized light and a 1 AU detection area, and obtain imaging results of the analyzed samples using this method. Results show that the resolution can be enhanced by 7% compared with that in image scanning microscopy with circularly polarized light, and is 1.54-fold higher than that in confocal microscopy with a pinhole of 1 AU. Additionally, the peak intensity of ISM is 1.54-fold higher than that of a confocal microscopy with a pinhole of 1 AU. In conclusion, the combination of the image scanning microscopy and the radially polarized light could improve the resolution, and it could realize high-resolution and high SNR imaging at the same time.

  17. CALDER: cryogenic light detector for rare events search

    CERN Document Server

    Pagnanini, L; Bellini, F; Calvo, M; Cardani, L; Casali, N; Castellano, M G; Colantoni, I; Coppolecchia, A; Cosmelli, C; Cruciani, A; De Bernardis, P; Di Domizio, S; D'Addabbo, A; Martinez, M; Masi, S; Tomei, C; Vignati, M

    2015-01-01

    The CALDER project aims at developing cryogenic light detectors with high sensitivity to UV and visible light, to be used for particle tagging in massive bolometers. Indeed the sensitivity of CUORE can be increased by a factor of 3, thanks to the reduction of the $\\alpha$-background, obtained by detecting the Cherenkov light (100 eV) emitted by $\\beta/\\gamma$ events. Currently used light detectors have not the features required to address this task, so we decided to develop a new light detector using Kinetic Inductance Detector as a sensor. This approach is very challenging and requires an intensive R$\\&$D to be satisfied. The first results of this activity are shown in the following.

  18. Optimization of light collection from crystal scintillators for cryogenic experiments

    Energy Technology Data Exchange (ETDEWEB)

    Danevich, F.A., E-mail: danevich@kinr.kiev.ua [Institute for Nuclear Research, MSP 03680, Kyiv (Ukraine); Kobychev, R.V. [Institute for Nuclear Research, MSP 03680, Kyiv (Ukraine); National Technical University of Ukraine “Kyiv Polytechnic Institute”, 03056 Kyiv (Ukraine); Kobychev, V.V. [Institute for Nuclear Research, MSP 03680, Kyiv (Ukraine); Kyungpook National University, Daegu 702-701 (Korea, Republic of); Kraus, H. [Department of Physics, University of Oxford, Keble Road, Oxford OX1 3RH (United Kingdom); Mikhailik, V.B. [Department of Physics, University of Oxford, Keble Road, Oxford OX1 3RH (United Kingdom); Diamond Light Source, Harwell Science Campus, Didcot, OX11 0DE (United Kingdom); Mokina, V.M. [Institute for Nuclear Research, MSP 03680, Kyiv (Ukraine)

    2014-04-21

    High light collection efficiency is an important requirement in any application of scintillation detectors. The purpose of this study is to investigate the possibility for improving this parameter in cryogenic scintillation bolometers, which can be considered as promising detectors in experiments investigating neutrinoless double beta decay and dark matter. Energy resolutions and relative pulse amplitudes of scintillation detectors using ZnWO{sub 4} scintillation crystals of different shapes (cylinder ∅ 20×20 mm and hexagonal prism with diagonal 20 mm and height 20 mm), reflector materials and shapes, optical contact and surface properties (polished and diffused) were measured at room temperature. Propagation of optical photons in these experimental conditions was simulated using Geant4 and ZEMAX codes. The results of the simulations are found to be in good agreement with each other and with direct measurements of the crystals. This could be applied to optimize the geometry of scintillation detectors used in the cryogenic experiments.

  19. Cryogenic-temperature electron microscopy direct imaging of carbon nanotubes and graphene solutions in superacids.

    Science.gov (United States)

    Kleinerman, O; Parra-Vasquez, A Nicholas G; Green, M J; Behabtu, N; Schmidt, J; Kesselman, E; Young, C C; Cohen, Y; Pasquali, M; Talmon, Y

    2015-07-01

    Cryogenic electron microscopy (cryo-EM) is a powerful tool for imaging liquid and semiliquid systems. While cryogenic transmission electron microscopy (cryo-TEM) is a standard technique in many fields, cryogenic scanning electron microscopy (cryo-SEM) is still not that widely used and is far less developed. The vast majority of systems under investigation by cryo-EM involve either water or organic components. In this paper, we introduce the use of novel cryo-TEM and cryo-SEM specimen preparation and imaging methodologies, suitable for highly acidic and very reactive systems. Both preserve the native nanostructure in the system, while not harming the expensive equipment or the user. We present examples of direct imaging of single-walled, multiwalled carbon nanotubes and graphene, dissolved in chlorosulfonic acid and oleum. Moreover, we demonstrate the ability of these new cryo-TEM and cryo-SEM methodologies to follow phase transitions in carbon nanotube (CNT)/superacid systems, starting from dilute solutions up to the concentrated nematic liquid-crystalline CNT phases, used as the 'dope' for all-carbon-fibre spinning. Originally developed for direct imaging of CNTs and graphene dissolution and self-assembly in superacids, these methodologies can be implemented for a variety of highly acidic systems, paving a way for a new field of nonaqueous cryogenic electron microscopy. © 2015 The Authors Journal of Microscopy © 2015 Royal Microscopical Society.

  20. Cryogenic transmission electron microscopy (cryo-TEM) for studying the morphology of colloidal drug delivery systems

    DEFF Research Database (Denmark)

    Kuntsche, Judith; Horst, Jennifer C; Bunjes, Heike

    2011-01-01

    Cryogenic transmission electron microscopy (cryo-TEM) has evolved into an indispensable tool for the characterization of colloidal drug delivery systems. It can be applied to study the size, shape and internal structure of nanoparticulate carrier systems as well as the overall colloidal composition...

  1. Cryogenic Electron Microscopy Studies: Structure and Formation of Self-assembled Nanostructures in Solution

    Science.gov (United States)

    Lee, Han Seung

    Cryogenic electron microscopy (Cryo-EM) techniques are among the most powerful to characterize self-assembling soft materials (colloids, polymers, and microemulsions, etc.) at the nanometer scale, without any need for implicit models or assumptions about the structure. We can even visualize structure under dynamic conditions, capturing each stage of development. In this thesis, cryo-EM has been used to investigate the formation and structure of a variety of self-assembling soft materials. Visualization is complemented by small angle X-ray scattering (SAXS), dynamic light scattering, and conductivity measurements. In each case, cryo-EM provides new insights, not otherwise available, into the nanostructure development. Self-assembly phenomena at the molecular level are critical to the performance of tremendous number of applied systems ranging from personal care products to industrial products. To evaluate these self-assembled materials, multiple characterization techniques are required. We investigated aggregation behavior of cesium dodecyl sulfate (CsDS) ionic surfactant in aqueous solution. Coupled with the real space data from cryogenic transmission electron microscopy (Cryo-TEM) and the inverse space data from SAXS, the experimental result of CsDS in aqueous solution gave a new insight in CsDS micellar structures and their development as a function of concentration. Cryo-TEM showed the presence of the liquid-like hydrocarbon core in the CsDS micelles and relatively thick shell structures at a low CsDS concentration. The core-shell sphere structure micelle shifted to core-shell cylindrical micelle structure at high concentration. The morphology and structure of paclitaxel silicate (PTX) prodrug, encapsulated with amphiphilic poly(ethylene glycol)-b-poly(lactic-co-glycolic acid) diblock copolymers were studied. The six different silicate PTX prodrug candidates were characterized with cryo-TEM. Direct imaging with cryo-TEM illustrated structure of prodrug

  2. Scanning SQUID microscopy in a cryogen-free refrigerator

    Science.gov (United States)

    Schaefer, Brian T.; Low, David; Prawiroatmodjo, Guenevere E. D. K.; Nangoi, J. Kevin; Kim, Jihoon; Nowack, Katja C.

    With helium prices rising and supply becoming increasingly uncertain, it has become attractive to use dry cryostats with cryocoolers rather than liquid helium to reach low temperatures. However, a cryocooler introduces vibrations at the sample stage, making scanning probe experiments more challenging. Here, we report our progress on a superconducting quantum interference device (SQUID) microscope implemented for the first time in a compact, cryogen-free 5 K system. Our microscope is designed to reach submicron spatial resolution and a flux sensitivity of approximately 1 μΦ0 /√{ Hz} , where Φ0 is the magnetic flux quantum. To enable height feedback while approaching and scanning samples, we mount the SQUID on a quartz tuning fork. Our system promises to meet the capabilities of similar systems implemented in helium cryostats.

  3. Light-Weight Injector Technology for Cryogenic Mars Ascent Engines

    Science.gov (United States)

    Trihn, Huu Phuoc; Cramer, John M.

    1998-01-01

    Preliminary mission studies for human exploration of Mars have been performed at Marshall Space Flight Center (MSFC). These studies indicate that for chemical rockets only a cryogenic propulsion system would provide high enough performance to be considered for a Mars ascent vehicle. Although the mission is possible with Earth-supplied propellants for this vehicle, utilization of in-situ propellants is highly attractive. This option would significantly reduce the overall mass of launch vehicles. Consequently, the cost of the mission would be greatly reduced because the number and size of the Earth launch vehicle(s) needed for the mission decrease. NASA/Johnson Space Center has initiated several concept studies of in-situ propellant production plants. Liquid oxygen (LOX) is the primary candidate for an in-situ oxidizer. In-situ fuel candidates include methane (CH4), ethylene (C2H4), and methanol (CH3OH). MSFC initiated a technology development program for a cryogenic propulsion system for the Mars human exploration mission in 1998. One part of this technology program is the effort described here: an evaluation of propellant injection concepts for a LOX/liquid methane Mars Ascent Engine (MAE) with an emphasis on light-weight, high efficiency, reliability, and thermal compatibility. In addition to the main objective, hot-fire tests of the subject injectors will be used to test other key technologies including light-weight combustion chamber materials and advanced ignition concepts. This state-of-the-art technology will then be applied to the development of a cryogenic propulsion system that will meet the requirements of the planned Mars sample return (MSR) mission. The current baseline propulsion system for the MSR mission uses a storable propellant combination [monomethyl hydrazine/mixed oxides of nitrogen-25. However, a mission option that incorporates in-situ propellant production and utilization for the ascent stage is being carefully considered as a subscale

  4. Cryogenic transmission electron microscopy of recombinant tuberculosis vaccine antigen with anionic liposomes reveals formation of flattened liposomes

    Directory of Open Access Journals (Sweden)

    Fox CB

    2014-03-01

    Full Text Available Christopher B Fox,1 Sean K Mulligan,2 Joyce Sung,2 Quinton M Dowling,1 H W Millie Fung,1 Thomas S Vedvick,1 Rhea N Coler1 1Infectious Disease Research Institute, Seattle, WA, USA; 2NanoImaging Services, La Jolla, CA, USA Abstract: Development of lipid-based adjuvant formulations to enhance the immunogenicity of recombinant vaccine antigens is a focus of modern vaccine research. Characterizing interactions between vaccine antigens and formulation excipients is important for establishing compatibility between the different components and optimizing vaccine stability and potency. Cryogenic transmission electron microscopy (TEM is a highly informative analytical technique that may elucidate various aspects of protein- and lipid-based structures, including morphology, size, shape, and phase structure, while avoiding artifacts associated with staining-based TEM. In this work, cryogenic TEM is employed to characterize a recombinant tuberculosis vaccine antigen, an anionic liposome formulation, and antigen–liposome interactions. By performing three-dimensional tomographic reconstruction analysis, the formation of a population of protein-containing flattened liposomes, not present in the control samples, was detected. It is shown that cryogenic TEM provides unique information regarding antigen–liposome interactions not detectable by light-scattering-based methods. Employing a suite of complementary analytical techniques is important to fully characterize interactions between vaccine components. Keywords: vaccine adjuvant; cryo-TEM; antigen-adjuvant interactions; vaccine physical characterization; vaccine formulation morphology; 3D tomographic reconstruction

  5. Cryogenic transmission electron microscopy of recombinant tuberculosis vaccine antigen with anionic liposomes reveals formation of flattened liposomes.

    Science.gov (United States)

    Fox, Christopher B; Mulligan, Sean K; Sung, Joyce; Dowling, Quinton M; Fung, H W Millie; Vedvick, Thomas S; Coler, Rhea N

    2014-01-01

    Development of lipid-based adjuvant formulations to enhance the immunogenicity of recombinant vaccine antigens is a focus of modern vaccine research. Characterizing interactions between vaccine antigens and formulation excipients is important for establishing compatibility between the different components and optimizing vaccine stability and potency. Cryogenic transmission electron microscopy (TEM) is a highly informative analytical technique that may elucidate various aspects of protein- and lipid-based structures, including morphology, size, shape, and phase structure, while avoiding artifacts associated with staining-based TEM. In this work, cryogenic TEM is employed to characterize a recombinant tuberculosis vaccine antigen, an anionic liposome formulation, and antigen-liposome interactions. By performing three-dimensional tomographic reconstruction analysis, the formation of a population of protein-containing flattened liposomes, not present in the control samples, was detected. It is shown that cryogenic TEM provides unique information regarding antigen-liposome interactions not detectable by light-scattering-based methods. Employing a suite of complementary analytical techniques is important to fully characterize interactions between vaccine components.

  6. Introduction to Modern Methods in Light Microscopy.

    Science.gov (United States)

    Ryan, Joel; Gerhold, Abby R; Boudreau, Vincent; Smith, Lydia; Maddox, Paul S

    2017-01-01

    For centuries, light microscopy has been a key method in biological research, from the early work of Robert Hooke describing biological organisms as cells, to the latest in live-cell and single-molecule systems. Here, we introduce some of the key concepts related to the development and implementation of modern microscopy techniques. We briefly discuss the basics of optics in the microscope, super-resolution imaging, quantitative image analysis, live-cell imaging, and provide an outlook on active research areas pertaining to light microscopy.

  7. CALDER: Cryogenic light detectors for background-free searches

    Science.gov (United States)

    Cardani, L.; Bellini, F.; Casali, N.; Castellano, M. G.; Colantoni, I.; Coppolecchia, A.; Cosmelli, C.; Cruciani, A.; Di Domizio, S.; Tomei, C.; Vignati, M.

    2015-08-01

    The development of background-free detectors is essential for experiments searching for rare events. Bolometers, that are among the most competitive devices for the study of neutrino-less double beta decay (0νDBD) and Dark Matter interactions, suffer from the absence of techniques that allow to identify the nature of the interacting particles. This limit can be overcome by coupling the bolometer to an independent device for the measurement of the light emitted by interactions, as the combined read-out of the bolometric and light signals allows to identify and reject particles different from those of interest. CUORE, the most advanced bolometric experiment for 0νDBD searches, could disentangle the electrons produced by 0νDBD from the dangerous background due to α particles, by measuring the (tiny) Cherenkov light emitted by electrons and not by α's. LUCIFER, a project based on ZnSe scintillating bolometers for the study of 82Se 0νDBD, would be competitive also in the search of Dark Matter interactions if equipped with light detectors that allow to distinguish and reject the background due to electrons and γ's. These advances require cryogenic detectors characterized by noise lower than 20 eV, large active area, wide temperature range of operation, high radio-purity and ease in fabricating hundreds of channels. The CALDER collaboration aims to develop such detectors by exploiting the superb energy resolution and natural multiplexed read-out provided by Kinetic Inductance Detectors.

  8. New application of superconductors: high sensitivity cryogenic light detectors

    CERN Document Server

    Cardani, L; Casali, N; Casellano, M G; Colantoni, I; Coppolecchia, A; Cosmelli, C; Cruciani, A; D'Addabbo, A; Di Domizio, S; Martinez, M; Tomei, C; Vignati, M

    2016-01-01

    In this paper we describe the current status of the CALDER project, which is developing ultra-sensitive light detectors based on superconductors for cryogenic applications. When we apply an AC current to a superconductor, the Cooper pairs oscillate and acquire kinetic inductance, that can be measured by inserting the superconductor in a LC circuit with high merit factor. Interactions in the superconductor can break the Cooper pairs, causing sizable variations in the kinetic inductance and, thus, in the response of the LC circuit. The continuous monitoring of the amplitude and frequency modulation allows to reconstruct the incident energy with excellent sensitivity. This concept is at the basis of Kinetic Inductance Detectors (KIDs), that are characterized by natural aptitude to multiplexed read-out (several sensors can be tuned to different resonant frequencies and coupled to the same line), resolution of few eV, stable behavior over a wide temperature range, and ease in fabrication. We present the results ob...

  9. Robust image alignment for cryogenic transmission electron microscopy.

    Science.gov (United States)

    McLeod, Robert A; Kowal, Julia; Ringler, Philippe; Stahlberg, Henning

    2017-03-01

    Cryo-electron microscopy recently experienced great improvements in structure resolution due to direct electron detectors with improved contrast and fast read-out leading to single electron counting. High frames rates enabled dose fractionation, where a long exposure is broken into a movie, permitting specimen drift to be registered and corrected. The typical approach for image registration, with high shot noise and low contrast, is multi-reference (MR) cross-correlation. Here we present the software package Zorro, which provides robust drift correction for dose fractionation by use of an intensity-normalized cross-correlation and logistic noise model to weight each cross-correlation in the MR model and filter each cross-correlation optimally. Frames are reliably registered by Zorro with low dose and defocus. Methods to evaluate performance are presented, by use of independently-evaluated even- and odd-frame stacks by trajectory comparison and Fourier ring correlation. Alignment of tiled sub-frames is also introduced, and demonstrated on an example dataset. Zorro source code is available at github.com/CINA/zorro. Copyright © 2016 Elsevier Inc. All rights reserved.

  10. IR microscopy utilizing intense supercontinuum light source

    DEFF Research Database (Denmark)

    Dupont, Sune; Petersen, Christian; Thøgersen, Jan;

    2012-01-01

    . The supercontinuum light source has a high brightness and spans the infrared region from 1400 nm to 4000 nm. This combination allows contact free high resolution hyper spectral infrared microscopy. The microscope is demonstrated by imaging an oil/water sample with 20 μm resolution.......Combining the molecular specificity of the infrared spectral region with high resolution microscopy has been pursued by researchers for decades. Here we demonstrate infrared supercontinuum radiated from an optical fiber as a promising new light source for infrared microspectroscopy...

  11. Polarized light microscopy: principles and practice.

    Science.gov (United States)

    Oldenbourg, Rudolf

    2013-11-01

    Polarized light microscopy provides unique opportunities for analyzing the molecular order in heterogeneous systems, such as living cells and tissues, without using exogenous dyes or labels. This article briefly discusses the theory of polarized light microscopy and elaborates on its practice using a traditional polarized light microscope and more specialized polarization microscopes such as the LC-PolScope, Oosight, or Abrio. The microscope components specific to analyzing the polarization of light, such as polarizer and compensator, are introduced, and quantitative techniques for measuring the birefringence of the specimen point by point using a traditional polarizing microscope are discussed. The new LC-PolScope greatly improves the analytic power of the technique, providing quantitative birefringence data simultaneously for every image point, thereby revealing molecular order with unprecedented sensitivity and at the highest resolution of the light microscope. Practical aspects discussed include the choice of optics, sample preparation, and combining polarized light with differential interference contrast and fluorescence microscopy. A glossary of polarization optical terms is also included to facilitate the discussion of observations made with a polarized light microscope.

  12. Core size determination and structural characterization of intravenous iron complexes by cryogenic transmission electron microscopy.

    Science.gov (United States)

    Wu, Yong; Petrochenko, Peter; Chen, Lynn; Wong, Sook Yee; Absar, Mohammad; Choi, Stephanie; Zheng, Jiwen

    2016-05-30

    Understanding physicochemical properties of intravenous (IV) iron drug products is essential to ensure the manufacturing process is consistent and streamlined. The history of physicochemical characterization of IV iron complex formulations stretches over several decades, with disparities in iron core size and particle morphology as the major source of debate. One of the main reasons for this controversy is room temperature sample preparation artifacts, which affect accurate determination of size, shape and agglomeration/aggregation of nanoscale iron particles. The present study is first to report the ultra-fine iron core structures of four IV iron complex formulations, sodium ferric gluconate, iron sucrose, low molecular weight iron dextran and ferumoxytol, using a cryogenic transmission electron microscopy (cryo-TEM) preservation technique, as opposed to the conventional room temperature (RT-TEM) technique. Our results show that room temperature preparation causes nanoparticle aggregation and deformation, while cryo-TEM preserves IV iron colloidal suspension in their native frozen-hydrated and undiluted state. In contrast to the current consensus in literature, all four IV iron colloids exhibit a similar morphology of their iron oxide cores with a spherical shape, narrow size distribution and an average size of 2nm. Moreover, out of the four tested formulations, ferumoxytol exhibits a cluster-like community of several iron carbohydrate particles which likely accounts for its large hydrodynamic size of 25nm, measured with dynamic light scattering. Our findings outline a suitable method for identifying colloidal nanoparticle core size in the native state, which is increasingly important for manufacturing and design control of complex drug formulations, such as IV iron drug products.

  13. New application of superconductors: High sensitivity cryogenic light detectors

    Science.gov (United States)

    Cardani, L.; Bellini, F.; Casali, N.; Castellano, M. G.; Colantoni, I.; Coppolecchia, A.; Cosmelli, C.; Cruciani, A.; D'Addabbo, A.; Di Domizio, S.; Martinez, M.; Tomei, C.; Vignati, M.

    2017-02-01

    In this paper we describe the current status of the CALDER project, which is developing ultra-sensitive light detectors based on superconductors for cryogenic applications. When we apply an AC current to a superconductor, the Cooper pairs oscillate and acquire kinetic inductance, that can be measured by inserting the superconductor in a LC circuit with high merit factor. Interactions in the superconductor can break the Cooper pairs, causing sizable variations in the kinetic inductance and, thus, in the response of the LC circuit. The continuous monitoring of the amplitude and frequency modulation allows to reconstruct the incident energy with excellent sensitivity. This concept is at the basis of Kinetic Inductance Detectors (KIDs) that are characterized by natural aptitude to multiplexed read-out (several sensors can be tuned to different resonant frequencies and coupled to the same line), resolution of few eV, stable behavior over a wide temperature range, and ease in fabrication. We present the results obtained by the CALDER collaboration with 2×2 cm2 substrates sampled by 1 or 4 Aluminum KIDs. We show that the performances of the first prototypes are already competitive with those of other commonly used light detectors, and we discuss the strategies for a further improvement.

  14. CALDER: Cryogenic light detectors for background-free searches

    Energy Technology Data Exchange (ETDEWEB)

    Cardani, L. [Dipartimento di Fisica - Sapienza Università di Roma, Roma - Italy and Physics Department, Princeton University, Princeton, NJ (United States); Bellini, F.; Casali, N.; Coppolecchia, A.; Cosmelli, C.; Cruciani, A.; Vignati, M. [Dipartimento di Fisica - Sapienza Università di Roma and INFN - Sezione di Roma, Roma - Italy (Italy); Castellano, M. G. [Istituto di Fotonica e Nanotecnologie - CNR, Roma - Italy (Italy); Colantoni, I. [Dipartimento di Fisica - Sapienza Università di Roma (Italy); Di Domizio, S. [Dipartimento di Fisica, Università di Genova, Genova - Italy and INFN Sezione di Genova, Genova - Italy (Italy); Tomei, C. [INFN - Sezione di Roma, Roma - Italy (Italy)

    2015-08-17

    The development of background-free detectors is essential for experiments searching for rare events. Bolometers, that are among the most competitive devices for the study of neutrino-less double beta decay (0νDBD) and Dark Matter interactions, suffer from the absence of techniques that allow to identify the nature of the interacting particles. This limit can be overcome by coupling the bolometer to an independent device for the measurement of the light emitted by interactions, as the combined read-out of the bolometric and light signals allows to identify and reject particles different from those of interest. CUORE, the most advanced bolometric experiment for 0νDBD searches, could disentangle the electrons produced by 0νDBD from the dangerous background due to α particles, by measuring the (tiny) Cherenkov light emitted by electrons and not by α’s. LUCIFER, a project based on ZnSe scintillating bolometers for the study of {sup 82}Se 0νDBD, would be competitive also in the search of Dark Matter interactions if equipped with light detectors that allow to distinguish and reject the background due to electrons and γ’s. These advances require cryogenic detectors characterized by noise lower than 20 eV, large active area, wide temperature range of operation, high radio-purity and ease in fabricating hundreds of channels. The CALDER collaboration aims to develop such detectors by exploiting the superb energy resolution and natural multiplexed read-out provided by Kinetic Inductance Detectors.

  15. Confocal multiview light-sheet microscopy

    Science.gov (United States)

    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

  16. Three axis vector magnet set-up for cryogenic scanning probe microscopy

    Energy Technology Data Exchange (ETDEWEB)

    Galvis, J. A. [Laboratorio de Bajas Temperaturas, Departamento de Física de la Materia Condensada, Instituto de Ciencia de Materiales Nicolás Cabrera, Condensed Matter Physics Center (IFIMAC), Facultad de Ciencias Universidad Autónoma de Madrid, 28049 Madrid (Spain); Departamento de Ciencias Naturales Facultad de Ingeniería Universidad Central, Bogotá (Colombia); Herrera, E.; Buendía, A. [Laboratorio de Bajas Temperaturas, Departamento de Física de la Materia Condensada, Instituto de Ciencia de Materiales Nicolás Cabrera, Condensed Matter Physics Center (IFIMAC), Facultad de Ciencias Universidad Autónoma de Madrid, 28049 Madrid (Spain); Guillamón, I.; Vieira, S.; Suderow, H. [Laboratorio de Bajas Temperaturas, Departamento de Física de la Materia Condensada, Instituto de Ciencia de Materiales Nicolás Cabrera, Condensed Matter Physics Center (IFIMAC), Facultad de Ciencias Universidad Autónoma de Madrid, 28049 Madrid (Spain); Unidad Asociada de Bajas Temperaturas y Altos Campos Magnéticos, UAM, CSIC, Cantoblanco, E-28049 Madrid (Spain); Azpeitia, J.; Luccas, R. F.; Munuera, C.; García-Hernandez, M. [Unidad Asociada de Bajas Temperaturas y Altos Campos Magnéticos, UAM, CSIC, Cantoblanco, E-28049 Madrid (Spain); Instituto de Ciencia de Materiales de Madrid, Consejo Superior de Investigaciones Científicas (ICMM-CSIC), Sor Juana Inés de la Cruz 3, 28049 Madrid (Spain); and others

    2015-01-15

    We describe a three axis vector magnet system for cryogenic scanning probe microscopy measurements. We discuss the magnet support system and the power supply, consisting of a compact three way 100 A current source. We obtain tilted magnetic fields in all directions with maximum value of 5T along z-axis and of 1.2T for XY-plane magnetic fields. We describe a scanning tunneling microscopy-spectroscopy (STM-STS) set-up, operating in a dilution refrigerator, which includes a new high voltage ultralow noise piezodrive electronics and discuss the noise level due to vibrations. STM images and STS maps show atomic resolution and the tilted vortex lattice at 150 mK in the superconductor β-Bi{sub 2}Pd. We observe a strongly elongated hexagonal lattice, which corresponds to the projection of the tilted hexagonal vortex lattice on the surface. We also discuss Magnetic Force Microscopy images in a variable temperature insert.

  17. Investigation of Neganov-Luke amplified cryogenic light-detectors for CRESST and EURECA

    Energy Technology Data Exchange (ETDEWEB)

    Ertl, Andreas; Guetlein, Achim; Lanfranchi, Jean-Come; Muenster, Andrea; Potzel, Walter; Roth, Sabine; Simon, Daniel; Scholl, Stephan; Sivers, Moritz von; Strauss, Raimund; Wawoczny, Stephan; Willers, Michael; Wuestrich, Marc; Zoeller, Andreas [Technische Universitaet Muenchen, Physik Department E15, Garching (Germany)

    2013-07-01

    Experiments for the direct detection of dark matter which employ the phonon-light technique like CRESST and the planned experiment EURECA rely heavily on the separation of the different nuclear-recoil bands at low energies for their background suppression. The CRESST experiment uses scintillating CaWO{sub 4} crystals as a target in the search for coherent WIMP-nucleon scattering. In the case of electron recoils, about 1% of the energy deposited in a CaWO{sub 4} crystal is detected as scintillation light in a separate cryogenic light-detector. For nuclear recoils the scintillation light is further quenched which motivates the need for very sensitive light-detectors. Neganov-Luke amplified cryogenic light-detectors offer a promising way to increase the sensitivity of cryogenic light-detectors by drifting photon induced electrons and holes in an applied electric field and thus amplifying the resulting phonon signal.

  18. Note: Development of a wideband amplifier for cryogenic scanning tunneling microscopy

    Science.gov (United States)

    Zhang, Chao; Jeon, Hoyeon; Oh, Myungchul; Lee, Minjun; Kim, Sungmin; Yi, Sunwouk; Lee, Hanho; Zoh, Inhae; Yoo, Yongchan; Kuk, Young

    2017-06-01

    A wideband cryogenic amplifier has been developed for low temperature scanning tunneling microscopy. The amplifier consisting of a wideband complementary metal oxide semiconductor field effect transistors operational amplifier together with a feedback resistor of 100 kΩ and a capacitor is mounted within a 4 K Dewar. This amplifier has a wide bandwidth and is successfully applied to scanning tunneling microscopy applications at low temperatures down to ˜7 K. The quality of the designed amplifier is validated by high resolution imaging. More importantly, the amplifier has also proved to be capable of performing scanning tunneling spectroscopy measurements, showing the detection of the Shockley surface state of the Au(111) surface and the superconducting gap of Nb(110).

  19. Metallothioneins for correlative light and electron microscopy.

    Science.gov (United States)

    Fernández de Castro, Isabel; Sanz-Sánchez, Laura; Risco, Cristina

    2014-01-01

    Structural biologists have been working for decades on new strategies to identify proteins in cells unambiguously. We recently explored the possibilities of using the small metal-binding protein, metallothionein (MT), as a tag to detect proteins in transmission electron microscopy. It had been reported that, when fused with a protein of interest and treated in vitro with gold salts, a single MT tag will build an electron-dense gold cluster ~1 nm in diameter; we provided proof of this principle by demonstrating that MT can be used to detect intracellular proteins in bacteria and eukaryotic cells. The method, which is compatible with a variety of sample processing techniques, allows specific detection of proteins in cells with exceptional sensitivity. We illustrated the applicability of the technique in a series of studies to visualize the intracellular distribution of bacterial and viral proteins. Immunogold labeling was fundamental to confirm the specificity of the MT-gold method. When proteins were double-tagged with green fluorescent protein and MT, direct correlative light and electron microscopy allowed visualization of the same macromolecular complexes with different spatial resolutions. MT-gold tagging might also become a useful tool for mapping proteins into the 3D-density maps produced by (cryo)-electron tomography. New protocols will be needed for double or multiple labeling of proteins, using different versions of MT with fluorophores of different colors. Further research is also necessary to render the MT-gold labeling procedure compatible with immunogold labeling on Tokuyasu cryosections and with cryo-electron microscopy of vitreous sections.

  20. Correlative light and electron microscopy : strategies and applications

    NARCIS (Netherlands)

    Driel, Linda Francina van

    2011-01-01

    Correlative light and electron microscopy (CLEM) refers to the observation of the same structures or ultrastructures with both light microscopy (LM) and electron microscopy (EM). LM provides an overview of the studied material, and enables the quick localization of structures that are fluorescently

  1. Electronic cameras for low-light microscopy.

    Science.gov (United States)

    Rasnik, Ivan; French, Todd; Jacobson, Ken; Berland, Keith

    2013-01-01

    This chapter introduces to electronic cameras, discusses the various parameters considered for evaluating their performance, and describes some of the key features of different camera formats. The chapter also presents the basic understanding of functioning of the electronic cameras and how these properties can be exploited to optimize image quality under low-light conditions. Although there are many types of cameras available for microscopy, the most reliable type is the charge-coupled device (CCD) camera, which remains preferred for high-performance systems. If time resolution and frame rate are of no concern, slow-scan CCDs certainly offer the best available performance, both in terms of the signal-to-noise ratio and their spatial resolution. Slow-scan cameras are thus the first choice for experiments using fixed specimens such as measurements using immune fluorescence and fluorescence in situ hybridization. However, if video rate imaging is required, one need not evaluate slow-scan CCD cameras. A very basic video CCD may suffice if samples are heavily labeled or are not perturbed by high intensity illumination. When video rate imaging is required for very dim specimens, the electron multiplying CCD camera is probably the most appropriate at this technological stage. Intensified CCDs provide a unique tool for applications in which high-speed gating is required. The variable integration time video cameras are very attractive options if one needs to acquire images at video rate acquisition, as well as with longer integration times for less bright samples. This flexibility can facilitate many diverse applications with highly varied light levels.

  2. Polarized light microscopy in mammalian oocytes.

    Science.gov (United States)

    Caamaño, J N; Muñoz, M; Diez, C; Gómez, E

    2010-06-01

    The meiotic spindle structure plays a key role in normal chromosome alignment and segregation during meiosis. Polarized light microscopy (PLM) allows non-invasive evaluation of the meiotic spindle of metaphase oocytes from different animal species. The purpose of this article is to review the use of PLM in animal reproduction, mainly in the assessment of the meiotic spindle in oocytes. A brief overview of the methods to assess the meiotic spindle is presented as well as the principles behind the PLM. The use of PLM to evaluate oocyte quality and spindle morphology is discussed and the results on the viability of the oocytes after being exposed to PLM are presented. Several researchers showed that PLM could be successfully implemented on cryopreservation, nuclear transfer and intracytoplasmic sperm injection procedures as a tool to improve the outcome of these procedures. In addition, PLM can be used to develop studies on oocyte maturation and spindle dynamics. However, the information on the practical use of this technology in farm animals is very limited and further studies are needed to assess the importance of PLM in animal reproduction.

  3. A quick guide to light microscopy in cell biology

    Science.gov (United States)

    Thorn, Kurt

    2016-01-01

    Light microscopy is a key tool in modern cell biology. Light microscopy has several features that make it ideally suited for imaging biology in living cells: the resolution is well-matched to the sizes of subcellular structures, a diverse range of available fluorescent probes makes it possible to mark proteins, organelles, and other structures for imaging, and the relatively nonperturbing nature of light means that living cells can be imaged for long periods of time to follow their dynamics. Here I provide a brief introduction to using light microscopy in cell biology, with particular emphasis on factors to be considered when starting microscopy experiments. PMID:26768859

  4. Direct observation of dipolar chains in ferrofluids in zero field using cryogenic electron microscopy

    CERN Document Server

    Butter, K; Frederik, P M; Vroege, G J; Philipse, A P

    2003-01-01

    The particle structure of ferrofluids is studied in situ, by cryogenic electron microscopy, on vitrified films of iron and magnetite dispersions. By means of synthesis of iron colloids with controlled particle size and different types of surfactant, dipolar particle interactions can be varied over a broad range, which significantly influences the ferrofluid particle structure. Our experiments on iron dispersions (in contrast to magnetite dispersions) for the first time demonstrate, in ferrofluids in zero field, a transition with increasing particle size from separate particles to linear chains of particles (Butter K, Bomans P H, Frederik P M, Vroege G J and Philipse A P 2003 Nature Mater. 2 88). These chains, already predicted theoretically by de Gennes and Pincus (de Gennes P G and Pincus P A 1970 Phys. Kondens. Mater. 11 189), very much resemble the fluctuating chains found in simulations of dipolar fluids (Weis J J 1998 Mol. Phys. 93 361, Chantrell R W, Bradbury A, Popplewell J and Charles S W 1982 J. Appl...

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

    Science.gov (United States)

    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.

  6. Light Microscopy: An ongoing contemporary revolution

    CERN Document Server

    Weisenburger, Siegfried

    2014-01-01

    Optical microscopy is one of the oldest scientific instruments that is still used in forefront research. Ernst Abbe's nineteenth century formulation of the resolution limit in microscopy let generations of scientists believe that optical studies of individual molecules and resolving sub-wavelength structures were not feasible. The Nobel Prize in 2014 for super-resolution fluorescence microscopy marks a clear recognition that the old beliefs have to be revisited. In this article, we present a critical overview of various recent developments in optical microscopy. In addition to the popular super-resolution fluorescence methods, we discuss the prospects of various other techniques and imaging contrasts and consider some of the fundamental and practical challenges that lie ahead.

  7. Light-sheet optimization for microscopy

    Science.gov (United States)

    Wilding, Dean; Pozzi, Paolo; Soloviev, Oleg; Vdovin, Gleb; Verhaegen, Michel

    2016-03-01

    Aberrations, scattering and absorption degrade the performance light-sheet fluorescence microscopes (LSFM). An adaptive optics system to correct for these artefacts and to optimize the light-sheet illumination is presented. This system allows a higher axial resolution to be recovered over the field-of-view of the detection objective. It is standard selective plane illumination microscope (SPIM) configuration modified with the addition of a spatial light modulator (SLM) and a third objective for the detection of transmitted light. Optimization protocols use this transmission light allowing the extension the depth-of-field and correction of aberrations whilst retaining a thin optical section.

  8. Effect of ambient humidity on light transmittance through skin phantoms during cryogen spray cooling

    Energy Technology Data Exchange (ETDEWEB)

    Ramirez-San-Juan C, Julio [Beckman Laser Institute and Medical Clinic, University of California, Irvine, CA 92612 (United States); Department of Optics, INAOE, AP 51 and 216, CP 72000 Puebla, Pue (Mexico); Choi, Bernard [Beckman Laser Institute and Medical Clinic, University of California, Irvine, CA 92612 (United States); Franco, Walfre [Department of Mechanical Engineering, University of California, Riverside, CA 92521 (United States); Nelson, J Stuart [Beckman Laser Institute and Medical Clinic, University of California, Irvine, CA 92612 (United States); Aguilar, Guillermo [Department of Mechanical Engineering, University of California, Riverside, CA 92521 (United States)

    2006-01-07

    Cryogen spray cooling (CSC) is a technique employed to reduce the risk of epidermal damage during dermatologic laser surgery. However, while CSC protects the epidermis from non-specific thermal damage, it might reduce the effective fluence reaching the target chromophore due to scattering of light by the spray droplets and subsequent water condensation/freezing on the skin surface. The objective of this work was to study the effect of ambient humidity ({omega}) on light transmittance during CSC. An integrating sphere was employed to measure the dynamics of light transmittance through a deformable agar phantom during CSC. The study included two representative CSC spurt patterns studied using four {omega}: 57, 40, 20 and 12%. Results show that during CSC, as {omega} increased, light transmittance decreased. For the highest humidity level (57%) studied, light transmittance reached a minimum of 55% approximately 30 ms after spurt termination. In a controlled environment with {omega} = 12%, light transmittance reached a minimum of 87% approximately 30 ms after spurt termination. The reduced light transmittance immediately after spurt termination was most likely because of scattering of light caused by condensation of water vapour due to aggressive cooling of ambient air in the wake of the cryogen spurt.

  9. Development of a Calibration System for Cryogenic Light Detectors in CUPID

    Science.gov (United States)

    Luo, Meng; Kolomensky, Yury; O'Donnell, Thomas; Schmidt, Benjamin; Cupid Collaboration

    2017-01-01

    If neutrino is a Majorana particle, it is possible to observe neutrinoless double beta decay (0 νββ), whose signature is a monochromatic line at the Q-value of the decay in the energy spectrum of the two electrons. Cryogenic Underground Observatory for Rare Events (CUORE) is an experiment which aims to search for 0 νββ in 130Te with TeO2 bolometers, whose background is dominated by α particles from natural radioactivity in the detector material. CUPID (CUORE Upgrade with Particle IDentification) is the next generation experiment proposed to distinguish 0 νββ events from those of α particles with Cherenkov radiation. An important part of CUPID R&D is to design, build and characterize a calibration system that can generate a known amount of light and transport that light to the dilution refrigerator at mK temperatures. We describe the design, implementation and performance of a calibration system developed for bolometric light detectors. Preparation work includes researching and selecting a light source (LED). A transport system (optical fiber) was developed to direct the light to the coldest part of the dilution refrigerator. Additionally, the light yield attenuation of optical fiber at cryogenic temperatures was measured. This project is supported by National Science Foundation and UC-Berkeley.

  10. Noise effects and filtering in controlled light exposure microscopy

    NARCIS (Netherlands)

    R.A. Hoebe; C.J.F. van Noorden; E.M.M. Manders

    2010-01-01

    Phototoxicity and photobleaching are major limitations of fluorescence live-cell microscopy. A straightforward way to limit phototoxicity and photobleaching is reduction of the excitation light dose, but this causes loss of image quality. In confocal fluorescence microscopy, the field of view is ill

  11. Simultaneous Correlative Light and Electron Microscopy of Samples in Liquid

    NARCIS (Netherlands)

    Liv, N.

    2014-01-01

    A combined use of fluorescence and light microscopy is a powerful approach to further increase our understanding in biological systems of structure-function relations at cellular and sub-cellular levels. The power of fluorescence microscopy (FM) is to spectrally resolve and visualize individual

  12. Impact of geometry on light collection efficiency of scintillation detectors for cryogenic rare event searches

    Energy Technology Data Exchange (ETDEWEB)

    Danevich, F.A.; Kobychev, V.V. [Institute for Nuclear Research, MSP 03680 Kyiv (Ukraine); Kobychev, R.V. [Institute for Nuclear Research, MSP 03680 Kyiv (Ukraine); National Technical University of Ukraine “Kyiv Polytechnic Institute”, 03056 Kyiv (Ukraine); Kraus, H. [Department of Physics, University of Oxford, Oxford OX1 3RH (United Kingdom); Mikhailik, V.B., E-mail: vmikhai@hotmail.com [Department of Physics, University of Oxford, Oxford OX1 3RH (United Kingdom); Diamond Light Source, Didcot OX11 0DE (United Kingdom); Mokina, V.M. [Institute for Nuclear Research, MSP 03680 Kyiv (Ukraine); Solsky, I.M. [Scientific Research Company CARAT, 79031 Lviv (Ukraine)

    2014-10-01

    Simulations of photon propagation in scintillation detectors were performed with the aim to find the optimal scintillator geometry, surface treatment, and shape of external reflector in order to achieve maximum light collection efficiency for detector configurations that avoid direct optical coupling, a situation that is commonly found in cryogenic scintillating bolometers in experimental searches for double beta decay and dark matter. To evaluate the light collection efficiency of various geometrical configurations we used the ZEMAX ray-tracing software. It was found that scintillators in the shape of a triangular prism with an external mirror shaped as truncated cone gives the highest light collection efficiency. The results of the simulations were confirmed by carrying out measurements of the light collection efficiencies of CaWO{sub 4} crystal scintillators. A comparison of simulated and measured values of light output shows good agreement.

  13. Correlative cryogenic tomography of cells using light and soft x-rays.

    Science.gov (United States)

    Smith, Elizabeth A; Cinquin, Bertrand P; Do, Myan; McDermott, Gerry; Le Gros, Mark A; Larabell, Carolyn A

    2014-08-01

    Correlated imaging is the process of imaging a specimen with two complementary modalities, and then combining the two data sets to create a highly informative, composite view. A recent implementation of this concept has been the combination of soft x-ray tomography (SXT) with fluorescence cryogenic microscopy (FCM). SXT-FCM is used to visualize cells that are held in a near-native, cryopreserved. The resultant images are, therefore, highly representative of both the cellular architecture and molecular organization in vivo. SXT quantitatively visualizes the cell and sub-cellular structures; FCM images the spatial distribution of fluorescently labeled molecules. Here, we review the characteristics of SXT-FCM, and briefly discuss how this method compares with existing correlative imaging techniques. We also describe how the incorporation of a cryo-rotation stage into a cryogenic fluorescence microscope allows acquisition of fluorescence cryogenic tomography (FCT) data. FCT is optimally suited for correlation with SXT, since both techniques image the specimen in 3-D, potentially with similar, isotropic spatial resolution.

  14. Humidity-controlled preparation of frozen-hydrated biological samples for cryogenic coherent x-ray diffraction microscopy

    Energy Technology Data Exchange (ETDEWEB)

    Takayama, Yuki; Nakasako, Masayoshi [Department of Physics, Faculty of Science and Technology, Keio University, 3-14-1 Hiyoshi, Kohoku-ku, Kanagawa 223-8522 (Japan); RIKEN Harima Institute/SPring-8, 1-1-1 Kouto, Mikaduki, Sayo, Hyogo 679-5148 (Japan)

    2012-05-15

    Coherent x-ray diffraction microscopy (CXDM) has the potential to visualize the structures of micro- to sub-micrometer-sized biological particles, such as cells and organelles, at high resolution. Toward advancing structural studies on the functional states of such particles, here, we developed a system for the preparation of frozen-hydrated biological samples for cryogenic CXDM experiments. The system, which comprised a moist air generator, microscope, micro-injector mounted on a micromanipulator, custom-made sample preparation chamber, and flash-cooling device, allowed for the manipulation of sample particles in the relative humidity range of 20%-94%rh at 293 K to maintain their hydrated and functional states. Here, we report the details of the system and the operation procedure, including its application to the preparation of a frozen-hydrated chloroplast sample. Sample quality was evaluated through a cryogenic CXDM experiment conducted at BL29XUL of SPring-8. Taking the performance of the system and the quality of the sample, the system was suitable to prepare frozen-hydrated biological samples for cryogenic CXDM experiments.

  15. High-resolution light microscopy of nanoforms

    Science.gov (United States)

    Vodyanoy, Vitaly; Pustovyy, Oleg; Vainrub, Arnold

    2007-09-01

    We developed a high resolution light imaging system. Diffraction gratings with 100 nm width lines as well as less than 100 nm size features of different-shaped objects are clearly visible on a calibrated microscope test slide (Vainrub et al., Optics Letters, 2006, 31, 2855). The two-point resolution increase results from a known narrowing of the central diffraction peak for the annular aperture. Better visibility and advanced contrast of the smallest features in the image are due to enhancement of high spatial frequencies in the optical transfer function. The imaging system is portable, low energy, and battery operated. It has been adapted to use in both transmitting and reflecting light. It is particularly applicable for motile nanoform systems where structure and functions can be depicted in real time. We have isolated micrometer and submicrometer particles, termed proteons, from human and animal blood. Proteons form by reversible seeded aggregation of proteins around proteon nucleating centers (PNCs). PNCs are comprised of 1-2nm metallic nanoclusters containing 40-300 atoms. Proteons are capable of spontaneous assembling into higher nanoform systems assuming structure of complicated topology. The arrangement of complex proteon system mimics the structure of a small biological cell. It has structures that imitate membrane and nucleolus or nuclei. Some of these nanoforms are motile. They interact and divide. Complex nanoform systems can spontaneously reduce to simple proteons. The physical properties of these nanoforms could shed some light on the properties of early life forms or forms at extreme conditions.

  16. Hyperspectral microscopy to identify foodborne bacteria with optimum lighting source

    Science.gov (United States)

    Hyperspectral microscopy is an emerging technology for rapid detection of foodborne pathogenic bacteria. Since scattering spectral signatures from hyperspectral microscopic images (HMI) vary with lighting sources, it is important to select optimal lights. The objective of this study is to compare t...

  17. Thermal Balloon Endometrial Ablation: Safety Aspects Evaluated by Serosal Temperature, Light Microscopy and Electron Microscopy

    DEFF Research Database (Denmark)

    Andersen, L F; Meinert, L; Junge, Jette

    1998-01-01

    subsequent hysterectomy the extent of thermal damage into the myometrium was assessed by light and electron microscopy. RESULTS: The highest temperature measured on the uterine serosa was 39.1 degrees C. Coagulation of the myometrium adjacent to the endometrium could be demonstrated by light microscopy...... in all patients, with a maximum depth of 11.5 mm. By electron microscopy no influence of heat could be demonstrated beyond 15 mm from the endometrial surface. CONCLUSION: Up to 16 min of thermal balloon endometrial ablation therapy can destroy the endometrium and the submucosal layers. The myometrium...

  18. Thermal Balloon Endometrial Ablation: Safety Aspects Evaluated by Serosal Temperature, Light Microscopy and Electron Microscopy

    DEFF Research Database (Denmark)

    Andersen, L F; Meinert, L; Junge, Jette

    1998-01-01

    subsequent hysterectomy the extent of thermal damage into the myometrium was assessed by light and electron microscopy. RESULTS: The highest temperature measured on the uterine serosa was 39.1 degrees C. Coagulation of the myometrium adjacent to the endometrium could be demonstrated by light microscopy...... in all patients, with a maximum depth of 11.5 mm. By electron microscopy no influence of heat could be demonstrated beyond 15 mm from the endometrial surface. CONCLUSION: Up to 16 min of thermal balloon endometrial ablation therapy can destroy the endometrium and the submucosal layers. The myometrium...

  19. Thermal balloon endometrial ablation: safety aspects evaluated by serosal temperature, light microscopy and electron microscopy

    DEFF Research Database (Denmark)

    Andersen, L F; Meinert, L; Rygaard, Carsten

    1998-01-01

    subsequent hysterectomy the extent of thermal damage into the myometrium was assessed by light and electron microscopy. RESULTS: The highest temperature measured on the uterine serosa was 39.1 degrees C. Coagulation of the myometrium adjacent to the endometrium could be demonstrated by light microscopy...... in all patients, with a maximum depth of 11.5 mm. By electron microscopy no influence of heat could be demonstrated beyond 15 mm from the endometrial surface. CONCLUSION: Up to 16 min of thermal balloon endometrial ablation therapy can destroy the endometrium and the submucosal layers. The myometrium...

  20. Near-infrared branding efficiently correlates light and electron microscopy.

    Science.gov (United States)

    Bishop, Derron; Nikić, Ivana; Brinkoetter, Mary; Knecht, Sharmon; Potz, Stephanie; Kerschensteiner, Martin; Misgeld, Thomas

    2011-06-05

    The correlation of light and electron microscopy of complex tissues remains a major challenge. Here we report near-infrared branding (NIRB), which facilitates such correlation by using a pulsed, near-infrared laser to create defined fiducial marks in three dimensions in fixed tissue. As these marks are fluorescent and can be photo-oxidized to generate electron contrast, they can guide re-identification of previously imaged structures as small as dendritic spines by electron microscopy.

  1. [Study of human leucine-rich amelogenin peptide and its regulation of mineralization by cryogenic transmission electron microscopy].

    Science.gov (United States)

    Kun, Tian; Xiaoyun, Feng; Qin, Du; Chuhang, Liao; Xiaohua, Ren

    2017-02-01

    Recombinant human leucine-rich amelogenin peptide (LRAP) was studied by cryogenic transmission electron microscopy (TEM); evaluation focused on its self-assembly and crystal growth in vitro. Human LRAP was recombined through prokaryotic expression vector pCold-SUMO and transformed into Escherichia coli BL21plys to acquire purified proteins. Cryogen TEM recorded assembly and self-assembling of LRAP from pH 3.5 to pH 8.0, and the hydroxyapatite crystal growth in the mixture of LRAP protein solution and artificial saliva was observed using TEM and selected area electron diffraction. More than 90% purity LRAP was expressed, purified and identified as described in methods. LRAP linked into oligomers, nanospheres, nanochains, and microribbons, whereas pH value increased from 3.5 to 8.0. Mature hydroxyapatite crystal growth was guided in artificial saliva filled with calcium phosphate. LRAP is simplified amelogenin functional domain and conserved the basic characters of amelogenin such as self-assembling and inducing crystallization along c axis. In the area of acellular synthesis of hydroxyapatite using extracellular enamel matrix protein, LRAP is one of candidate repair materials for irregular hard tissue defection.
.

  2. eduSPIM: Light Sheet Microscopy in the Museum

    Science.gov (United States)

    Schmid, Benjamin; Weber, Michael; Huisken, Jan

    2016-01-01

    Light Sheet Microscopy in the Museum Light sheet microscopy (or selective plane illumination microscopy) is an important imaging technique in the life sciences. At the same time, this technique is also ideally suited for community outreach projects, because it produces visually appealing, highly dynamic images of living organisms and its working principle can be understood with basic optics knowledge. Still, the underlying concepts are widely unknown to the non-scientific public. On the occasion of the UNESCO International Year of Light, a technical museum in Dresden, Germany, launched a special, interactive exhibition. We built a fully functional, educational selective plane illumination microscope (eduSPIM) to demonstrate how developments in microscopy promote discoveries in biology. Design Principles of an Educational Light Sheet Microscope To maximize educational impact, we radically reduced a standard light sheet microscope to its essential components without compromising functionality and incorporated stringent safety concepts beyond those needed in the lab. Our eduSPIM system features one illumination and one detection path and a sealed sample chamber. We image fixed zebrafish embryos with fluorescent vasculature, because the structure is meaningful to laymen and visualises the optical principles of light sheet microscopy. Via a simplified interface, visitors acquire fluorescence and transmission data simultaneously. The eduSPIM Design Is Tailored Easily to Fit Numerous Applications The universal concepts presented here may also apply to other scientific approaches that are communicated to laymen in interactive settings. The specific eduSPIM design is adapted easily for various outreach and teaching activities. eduSPIM may even prove useful for labs needing a simple SPIM. A detailed parts list and schematics to rebuild eduSPIM are provided. PMID:27560188

  3. A Stochastic Hill Climbing Approach for Simultaneous 2D Alignment and Clustering of Cryogenic Electron Microscopy Images.

    Science.gov (United States)

    Reboul, Cyril F; Bonnet, Frederic; Elmlund, Dominika; Elmlund, Hans

    2016-06-07

    A critical step in the analysis of novel cryogenic electron microscopy (cryo-EM) single-particle datasets is the identification of homogeneous subsets of images. Methods for solving this problem are important for data quality assessment, ab initio 3D reconstruction, and analysis of population diversity due to the heterogeneous nature of macromolecules. Here we formulate a stochastic algorithm for identification of homogeneous subsets of images. The purpose of the method is to generate improved 2D class averages that can be used to produce a reliable 3D starting model in a rapid and unbiased fashion. We show that our method overcomes inherent limitations of widely used clustering approaches and proceed to test the approach on six publicly available experimental cryo-EM datasets. We conclude that, in each instance, ab initio 3D reconstructions of quality suitable for initialization of high-resolution refinement are produced from the cluster centers.

  4. New resolving power for light microscopy: applications to neurobiology.

    Science.gov (United States)

    Dani, Adish; Huang, Bo

    2010-10-01

    The recent invention of super-resolution fluorescence microscopy brings more than an order of magnitude gain in the spatial resolution of light microscopy. New opportunities keep emerging with the multicolor, three-dimensional, and live imaging functionalities gained in the past three years. The power of this technology has been demonstrated by imaging the organization of organelles and molecular complexes, with recent applications increasingly showing its potential in neurobiology. These developments are exemplified by the visualization of components inside dendritic spines to fine morphologies of neurons. In combination with correlative electron microscopy, functional imaging, and electrical/optogenetic stimulation tools, super-resolution fluorescence microscopy has the potential to provide further insights ranging from the molecular details of neurons up to the functional mechanisms of neuronal circuits.

  5. Miniaturized modules for light sheet microscopy with low chromatic aberration.

    Science.gov (United States)

    Bruns, T; Bauer, M; Bruns, S; Meyer, H; Kubin, D; Schneckenburger, H

    2016-12-01

    Two miniaturized fibre-coupled modules for light sheet-based microscopy are described and compared with respect to image quality, chromatic aberration and beam alignment. Whereas in one module the light sheet is created by an achromatic cylindrical lens, reflection by a spherical mirror and concomitant astigmatic distortion are used to create the light sheet in the second module. Test experiments with fluorescent dyes in solution and multicellular tumour spheroids are reported, and some details on construction are given for both systems. Both modules are optimized for imaging individual cell layers of 3D biological samples and can be adapted to fit commercial microscopes.

  6. Fundamentals of fluorescence microscopy exploring life with light

    CERN Document Server

    Mondal, Partha Pratim

    2014-01-01

    This book starts at an introductory level and leads reader to the most advanced developments in fluorescence imaging and super-resolution techniques that have enabled the emergence of new disciplines such as nanobioimaging, multiphoton microscopy, photodynamic therapy, nanometrology and nanosensors. The interdisciplinary subject of fluorescence microscopy and imaging requires complete knowledge of imaging optics and molecular physics. So, this book approaches the subject by introducing optical imaging concepts before going deep into the advanced imaging systems and their applications. Molecular orbital theory forms the basis for understanding fluorescent molecules and thereby facilitates complete explanation of light-matter interaction at the geometrical focus. The two disciplines have some overlap since light controls the states of molecules and conversely, molecular states control the emitted light. These two mechanisms together determine essential fluorescence  factors and phenomena such as, molecular cro...

  7. Imaging of Active Microwave Devices at Cryogenic Temperatures using Scanning Near-Field Microwave Microscopy

    Science.gov (United States)

    Thanawalla, Ashfaq S.; Dutta, S. K.; Vlahacos, C. P.; Steinhauer, D. E.; Feenstra, B. J.; Anlage, Steven M.; Wellstood, F. C.

    1998-03-01

    The ability to image electric fields in operating microwave devices is interesting both from the fundamental point of view and for diagnostic purposes. To that end we have constructed a scanning near-field microwave microscope which uses an open-ended coaxial probe and operates at cryogenic temperatures.(For related publications see: C. P. Vlahacos, R. C. Black, S. M. Anlage, A. Amar and F. C. Wellstood, Appl. Phys. Lett. 69), 3274 (1996) and S. M. Anlage, C. P. Vlahacos, Sudeep Dutta and F. C. Wellstood, IEEE Trans. Appl. Supercond. 7, 3686 (1997). Using this system we have imaged electric fields generated by both normal metal and superconducting microstrip resonators at temperatures ranging from 77 K to 300 K. We will present images and discuss our results including observations of clear standing wave patterns at the fundamental resonant frequency and an increased quality factor of the resonators at low temperatures.

  8. Contributed review: Review of integrated correlative light and electron microscopy.

    Science.gov (United States)

    Timmermans, F J; Otto, C

    2015-01-01

    New developments in the field of microscopy enable to acquire increasing amounts of information from large sample areas and at an increased resolution. Depending on the nature of the technique, the information may reveal morphological, structural, chemical, and still other sample characteristics. In research fields, such as cell biology and materials science, there is an increasing demand to correlate these individual levels of information and in this way to obtain a better understanding of sample preparation and specific sample properties. To address this need, integrated systems were developed that combine nanometer resolution electron microscopes with optical microscopes, which produce chemically or label specific information through spectroscopy. The complementary information from electron microscopy and light microscopy presents an opportunity to investigate a broad range of sample properties in a correlated fashion. An important part of correlating the differences in information lies in bridging the different resolution and image contrast features. The trend to analyse samples using multiple correlated microscopes has resulted in a new research field. Current research is focused, for instance, on (a) the investigation of samples with nanometer scale distribution of inorganic and organic materials, (b) live cell analysis combined with electron microscopy, and (c) in situ spectroscopic and electron microscopy analysis of catalytic materials, but more areas will benefit from integrated correlative microscopy.

  9. Contributed Review: Review of integrated correlative light and electron microscopy

    Energy Technology Data Exchange (ETDEWEB)

    Timmermans, F. J.; Otto, C. [Medical Cell Biophysics Group, MIRA Institute, University of Twente, P.O. Box 217, 7500 AE Enschede (Netherlands)

    2015-01-15

    New developments in the field of microscopy enable to acquire increasing amounts of information from large sample areas and at an increased resolution. Depending on the nature of the technique, the information may reveal morphological, structural, chemical, and still other sample characteristics. In research fields, such as cell biology and materials science, there is an increasing demand to correlate these individual levels of information and in this way to obtain a better understanding of sample preparation and specific sample properties. To address this need, integrated systems were developed that combine nanometer resolution electron microscopes with optical microscopes, which produce chemically or label specific information through spectroscopy. The complementary information from electron microscopy and light microscopy presents an opportunity to investigate a broad range of sample properties in a correlated fashion. An important part of correlating the differences in information lies in bridging the different resolution and image contrast features. The trend to analyse samples using multiple correlated microscopes has resulted in a new research field. Current research is focused, for instance, on (a) the investigation of samples with nanometer scale distribution of inorganic and organic materials, (b) live cell analysis combined with electron microscopy, and (c) in situ spectroscopic and electron microscopy analysis of catalytic materials, but more areas will benefit from integrated correlative microscopy.

  10. Light Microscopy and Polarized Microscopy: A Dermatological Tool to Diagnose Gray Hair Syndromes.

    Science.gov (United States)

    Chandravathi, P L; Karani, Hetal Deepak; Siddaiahgari, Sirisha Rani; Lingappa, Lokesh

    2017-01-01

    Gray hair syndromes are rare syndromes which have an autosomal recessive inheritance and are characterized by pigmentary dilution of skin and hair, defects in immunological function, and nervous system defects. They comprise three disorders namely Chediak-Higashi syndrome (CHS), Griscelli syndrome (GPS), and Elejalde syndrome. Clinically, it is difficult to distinguish these disorders as their clinical features may overlap. Hence, to make a correct diagnosis and differentiate between CHS and GPS light microscopic examination of skin and hair shafts as well as peripheral blood smear evaluations should be done. In cases where the diagnosis is not possible chromosomal analysis for specific mutations can be done. In resource-poor settings where chromosomal analysis is not possible, and light microscopy findings are inconclusive, polarized microscopy can serve as a useful tool to distinguish between CHS and GPS. We report three cases with gray hair syndromes where the diagnosis on light microscopy and polarized microscopy of hair shaft correlated with the bone marrow examination findings and chromosomal analysis, thus emphasizing the importance of a noninvasive, cost-effective, and time-saving alternative in the diagnosis of these syndromes.

  11. High-resolution imaging by scanning electron microscopy of semithin sections in correlation with light microscopy.

    Science.gov (United States)

    Koga, Daisuke; Kusumi, Satoshi; Shodo, Ryusuke; Dan, Yukari; Ushiki, Tatsuo

    2015-12-01

    In this study, we introduce scanning electron microscopy (SEM) of semithin resin sections. In this technique, semithin sections were adhered on glass slides, stained with both uranyl acetate and lead citrate, and observed with a backscattered electron detector at a low accelerating voltage. As the specimens are stained in the same manner as conventional transmission electron microscopy (TEM), the contrast of SEM images of semithin sections was similar to TEM images of ultrathin sections. Using this technique, wide areas of semithin sections were also observed by SEM, without the obstruction of grids, which was inevitable for traditional TEM. This study also applied semithin section SEM to correlative light and electron microscopy. Correlative immunofluorescence microscopy and immune-SEM were performed in semithin sections of LR white resin-embedded specimens using a FluoroNanogold-labeled secondary antibody. Because LR white resin is hydrophilic and electron stable, this resin is suitable for immunostaining and SEM observation. Using correlative microscopy, the precise localization of the primary antibody was demonstrated by fluorescence microscopy and SEM. This method has great potential for studies examining the precise localization of molecules, including Golgi- and ER-associated proteins, in correlation with LM and SEM.

  12. Light microscopy mapping of connections in the intact brain.

    Science.gov (United States)

    Kim, Sung-Yon; Chung, Kwanghun; Deisseroth, Karl

    2013-12-01

    Mapping of neural connectivity across the mammalian brain is a daunting and exciting prospect. Current approaches can be divided into three classes: macroscale, focusing on coarse inter-regional connectivity; mesoscale, involving a finer focus on neurons and projections; and microscale, reconstructing full details of all synaptic contacts. It remains to be determined how to bridge the datasets or insights from the different levels of study. Here we review recent light-microscopy-based approaches that may help in integration across scales.

  13. Structured light sheet fluorescence microscopy based on four beam interference.

    Science.gov (United States)

    Lei, Ming; Zumbusch, Andreas

    2010-08-30

    A 3D structured light sheet microscope using a four-faceted symmetric pyramid is presented. The sample is illuminated by the resulting four beam interference field. This approach combines advantages of standing wave and structured illumination microscopy. Examples of micrographs of fluorescently labeled Chinese hamster ovary (CHO) cells as well as of the compound eyes of drosophila are shown and the optical sectioning ability of our system is demonstrated. The capabilities and the limitations of the scheme are discussed.

  14. Visualizing aquatic bacteria by light and transmission electron microscopy.

    Science.gov (United States)

    Silva, Thiago P; Noyma, Natália P; Duque, Thabata L A; Gamalier, Juliana P; Vidal, Luciana O; Lobão, Lúcia M; Chiarini-Garcia, Hélio; Roland, Fábio; Melo, Rossana C N

    2014-01-01

    The understanding of the functional role of aquatic bacteria in microbial food webs is largely dependent on methods applied to the direct visualization and enumeration of these organisms. While the ultrastructure of aquatic bacteria is still poorly known, routine observation of aquatic bacteria by light microscopy requires staining with fluorochromes, followed by filtration and direct counting on filter surfaces. Here, we used a new strategy to visualize and enumerate aquatic bacteria by light microscopy. By spinning water samples from varied tropical ecosystems in a cytocentrifuge, we found that bacteria firmly adhere to regular slides, can be stained by fluorochoromes with no background formation and fast enumerated. Significant correlations were found between the cytocentrifugation and filter-based methods. Moreover, preparations through cytocentrifugation were more adequate for bacterial viability evaluation than filter-based preparations. Transmission electron microscopic analyses revealed a morphological diversity of bacteria with different internal and external structures, such as large variation in the cell envelope and capsule thickness, and presence or not of thylakoid membranes. Our results demonstrate that aquatic bacteria represent an ultrastructurally diverse population and open avenues for easy handling/quantification and better visualization of bacteria by light microscopy without the need of filter membranes.

  15. Cryogenic refrigeration requirements for superconducting insertion devices in a light source

    Energy Technology Data Exchange (ETDEWEB)

    Green, Michael A.; Green, Michael A.; Green, Michael A.

    2003-08-15

    This report discusses cryogenic cooling superconducting insertion devices for modern light sources. The introductory part of the report discusses the difference between wiggler and undulators and how the bore temperature may affect the performance of the magnets. The steps one would take to reduce the gap between the cold magnet pole are discussed. One section of the report is devoted to showing how one would calculate the heat that enters the device. Source of heat include, heat entering through the vacuum chamber, heating due to stray electrons and synchrotron radiation, heating due to image current on the bore, heat flow by conduction and radiation, and heat transfer into the cryostat through the magnet leads. A section of the report is devoted to cooling options such as small cryo-cooler and larger conventional helium refrigerators. This section contains a discussion as to when it is appropriate to use small coolers that do not have J-T circuits. Candidate small cryo-coolers are discussed in this section of the report. Cooling circuits for cooling with a conventional refrigerator are also discussed. A section of the report is devoted to vibration isolation and how this may affect how the cooling is attached to the device. Vibration isolation using straps is compared to vibration isolation using helium heat pipes. The vibration isolation of a conventional refrigeration system is also discussed. Finally, the cool down of an insertion device is discussed. The device can either be cooled down using liquid cryogenic nitrogen and liquid helium or by using the cooler used to keep the devices cold over the long haul.

  16. Cryogenic x-ray diffraction microscopy utilizing high-pressure cryopreservation.

    Science.gov (United States)

    Lima, Enju; Chushkin, Yuriy; van der Linden, Peter; Kim, Chae Un; Zontone, Federico; Carpentier, Philippe; Gruner, Sol M; Pernot, Petra

    2014-10-01

    We present cryo x-ray diffraction microscopy of high-pressure-cryofixed bacteria and report high-convergence imaging with multiple image reconstructions. Hydrated D. radiodurans cells were cryofixed at 200 MPa pressure into ∼10-μm-thick water layers and their unstained, hydrated cellular environments were imaged by phasing diffraction patterns, reaching sub-30-nm resolutions with hard x-rays. Comparisons were made with conventional ambient-pressure-cryofixed samples, with respect to both coherent small-angle x-ray scattering and the image reconstruction. The results show a correlation between the level of background ice signal and phasing convergence, suggesting that phasing difficulties with frozen-hydrated specimens may be caused by high-background ice scattering.

  17. Topography and refractometry of nanostructures using spatial light interference microscopy.

    Science.gov (United States)

    Wang, Zhuo; Chun, Ik Su; Li, Xiuling; Ong, Zhun-Yong; Pop, Eric; Millet, Larry; Gillette, Martha; Popescu, Gabriel

    2010-01-15

    Spatial light interference microscopy (SLIM) is a novel method developed in our laboratory that provides quantitative phase images of transparent structures with a 0.3 nm spatial and 0.03 nm temporal accuracy owing to the white light illumination and its common path interferometric geometry. We exploit these features and demonstrate SLIM's ability to perform topography at a single atomic layer in graphene. Further, using a decoupling procedure that we developed for cylindrical structures, we extract the axially averaged refractive index of semiconductor nanotubes and a neurite of a live hippocampal neuron in culture. We believe that this study will set the basis for novel high-throughput topography and refractometry of man-made and biological nanostructures.

  18. White-light interferometric microscopy for wafer defect inspection

    Science.gov (United States)

    Zhou, Renjie; Edwards, Christopher; Bryniarski, Casey; Dallmann, Marjorie F.; Popescu, Gabriel; Goddard, Lynford L.

    2015-03-01

    White-light imaging systems are free of laser-speckle. Thus, they offer high sensitivity for optical defect metrology, especially when used with interferometry based quantitative phase imaging. This can be a potential solution for wafer inspection beyond the 9 nm node. Recently, we built a white-light epi-illumination diffraction phase microscopy (epi-wDPM) for wafer defect inspection. The system is also equipped with an XYZ scanning stage and real-time processing. Preliminary results have demonstrated detection of 15 nm by 90 nm in a 9 nm node densely patterned wafer with bright-field imaging. Currently, we are implementing phase imaging with epi-wDPM for additional sensitivity.

  19. Sea Spray Aerosol Structure and Composition Using Cryogenic Transmission Electron Microscopy

    Energy Technology Data Exchange (ETDEWEB)

    Patterson, Joseph P.; Collins, Douglas B.; Michaud, Jennifer M.; Axson, Jessica L.; Sultana, Camile M.; Moser, Trevor; Dommer, Abigail C.; Conner, Jack; Grassian, Vicki H.; Stokes, M. Dale; Deane, Grant B.; Evans, James E.; Burkart, Michael D.; Prather, Kimberly A.; Gianneschi, Nathan C.

    2016-01-27

    The surface properties of atmospheric aerosol particles largely control their impact on climate by affecting their ability to uptake water, react heterogeneously, and nucleate ice in clouds. However, in the vacuum of a conventional electron microscope, the native surface structure often undergoes chemical rearrangement resulting in surfaces that are quite different from their atmospheric configurations. Herein, we report the development of a cryo-TEM approach where sea spray aerosol particles are flash frozen in their native state and then probed by electron microscopy. This unique approach allows for the detection of not only mixed salts, but also soft materials including whole hydrated bacteria, diatoms, virus particles, marine vesicles, as well as gel networks within hydrated salt droplets. We anticipate this method will open up a new avenue of analysis for aerosol particles, not only for ocean-derived aerosols, but for those produced from other sources where there is interest in the transfer of organic or biological species from the biosphere to the atmosphere.

  20. Invited Review Article: Advanced light microscopy for biological space research

    Science.gov (United States)

    De Vos, Winnok H.; Beghuin, Didier; Schwarz, Christian J.; Jones, David B.; van Loon, Jack J. W. A.; Bereiter-Hahn, Juergen; Stelzer, Ernst H. K.

    2014-10-01

    As commercial space flights have become feasible and long-term extraterrestrial missions are planned, it is imperative that the impact of space travel and the space environment on human physiology be thoroughly characterized. Scrutinizing the effects of potentially detrimental factors such as ionizing radiation and microgravity at the cellular and tissue level demands adequate visualization technology. Advanced light microscopy (ALM) is the leading tool for non-destructive structural and functional investigation of static as well as dynamic biological systems. In recent years, technological developments and advances in photochemistry and genetic engineering have boosted all aspects of resolution, readout and throughput, rendering ALM ideally suited for biological space research. While various microscopy-based studies have addressed cellular response to space-related environmental stressors, biological endpoints have typically been determined only after the mission, leaving an experimental gap that is prone to bias results. An on-board, real-time microscopical monitoring device can bridge this gap. Breadboards and even fully operational microscope setups have been conceived, but they need to be rendered more compact and versatile. Most importantly, they must allow addressing the impact of gravity, or the lack thereof, on physiologically relevant biological systems in space and in ground-based simulations. In order to delineate the essential functionalities for such a system, we have reviewed the pending questions in space science, the relevant biological model systems, and the state-of-the art in ALM. Based on a rigorous trade-off, in which we recognize the relevance of multi-cellular systems and the cellular microenvironment, we propose a compact, but flexible concept for space-related cell biological research that is based on light sheet microscopy.

  1. Invited Review Article: Advanced light microscopy for biological space research

    Energy Technology Data Exchange (ETDEWEB)

    De Vos, Winnok H., E-mail: winnok.devos@uantwerpen.be [Laboratory of Cell Biology and Histology, Department of Veterinary Sciences, University of Antwerp, Antwerp (Belgium); Cell Systems and Imaging Research Group, Department of Molecular Biotechnology, Ghent University, Ghent (Belgium); Beghuin, Didier [Lambda-X, Nivelles (Belgium); Schwarz, Christian J. [European Space Agency (ESA), ESTEC, TEC-MMG, Noordwijk (Netherlands); Jones, David B. [Institute for Experimental Orthopaedics and Biomechanics, Philipps University, Marburg (Germany); Loon, Jack J. W. A. van [Department of Oral and Maxillofacial Surgery/Oral Pathology, VU University Medical Center and Department of Oral Cell Biology, Academic Centre for Dentistry Amsterdam, Amsterdam (Netherlands); Bereiter-Hahn, Juergen; Stelzer, Ernst H. K. [Physical Biology, BMLS (FB15, IZN), Goethe University, Frankfurt am Main (Germany)

    2014-10-15

    As commercial space flights have become feasible and long-term extraterrestrial missions are planned, it is imperative that the impact of space travel and the space environment on human physiology be thoroughly characterized. Scrutinizing the effects of potentially detrimental factors such as ionizing radiation and microgravity at the cellular and tissue level demands adequate visualization technology. Advanced light microscopy (ALM) is the leading tool for non-destructive structural and functional investigation of static as well as dynamic biological systems. In recent years, technological developments and advances in photochemistry and genetic engineering have boosted all aspects of resolution, readout and throughput, rendering ALM ideally suited for biological space research. While various microscopy-based studies have addressed cellular response to space-related environmental stressors, biological endpoints have typically been determined only after the mission, leaving an experimental gap that is prone to bias results. An on-board, real-time microscopical monitoring device can bridge this gap. Breadboards and even fully operational microscope setups have been conceived, but they need to be rendered more compact and versatile. Most importantly, they must allow addressing the impact of gravity, or the lack thereof, on physiologically relevant biological systems in space and in ground-based simulations. In order to delineate the essential functionalities for such a system, we have reviewed the pending questions in space science, the relevant biological model systems, and the state-of-the art in ALM. Based on a rigorous trade-off, in which we recognize the relevance of multi-cellular systems and the cellular microenvironment, we propose a compact, but flexible concept for space-related cell biological research that is based on light sheet microscopy.

  2. Invited review article: Advanced light microscopy for biological space research.

    Science.gov (United States)

    De Vos, Winnok H; Beghuin, Didier; Schwarz, Christian J; Jones, David B; van Loon, Jack J W A; Bereiter-Hahn, Juergen; Stelzer, Ernst H K

    2014-10-01

    As commercial space flights have become feasible and long-term extraterrestrial missions are planned, it is imperative that the impact of space travel and the space environment on human physiology be thoroughly characterized. Scrutinizing the effects of potentially detrimental factors such as ionizing radiation and microgravity at the cellular and tissue level demands adequate visualization technology. Advanced light microscopy (ALM) is the leading tool for non-destructive structural and functional investigation of static as well as dynamic biological systems. In recent years, technological developments and advances in photochemistry and genetic engineering have boosted all aspects of resolution, readout and throughput, rendering ALM ideally suited for biological space research. While various microscopy-based studies have addressed cellular response to space-related environmental stressors, biological endpoints have typically been determined only after the mission, leaving an experimental gap that is prone to bias results. An on-board, real-time microscopical monitoring device can bridge this gap. Breadboards and even fully operational microscope setups have been conceived, but they need to be rendered more compact and versatile. Most importantly, they must allow addressing the impact of gravity, or the lack thereof, on physiologically relevant biological systems in space and in ground-based simulations. In order to delineate the essential functionalities for such a system, we have reviewed the pending questions in space science, the relevant biological model systems, and the state-of-the art in ALM. Based on a rigorous trade-off, in which we recognize the relevance of multi-cellular systems and the cellular microenvironment, we propose a compact, but flexible concept for space-related cell biological research that is based on light sheet microscopy.

  3. 3D correlative light and electron microscopy of cultured cells using serial blockface scanning electron microscopy

    Science.gov (United States)

    Lerner, Thomas R.; Burden, Jemima J.; Nkwe, David O.; Pelchen-Matthews, Annegret; Domart, Marie-Charlotte; Durgan, Joanne; Weston, Anne; Jones, Martin L.; Peddie, Christopher J.; Carzaniga, Raffaella; Florey, Oliver; Marsh, Mark; Gutierrez, Maximiliano G.

    2017-01-01

    ABSTRACT The processes of life take place in multiple dimensions, but imaging these processes in even three dimensions is challenging. Here, we describe a workflow for 3D correlative light and electron microscopy (CLEM) of cell monolayers using fluorescence microscopy to identify and follow biological events, combined with serial blockface scanning electron microscopy to analyse the underlying ultrastructure. The workflow encompasses all steps from cell culture to sample processing, imaging strategy, and 3D image processing and analysis. We demonstrate successful application of the workflow to three studies, each aiming to better understand complex and dynamic biological processes, including bacterial and viral infections of cultured cells and formation of entotic cell-in-cell structures commonly observed in tumours. Our workflow revealed new insight into the replicative niche of Mycobacterium tuberculosis in primary human lymphatic endothelial cells, HIV-1 in human monocyte-derived macrophages, and the composition of the entotic vacuole. The broad application of this 3D CLEM technique will make it a useful addition to the correlative imaging toolbox for biomedical research. PMID:27445312

  4. 3D correlative light and electron microscopy of cultured cells using serial blockface scanning electron microscopy.

    Science.gov (United States)

    Russell, Matthew R G; Lerner, Thomas R; Burden, Jemima J; Nkwe, David O; Pelchen-Matthews, Annegret; Domart, Marie-Charlotte; Durgan, Joanne; Weston, Anne; Jones, Martin L; Peddie, Christopher J; Carzaniga, Raffaella; Florey, Oliver; Marsh, Mark; Gutierrez, Maximiliano G; Collinson, Lucy M

    2017-01-01

    The processes of life take place in multiple dimensions, but imaging these processes in even three dimensions is challenging. Here, we describe a workflow for 3D correlative light and electron microscopy (CLEM) of cell monolayers using fluorescence microscopy to identify and follow biological events, combined with serial blockface scanning electron microscopy to analyse the underlying ultrastructure. The workflow encompasses all steps from cell culture to sample processing, imaging strategy, and 3D image processing and analysis. We demonstrate successful application of the workflow to three studies, each aiming to better understand complex and dynamic biological processes, including bacterial and viral infections of cultured cells and formation of entotic cell-in-cell structures commonly observed in tumours. Our workflow revealed new insight into the replicative niche of Mycobacterium tuberculosis in primary human lymphatic endothelial cells, HIV-1 in human monocyte-derived macrophages, and the composition of the entotic vacuole. The broad application of this 3D CLEM technique will make it a useful addition to the correlative imaging toolbox for biomedical research. © 2017. Published by The Company of Biologists Ltd.

  5. Structure Prediction of Self-Assembled Dye Aggregates from Cryogenic Transmission Electron Microscopy, Molecular Mechanics, and Theory of Optical Spectra.

    Science.gov (United States)

    Friedl, Christian; Renger, Thomas; Berlepsch, Hans V; Ludwig, Kai; Schmidt Am Busch, Marcel; Megow, Jörg

    2016-09-01

    Cryogenic transmission electron microscopy (cryo-TEM) studies suggest that TTBC molecules self-assemble in aqueous solution to form single-walled tubes with a diameter of about 35 Å. In order to reveal the arrangement and mutual orientations of the individual molecules in the tube, we combine information from crystal structure data of this dye with a calculation of linear absorbance and linear dichroism spectra and molecular dynamics simulations. We start with wrapping crystal planes in different directions to obtain tubes of suitable diameter. This set of tube models is evaluated by comparing the resulting optical spectra with experimental data. The tubes that can explain the spectra are investigated further by molecular dynamics simulations, including explicit solvent molecules. From the trajectories of the most stable tube models, the short-range ordering of the dye molecules is extracted and the optimization of the structure is iteratively completed. The final structural model is a tube of rings with 6-fold rotational symmetry, where neighboring rings are rotated by 30° and the transition dipole moments of the chromophores form an angle of 74° with respect to the symmetry axis of the tube. This model is in agreement with cryo-TEM images and can explain the optical spectra, consisting of a sharp red-shifted J-band that is polarized parallel to to the symmetry axis of the tube and a broad blue-shifted H-band polarized perpendicular to this axis. The general structure of the homogeneous spectrum of this hybrid HJ-aggregate is described by an analytical model that explains the difference in redistribution of oscillator strength inside the vibrational manifolds of the J- and H-bands and the relative intensities and excitation energies of those bands. In addition to the particular system investigated here, the present methodology can be expected to aid the structure prediction for a wide range of self-assembled dye aggregates.

  6. Coherent imaging with incoherent light in digital holographic microscopy

    Science.gov (United States)

    Chmelik, Radim

    2012-01-01

    Digital holographic microscope (DHM) allows for imaging with a quantitative phase contrast. In this way it becomes an important instrument, a completely non-invasive tool for a contrast intravital observation of living cells and a cell drymass density distribution measurement. A serious drawback of current DHMs is highly coherent illumination which makes the lateral resolution worse and impairs the image quality by a coherence noise and a parasitic interference. An uncompromising solution to this problem can be found in the Leith concept of incoherent holography. An off-axis hologram can be formed with arbitrary degree of light coherence in systems equipped with an achromatic interferometer and thus the resolution and the image quality typical for an incoherent-light wide-field microscopy can be achieved. In addition, advanced imaging modes based on limited coherence can be utilized. The typical example is a coherence-gating effect which provides a finite axial resolution and makes DHM image similar to that of a confocal microscope. These possibilities were described theoretically using the formalism of three-dimensional coherent transfer functions and proved experimentally by the coherence-controlled holographic microscope which is DHM based on the Leith achromatic interferometer. Quantitative-phase-contrast imaging is demonstrated with incoherent light by the living cancer cells observation and their motility evaluation. The coherence-gating effect was proved by imaging of model samples through a scattering layer and living cells inside an opalescent medium.

  7. Vacuum ultra-violet and ultra-violet scintillation light detection by means of silicon photomultipliers at cryogenic temperature

    Energy Technology Data Exchange (ETDEWEB)

    Falcone, A., E-mail: andrea.falcone@pv.infn.it [University of Pavia, via Bassi, 6, 27100 Pavia (Italy); INFN Sezione di Pavia, via Bassi, 6, 27100 Pavia (Italy); Bertoni, R. [INFN Sezione di Milano Bicocca, Piazza della Scienza, 3, 20126 Milano (Italy); Boffelli, F. [University of Pavia, via Bassi, 6, 27100 Pavia (Italy); INFN Sezione di Pavia, via Bassi, 6, 27100 Pavia (Italy); Bonesini, M. [INFN Sezione di Milano Bicocca, Piazza della Scienza, 3, 20126 Milano (Italy); Cervi, T. [University of Pavia, via Bassi, 6, 27100 Pavia (Italy); Menegolli, A. [University of Pavia, via Bassi, 6, 27100 Pavia (Italy); INFN Sezione di Pavia, via Bassi, 6, 27100 Pavia (Italy); Montanari, C.; Prata, M.C.; Rappoldi, A.; Raselli, G.L.; Rossella, M.; Simonetta, M. [INFN Sezione di Pavia, via Bassi, 6, 27100 Pavia (Italy); Spanu, M. [University of Pavia, via Bassi, 6, 27100 Pavia (Italy); Torti, M. [University of Pavia, via Bassi, 6, 27100 Pavia (Italy); INFN Sezione di Pavia, via Bassi, 6, 27100 Pavia (Italy); Zani, A. [INFN Sezione di Pavia, via Bassi, 6, 27100 Pavia (Italy)

    2015-07-01

    We tested the performance of two types of silicon photomultipliers, AdvanSiD ASD-NUV-SiPM3S-P and Hamamatsu 3×3 MM-50 UM VUV2, both at room (300 K) and at liquid nitrogen (77 K) temperature: breakdown voltage, quenching resistance, signal shape, gain and dark counts rate have been studied as function of temperature. The response of the devices to ultra-violet light is also studied. - Highlights: • We tested 2 SiPMs both at room and at cryogenic temperature. • Breakdown voltage, quenching resistance, gain and dark rate were measured. • Efficiency for VUV light detection was measured.

  8. Advanced light microscopy core facilities: Balancing service, science and career.

    Science.gov (United States)

    Ferrando-May, Elisa; Hartmann, Hella; Reymann, Jürgen; Ansari, Nariman; Utz, Nadine; Fried, Hans-Ulrich; Kukat, Christian; Peychl, Jan; Liebig, Christian; Terjung, Stefan; Laketa, Vibor; Sporbert, Anje; Weidtkamp-Peters, Stefanie; Schauss, Astrid; Zuschratter, Werner; Avilov, Sergiy

    2016-06-01

    Core Facilities (CF) for advanced light microscopy (ALM) have become indispensable support units for research in the life sciences. Their organizational structure and technical characteristics are quite diverse, although the tasks they pursue and the services they offer are similar. Therefore, throughout Europe, scientists from ALM-CFs are forming networks to promote interactions and discuss best practice models. Here, we present recommendations for ALM-CF operations elaborated by the workgroups of the German network of ALM-CFs, German Bio-Imaging (GerBI). We address technical aspects of CF planning and instrument maintainance, give advice on the organization and management of an ALM-CF, propose a scheme for the training of CF users, and provide an overview of current resources for image processing and analysis. Further, we elaborate on the new challenges and opportunities for professional development and careers created by CFs. While some information specifically refers to the German academic system, most of the content of this article is of general interest for CFs in the life sciences. Microsc. Res. Tech. 79:463-479, 2016. © 2016 THE AUTHORS MICROSCOPY RESEARCH AND TECHNIQUE PUBLISHED BY WILEY PERIODICALS, INC.

  9. Advanced light microscopy core facilities: Balancing service, science and career

    Science.gov (United States)

    Hartmann, Hella; Reymann, Jürgen; Ansari, Nariman; Utz, Nadine; Fried, Hans‐Ulrich; Kukat, Christian; Peychl, Jan; Liebig, Christian; Terjung, Stefan; Laketa, Vibor; Sporbert, Anje; Weidtkamp‐Peters, Stefanie; Schauss, Astrid; Zuschratter, Werner; Avilov, Sergiy

    2016-01-01

    ABSTRACT Core Facilities (CF) for advanced light microscopy (ALM) have become indispensable support units for research in the life sciences. Their organizational structure and technical characteristics are quite diverse, although the tasks they pursue and the services they offer are similar. Therefore, throughout Europe, scientists from ALM‐CFs are forming networks to promote interactions and discuss best practice models. Here, we present recommendations for ALM‐CF operations elaborated by the workgroups of the German network of ALM‐CFs, German Bio‐Imaging (GerBI). We address technical aspects of CF planning and instrument maintainance, give advice on the organization and management of an ALM‐CF, propose a scheme for the training of CF users, and provide an overview of current resources for image processing and analysis. Further, we elaborate on the new challenges and opportunities for professional development and careers created by CFs. While some information specifically refers to the German academic system, most of the content of this article is of general interest for CFs in the life sciences. Microsc. Res. Tech. 79:463–479, 2016. © 2016 THE AUTHORS MICROSCOPY RESEARCH AND TECHNIQUE PUBLISHED BY WILEY PERIODICALS, INC. PMID:27040755

  10. Emission characteristics of light-emitting diodes by confocal microscopy

    Science.gov (United States)

    Cheung, W. S.; Choi, H. W.

    2016-03-01

    The emission profiles of light-emitting diodes have typically be measured by goniophotometry. However this technique suffers from several drawbacks, including the inability to generate three-dimensional intensity profiles as well as poor spatial resolution. These limitations are particularly pronounced when the technique is used to compared devices whose emission patterns have been modified through surface texturing at the micrometer and nanometer scales,. In view of such limitations, confocal microscopy has been adopted for the study of emission characteristics of LEDs. This enables three-dimensional emission maps to be collected, from which two-dimensional cross-sectional emission profiles can be generated. Of course, there are limitations associated with confocal microscopy, including the range of emission angles that can be measured due to the limited acceptance angle of the objective. As an illustration, the technique has been adopted to compare the emission profiles of LEDs with different divergence angles using an objective with a numerical aperture of 0.8. It is found that the results are consistent with those obtained by goniophotometry when the divergence angle is less that the acceptance angle of the objective.

  11. Use of polarized light microscopy in porcine reproductive technologies.

    Science.gov (United States)

    Caamaño, J N; Maside, C; Gil, M A; Muñoz, M; Cuello, C; Díez, C; Sánchez-Osorio, J R; Martín, D; Gomis, J; Vazquez, J M; Roca, J; Carrocera, S; Martinez, E A; Gómez, E

    2011-09-01

    The meiotic spindle in the oocyte is composed of microtubules and plays an important role during chromosome alignment and separation at meiosis. Polarized light microscopy (PLM) could be useful for a non-invasive evaluation of the meiotic spindle and may allow removal of nuclear structures without fluorochrome staining and ultraviolet exposure. In this study, PLM was used to assess its potential application in porcine reproductive technologies. The objectives of the present study were to assess the efficiency of PLM to detect microtubule-polymerized protein in in vitro-matured porcine oocytes; to examine its effects on the oocyte developmental competence; to select oocytes based on the presence of the meiotic spindle detected by PLM; and to assess the efficiency oocyte enucleation assisted with PLM. In the first experiment, the presence of microtubule-polymerized protein was assessed and confirmed in oocytes (n = 117) by immunostaining and chromatin detection. In the second experiment, oocytes (n = 160) were exposed or not (controls) to PLM for 10 minutes, and then parthenogenetically activated and cultured in vitro. In the third experiment, development competence of oocytes with a positive or negative signal to PLM was analyzed after in vitro fertilization. Finally, oocytes (n = 54) were enucleated using PLM as a tool to remove the meiotic spindle. A positive PLM signal was detected in 98.2 % of the oocytes, which strongly correlated (r = 1; p PLM did not differ significantly from controls on cleavage, total blastocyst, expanded blastocyst rates and total cell numbers. The percentage of oocytes at the MII stage and blastocyst formation rate in the negative PLM group significantly differed from control and PLM positive groups. Overall efficiency of spindle removal using the PLM-Oosight system was 92.6%. These results suggest that polarized light microscopy is an efficient system to detect microtubule-polymerized protein in in vitro-matured porcine oocytes and does

  12. Light sheet microscopy reveals more gradual light attenuation in light green versus dark green soybean leaves

    Science.gov (United States)

    Light wavelengths preferentially absorbed by chlorophyll (chl) often display steep absorption gradients. This oversaturates photosynthesis in upper chloroplasts and deprives lower chloroplasts of blue and red light, causing a steep gradient in carbon fixation. Reducing chl content could create a mor...

  13. Spermatozoa characteristics in six psittacine species using light microscopy.

    Science.gov (United States)

    Stelzer, G; Schmidt, V; Sobiraj, A; Krautwald-Junghanns, M-E

    2009-12-01

    Even though breeding of companion birds has increased continuously for years, the fecundity assessment of birds has hardly been acknowledged. Knowledge of the structure of spermatozoa is crucial for evaluation of the basic reproductive biology of any species as well as for phylogenetic research and cladistic analyses of internal relationships. Spermatozoa of six different psittacine species (Nymphicus hollandicus, Myiopsitta monachus, Agapornis roseicollis, Melopsittacus undulatus, Tanygnathus lucionensis, Guarouba guarouba) were examined using light microscopy. Head length (nucleus including acrosome), head width, midpiece length and tail length were measured and documented. Significant differences were obvious among almost all of the species for almost all four parameters. However, in all the six species a significant moderate correlation between spermatozoa midpiece lengths and tail lengths (r=0.535, p

  14. Breast cancer diagnosis using spatial light interference microscopy

    Science.gov (United States)

    Majeed, Hassaan; Kandel, Mikhail E.; Han, Kevin; Luo, Zelun; Macias, Virgilia; Tangella, Krishnarao; Balla, Andre; Popescu, Gabriel

    2015-11-01

    The standard practice in histopathology of breast cancers is to examine a hematoxylin and eosin (H&E) stained tissue biopsy under a microscope to diagnose whether a lesion is benign or malignant. This determination is made based on a manual, qualitative inspection, making it subject to investigator bias and resulting in low throughput. Hence, a quantitative, label-free, and high-throughput diagnosis method is highly desirable. We present here preliminary results showing the potential of quantitative phase imaging for breast cancer screening and help with differential diagnosis. We generated phase maps of unstained breast tissue biopsies using spatial light interference microscopy (SLIM). As a first step toward quantitative diagnosis based on SLIM, we carried out a qualitative evaluation of our label-free images. These images were shown to two pathologists who classified each case as either benign or malignant. This diagnosis was then compared against the diagnosis of the two pathologists on corresponding H&E stained tissue images and the number of agreements were counted. The agreement between SLIM and H&E based diagnosis was 88% for the first pathologist and 87% for the second. Our results demonstrate the potential and promise of SLIM for quantitative, label-free, and high-throughput diagnosis.

  15. Quantitative polarized light microscopy of unstained mammalian cochlear sections

    Science.gov (United States)

    Kalwani, Neil M.; Ong, Cheng Ai; Lysaght, Andrew C.; Haward, Simon J.; McKinley, Gareth H.; Stankovic, Konstantina M.

    2013-02-01

    Hearing loss is the most common sensory deficit in the world, and most frequently it originates in the inner ear. Yet, the inner ear has been difficult to access for diagnosis because of its small size, delicate nature, complex three-dimensional anatomy, and encasement in the densest bone in the body. Evolving optical methods are promising to afford cellular diagnosis of pathologic changes in the inner ear. To appropriately interpret results from these emerging technologies, it is important to characterize optical properties of cochlear tissues. Here, we focus on that characterization using quantitative polarized light microscopy (qPLM) applied to unstained cochlear sections of the mouse, a common animal model of human hearing loss. We find that the most birefringent cochlear materials are collagen fibrils and myelin. Retardance of the otic capsule, the spiral ligament, and the basilar membrane are substantially higher than that of other cochlear structures. Retardance of the spiral ligament and the basilar membrane decrease from the cochlear base to the apex, compared with the more uniform retardance of other structures. The intricate structural details revealed by qPLM of unstained cochlear sections ex vivo strongly motivate future application of polarization-sensitive optical coherence tomography to human cochlea in vivo.

  16. Light sheet microscopy reveals more gradual light attenuation in light-green versus dark-green soybean leaves.

    Science.gov (United States)

    Slattery, Rebecca A; Grennan, Aleel K; Sivaguru, Mayandi; Sozzani, Rosangela; Ort, Donald R

    2016-08-01

    Light wavelengths preferentially absorbed by chlorophyll (chl) often display steep absorption gradients. This over-saturates photosynthesis in upper chloroplasts and deprives lower chloroplasts of blue and red light. Reducing chl content could create a more even leaf light distribution and thereby increase leaf light-use efficiency and overall canopy photosynthesis. This was tested on soybean cultivar 'Clark' (WT) and a near-isogenic chl b deficient mutant, Y11y11, grown in controlled environment chambers and in the field. Light attenuation was quantified using a novel approach involving light sheet microscopy. Leaf adaxial and abaxial surfaces were illuminated separately with blue, red, and green wavelengths, and chl fluorescence was detected orthogonally to the illumination plane. Relative fluorescence was significantly greater in deeper layers of the Y11y11 mesophyll than in WT, with the greatest differences in blue, then red, and finally green light when illuminated from the adaxial surface. Modeled relative photosynthesis based on chlorophyll profiles and Beer's Law predicted less steep gradients in mutant relative photosynthesis rates compared to WT. Although photosynthetic light-use efficiency was greater in the field-grown mutant with ~50% lower chl, light-use efficiency was lower in the mutant when grown in chambers where chl was ~80% reduced. This difference is probably due to pleiotropic effects of the mutation that accompany very severe reductions in chlorophyll and may warrant further testing in other low-chl lines.

  17. Seeing the forest tree by tree: super-resolution light microscopy meets the neurosciences.

    Science.gov (United States)

    Maglione, Marta; Sigrist, Stephan J

    2013-07-01

    Light microscopy can be applied in vivo and can sample large tissue volumes, features crucial for the study of single neurons and neural circuits. However, light microscopy per se is diffraction-limited in resolution, and the substructure of core signaling compartments of neuronal circuits--axons, presynaptic active zones, postsynaptic densities and dendritic spines-can be only insufficiently characterized by standard light microscopy. Recently, several forms of super-resolution light microscopy breaking the diffraction-imposed resolution limit have started to allow highly resolved, dynamic imaging in the cell-biologically highly relevant 10-100 nanometer range ('mesoscale'). New, sometimes surprising answers concerning how protein mobility and protein architectures shape neuronal communication have already emerged. Here we start by briefly introducing super-resolution microscopy techniques, before we describe their use in the analysis of neuronal compartments. We conclude with long-term prospects for super-resolution light microscopy in the molecular and cellular neurosciences.

  18. Background suppression in TeO{sub 2} bolometers with Neganov-Luke amplified cryogenic light detectors

    Energy Technology Data Exchange (ETDEWEB)

    Willers, Michael; Lanfranchi, Jean-Come; Oberauer, Lothar; Schoenert, Stefan [Technische Universitaet Muenchen, Physik Department E15, James Franck Strasse, 85748 Garching (Germany); Excellence Cluster Universe, Technische Universitaet Muenchen, Boltzmannstr. 2, 85748, Garching (Germany); Muenster, Andrea; Potzel, Walter; Roth, Sabine; Wawoczny, Stephan; Zoeller, Andreas [Technische Universitaet Muenchen, Physik Department E15, James Franck Strasse, 85748 Garching (Germany); Giuliani, Andrea [Centre de Sciences Nucleaires et de Sciences de la Matiere, 91405 Orsay Campus (France)

    2015-07-01

    The Neganov-Luke (NL) effect offers a promising way to increase the sensitivity of cryogenic light detectors at low energies. In this talk we show that a highly efficient discrimination between α and e{sup -}/γ induced events in TeO{sub 2} crystals (used in the search for the neutrinoless double beta decay) can be achieved by measuring the Cherenkov radiation emitted by high-energetic electrons within the crystal. By using NL amplified light detectors, a suppression of ∝99% of α-induced events with energies close to the Q-value of {sup 130}Te at ∝2.5 MeV has been achieved for the first time while simultaneously accepting 99.8% of all e{sup -}/γ-induced events.

  19. Fluorescence Efficiency and Stability of Radio-Pure Tetraphenyl-butadiene Based Coatings for VUV Light Detection in Cryogenic Environments

    CERN Document Server

    Baudis, Laura; Dressler, Rugard; Piastra, Francesco; Usoltsev, Ilya; Walter, Manuel

    2015-01-01

    The detection of VUV scintillation light, e.g. in (liquid) argon detectors, commonly includes a reflector with a fluorescent coating, converting UV photons to visible light. The light yield of these detectors depends directly on the conversion efficiency. Several coating/reflector combinations were produced using VM2000, a specular reflecting multi layer polymer, and Tetratex, a diffuse reflecting PTFE fabric, as reflector foils. The efficiency of these coatings was optimised and has been measured in a dedicated liquid argon setup built at the University of Zurich. It employs a small, 1.3 kg LAr cell viewed by a 3-inch, low radioactivity PMT of type R11065-10 from Hamamatsu. The cryogenic stability of these coatings was additionally studied. The optimum reflector/coating combination was found to be Tetratex dip coated with Tetraphenyl-butadiene with a thickness of 0.9 mg/cm$^2$ resulting in a 3.6 times higher light yield compared to uncoated VM2000. Its performance was stable in long term measurements, ran up...

  20. Observation Platform for Dynamic Biomedical and Biotechnology Experiments using the ISS Light Microscopy Module Project

    Data.gov (United States)

    National Aeronautics and Space Administration — The proposed "Observation platform for dynamic biomedical and biotechnology experiments using the ISS Light Microscopy Module" consists of a platen sized to fit the...

  1. Optimizing and extending light-sculpting microscopy for fast functional imaging in neuroscience

    CERN Document Server

    Rupprecht, Peter; Groessl, Florian; Haubensak, Wulf E; Vaziri, Alipasha

    2015-01-01

    A number of questions in systems biology such as understanding how dynamics of neuronal networks are related to brain function require the ability to capture the functional dynamics of large cellular populations at high speed. Recently, this has driven the development of a number of parallel and high speed imaging techniques such as light-sculpting microscopy, which has been used to capture neuronal dynamics at the whole brain and single cell level in small model organism. However, the broader applicability of light-sculpting microscopy is limited by the size of volumes for which high speed imaging can be obtained and scattering in brain tissue. Here, we present strategies for optimizing the present tradeoffs in light-sculpting microscopy. Various scanning modalities in light-sculpting microscopy are theoretically and experimentally evaluated, and strategies to maximize the obtainable volume speeds, and depth penetration in brain tissue using different laser systems are provided. Design-choices, important par...

  2. Observation Platform for Dynamic Biomedical and Biotechnology Experiments using the ISS Light Microscopy Module Project

    Data.gov (United States)

    National Aeronautics and Space Administration — The objective of the research is the completion of an observation platform for the ISS Light Microscopy Module (LMM) as it currently resides on the US Fluids...

  3. Quantification of shrinkage microcracking in young mortar with fluorescence light microscopy and ESEM

    NARCIS (Netherlands)

    Bisschop, J.; Van Mier, J.C.M.

    1999-01-01

    In this paper a method is described to quantify shrinkage microcracking in young mortar by means of crack mapping. Visualisation of the microcracks is realised with two techniques: Fluorescence Light Microscopy (FLM) and Environmental Scanning Electron Microscopy (ESEM). The preliminary results obta

  4. CHROMATIN TEXTURE OF MELANOCYTIC NUCLEI - CORRELATION BETWEEN LIGHT AND ELECTRON-MICROSCOPY

    NARCIS (Netherlands)

    ABMAYR, W; STOLZ, W; KORHERR, S; WILD, W; SCHMOECKEL, C

    1987-01-01

    Cells of a benign pigmented mole and a malignant melanoma were used to compare electron microscopy (EM) and light microscopy (LM) with high-resolution TV-scanning and multivariate analysis methods. Special emphasis was placed on different kinds of chromatin texture features and their discriminating

  5. Deep focus; a digital image processing technique to produce improved focal depth in light microscopy:

    OpenAIRE

    Goldsmith, Noel T.

    2000-01-01

    In light microscopy, the spatial transverse resolution is a function of the wavelength and numerical aperture. The depth resolution is another function of these parameters. The factors that enable the detection of fine detail, make the sharp focusing of more than a thin slice of the depth in an object impossible. When the examination of fracture surfaces is attempted using light reflection microscopy, the roughness will often restrict the in-focus parts of an image to a small portion of the f...

  6. Synchrotron radiation as a light source in confocal microscopy

    Energy Technology Data Exchange (ETDEWEB)

    van der Oord, C.J.R.; Gerritsen, H.C.; Levine, Y.K. (University of Utrecht, P.O. Box 80.000, 3508 TA Utrecht (Netherlands)); Myring, W.J.; Jones, G.R.; Munro, I.H. (Daresbury Laboratory (United Kingdom))

    1992-01-01

    The optical properties of a confocal scanning microscope that was designed to utilize a synchrotron as light source are presented. The usable spectral range is from 200 nm up to 700 nm. Using 325-nm laser light, it is shown that the lateral resolution is about 125 nm, and the axial resolution better than 250 nm. After transport of the microscope from Utrecht to the Daresbury Synchrotron Source, 200-nm excitation can be applied, and the lateral resolution will drop to below 100 nm.

  7. Analysis of confocal microscopy under ultrashort light-pulse illumination

    Energy Technology Data Exchange (ETDEWEB)

    Kempe, M.; Rudolph, W. (Univ. of New Mexico, Albuquerque (United States))

    1993-02-01

    The resolution of confocal laser scanning microscopes is analyzed if they are used in measurements that are to combine high spatial and high temporal resoltuion. A generalized Fourier-optical treatment is developed in which the system characteristics contain all necessary information regarding the optical arrangement and the illuminating light pulses. Coherent and incoherent imaging are considered in detail. 10 refs., 8 figs.

  8. Non-linear optical microscopy sheds light on cardiovascular disease.

    Directory of Open Access Journals (Sweden)

    Valentina Caorsi

    Full Text Available Many cardiac diseases have been associated with increased fibrosis and changes in the organization of fibrillar collagen. The degree of fibrosis is routinely analyzed with invasive histological and immunohistochemical methods, giving a limited and qualitative understanding of the tissue's morphological adaptation to disease. Our aim is to quantitatively evaluate the increase in fibrosis by three-dimensional imaging of the collagen network in the myocardium using the non-linear optical microscopy techniques Two-Photon Excitation microscopy (TPE and Second Harmonic signal Generation (SHG. No sample staining is needed because numerous endogenous fluorophores are excited by a two-photon mechanism and highly non-centrosymmetric structures such as collagen generate strong second harmonic signals. We propose for the first time a 3D quantitative analysis to carefully evaluate the increased fibrosis in tissue from a rat model of heart failure post myocardial infarction. We show how to measure changes in fibrosis from the backward SHG (B(SHG alone, as only backward-propagating SHG is accessible for true in vivo applications. A 5-fold increase in collagen I fibrosis is detected in the remote surviving myocardium measured 20 weeks after infarction. The spatial distribution is also shown to change markedly, providing insight into the morphology of disease progression.

  9. Non-Linear Optical Microscopy Sheds Light on Cardiovascular Disease

    Science.gov (United States)

    Caorsi, Valentina; Toepfer, Christopher; Sikkel, Markus B.; Lyon, Alexander R.; MacLeod, Ken; Ferenczi, Mike A.

    2013-01-01

    Many cardiac diseases have been associated with increased fibrosis and changes in the organization of fibrillar collagen. The degree of fibrosis is routinely analyzed with invasive histological and immunohistochemical methods, giving a limited and qualitative understanding of the tissue's morphological adaptation to disease. Our aim is to quantitatively evaluate the increase in fibrosis by three-dimensional imaging of the collagen network in the myocardium using the non-linear optical microscopy techniques Two-Photon Excitation microscopy (TPE) and Second Harmonic signal Generation (SHG). No sample staining is needed because numerous endogenous fluorophores are excited by a two-photon mechanism and highly non-centrosymmetric structures such as collagen generate strong second harmonic signals. We propose for the first time a 3D quantitative analysis to carefully evaluate the increased fibrosis in tissue from a rat model of heart failure post myocardial infarction. We show how to measure changes in fibrosis from the backward SHG (BSHG) alone, as only backward-propagating SHG is accessible for true in vivo applications. A 5-fold increase in collagen I fibrosis is detected in the remote surviving myocardium measured 20 weeks after infarction. The spatial distribution is also shown to change markedly, providing insight into the morphology of disease progression. PMID:23409139

  10. Postnatal development of the dog pineal gland. Light microscopy

    OpenAIRE

    Calvo, J.L.; Boya, J; García-Mauriño, A.; López Carbonell, A.

    1990-01-01

    The light microscopical morphology of the dog pineal gland from the first postnatal day to maturity is described. In the first postnatal week, the pineal parenchyma shows immature cells and many mitotic figures. In this week, pigmented cells are obsemed for the first time, both in the pineal gland and in extrapineal nodules. Throughout the second week, the pineal parenchyma shows a cordonal pattern that disappears progressively in the following stages. From the...

  11. Spectrally And Temporally Resolved Low-Light Level Video Microscopy

    Science.gov (United States)

    Wampler, John E.; Furukawa, Ruth; Fechheimer, Marcus

    1989-12-01

    The IDG law-light video microscope system was designed to aid studies of localization of subcellular luminescence sources and stimulus/response coupling in single living cells using luminescent probes. Much of the motivation for design of this instrument system came from the pioneering efforts of Dr. Reynolds (Reynolds, Q. Rev. Biophys. 5, 295-347; Reynolds and Taylor, Bioscience 30, 586-592) who showed the value of intensified video camera systems for detection and localizion of fluorescence and bioluminescence signals from biological tissues. Our instrument system has essentially two roles, 1) localization and quantitation of very weak bioluminescence signals and 2) quantitation of intracellular environmental characteristics such as pH and calcium ion concentrations using fluorescent and bioluminescent probes. The instrument system exhibits over one million fold operating range allowing visualization and enhancement of quantum limited images with quantum limited response, spectral analysis of fluorescence signals, and transmitted light imaging. The computer control of the system implements rapid switching between light regimes, spatially resolved spectral scanning, and digital data processing for spectral shape analysis and for detailed analysis of the statistical distribution of single cell measurements. The system design and software algorithms used by the system are summarized. These design criteria are illustrated with examples taken from studies of bioluminescence, applications of bioluminescence to study developmental processes and gene expression in single living cells, and applications of fluorescent probes to study stimulus/response coupling in living cells.

  12. Particle Shape Characterization of Lunar Regolith using Reflected Light Microscopy

    Science.gov (United States)

    McCarty, C. B.; Garcia, G. C.; Rickman, D.

    2014-12-01

    Automated identification of particles in lunar thin sections is necessary for practical measurement of particle shape, void characterization, and quantitative characterization of sediment fabric. This may be done using image analysis, but several aspects of the lunar regolith make such automations difficult. For example, many of the particles are shattered; others are aggregates of smaller particles. Sieve sizes of the particles span 5 orders of magnitude. The physical thickness of a thin section, at a nominal 30 microns, is large compared to the size of many of the particles. Image acquisition modes, such as SEM and reflected light, while superior to transmitted light, still have significant ambiguity as to the volume being sampled. It is also desirable to have a technique that is inexpensive, not resource intensive, and analytically robust. To this end, we have developed an image acquisition and processing protocol that identifies and delineates resolvable particles on the front surface of a lunar thin section using a petrographic microscope in reflected light. For a polished thin section, a grid is defined covering the entire thin section. The grid defines discrete images taken with 20% overlap, minimizing the number of particles that intersect image boundaries. In reflected light mode, two images are acquired at each grid location, with a closed aperture diaphragm. One image, A, is focused precisely on the front surface of the thin section. The second image, B, is made after the stage is brought toward the objective lens just slightly. A bright fringe line, analogous to a Becke line, appears inside all transparent particles at the front surface of the section in the second image. The added light in the bright line corresponds to a deficit around the particles. Particle identification is done using ImageJ and uses multiple steps. A hybrid 5x5 median filter is used to make images Af and Bf. This primarily removes very small particles just below the front surface

  13. Making microscopy count: quantitative light microscopy of dynamic processes in living plants.

    Science.gov (United States)

    Fricker, Mark D; Moger, Julian; Littlejohn, George R; Deeks, Michael J

    2016-08-01

    Cell theory has officially reached 350 years of age as the first use of the word 'cell' in a biological context can be traced to a description of plant material by Robert Hooke in his historic publication 'Micrographia: or some physiological definitions of minute bodies'. The 2015 Royal Microscopical Society Botanical Microscopy meeting was a celebration of the streams of investigation initiated by Hooke to understand at the subcellular scale how plant cell function and form arises. Much of the work presented, and Honorary Fellowships awarded, reflected the advanced application of bioimaging informatics to extract quantitative data from micrographs that reveal dynamic molecular processes driving cell growth and physiology. The field has progressed from collecting many pixels in multiple modes to associating these measurements with objects or features that are meaningful biologically. The additional complexity involves object identification that draws on a different type of expertise from computer science and statistics that is often impenetrable to biologists. There are many useful tools and approaches being developed, but we now need more interdisciplinary exchange to use them effectively. In this review we show how this quiet revolution has provided tools available to any personal computer user. We also discuss the oft-neglected issue of quantifying algorithm robustness and the exciting possibilities offered through the integration of physiological information generated by biosensors with object detection and tracking. © 2016 The Authors Journal of Microscopy © 2016 Royal Microscopical Society.

  14. Preparation of Drosophila S2 cells for Light Microscopy

    Science.gov (United States)

    Buster, Daniel W.; Nye, Jonathan; Klebba, Joseph E.; Rogers, Gregory C.

    2010-01-01

    The ideal experimental system would be cheap and easy to maintain, amenable to a variety of techniques, and would be supported by an extensive literature and genome sequence database. Cultured Drosophila S2 cells, the product of disassociated 20-24 hour old embryos1, possess all these properties. Consequently, S2 cells are extremely well-suited for the analysis of cellular processes, including the discovery of the genes encoding the molecular components of the process or mechanism of interest. The features of S2 cells that are most responsible for their utility are the ease with which they are maintained, their exquisite sensitivity to double-stranded (ds)RNA-mediated interference (RNAi), and their tractability to fluorescence microscopy as either live or fixed cells. S2 cells can be grown in a variety of media, including a number of inexpensive, commercially-available, fully-defined, serum-free media2. In addition, they grow optimally and quickly at 21-24°C and can be cultured in a variety of containers. Unlike mammalian cells, S2 cells do not require a regulated atmosphere, but instead do well with normal air and can even be maintained in sealed flasks. Complementing the ease of RNAi in S2 cells is the ability to readily analyze experimentally-induced phenotypes by phase or fluorescence microscopy of fixed or live cells. S2 cells grow in culture as a single monolayer but do not display contact inhibition. Instead, cells tend to grow in colonies in dense cultures. At low density, S2 cultures grown on glass or tissue culture-treated plastic are round and loosely-attached. However, the cytology of S2 cells can be greatly improved by inducing them to flatten extensively by briefly culturing them on a surface coated with the lectin, concanavalin A (ConA)3. S2 cells can also be stably transfected with fluorescently-tagged markers to label structures or organelles of interest in live or fixed cells. Therefore, the usual scenario for the microscopic analysis of cells is

  15. An application of fast response Polarized Light Microscopy

    Science.gov (United States)

    Kantha, Deependra; van Winkle, David

    2007-03-01

    A fast response polarized light microscope was designed based on the algorithm by Shribak et. al (Applied Optics, vol. 42, 3009-3017). A pulsed laser beam was passed through two Pockels cells aligned at different angles with respect to optical axis. The retardance of the Pockels cell was controlled by external switches and power supplies. The electronics circuit in the system allows change of the retardance of the Pockels cell each millisecond for four milliseconds. In four milliseconds, four images of a birefringent sample, formed by different states of polarized light are recorded. The images are added appropriately to calculate retardence amplitude and phase by using codes written in imageJ software. The microscope was used to show the retardance and phase of a rabbit muscle fiber. Recordings were also taken of the contraction of Vorticella convallaria but the changes were too fast to yield retardance images. This type of microscope can be used to study different kinds of biological functions that change on a timescale slower than four milliseconds but faster than two seconds.

  16. Light-sheet microscopy for quantitative ovarian folliculometry

    Science.gov (United States)

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

    2017-02-01

    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. Owing to the large field of view and fast acquisition speed of our newly developed SPIM system, volumetric image stacks from entire intact samples of pig ovaries have been rendered demonstrating clearly discernible follicular features like follicle diameters (70 μm - 2.5 mm), size of developing Cumulus oophorus complexes (COC ) (40 μm - 110 μm), and follicular wall thicknesses (90 μm-120 μm). The observation of clearly distinguishable COCs protruding into the follicular antrum was also shown possible, and correlation with the developmental stage of the follicles was determined. Follicles of all developmental stages were identified, and even the small primordial follicle clusters forming the egg nest could be observed. 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.

  17. Polarized Light Microscopy in Reproductive and Developmental Biology

    Science.gov (United States)

    KOIKE-TANI, MAKI; TANI, TOMOMI; MEHTA, SHALIN B.; VERMA, AMITABH; OLDENBOURG, RUDOLF

    2016-01-01

    SUMMARY The polarized light microscope reveals orientational order in native molecular structures inside living cells, tissues, and whole organisms. It is a powerful tool used to monitor and analyze the early developmental stages of organisms that lend themselves to microscopic observations. In this article, we briefly discuss the components specific to a traditional polarizing microscope and some historically important observations on: chromosome packing in the sperm head, the first zygote division of the sea urchin, and differentiation initiated by the first asymmetric cell division in the sand dollar. We then introduce the LC-PolScope and describe its use for measuring birefringence and polarized fluorescence in living cells and tissues. Applications range from the enucleation of mouse oocytes to analyzing the polarized fluorescence of the water strider acrosome. We end with new results on the birefringence of the developing chick brain, which we analyzed between developmental stages of days 12–20. PMID:23901032

  18. Correlative cryo-fluorescence light microscopy and cryo-electron tomography of Streptomyces.

    Science.gov (United States)

    Koning, Roman I; Celler, Katherine; Willemse, Joost; Bos, Erik; van Wezel, Gilles P; Koster, Abraham J

    2014-01-01

    Light microscopy and electron microscopy are complementary techniques that in a correlative approach enable identification and targeting of fluorescently labeled structures in situ for three-dimensional imaging at nanometer resolution. Correlative imaging allows electron microscopic images to be positioned in a broader temporal and spatial context. We employed cryo-correlative light and electron microscopy (cryo-CLEM), combining cryo-fluorescence light microscopy and cryo-electron tomography, on vitrified Streptomyces bacteria to study cell division. Streptomycetes are mycelial bacteria that grow as long hyphae and reproduce via sporulation. On solid media, Streptomyces subsequently form distinct aerial mycelia where cell division leads to the formation of unigenomic spores which separate and disperse to form new colonies. In liquid media, only vegetative hyphae are present divided by noncell separating crosswalls. Their multicellular life style makes them exciting model systems for the study of bacterial development and cell division. Complex intracellular structures have been visualized with transmission electron microscopy. Here, we describe the methods for cryo-CLEM that we applied for studying Streptomyces. These methods include cell growth, fluorescent labeling, cryo-fixation by vitrification, cryo-light microscopy using a Linkam cryo-stage, image overlay and relocation, cryo-electron tomography using a Titan Krios, and tomographic reconstruction. Additionally, methods for segmentation, volume rendering, and visualization of the correlative data are described.

  19. Shedding new light on viruses: super-resolution microscopy for studying human immunodeficiency virus.

    Science.gov (United States)

    Müller, Barbara; Heilemann, Mike

    2013-10-01

    For more than 70 years electron microscopy (EM) techniques have played an important role in investigating structures of enveloped viruses. By contrast, use of fluorescence microscopy (FM) methods for this purpose was limited by the fact that the size of virus particles is generally around or below the diffraction limit of light microscopy. Various super-resolution (SR) fluorescence imaging techniques developed over the past two decades bypass the diffraction limit of light microscopy, allowing visualization of subviral details and bridging the gap between conventional FM and EM methods. We summarize here findings on human immunodeficiency virus (HIV-1) obtained using SR-FM techniques. Although the number of published studies is currently limited and some of the pioneering analyses also covered methodological or descriptive aspects, recent publications clearly indicate the potential to approach open questions in HIV-1 replication from a new angle.

  20. Cryogenic Wide-Area Light Detectors for Neutrino and Dark Matter Searches

    Science.gov (United States)

    Di Domizio, S.; Bagni, R.; Battistelli, E. S.; Bellini, F.; Bucci, C.; Calvo, M.; Cardani, L.; Castellano, M. G.; Coppolecchia, A.; Cosmelli, C.; Cruciani, A.; D'Addabbo, A.; de Bernardis, P.; Masi, S.; Pinci, D.; Vignati, M.

    2014-09-01

    Large-mass arrays of bolometers proved to be good detectors for neutrinoless double beta decay (0DBD) and dark matter searches. CUORE and LUCIFER are bolometric 0DBD experiments that will start to take data in 2015 at Laboratori Nazionali del Gran Sasso in Italy. The sensitivity of CUORE could be increased by removing the background due to particles, by detecting the small amount of Čerenkov light (100 eV) emitted by the s' signal and not by s. LUCIFER could be extended to detect also dark matter, provided that the background from / particles (100 eV of scintillation light) is discriminated from nuclear recoils of about 10 keV energy (no light). We have recently started to develop light detectors for CUORE, LUCIFER and similar bolometric experiments. The aim is to obtain detectors with an active area of (the face of bolometric crystals), operating at 10 mK, and with an energy resolution at the baseline below 20 eV RMS. We have chosen to develop phonon-mediated detectors with KID sensors. We are currently testing the first prototypes.

  1. Morphological aspects of Angiostrongylus costaricensis by light and scanning electron microscopy.

    Science.gov (United States)

    Rebello, Karina M; Menna-Barreto, Rubem F S; Chagas-Moutinho, Vanessa A; Mota, Ester M; Perales, Jonas; Neves-Ferreira, Ana Gisele C; Oliveira-Menezes, Aleksandra; Lenzi, Henrique

    2013-09-01

    Angiostrongylus costaricensis is a parasitic nematode that can cause severe gastrointestinal disease, known as abdominal angiostrongiliasis, in humans. This paper presents the characterization of first- and third-stage larvae and male and female adult worms of A. costaricensis by scanning electron and light microscopy. Several novel anatomical structures were identified by scanning electron microscopy, including details of the cuticular striations of the spicules in male worms and a protective flap of the cuticle covering the vulvar aperture in female worms. Other taxonomic features revealed by light microscopy include the gubernaculum and the esophageal-intestinal valve. The use of two microscopy techniques allowed a detailed characterization of the morphology of this nematode. A number of previously identified taxonomic features, such as the striated nature of the spicules and the lateral alae were confirmed; however, the use of scanning electron microscopy resulted in a reassessment of the correct number of papillae distributed around the oral opening and behind the cloacal opening. These observations, in combination with light microscopy-based characterization of the gubernaculum and esophageal valves, have allowed a more detailed description of this nematode taxonomy.

  2. Structured light optical microscopy for three-dimensional reconstruction of technical surfaces

    Science.gov (United States)

    Kettel, Johannes; Reinecke, Holger; Müller, Claas

    2016-04-01

    In microsystems technology quality control of micro structured surfaces with different surface properties is playing an ever more important role. The process of quality control incorporates three-dimensional (3D) reconstruction of specularand diffusive reflecting technical surfaces. Due to the demand on high measurement accuracy and data acquisition rates, structured light optical microscopy has become a valuable solution to solve this problem providing high vertical and lateral resolution. However, 3D reconstruction of specular reflecting technical surfaces still remains a challenge to optical measurement principles. In this paper we present a measurement principle based on structured light optical microscopy which enables 3D reconstruction of specular- and diffusive reflecting technical surfaces. It is realized using two light paths of a stereo microscope equipped with different magnification levels. The right optical path of the stereo microscope is used to project structured light onto the object surface. The left optical path is used to capture the structured illuminated object surface with a camera. Structured light patterns are generated by a Digital Light Processing (DLP) device in combination with a high power Light Emitting Diode (LED). Structured light patterns are realized as a matrix of discrete light spots to illuminate defined areas on the object surface. The introduced measurement principle is based on multiple and parallel processed point measurements. Analysis of the measured Point Spread Function (PSF) by pattern recognition and model fitting algorithms enables the precise calculation of 3D coordinates. Using exemplary technical surfaces we demonstrate the successful application of our measurement principle.

  3. Topography and refractometry of nanostructures using spatial light interference microscopy (SLIM)

    OpenAIRE

    2010-01-01

    Spatial Light Interference Microscopy (SLIM) is a novel method developed in our laboratory that provides quantitative phase images of transparent structures with 0.3 nm spatial and 0.03 nm temporal accuracy owing to the white light illumination and its common path interferometric geometry. We exploit these features and demonstrate SLIM's ability to perform topography at a single atomic layer in graphene. Further, using a decoupling procedure that we developed for cylindrical structures, we ex...

  4. Discrimination of Dendrobium officinale and Its Common Adulterants by Combination of Normal Light and Fluorescence Microscopy

    Directory of Open Access Journals (Sweden)

    Chu Chu

    2014-03-01

    Full Text Available The stems of Dendrobium officinale Kimura et Migo, named Tie-pi-shi-hu, is one of the most endangered and precious species in China. Because of its various pharmacodynamic effects, D. officinale is widely recognized as a high-quality health food in China and other countries in south and south-east Asia. With the rising interest of D. officinale, its products have a high price due to a limited supply. This high price has led to the proliferation of adulterants in the market. To ensure the safe use of D. officinale, a fast and convenient method combining normal and fluorescence microscopy was applied in the present study to distinguish D. officinale from three commonly used adulterants including Zi-pi-shi-hu (D. devonianum, Shui-cao-shi-hu (D. aphyllum, Guang-jie-shi-hu (D. gratiosissimum. The result demonstrated that D. officinale could be identified by the characteristic “two hat-shaped” vascular bundle sheath observed under the fluorescence microscopy and the distribution of raphides under normal light microscopy. The other three adulterants could be discriminated by the vascular bundle differences and the distribution of raphides under normal light microscopy. This work indicated that combination of normal light and fluorescence microscopy is a fast and efficient technique to scientifically distinguish D. officinale from the commonly confused species.

  5. Correlative light/electron microscopy for the investigation of microbial mats from Black Sea Cold Seeps.

    Science.gov (United States)

    Wrede, Christoph; Heller, Christina; Reitner, Joachim; Hoppert, Michael

    2008-05-01

    In several fields of cell biology, correlative microscopy is applied to compare the structure of objects at high resolution under the electron microscope with low resolution light microscopy images of the same sample. It is, however, difficult to prepare samples and marker systems that are applicable for both microscopic techniques for the same specimen at the same time. In our studies, we used microbial mats from Cold Seep communities for a simple and rapid correlative microscopy method. The mats consist of bacterial and archaeal microorganisms, coupling reverse methanogenesis to the reduction of sulfate. The reverse methanogenic pathway also generates carbonates that precipitate inside the mat and may be the main reason for the formation of a microbial reef. The mat shows highly differentiated aggregates of various organisms, tightly interconnected by extracellular polysaccharides. In order to investigate the role of EPS as adhesive mucilage for the biofilm and as a precipitation matrix for carbonate minerals, samples were embedded in a hydrophilic resin (Lowicryl K4 M). Sections were suitable for light as well as electron microscopy in combination with lectins, either labeled with a fluorescent marker or with colloidal gold. This allows lectin mapping at low resolution for light microscopy in direct comparison with a highly resolved electron microscopic image.

  6. Discrimination of Dendrobium officinale and its common adulterants by combination of normal light and fluorescence microscopy.

    Science.gov (United States)

    Chu, Chu; Yin, Huimin; Xia, Li; Cheng, Dongping; Yan, Jizhong; Zhu, Lin

    2014-03-24

    The stems of Dendrobium officinale Kimura et Migo, named Tie-pi-shi-hu, is one of the most endangered and precious species in China. Because of its various pharmacodynamic effects, D. officinale is widely recognized as a high-quality health food in China and other countries in south and south-east Asia. With the rising interest of D. officinale, its products have a high price due to a limited supply. This high price has led to the proliferation of adulterants in the market. To ensure the safe use of D. officinale, a fast and convenient method combining normal and fluorescence microscopy was applied in the present study to distinguish D. officinale from three commonly used adulterants including Zi-pi-shi-hu (D. devonianum), Shui-cao-shi-hu (D. aphyllum), Guang-jie-shi-hu (D. gratiosissimum). The result demonstrated that D. officinale could be identified by the characteristic "two hat-shaped" vascular bundle sheath observed under the fluorescence microscopy and the distribution of raphides under normal light microscopy. The other three adulterants could be discriminated by the vascular bundle differences and the distribution of raphides under normal light microscopy. This work indicated that combination of normal light and fluorescence microscopy is a fast and efficient technique to scientifically distinguish D. officinale from the commonly confused species.

  7. Developments in application of light and scanning electron microscopy techniques for cell wall degradation studies.

    NARCIS (Netherlands)

    Engels, F.M.

    1996-01-01

    The results of recent technological developments in light and scanning electron microscopy closely used for research on forage cell wall degradation in ruminants, are reviewed. The indigestibility of forages by rumen microorganisms used to be ascribed mainly to an overall presence of lignin in the p

  8. Modeling optical behavior of birefringent biological tissues for evaluation of quantitative polarized light microscopy

    NARCIS (Netherlands)

    Turnhout, van M.C.; Kranenbarg, S.; Leeuwen, van J.L.

    2009-01-01

    Quantitative polarized light microscopy (qPLM) is a popular tool for the investigation of birefringent architectures in biological tissues. Collagen, the most abundant protein in mammals, is such a birefringent material. Interpretation of results of qPLM in terms of collagen network architecture and

  9. Morphology of Ichthyophonus hoferi assessed by light and scanning electron microscopy

    DEFF Research Database (Denmark)

    Spanggaard, Bettina; Huss, Hans Henrik; Bresciani, J.

    1995-01-01

    The morphology of Ichthyophonus hoferi in vitro at pH 3.5 and 7.0 is described using light and scanning electron microscopy. Only vegetative growth was observed. At pH 3.5, hyphal growth was seen. The hyphae of I. hoferi are characterized by evacuated hyphal walls with the cytoplasm migrating...

  10. Structure of the light harvesting antenna from Rhodospirillum molischianum studied by electron microscopy

    NARCIS (Netherlands)

    Boonstra, Arjen F.; Germeroth, Lothar; Boekema, Egbert J.

    1994-01-01

    The structure of two types of isolated light-harvesting antenna complexes from Rhodospirillum molischianum was studied by electron microscopy and image analysis. The B870 reaction center complex forms an almost circular particle with a diameter in the plane of the membrane of about 10.7-11.2 nm. A

  11. Live imaging of nervous system development and function using light-sheet microscopy.

    Science.gov (United States)

    Lemon, William C; Keller, Philipp J

    2015-01-01

    In vivo imaging applications typically require carefully balancing conflicting parameters. Often it is necessary to achieve high imaging speed, low photo-bleaching, and photo-toxicity, good three-dimensional resolution, high signal-to-noise ratio, and excellent physical coverage at the same time. Light-sheet microscopy provides good performance in all of these categories, and is thus emerging as a particularly powerful live imaging method for the life sciences. We see an outstanding potential for applying light-sheet microscopy to the study of development and function of the early nervous system in vertebrates and higher invertebrates. Here, we review state-of-the-art approaches to live imaging of early development, and show how the unique capabilities of light-sheet microscopy can further advance our understanding of the development and function of the nervous system. We discuss key considerations in the design of light-sheet microscopy experiments, including sample preparation and fluorescent marker strategies, and provide an outlook for future directions in the field.

  12. Looking at glass from a different angle: New insights into fracture patterns through transmitted light microscopy

    NARCIS (Netherlands)

    Van der Velde, O.; Copuroglu, O.; Veer, F.A.

    2015-01-01

    This paper shows the benefit of using transmitted light microscopy together with a Z-scanning software in fractographical analyses of glass. The strength of glass is largely dependent on processes that happen at the microscale. In this research, 52 plates were fractured in a biaxial tensile test. Th

  13. SPED Light Sheet Microscopy: Fast Mapping of Biological System Structure and Function.

    Science.gov (United States)

    Tomer, Raju; Lovett-Barron, Matthew; Kauvar, Isaac; Andalman, Aaron; Burns, Vanessa M; Sankaran, Sethuraman; Grosenick, Logan; Broxton, Michael; Yang, Samuel; Deisseroth, Karl

    2015-12-17

    The goal of understanding living nervous systems has driven interest in high-speed and large field-of-view volumetric imaging at cellular resolution. Light sheet microscopy approaches have emerged for cellular-resolution functional brain imaging in small organisms such as larval zebrafish, but remain fundamentally limited in speed. Here, we have developed SPED light sheet microscopy, which combines large volumetric field-of-view via an extended depth of field with the optical sectioning of light sheet microscopy, thereby eliminating the need to physically scan detection objectives for volumetric imaging. SPED enables scanning of thousands of volumes-per-second, limited only by camera acquisition rate, through the harnessing of optical mechanisms that normally result in unwanted spherical aberrations. We demonstrate capabilities of SPED microscopy by performing fast sub-cellular resolution imaging of CLARITY mouse brains and cellular-resolution volumetric Ca(2+) imaging of entire zebrafish nervous systems. Together, SPED light sheet methods enable high-speed cellular-resolution volumetric mapping of biological system structure and function.

  14. Imaging of human differentiated 3D neural aggregates using light sheet fluorescence microscopy

    Directory of Open Access Journals (Sweden)

    Emilio J Gualda

    2014-08-01

    Full Text Available The development of three dimensional cell cultures represents a big step for the better understanding of cell behavior and disease in a more natural like environment, providing not only single but multiple cell type interactions in a complex three dimensional matrix, highly resembling physiological conditions. Light sheet fluorescence microscopy is becoming an excellent tool for fast imaging of such three-dimensional biological structures. We demonstrate the potential of this technique for the imaging of human differentiated 3D neural aggregates in fixed and live samples, namely calcium imaging and cell death processes, showing the power of imaging modality compared with traditional microscopy. The combination of light sheet microscopy and 3D neural cultures will open the door to more challenging experiments involving drug testing at large scale as well as a better understanding of relevant biological processes in a more realistic environment.

  15. Doubling the resolution of spatial-light-modulator-based differential interference contrast microscopy by structured illumination.

    Science.gov (United States)

    Chen, Jianling; Lv, Xiaohua; Zeng, Shaoqun

    2013-09-01

    Recently developed spatial light modulator (SLM)-based differential interference contrast (DIC) microscopy [Opt. Lett. 34, 2988 (2009)] reveals flexibility on the implementation of DIC imaging. However, its numerical aperture (spatial resolution) is limited to maintain sufficient interference contrast, because it requires two beams to interfere. We present a structured illumination (SI) SLM-based DIC microscopy to effectively improve the lateral resolution of the SLM-based DIC microscopy. The SI field is generated and controlled by an adjustable grating displayed on an SLM. The SI SLM-based DIC expands the bandwidth of the coherent transfer function of the SLM-based DIC imaging system, thus improving the spatial resolution. The reconstructed SI SLM-based DIC image exhibits lateral resolution of approximately 208 nm, doubling that of the common SLM-based DIC image (approximately 415 nm). SI SLM-based DIC microscopy has the potential for achieving high-resolution quantitative phase images.

  16. Optical near-field microscopy of light focusing through a photonic crystal flat lens.

    Science.gov (United States)

    Fabre, Nathalie; Lalouat, Loïc; Cluzel, Benoit; Mélique, Xavier; Lippens, Didier; de Fornel, Frédérique; Vanbésien, Olivier

    2008-08-15

    We report here the direct observation by using a scanning near-field microscopy technique of the light focusing through a photonic crystal flat lens designed and fabricated to operate at optical frequencies. The lens is fabricated using a III-V semiconductor slab, and we directly visualize the propagation of the electromagnetic waves by using a scanning near-field optical microscope. We directly evidence spatially, as well as spectrally, the focusing operating regime of the lens. At last, in light of the experimental scanning near-field optical microscope pictures, we discuss the lens ability to focus light at a subwavelength scale.

  17. Enhanced light microscopy visualization of virus particles from Zika virus to filamentous ebolaviruses.

    Science.gov (United States)

    Daaboul, George G; Freedman, David S; Scherr, Steven M; Carter, Erik; Rosca, Alexandru; Bernstein, David; Mire, Chad E; Agans, Krystle N; Hoenen, Thomas; Geisbert, Thomas W; Ünlü, M Selim; Connor, John H

    2017-01-01

    Light microscopy is a powerful tool in the detection and analysis of parasites, fungi, and prokaryotes, but has been challenging to use for the detection of individual virus particles. Unlabeled virus particles are too small to be visualized using standard visible light microscopy. Characterization of virus particles is typically performed using higher resolution approaches such as electron microscopy or atomic force microscopy. These approaches require purification of virions away from their normal millieu, requiring significant levels of expertise, and can only enumerate small numbers of particles per field of view. Here, we utilize a visible light imaging approach called Single Particle Interferometric Reflectance Imaging Sensor (SP-IRIS) that allows automated counting and sizing of thousands of individual virions. Virions are captured directly from complex solutions onto a silicon chip and then detected using a reflectance interference imaging modality. We show that the use of different imaging wavelengths allows the visualization of a multitude of virus particles. Using Violet/UV illumination, the SP-IRIS technique is able to detect individual flavivirus particles (~40 nm), while green light illumination is capable of identifying and discriminating between vesicular stomatitis virus and vaccinia virus (~360 nm). Strikingly, the technology allows the clear identification of filamentous infectious ebolavirus particles and virus-like particles. The ability to differentiate and quantify unlabeled virus particles extends the usefulness of traditional light microscopy and can be embodied in a straightforward benchtop approach allowing widespread applications ranging from rapid detection in biological fluids to analysis of virus-like particles for vaccine development and production.

  18. Comparative analysis of clastogen-induced chromosome aberrations observed with light microscopy and by means of atomic force microscopy.

    Science.gov (United States)

    Koleva, Vanya Petrova; Dragoeva, Asya Pencheva; Andreeva, Andreana Ivanova; Burova, Marina Todorova; Georgiev, Sevdalin; Enchev, Dobromir Dimitrov

    2013-04-30

    Different types of chromosome aberration were observed in mouse bone-marrow cells after treatment with 4-bromo-N,N-diethyl-5,5-dimethyl-2,5-dihydro-1,2-oxaphosphol-2-amine 2-oxide (Br-oxaphosphole, Br-oxph) in a previous study. The aim of the present study is to perform a comparative analysis of these chromosomal damages observed with light microscopy (LM) and by means of atomic force microscopy (AFM). The kinds of aberrations scored by LM were substantially corrected by images at the ultrastructural level. The AFM analysis excluded 29.0% of gaps and 33.3% of fusion-type aberrations. On the other hand, AFM revealed the presence of aberrations that were not visible under the LM. This indicates that only AFM images would provide precise information about the real nature of chromosomal damages. The results of our study revealed that the 'real gaps' represented about 50% of all the gaps visible under LM. Excluded 'false gaps' were detected via AFM as breaks or decondensed chromosome regions. These results would support the statement that gaps must be included when testing genotoxicity. The ultrastructural analysis also confirmed the validity of using LM in the mouse bone-marrow chromosome aberration test, as a tool for detecting genotoxicity of chemicals in routine studies. When there is a need for precise evaluation of chromosome damage, only AFM images can provide information on specific genotoxic effects.

  19. Ultra-light weight undamped tuned dynamic absorber for cryogenically cooled infrared electro-optic payload

    Science.gov (United States)

    Veprik, Alexander; Babitsky, Vladimir

    2017-04-01

    Attenuation of tonal cryocooler induced vibration in infrared electro-optical payloads may be achieved by using of Tuned Dynamic Absorber (TDA) which is, generally speaking, a passive, weakly damped mass-spring system the resonant frequency of which is precisely matched with the driving frequency. Added TDA results in a favorable modification of the frequency response functions of combined structure. In particular, a favorable antiresonant notch appears at the frequency of tonal excitation along with the adjacent secondary resonance, the width and depth of which along with its closeness to the secondary resonance are strongly dependent on the mass and damping ratios. Using heavier TDA favorably results in wider and deeper antiresonant notch along with increased gap between antiresonant and resonant frequencies. Lowering damping in TDA favorably results in deepening the antiresonant notch. The weight of TDA is usually subjected to tight design constrains. Use of lightweight TDA not only diminishes the attainable performance but also complicates the procedure of frequency matching. Along these lines, even minor frequency deviations may negate the TDA performance and even result in TDA failure in case of resonant build up. The authors are presenting theoretical and practical aspects of designing and constructing ultra-light weight TDA in application to vibration attenuation of electro-optical infrared payload relying on Split Stirling linear cryocooler, the driving frequency of which is fixed and may be accurately tuned and maintained using a digital controller over the entire range of working conditions and lifetime; the lack of mass ratio is compensated by minimizing the damping ratio. In one particular case, in excess of 100-fold vibration attenuation has been achieved by adding as little as 5% to the payload weight.

  20. Helium cryogenics

    CERN Document Server

    Van Sciver, Steven W

    2012-01-01

    Twenty five years have elapsed since the original publication of Helium Cryogenics. During this time, a considerable amount of research and development involving helium fluids has been carried out culminating in several large-scale projects. Furthermore, the field has matured through these efforts so that there is now a broad engineering base to assist the development of future projects. Helium Cryogenics, 2nd edition brings these advances in helium cryogenics together in an updated form. As in the original edition, the author's approach is to survey the field of cryogenics with emphasis on helium fluids. This approach is more specialized and fundamental than that contained in other cryogenics books, which treat the associated range of cryogenic fluids. As a result, the level of treatment is more advanced and assumes a certain knowledge of fundamental engineering and physics principles, including some quantum mechanics. The goal throughout the work is to bridge the gap between the physics and engineering aspe...

  1. Breaking the spatial resolution barrier via iterative sound-light interaction in deep tissue microscopy.

    Science.gov (United States)

    Si, Ke; Fiolka, Reto; Cui, Meng

    2012-01-01

    Optical microscopy has so far been restricted to superficial layers, leaving many important biological questions unanswered. Random scattering causes the ballistic focus, which is conventionally used for image formation, to decay exponentially with depth. Optical imaging beyond the ballistic regime has been demonstrated by hybrid techniques that combine light with the deeper penetration capability of sound waves. Deep inside highly scattering media, the sound focus dimensions restrict the imaging resolutions. Here we show that by iteratively focusing light into an ultrasound focus via phase conjugation, we can fundamentally overcome this resolution barrier in deep tissues and at the same time increase the focus to background ratio. We demonstrate fluorescence microscopy beyond the ballistic regime of light with a threefold improved resolution and a fivefold increase in contrast. This development opens up practical high resolution fluorescence imaging in deep tissues.

  2. Breaking the spatial resolution barrier via iterative sound-light interaction in deep tissue microscopy

    CERN Document Server

    Si, Ke; Cui, Meng

    2012-01-01

    Optical microscopy has so far been restricted to superficial layers, leaving many important biological questions unanswered. Random scattering causes the ballistic focus, which is conventionally used for image formation, to decay exponentially with depth. Optical imaging beyond the ballistic regime has been demonstrated by hybrid techniques that combine light with the deeper penetration capability of sound waves. Deep inside highly scattering media, the sound focus dimensions restrict the imaging resolutions. Here we show that by iteratively focusing light into an ultrasound focus via phase conjugation, we can fundamentally overcome this resolution barrier in deep tissues and at the same time increase the focus to background ratio. We demonstrate fluorescence microscopy beyond the ballistic regime of light with a threefold improved resolution and a fivefold increase in contrast. This development opens up practical high resolution fluorescence imaging in deep tissues.

  3. Cryogenic exciter

    Science.gov (United States)

    Bray, James William [Niskayuna, NY; Garces, Luis Jose [Niskayuna, NY

    2012-03-13

    The disclosed technology is a cryogenic static exciter. The cryogenic static exciter is connected to a synchronous electric machine that has a field winding. The synchronous electric machine is cooled via a refrigerator or cryogen like liquid nitrogen. The static exciter is in communication with the field winding and is operating at ambient temperature. The static exciter receives cooling from a refrigerator or cryogen source, which may also service the synchronous machine, to selected areas of the static exciter and the cooling selectively reduces the operating temperature of the selected areas of the static exciter.

  4. Light sheet microscopy reveals more gradual light attenuation in light-green versus dark-green soybean leaves

    OpenAIRE

    Slattery, Rebecca A.; Grennan, Aleel K; Sivaguru, Mayandi; Sozzani, Rosangela; Ort, Donald R.

    2016-01-01

    Light wavelengths preferentially absorbed by chlorophyll (chl) often display steep absorption gradients. This over-saturates photosynthesis in upper chloroplasts and deprives lower chloroplasts of blue and red light. Reducing chl content could create a more even leaf light distribution and thereby increase leaf light-use efficiency and overall canopy photosynthesis. This was tested on soybean cultivar ‘Clark’ (WT) and a near-isogenic chl b deficient mutant, Y11y11, grown in controlled environ...

  5. Microstructure formation of lithium-ion battery electrodes during drying - An ex-situ study using cryogenic broad ion beam slope-cutting and scanning electron microscopy (Cryo-BIB-SEM)

    Science.gov (United States)

    Jaiser, Stefan; Kumberg, Jana; Klaver, Jop; Urai, Janos L.; Schabel, Wilhelm; Schmatz, Joyce; Scharfer, Philip

    2017-03-01

    Properties of lithium-ion battery electrodes relate to the complex microstructure that develops during solvent removal. We use cryogenic scanning electron microscopy in combination with broad ion beam slope-cutting (Cryo-BIB-SEM) for the ex-situ imaging of film formation in battery electrodes. Drying of anode films is quenched by cryo-preservation in slushy nitrogen at systematically increasing drying times, followed by SEM imaging under cryogenic conditions. Energy dispersive x-ray spectroscopy (EDS) and image processing of segmented cross-sections are used to analyze the development of component gradients with time. We find electrode films to shrink homogeneously and not in a top-down consolidation process as previously hypothesized. Binder gradients evolve in the liquid phase and initiate solvent diffusion from the bulk to the surface, thereby dragging binder towards the surface. Capillary transport is identified as a fundamental process that directly impacts drying kinetics and binder distribution.

  6. A novel fibrous duct structure discovered in the brain meninges by using polarized light microscopy

    Science.gov (United States)

    Nam, Min-Ho; Jung, Sharon Jiyoon; Soh, Kwang-Sup; Lim, Jaekwan; Seo, Eunseok; Lim, Jun; Baek, Miok; Lee, Sang Joon

    2016-05-01

    We have previously reported the discovery of a novel fibrous structure (NFS) consisting of unidirectionally arranged collagen fibers in the spinal pia mater. Due to its unique structure, it was easily detected using polarized light microscopy. In the current study, we describe the discovery of a similar NFS in the brain meninges of rats by using polarized light microscopy. This NFS is located beneath the superior sagittal sinus. Initially, we systemically analyzed the polarization properties of the NFS. The change in the light intensity of the NFS, with respect to the polarization angle, was eight times greater than that of blood vessels, showing that the collagen fibers are oriented in a particular direction with almost perfect parallelism (0.99). The orientation angle of the polarization ellipse confirmed the orientation of the collagen fibers in the NFS. Histological studies further confirmed that the unidirectionally arranged collagen fibers were responsible for this distinct polarization property. Surprisingly, X-ray microtomography and 3D confocal imaging revealed that the NFS contains within it a duct structure, a putative primo vessel. In conclusion, we report a NFS in the brain meninges, detected by using polarized light microscopy, that provides space for a putative primo vessel, not a blood vessel.

  7. Live imaging of Tribolium castaneum embryonic development using light-sheet-based fluorescence microscopy.

    Science.gov (United States)

    Strobl, Frederic; Schmitz, Alexander; Stelzer, Ernst H K

    2015-10-01

    Tribolium castaneum has become an important insect model organism for evolutionary developmental biology, genetics and biotechnology. However, few protocols for live fluorescence imaging of Tribolium have been reported, and little image data is available. Here we provide a protocol for recording the development of Tribolium embryos with light-sheet-based fluorescence microscopy. The protocol can be completed in 4-7 d and provides procedural details for: embryo collection, microscope configuration, embryo preparation and mounting, noninvasive live imaging for up to 120 h along multiple directions, retrieval of the live embryo once imaging is completed, and image data processing, for which exemplary data is provided. Stringent quality control criteria for developmental biology studies are also discussed. Light-sheet-based fluorescence microscopy complements existing toolkits used to study Tribolium development, can be adapted to other insect species, and requires no advanced imaging or sample preparation skills.

  8. 3D print customized sample holders for live light sheet microscopy.

    Science.gov (United States)

    Jeandupeux, Emeric; Lobjois, Valérie; Ducommun, Bernard

    2015-08-01

    A major hurdle to the widespread application of light sheet microscopy is the lack of versatile and non-intrusive sample holders that are adaptable to a variety of biological samples for live imaging. To overcome this limitation, we present herein the application of 3D printing to the fabrication of a fully customizable casting kit. 3D printing enables facile preparation of hydrogel sample holders adaptable to any shape and number of specimen. As an example, we present the use of this device to produce a four-sample holder adapted to parallel live monitoring of multicellular tumor spheroid growth. To share our solution with the light sheet microscopy community, all files necessary to produce or customize sample holders are freely available online.

  9. LED arrays as cost effective and efficient light sources for widefield microscopy.

    Directory of Open Access Journals (Sweden)

    Dinu F Albeanu

    Full Text Available New developments in fluorophores as well as in detection methods have fueled the rapid growth of optical imaging in the life sciences. Commercial widefield microscopes generally use arc lamps, excitation/emission filters and shutters for fluorescence imaging. These components can be expensive, difficult to maintain and preclude stable illumination. Here, we describe methods to construct inexpensive and easy-to-use light sources for optical microscopy using light-emitting diodes (LEDs. We also provide examples of its applicability to biological fluorescence imaging.

  10. Correlative Light and Scanning X-Ray Scattering Microscopy of Healthy and Pathologic Human Bone Sections

    Science.gov (United States)

    Giannini, C.; Siliqi, D.; Bunk, O.; Beraudi, A.; Ladisa, M.; Altamura, D.; Stea, S.; Baruffaldi, F.

    2012-01-01

    Scanning small and wide angle X-ray scattering (scanning SWAXS) experiments were performed on healthy and pathologic human bone sections. Via crystallographic tools the data were transformed into quantitative images and as such compared with circularly polarized light (CPL) microscopy images. SWAXS and CPL images allowed extracting information of the mineral nanocrystalline phase embedded, with and without preferred orientation, in the collagen fibrils, mapping local changes at sub-osteon resolution. This favorable combination has been applied for the first time to biopsies of dwarfism syndrome and Paget's disease to shed light onto the cortical structure of natural bone in healthy and pathologic sections. PMID:22666538

  11. Low-cost multimodal light sheet microscopy for optically cleared tissues and living specimens.

    Science.gov (United States)

    Rouger, Vincent; Alchini, Ricardo; Kazarine, Alexei; Gopal, Angelica A; Girouard, Marie-Pier; Fournier, Alyson E; Wiseman, Paul W

    2016-12-01

    Light sheet microscopy techniques have expanded with designs to address many new applications. Due to rapid advancements in computing power, camera/detector technologies, and tissue clearing techniques, light sheet methods are becoming increasingly popular for biomedical imaging applications at the cellular and tissue levels. Light sheet imaging modalities couple rapid imaging rates, low-levels of phototoxicity, and excellent signal to noise ratios, contributing to their popularity for experimental biology. However, the current major limitation of light sheet microscopy arises from optical aberrations, with the main drawback being the defocusing introduced by refractive index variations that accompany clearing techniques. Here, we propose an inexpensive and easy to build light sheet based instrumentation to overcome this limitation by optomechanically decoupling the sample scanning movement from the detection step. Our solution is relatively simple to implement and also provides increased modularity by using a swappable excitation arm. This expands the range of samples we can image on a single system, from high resolution for single cells at ? m spatial resolution, to tissues with mm spatial resolution. We demonstrate our approach, using the system to image iDISCO cleared embryos and sciatic nerves, and provide the full three-dimensional reconstruction of these objects in minutes.

  12. Acoustic Microscopy at Cryogenic Temperatures.

    Science.gov (United States)

    1982-01-01

    of counteracting the formation of shock waves in the liquid. The nonlinear distortion of the wave is equivalent to the generation of higher harmonics...z) tipo • .-0 dz (18) From (17) we see that the second harmonic beam has the amplitude cross section and phase curvature of a gaussian focused beam

  13. Wide-field imaging through scattering media by scattered light fluorescence microscopy

    Science.gov (United States)

    Zhou, Yulan; Li, Xun

    2017-08-01

    To obtain images through scattering media, scattered light fluorescence (SLF) microscopy that utilizes the optical memory effect has been developed. However, the small field of view (FOV) of SLF microscopy limits its application. In this paper, we have introduced a re-modulation method to achieve wide-field imaging through scattering media by SLF microscopy. In the re-modulation method, to raster scan the focus across the object plane, the incident wavefront is re-modulated via a spatial light modulator (SLM) in the updated phase compensation calculated using the optimized iterative algorithm. Compared with the conventional optical memory effect method, the re-modulation method can greatly increase the FOV of a SLF microscope. With the phase compensation theoretically calculated, the process of updating the phase compensation of a high speed SLM is fast. The re-modulation method does not increase the imaging time. The re-modulation method is, therefore, expected to make SLF microscopy have much wider applications in biology, medicine and physiology.

  14. Breaking the spatial resolution barrier via iterative sound-light interaction in deep tissue microscopy

    OpenAIRE

    Ke Si; Reto Fiolka; Meng Cui

    2012-01-01

    Optical microscopy has so far been restricted to superficial layers, leaving many important biological questions unanswered. Random scattering causes the ballistic focus, which is conventionally used for image formation, to decay exponentially with depth. Optical imaging beyond the ballistic regime has been demonstrated by hybrid techniques that combine light with the deeper penetration capability of sound waves. Deep inside highly scattering media, the sound focus dimensions restrict the ima...

  15. Correlative Light and Scanning X-Ray Scattering Microscopy of Healthy and Pathologic Human Bone Sections

    OpenAIRE

    Giannini, C.; D. Siliqi; Bunk, O.; Beraudi, A.; Ladisa, M.; Altamura, D.; Stea, S.; Baruffaldi, F.

    2012-01-01

    Scanning small and wide angle X-ray scattering (scanning SWAXS) experiments were performed on healthy and pathologic human bone sections. Via crystallographic tools the data were transformed into quantitative images and as such compared with circularly polarized light (CPL) microscopy images. SWAXS and CPL images allowed extracting information of the mineral nanocrystalline phase embedded, with and without preferred orientation, in the collagen fibrils, mapping local changes at sub-osteon res...

  16. Gastric Tissue Damage Analysis Generated by Ischemia: Bioimpedance, Confocal Endomicroscopy, and Light Microscopy

    Science.gov (United States)

    Beltran, Nohra E.; Garcia, Laura E.; Garcia-Lorenzana, Mario

    2013-01-01

    The gastric mucosa ischemic tissular damage plays an important role in critical care patients' outcome, because it is the first damaged tissue by compensatory mechanism during shock. The aim of the study is to relate bioimpedance changes with tissular damage level generated by ischemia by means of confocal endomicroscopy and light microscopy. Bioimpedance of the gastric mucosa and confocal images were obtained from Wistar male rats during basal and ischemia conditions. They were anesthetized, and stain was applied (fluorescein and/or acriflavine). The impedance spectroscopy catheter was inserted and then confocal endomicroscopy probe. After basal measurements and biopsy, hepatic and gastric arteries clamping induced ischemia. Finally, pyloric antrum tissue was preserved in buffered formaldehyde (10%) for histology processing using light microscopy. Confocal images were equalized, binarized, and boundary defined, and infiltrations were quantified. Impedance and infiltrations increased with ischemia showing significant changes between basal and ischemia conditions (P < 0.01). Light microscopy analysis allows detection of general alterations in cellular and tissular integrity, confirming gastric reactance and confocal images quantification increments obtained during ischemia. PMID:23841094

  17. Gastric Tissue Damage Analysis Generated by Ischemia: Bioimpedance, Confocal Endomicroscopy, and Light Microscopy

    Directory of Open Access Journals (Sweden)

    Nohra E. Beltran

    2013-01-01

    Full Text Available The gastric mucosa ischemic tissular damage plays an important role in critical care patients’ outcome, because it is the first damaged tissue by compensatory mechanism during shock. The aim of the study is to relate bioimpedance changes with tissular damage level generated by ischemia by means of confocal endomicroscopy and light microscopy. Bioimpedance of the gastric mucosa and confocal images were obtained from Wistar male rats during basal and ischemia conditions. They were anesthetized, and stain was applied (fluorescein and/or acriflavine. The impedance spectroscopy catheter was inserted and then confocal endomicroscopy probe. After basal measurements and biopsy, hepatic and gastric arteries clamping induced ischemia. Finally, pyloric antrum tissue was preserved in buffered formaldehyde (10% for histology processing using light microscopy. Confocal images were equalized, binarized, and boundary defined, and infiltrations were quantified. Impedance and infiltrations increased with ischemia showing significant changes between basal and ischemia conditions (. Light microscopy analysis allows detection of general alterations in cellular and tissular integrity, confirming gastric reactance and confocal images quantification increments obtained during ischemia.

  18. Multiphoton microscopy and image guided light activated therapy using nanomaterials (Conference Presentation)

    Science.gov (United States)

    Prasad, Paras N.

    2017-02-01

    This talk will focus on design and applications of nanomaterials exhibiting strong multiphoton upconversion for multiphoton microscopy as well as for image-guided and light activated therapy .1-3 Such processes can occur by truly nonlinear optical interactions proceeding through virtual intermediate states or by stepwise coupled linear excitations through real intermediate states. Multiphoton processes in biocompatible multifunctional nanoparticles allow for 3D deep tissue imaging. In addition, they can produce in-situ photon conversion of deep tissue penetrating near IR light into a needed shorter wavelength light for photo-activated therapy at a targeted site, thus overcoming the limited penetration of UV or visible light into biological media. We are using near IR emitters such as silicon quantum dots which also exhibit strong multiphoton excitation for multiphoton microscopy. Another approach involves nonlinear nanocrystals such as ZnO which can produce four wave mixing, sum frequency generation as well as second harmonic generation to convert a deep tissue penetrating Near IR light at the targeted biological site to a desired shorter wavelength light suitable for bio imaging or activation of a therapy. We have utilized this approach to activate a photosensitizer for photodynamic therapy. Yet another type of upconversion materials is rare-earth ion doped optical nanotransformers which transform a Near IR (NIR) light from an external source by sequential single photon absorption, in situ and on demand, to a needed wavelength. Applications of these nanotransformers in multiphoton photoacoustic imaging will also be presented. An exciting direction pursued by us using these multiphoton nanoparticles, is functional imaging of brain. Simultaneously, they can effect optogenetics for regioselective stimulation of neurons for providing an effective intervention/augmentation strategy to enhance the cognitive state and lead to a foundation for futuristic vision of super

  19. Automatic and adaptive heterogeneous refractive index compensation for light-sheet microscopy.

    Science.gov (United States)

    Ryan, Duncan P; Gould, Elizabeth A; Seedorf, Gregory J; Masihzadeh, Omid; Abman, Steven H; Vijayaraghavan, Sukumar; Macklin, Wendy B; Restrepo, Diego; Shepherd, Douglas P

    2017-09-20

    Optical tissue clearing has revolutionized researchers' ability to perform fluorescent measurements of molecules, cells, and structures within intact tissue. One common complication to all optically cleared tissue is a spatially heterogeneous refractive index, leading to light scattering and first-order defocus. We designed C-DSLM (cleared tissue digital scanned light-sheet microscopy) as a low-cost method intended to automatically generate in-focus images of cleared tissue. We demonstrate the flexibility and power of C-DSLM by quantifying fluorescent features in tissue from multiple animal models using refractive index matched and mismatched microscope objectives. This includes a unique measurement of myelin tracks within intact tissue using an endogenous fluorescent reporter where typical clearing approaches render such structures difficult to image. For all measurements, we provide independent verification using standard serial tissue sectioning and quantification methods. Paired with advancements in volumetric image processing, C-DSLM provides a robust methodology to quantify sub-micron features within large tissue sections.Optical clearing of tissue has enabled optical imaging deeper into tissue due to significantly reduced light scattering. Here, Ryan et al. tackle first-order defocus, an artefact of a non-uniform refractive index, extending light-sheet microscopy to partially cleared samples.

  20. Interaction of light and surface plasmon polaritons in Ag islands studied by nonlinear photoemission microscopy.

    Science.gov (United States)

    Buckanie, N M; Kirschbaum, P; Sindermann, S; Meyer zu Heringdorf, F-J

    2013-07-01

    Two photon photoemission microscopy was used to study the interaction of femtosecond laser pulses with Ag islands prepared using different strategies on Si(111) and SiO₂. The femtosecond laser pulses initiate surface plasmon polariton (SPP) waves at the edges of the island. The superposition of the electrical fields of the femtosecond laser pulses with the electrical fields of the SPP results in a moiré pattern that is comparable despite the rather different methods of preparation and that gives access to the wavelength and direction of the SPP waves. If the SPPs reach edges of the Ag islands, they can be converted back into light waves. The incident and refracted light waves result in an interference pattern that can again be described with a moiré pattern, demonstrating that Ag islands can be used as plasmonic beam deflectors for light.

  1. Macro-optical trapping for sample confinement in light sheet microscopy.

    Science.gov (United States)

    Yang, Zhengyi; Piksarv, Peeter; Ferrier, David E K; Gunn-Moore, Frank J; Dholakia, Kishan

    2015-08-01

    Light sheet microscopy is a powerful approach to construct three-dimensional images of large specimens with minimal photo-damage and photo-bleaching. To date, the specimens are usually mounted in agents such as agarose, potentially restricting the development of live samples, and also highly mobile specimens need to be anaesthetized before imaging. To overcome these problems, here we demonstrate an integrated light sheet microscope which solely uses optical forces to trap and hold the sample using a counter-propagating laser beam geometry. Specifically, tobacco plant cells and living Spirobranchus lamarcki larvae were successfully trapped and sectional images acquired. This novel approach has the potential to significantly expand the range of applications for light sheet imaging.

  2. Two-photon microscopy with diffractive optical elements and spatial light modulators

    Directory of Open Access Journals (Sweden)

    Brendon O Watson

    2010-09-01

    Full Text Available Two-photon microscopy is often performed at slow frame rates, due to the need to serially scan all points in a field of view with a single laser beam. To overcome this problem, we have developed two optical methods that split and multiplex a laser beam across the sample. In the first method a diffractive optical element (DOE generates a fixed number of beamlets that are scanned in parallel, resulting in a corresponding increase in speed, or in signal-to-noise ratio, in time-lapse measurements. The second method uses a computer-controlled spatial light modulator (SLM, to generate any arbitrary spatio-temporal light pattern. With an SLM one can image or photostimulate any predefined region of the image, such as neurons or dendritic spines. In addition, SLMs can be used to mimic a large number of optical transfer functions, including light path corrections or as adaptive optical devices.

  3. Enhanced image reconstruction of three-dimensional fluorescent assays by subtractive structured-light illumination microscopy.

    Science.gov (United States)

    Choi, Jong-ryul; Kim, Donghyun

    2012-10-01

    We investigate improved image reconstruction of structured light illumination for high-resolution imaging of three-dimensional (3D) cell-based assays. For proof of concept, an in situ fluorescence optical detection system was built with a digital micromirror device as a spatial light modulator, for which phase and tilting angle in a grid pattern were varied to implement specific image reconstruction schemes. Subtractive reconstruction algorithms based on structured light illumination were used to acquire images of fluorescent microbeads deposited as a two-dimensional monolayer or in 3D alginate matrix. We have confirmed that an optical subtraction algorithm improves axial and lateral resolution by effectively removing out-of-focus fluorescence. The results suggest that subtractive image reconstruction can be useful for structured illumination microscopy of broad types of cell-based assays with high image resolution.

  4. High resolution x-ray Thomson scattering measurements from cryogenic hydrogen jets using the linac coherent light source.

    Science.gov (United States)

    Fletcher, L B; Zastrau, U; Galtier, E; Gamboa, E J; Goede, S; Schumaker, W; Ravasio, A; Gauthier, M; MacDonald, M J; Chen, Z; Granados, E; Lee, H J; Fry, A; Kim, J B; Roedel, C; Mishra, R; Pelka, A; Kraus, D; Barbrel, B; Döppner, T; Glenzer, S H

    2016-11-01

    We present the first spectrally resolved measurements of x-rays scattered from cryogenic hydrogen jets in the single photon counting limit. The 120 Hz capabilities of the LCLS, together with a novel hydrogen jet design [J. B. Kim et al., Rev. Sci. Instrum. (these proceedings)], allow for the ability to record a near background free spectrum. Such high-dynamic-range x-ray scattering measurements enable a platform to study ultra-fast, laser-driven, heating dynamics of hydrogen plasmas. This measurement has been achieved using two highly annealed pyrolytic graphite crystal spectrometers to spectrally resolve 5.5 keV x-rays elastically and inelastically scattered from cryogenic hydrogen and focused on Cornell-SLAC pixel array detectors [S. Herrmann et al., Nucl. Instrum. Methods Phys. Res., Sect. A 718, 550 (2013)].

  5. Making Mass Spectrometry See the Light: The Promises and Challenges of Cryogenic Infrared Ion Spectroscopy as a Bioanalytical Technique.

    Science.gov (United States)

    Cismesia, Adam P; Bailey, Laura S; Bell, Matthew R; Tesler, Larry F; Polfer, Nicolas C

    2016-05-01

    The detailed chemical information contained in the vibrational spectrum of a cryogenically cooled analyte ion would, in principle, make infrared (IR) ion spectroscopy a gold standard technique for molecular identification in mass spectrometry. Despite this immense potential, there are considerable challenges in both instrumentation and methodology to overcome before the technique is analytically useful. Here, we discuss the promise of IR ion spectroscopy for small molecule analysis in the context of metabolite identification. Experimental strategies to address sensitivity constraints, poor overall duty cycle, and speed of the experiment are intimately tied to the development of a mass-selective cryogenic trap. Therefore, the most likely avenues for success, in the authors' opinion, are presented here, alongside alternative approaches and some thoughts on data interpretation.

  6. High resolution x-ray Thomson scattering measurements from cryogenic hydrogen jets using the linac coherent light source

    Science.gov (United States)

    Fletcher, L. B.; Zastrau, U.; Galtier, E.; Gamboa, E. J.; Goede, S.; Schumaker, W.; Ravasio, A.; Gauthier, M.; MacDonald, M. J.; Chen, Z.; Granados, E.; Lee, H. J.; Fry, A.; Kim, J. B.; Roedel, C.; Mishra, R.; Pelka, A.; Kraus, D.; Barbrel, B.; Döppner, T.; Glenzer, S. H.

    2016-11-01

    We present the first spectrally resolved measurements of x-rays scattered from cryogenic hydrogen jets in the single photon counting limit. The 120 Hz capabilities of the LCLS, together with a novel hydrogen jet design [J. B. Kim et al., Rev. Sci. Instrum. (these proceedings)], allow for the ability to record a near background free spectrum. Such high-dynamic-range x-ray scattering measurements enable a platform to study ultra-fast, laser-driven, heating dynamics of hydrogen plasmas. This measurement has been achieved using two highly annealed pyrolytic graphite crystal spectrometers to spectrally resolve 5.5 keV x-rays elastically and inelastically scattered from cryogenic hydrogen and focused on Cornell-SLAC pixel array detectors [S. Herrmann et al., Nucl. Instrum. Methods Phys. Res., Sect. A 718, 550 (2013)].

  7. High resolution x-ray Thomson scattering measurements from cryogenic hydrogen jets using the linac coherent light source

    Energy Technology Data Exchange (ETDEWEB)

    Fletcher, L. B., E-mail: lbfletch@slac.stanford.edu; Galtier, E.; Gamboa, E. J.; Schumaker, W.; Gauthier, M.; Granados, E.; Lee, H. J.; Fry, A.; Kim, J. B.; Roedel, C.; Mishra, R.; Glenzer, S. H. [SLAC National Accelerator Laboratory, Menlo Park, California 94025 (United States); Zastrau, U. [European XFEL, Schenefeld (Germany); Goede, S. [SLAC National Accelerator Laboratory, Menlo Park, California 94025 (United States); European XFEL, Schenefeld (Germany); Ravasio, A. [SLAC National Accelerator Laboratory, Menlo Park, California 94025 (United States); Laboratoire pour l’Utilisation des Lasers Intenses, Palaiseau Cedex (France); MacDonald, M. J. [SLAC National Accelerator Laboratory, Menlo Park, California 94025 (United States); University of Michigan, Ann Arbor, Michigan 48109 (United States); Chen, Z. [SLAC National Accelerator Laboratory, Menlo Park, California 94025 (United States); University of Alberta, Edmonton, Alberta T6G 2R3 (Canada); Pelka, A. [Helmholtz Zentrum Dresden-Rossendorf, Dresden (Germany); Kraus, D. [Helmholtz Zentrum Dresden-Rossendorf, Dresden (Germany); Department of Physics, University of California Berkeley, Berkeley, California 94720 (United States); Barbrel, B. [Department of Physics, University of California Berkeley, Berkeley, California 94720 (United States); and others

    2016-11-15

    We present the first spectrally resolved measurements of x-rays scattered from cryogenic hydrogen jets in the single photon counting limit. The 120 Hz capabilities of the LCLS, together with a novel hydrogen jet design [J. B. Kim et al., Rev. Sci. Instrum. (these proceedings)], allow for the ability to record a near background free spectrum. Such high-dynamic-range x-ray scattering measurements enable a platform to study ultra-fast, laser-driven, heating dynamics of hydrogen plasmas. This measurement has been achieved using two highly annealed pyrolytic graphite crystal spectrometers to spectrally resolve 5.5 keV x-rays elastically and inelastically scattered from cryogenic hydrogen and focused on Cornell-SLAC pixel array detectors [S. Herrmann et al., Nucl. Instrum. Methods Phys. Res., Sect. A 718, 550 (2013)].

  8. Axial Phase-Darkfield-Contrast (APDC), a new technique for variable optical contrasting in light microscopy.

    Science.gov (United States)

    Piper, T; Piper, J

    2012-09-01

    Axial phase-darkfield-contrast (APDC) has been developed as an illumination technique in light microscopy which promises significant improvements and a higher variability in imaging of several transparent 'problem specimens'. With this method, a phase contrast image is optically superimposed on an axial darkfield image so that a partial image based on the principal zeroth order maximum (phase contrast) interferes with an image, which is based on the secondary maxima (axial darkfield). The background brightness and character of the resulting image can be continuously modulated from a phase contrast-dominated to a darkfield-dominated character. In order to achieve this illumination mode, normal objectives for phase contrast have to be fitted with an additional central light stopper needed for axial (central) darkfield illumination. In corresponding condenser light masks, a small perforation has to be added in the centre of the phase contrast providing light annulus. These light modulating elements are properly aligned when the central perforation is congruent with the objective's light stop and the light annulus is conjugate with the phase ring. The breadth of the condenser light annulus and thus the intensity of the phase contrast partial image can be regulated with the aperture diaphragm. Additional contrast effects can be achieved when both illuminating light components are filtered at different colours. In this technique, the axial resolution (depth of field) is significantly enhanced and the specimen's three-dimensional appearance is accentuated with improved clarity as well as fine details at the given resolution limit. Typical artefacts associated with phase contrast and darkfield illumination are reduced in our methods.

  9. Correlation of morphological alterations of light and electron microscopy in chronic type B and C hepatitis.

    Science.gov (United States)

    Kasprzak, A; Biczysko, W; Adamek, A; Zabel, M; Surdyk-Zasada, J

    2001-05-01

    Chronic type B and C hepatitis involves inflammatory lesions of a variable intensity and variably advanced fibrosis. Considering current, progressively growing requirements for correct evaluation of lesions in liver biopsies, an attempt was made to appraise suitability of selected techniques for a broadened histopathological diagnosis. The lesions were evaluated at the level of light and electron microscopy. Material for the study consisted of liver biopsies obtained from adults and children (n = 60) with serological markers of chronic type B or type C hepatitis. Routine techniques of staining for light and electron microscopy, as well as the techniques of Brachet and Feulgen, were applied. HBcAg expression and HBV-DNA detection in children with chronic type B hepatitis were studied employing the avidin-biotin peroxidase complex (ABC) technique and in situ hybridisation with the ImmunoMax signal amplification. Slight or moderately intense inflammatory lesions (grading of 1 to 2 points) and a low level of fibrosis advancement (staging of 1 to 2 points) prevailed in the material, independently of the etiologic agent involved and age of the patient. Both in children and in adults, extensive lesions in the nuclear chromatin represented the common trait of chronic type B and type C hepatitis examined by light microscopy. Ultrastructural patterns confirmed the lesions and demonstrated virus-resembling particles in the cell nuclei. In HCV infection, hepatocyte cytoplasm contained tubular and horseshoe-shaped structures with lesions of mitochondria, while in HBV infection Dane's particles and tubular forms of HBsAg were detected. For cognitive reasons and due to frequently equivocal literature data, our data on ultrastructural lesions in chronic type C hepatitis seem to be of particular interest. Using the ImmunoMax signal amplification, we were able to diagnose HBV infection under light microscope and to define stage of the infection. Their sensitivity, specificity and

  10. Application of the fluorescence light microscopy in the textural study of portland cement clinker

    Directory of Open Access Journals (Sweden)

    Montoto San Miguel, Modesto

    1987-03-01

    Full Text Available The application of fluorescence light microscopy in the textural study of Portland cement clinker, specially its porosity, is presented. Principles and types of the technique are commented and the suggested sample preparation method is described. The use of fluorescence microscopy allows an easier study of the clinker porosity, and very proper images for automated quantification can be obtained. Besides, the samples can also be observed by reflected-light polarizing microscopy.

    Se presenta la utilidad de la microscopía óptica de fluorescencia para el estudio textural del clínker de cemento Portland, especialmente su porosidad. Se comentan los fundamentos y modalidades de la técnica, y se describe el método recomendado de preparación de muestras. La utilización de la microscopía de fluorescencia permite un estudio más fácil de la porosidad, obteniéndose imágenes muy apropiadas para su cuantificación mediante técnicas automatizadas. Además, las muestras para fluorescencia pueden ser estudiadas complementariamente por microscopía óptica de polarización por luz reflejada.

  11. Light microscopy morphological characteristics of the sperm flagellum may be related to axonemal abnormalities.

    Science.gov (United States)

    Mitchell, V; Sigala, J; Ballot, C; Jumeau, F; Barbotin, A L; Duhamel, A; Rives, N; Rigot, J M; Escalier, D; Peers, M C

    2015-03-01

    Although electron microscopy provides a detailed analysis of ultrastructural abnormalities, this technique is not available in all laboratories. We sought to determine whether certain characteristics of the flagellum as assessed by light microscopy were related to axonemal abnormalities. Forty-one patients with an absence of outer dynein arms (type I), a lack of a central complex (type III) and an absence of peripheral doublets (type IV) were studied. Sperm morphology was scored according to David's modified classification. Flagella with an irregular thickness were classified as being of normal length, short or broken. There were correlations between missing outer dynein arms and abnormal, short or coiled flagellum. Type III patients showed the highest flagellar defects (a short (P = 0.0027) or an absent flagellum (P = 0.011)). Just over 68% of the irregular flagella were short in Type III patients, whereas this value was only 34.5% in type I and 26.4% in type IV (P = 0.002). There was a negative correlation between misassembly and spermatozoa of irregular flagella (r = -0.79; P = 0.019). It is concluded that light microscopy analysis of flagellum abnormalities may help provide a correct diagnosis, identify sperm abnormalities with fertility potentials and outcomes in assisted reproduction technologies and assess the genetic risk.

  12. Developing methods based on light sheet fluorescence microscopy for biophysical investigations of larval zebrafish

    Science.gov (United States)

    Taormina, Michael J.

    Adapting the tools of optical microscopy to the large-scale dynamic systems encountered in the development of multicellular organisms provides a path toward understanding the physical processes necessary for complex life to form and function. Obtaining quantitatively meaningful results from such systems has been challenging due to difficulty spanning the spatial and temporal scales representative of the whole, while also observing the many individual members from which complex and collective behavior emerges. A three-dimensional imaging technique known as light sheet fluorescence microscopy provides a number of significant benefits for surmounting these challenges and studying developmental systems. A thin plane of fluorescence excitation light is produced such that it coincides with the focal plane of an imaging system, providing rapid acquisition of optically sectioned images that can be used to construct a three-dimensional rendition of a sample. I discuss the implementation of this technique for use in larva of the model vertebrate Danio rerio (zebrafish). The nature of light sheet imaging makes it especially well suited to the study of large systems while maintaining good spatial resolution and minimizing damage to the specimen from excessive exposure to excitation light. I show the results from a comparative study that demonstrates the ability to image certain developmental processes non-destructively, while in contrast confocal microscopy results in abnormal growth due to phototoxicity. I develop the application of light sheet microscopy to the study of a previously inaccessible system: the bacterial colonization of a host organism. Using the technique, we are able to obtain a survey of the intestinal tract of a larval zebrafish and observe the location of microbes as they grow and establish a stable population in an initially germ free fish. Finally, I describe a new technique to measure the fluid viscosity of this intestinal environment in vivo using

  13. Interaction of light and surface plasmon polaritons in Ag Islands studied by nonlinear photoemission microscopy

    Energy Technology Data Exchange (ETDEWEB)

    Buckanie, N.M.; Kirschbaum, P.; Sindermann, S.; Heringdorf, F.-J. Meyer zu, E-mail: meyerzh@uni-due.de

    2013-07-15

    Two photon photoemission microscopy was used to study the interaction of femtosecond laser pulses with Ag islands prepared using different strategies on Si(111) and SiO{sub 2}. The femtosecond laser pulses initiate surface plasmon polariton (SPP) waves at the edges of the island. The superposition of the electrical fields of the femtosecond laser pulses with the electrical fields of the SPP results in a moiré pattern that is comparable despite the rather different methods of preparation and that gives access to the wavelength and direction of the SPP waves. If the SPPs reach edges of the Ag islands, they can be converted back into light waves. The incident and refracted light waves result in an interference pattern that can again be described with a moiré pattern, demonstrating that Ag islands can be used as plasmonic beam deflectors for light. - Highlights: • Surface plasmon polaritons were studied on Ag islands in two photon photoemission microscopy. • Ag islands were prepared using self-assembly, electron beam lithography, and a focused ion beam. • The SPP pattern on Ag islands can be described with a simple moiré concept. • SPP output coupling results in a pattern that can again be described by the moiré effect.

  14. Balamuthia mandrillaris: Further morphological observations of trophozoites by light, scanning and transmission electron microscopy.

    Science.gov (United States)

    González-Robles, Arturo; Lares-Villa, Fernando; Lares-Jiménez, Luis Fernando; Omaña-Molina, Maritza; Salazar-Villatoro, Lizbeth; Martínez-Palomo, Adolfo

    2015-10-01

    Additional morphological features of Balamuthia mandrillaris observed by light and electron microscopy are reported. Trophozoites were extremely pleomorphic: their cell shapes ranged from rounded to elongated and sometimes they appeared exceptionally stretched out and branched. By transmission electron microscopy it was possible to observe two different cytoplasmic areas, the ectoplasm and the endoplasm and often sections of rough endoplasmic reticulum were found in the transition zone. The cytoplasm was very fibrogranular and most of the organelles typically found in eukaryotic cells were observed. A particular finding was the presence of numerous mitochondria with a different structure from those of other free-living amoebae. The observations reported here may reinforce the morphological knowledge of this amoeba and provide a background for further analyses.

  15. Lipid vesicle shape analysis from populations using light video microscopy and computer vision.

    Directory of Open Access Journals (Sweden)

    Jernej Zupanc

    Full Text Available We present a method for giant lipid vesicle shape analysis that combines manually guided large-scale video microscopy and computer vision algorithms to enable analyzing vesicle populations. The method retains the benefits of light microscopy and enables non-destructive analysis of vesicles from suspensions containing up to several thousands of lipid vesicles (1-50 µm in diameter. For each sample, image analysis was employed to extract data on vesicle quantity and size distributions of their projected diameters and isoperimetric quotients (measure of contour roundness. This process enables a comparison of samples from the same population over time, or the comparison of a treated population to a control. Although vesicles in suspensions are heterogeneous in sizes and shapes and have distinctively non-homogeneous distribution throughout the suspension, this method allows for the capture and analysis of repeatable vesicle samples that are representative of the population inspected.

  16. Morphology of the dentin structure of sloths Bradypus tridactylus: a light and scanning electron microscopy investigation.

    Science.gov (United States)

    Santana, L N S; Barbosa, L V M; Teixeira, F B; Costa, A M P; Fernandes, L M P; Lima, R R

    2013-12-01

    The aim of this study was to describe the dentine morphology of sloths (Bradypus tridactylus). The sloth teeth were removed and prepared for light microscopy (LM) and scanning electron microscopy analyses (SEM). LM revealed two patterns of tubular dentins: an outer with dentinary tubules over the all tooth length and one in the inner part with larger diameter and more spaced tubules, when compared to those present in the outer dentine. These findings were confirmed by SEM, which revealed a tubular pattern in the outer dentine like in humans. The inner dentine displayed pared grouped tubules that were characterized as vascular channels. It can be concluded that this sloth species present two types of dentins: an inner dentin (ortodentin) and an outer dentin characterized as a vascular dentin. This suggests a partial evolutive/adaptive process of this dental tissue, as compared to other mammalian species.

  17. Adaptive light-sheet microscopy for long-term, high-resolution imaging in living organisms.

    Science.gov (United States)

    Royer, Loïc A; Lemon, William C; Chhetri, Raghav K; Wan, Yinan; Coleman, Michael; Myers, Eugene W; Keller, Philipp J

    2016-12-01

    Optimal image quality in light-sheet microscopy requires a perfect overlap between the illuminating light sheet and the focal plane of the detection objective. However, mismatches between the light-sheet and detection planes are common owing to the spatiotemporally varying optical properties of living specimens. Here we present the AutoPilot framework, an automated method for spatiotemporally adaptive imaging that integrates (i) a multi-view light-sheet microscope capable of digitally translating and rotating light-sheet and detection planes in three dimensions and (ii) a computational method that continuously optimizes spatial resolution across the specimen volume in real time. We demonstrate long-term adaptive imaging of entire developing zebrafish (Danio rerio) and Drosophila melanogaster embryos and perform adaptive whole-brain functional imaging in larval zebrafish. Our method improves spatial resolution and signal strength two to five-fold, recovers cellular and sub-cellular structures in many regions that are not resolved by non-adaptive imaging, adapts to spatiotemporal dynamics of genetically encoded fluorescent markers and robustly optimizes imaging performance during large-scale morphogenetic changes in living organisms.

  18. Tissue and cellular localization of tannins in Tunisian dates (Phoenix dactylifera L.) by light and transmission electron microscopy.

    Science.gov (United States)

    Hammouda, Hédi; Alvarado, Camille; Bouchet, Brigitte; Kalthoum-Chérif, Jamila; Trabelsi-Ayadi, Malika; Guyot, Sylvain

    2014-07-16

    A histological approach including light microscopy and transmission electron microscopy (TEM) was used to provide accurate information on the localization of condensed tannins in the edible tissues and in the stone of date fruits (Phoenix dactylifera L.). Light microscopy was carried out on fresh tissues after staining by 4-dimethylaminocinnamaldehyde (DMACA) for a specific detection of condensed tannins. Thus, whether under light microscopy or transmission electron microscopy (TEM), results showed that tannins are not located in the epidermis but more deeply in the mesocarp in the vacuole of very large cells. Regarding the stones, tannins are found in a specific cell layer located at 50 μm from the sclereid cells of the testa.

  19. Comparing the use of virtual and conventional light microscopy in practical sessions: Virtual reality in Tabuk University

    Directory of Open Access Journals (Sweden)

    Ayman F.A. Foad, MD

    2017-04-01

    We randomly assigned two groups of second-year medical students from the University of Tabuk in KSA to use either conventional light or virtual microscopy practical sessions. The students' perceptions were assessed by written and practical exams. Students in the virtual microscopy group performed better than those in the light microscopy group in both practical and written exams, as reflected by their more-uniform performance and less-scattered grades. The virtual microscopy group had the advantage of optional online off-campus access to study materials, which they spent an average of 2.5 h reviewing. Virtual microscopy is a valid educational tool that can augment conventional microscopy in pathology practical sessions, and its application is convenient for both students and staff.

  20. Simultaneous whole-animal 3D-imaging of neuronal activity using light field microscopy

    CERN Document Server

    Prevedel, R; Hoffmann, M; Pak, N; Wetzstein, G; Kato, S; Schrödel, T; Raskar, R; Zimmer, M; Boyden, E S; Vaziri, A

    2014-01-01

    3D functional imaging of neuronal activity in entire organisms at single cell level and physiologically relevant time scales faces major obstacles due to trade-offs between the size of the imaged volumes, and spatial and temporal resolution. Here, using light-field microscopy in combination with 3D deconvolution, we demonstrate intrinsically simultaneous volumetric functional imaging of neuronal population activity at single neuron resolution for an entire organism, the nematode Caenorhabditis elegans. The simplicity of our technique and possibility of the integration into epi-fluoresence microscopes makes it an attractive tool for high-speed volumetric calcium imaging.

  1. Calcium imaging of neural circuits with extended depth-of-field light-sheet microscopy.

    Science.gov (United States)

    Quirin, Sean; Vladimirov, Nikita; Yang, Chao-Tsung; Peterka, Darcy S; Yuste, Rafael; Ahrens, Misha B

    2016-03-01

    Increasing the volumetric imaging speed of light-sheet microscopy will improve its ability to detect fast changes in neural activity. Here, a system is introduced for brain-wide imaging of neural activity in the larval zebrafish by coupling structured illumination with cubic phase extended depth-of-field (EDoF) pupil encoding. This microscope enables faster light-sheet imaging and facilitates arbitrary plane scanning-removing constraints on acquisition speed, alignment tolerances, and physical motion near the sample. The usefulness of this method is demonstrated by performing multi-plane calcium imaging in the fish brain with a 416×832×160  μm field of view at 33 Hz. The optomotor response behavior of the zebrafish is monitored at high speeds, and time-locked correlations of neuronal activity are resolved across its brain.

  2. Topography and refractometry of nanostructures using spatial light interference microscopy (SLIM)

    Science.gov (United States)

    Wang, Zhuo; Chun, Ik Su; Li, Xiuling; Ong, Zhun-Yong; Pop, Eric; Millet, Larry; Gillette, Martha; Popescu, Gabriel

    2010-01-01

    Spatial Light Interference Microscopy (SLIM) is a novel method developed in our laboratory that provides quantitative phase images of transparent structures with 0.3 nm spatial and 0.03 nm temporal accuracy owing to the white light illumination and its common path interferometric geometry. We exploit these features and demonstrate SLIM's ability to perform topography at a single atomic layer in graphene. Further, using a decoupling procedure that we developed for cylindrical structures, we extract the axially-averaged refractive index of semiconductor nanotubes and a neurite of a live hippocampal neuron in culture. We believe that this study will set the basis for novel high-throughput topography and refractometry of man-made and biological nanostructures. PMID:20081970

  3. Feedback phase correction of Bessel beams in confocal line light-sheet microscopy: a simulation study.

    Science.gov (United States)

    Moosavi, S Hoda; Gohn-Kreuz, Cristian; Rohrbach, Alexander

    2013-08-10

    Confocal line detection has been shown to improve contrast in light-sheet-based microscopy especially when illuminating the sample by Bessel beams. Besides their self-reconstructing capability, the stability in propagation direction of Bessel beams allows to block the unwanted emission light from the Bessel beam's ring system. However, due to phase aberrations induced especially at the border of the specimen, Bessel beams may not propagate along lines parallel to the slit detector. Here we present a concept of how to correct the phase of each incident Bessel beam such that the efficiency of confocal line detection is improved by up to 200%-300%. The applicability of the method is verified by the results obtained from numerical simulations based on the beam propagation method.

  4. Precise measurement of the resolution in light microscopy using Fourier transform

    Science.gov (United States)

    Vainrub, Arnold

    2008-04-01

    The resolution power of light microscope has been accurately measured (±5%) by Fourier transform of various object images and further evaluation of the highest spatial frequency in Fourier spectrum. Any unknown shape plane object with a shape feature's size smaller than the resolution to be measured was shown to provide a reliable resolution test. This simple method gives a direct measurement of the resolution power as defined by Abbe [Archiv. F. Mikroskopische Anat. 9, 413 (1873)]. The results have been justified by comparison to a standard resolution measurement by using calibrated periodic line patterns. Notably, the approach is applicable in super-resolution light microscopy (transmission, reflection, and fluorescence), where calibrated resolution targets do not occur. It was conveniently implemented by using a compact disk as a test object and free IMAGEJ imaging software.

  5. In situ transmission electron microscopy of light-induced photocatalytic reactions.

    Science.gov (United States)

    Cavalca, F; Laursen, A B; Kardynal, B E; Dunin-Borkowski, R E; Dahl, S; Wagner, J B; Hansen, T W

    2012-02-24

    Transmission electron microscopy (TEM) makes it possible to obtain insight into the structure, composition and reactivity of photocatalysts, which are of fundamental interest for sustainable energy research. Such insight can be used for further material optimization. Here, we combine conventional TEM analysis of photocatalysts with environmental TEM (ETEM) and photoactivation using light. Two novel types of TEM specimen holder that enable in situ illumination are developed to study light-induced phenomena in photoactive materials, systems and photocatalysts at the nanoscale under working conditions. The technological development of the holders is described and two representative photo-induced phenomena are studied: the photodegradation of Cu₂O and the photodeposition of Pt onto a GaN:ZnO photocatalyst.

  6. Cryogenic Systems

    Science.gov (United States)

    Hosoyama, Kenji

    2002-02-01

    In this lecture we discuss the principle of method of cooling to a very low temperature, i.e. cryogenic. The "gas molecular model" will be introduced to explain the mechanism cooling by the expansion engine and the Joule-Thomson expansion valve. These two expansion processes are normally used in helium refrigeration systems to cool the process gas to cryogenic temperature. The reverse Carnot cycle will be discussed in detail as an ideal refrigeration cycle. First the fundamental process of liquefaction and refrigeration cycles will be discussed, and then the practical helium refrigeration system. The process flow of the system and the key components; -compressor, expander, and heat exchanger- will be discussed. As an example of an actual refrigeration system, we will use the cryogenic system for the KEKB superconducting RF cavity. We will also discuss the liquid helium distribution system, which is very important, especially for the cryogenic systems used in accelerator applications. 1 Principles of Cooling and Fundamental Cooling Cycle 2 Expansion engine, Joule-Thomson expansion, kinetic molecular theory, and enthalpy 3 Liquefaction Systems 4 Refrigeration Systems 5 Practical helium liquefier/refrigeration system 6 Cryogenic System for TRISTAN Superconducting RF Cavity

  7. Enhanced light element imaging in atomic resolution scanning transmission electron microscopy

    Energy Technology Data Exchange (ETDEWEB)

    Findlay, S.D., E-mail: scott.findlay@monash.edu [School of Physics, Monash University, Victoria 3800 (Australia); Kohno, Y. [JEOL Ltd., Tokyo 196-8558 (Japan); Cardamone, L.A. [School of Physics, Monash University, Victoria 3800 (Australia); Ikuhara, Y. [Institute of Engineering Innovation, School of Engineering, University of Tokyo, Tokyo 113-8656 (Japan); Nanostructures Research Laboratory, Japan Fine Ceramics Center, Nagoya 456-8587 (Japan); WPI Advanced Institute for Materials Research, Tohoku University, Sendai 980-8577 (Japan); Shibata, N. [Institute of Engineering Innovation, School of Engineering, University of Tokyo, Tokyo 113-8656 (Japan); PRESTO, Japan Science and Technology Agency, Saitama 332-0012 (Japan)

    2014-01-15

    We show that an imaging mode based on taking the difference between signals recorded from the bright field (forward scattering region) in atomic resolution scanning transmission electron microscopy provides an enhancement of the detectability of light elements over existing techniques. In some instances this is an enhancement of the visibility of the light element columns relative to heavy element columns. In all cases explored it is an enhancement in the signal-to-noise ratio of the image at the light column site. The image formation mechanisms are explained and the technique is compared with earlier approaches. Experimental data, supported by simulation, are presented for imaging the oxygen columns in LaAlO{sub 3}. Case studies looking at imaging hydrogen columns in YH{sub 2} and lithium columns in Al{sub 3}Li are also explored through simulation, particularly with respect to the dependence on defocus, probe-forming aperture angle and detector collection aperture angles. - Author-Highlights: • We present a method for enhancing the visibility and reliability of imaging light elements in STEM. • The method involves taking the difference between signals on separate bright field detectors. • Experimental data for LaAlO{sub 3} are presented, and are shown to compare favourably with simulation. • Optimum imaging parameters are explored through simulation.

  8. Direct imaging of phase objects enables conventional deconvolution in bright field light microscopy.

    Directory of Open Access Journals (Sweden)

    Carmen Noemí Hernández Candia

    Full Text Available In transmitted optical microscopy, absorption structure and phase structure of the specimen determine the three-dimensional intensity distribution of the image. The elementary impulse responses of the bright field microscope therefore consist of separate absorptive and phase components, precluding general application of linear, conventional deconvolution processing methods to improve image contrast and resolution. However, conventional deconvolution can be applied in the case of pure phase (or pure absorptive objects if the corresponding phase (or absorptive impulse responses of the microscope are known. In this work, we present direct measurements of the phase point- and line-spread functions of a high-aperture microscope operating in transmitted bright field. Polystyrene nanoparticles and microtubules (biological polymer filaments serve as the pure phase point and line objects, respectively, that are imaged with high contrast and low noise using standard microscopy plus digital image processing. Our experimental results agree with a proposed model for the response functions, and confirm previous theoretical predictions. Finally, we use the measured phase point-spread function to apply conventional deconvolution on the bright field images of living, unstained bacteria, resulting in improved definition of cell boundaries and sub-cellular features. These developments demonstrate practical application of standard restoration methods to improve imaging of phase objects such as cells in transmitted light microscopy.

  9. Electron and Light Microscopy Techniques Suitable for Studying Fatigue Damage in a Crystallized Glass Ceramic

    Science.gov (United States)

    Harrell, Shelley; Zaretsky, Erwin V.

    1961-01-01

    The crystals of Pyroceram are randomly oriented and highly reflective so that standard microscopy techniques are not satisfactory for studying this material. Standard replicating procedures proved difficult to use. New microscopy techniques and procedures have therefore been developed. A method for locating, orienting, and identifying specific areas to be viewed with an electron microscope is described. This method not require any special equipment. Plastic replicas were found to be unsatisfactory because of their tendency to adhere to Pryoceram. This caused them to tear when released or resulted in artifacts. Preshadowed silicon monoxide replicas were satisfactory but required a releasing agent. A method of depositing the releasing agent is described. To polish specimens without evidence of fire-polishing, it was found necessary to use a vibratory polishing technique. Chrome oxide was used as the abrasive and either water or kerosene as the lubricant. Vibratory polishing is extremely slow, but surfaces so polished show no evidence of fire polishing, even when examined by electron microscopy. The most satisfactory etching process used for Pyroceram 9608 consisted of a primary etch of 5 milliliters of hydrochloric acid (concentrated), 5 milliliters of hydrogen fluoride (45 percent), and 45 milliliters of water, and a secondary etch with methyl alcohol replacing the water. Best results were obtained with total etching times from 25 to 30 seconds. Staining of the Pyroceram surface with a Sanford's marker was found to be an expedient way to reduce the glare of reflected light.

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

    Science.gov (United States)

    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.

  11. Imaging of Scleral Collagen Deformation Using Combined Confocal Raman Microspectroscopy and Polarized Light Microscopy Techniques.

    Science.gov (United States)

    Chakraborty, Nilay; Wang, Mian; Solocinski, Jason; Kim, Wonsuk; Argento, Alan

    2016-01-01

    This work presents an optospectroscopic characterization technique for soft tissue microstructure using site-matched confocal Raman microspectroscopy and polarized light microscopy. Using the technique, the microstructure of soft tissue samples is directly observed by polarized light microscopy during loading while spatially correlated spectroscopic information is extracted from the same plane, verifying the orientation and arrangement of the collagen fibers. Results show the response and orientation of the collagen fiber arrangement in its native state as well as during tensile and compressive loadings in a porcine sclera model. An example is also given showing how the data can be used with a finite element program to estimate the strain in individual collagen fibers. The measurements demonstrate features that indicate microstructural reorganization and damage of the sclera's collagen fiber arrangement under loading. The site-matched confocal Raman microspectroscopic characterization of the tissue provides a qualitative measure to relate the change in fibrillar arrangement with possible chemical damage to the collagen microstructure. Tests and analyses presented here can potentially be used to determine the stress-strain behavior, and fiber reorganization of the collagen microstructure in soft tissue during viscoelastic response.

  12. Virtual unfolding of light sheet fluorescence microscopy dataset for quantitative analysis of the mouse intestine

    Science.gov (United States)

    Candeo, Alessia; Sana, Ilenia; Ferrari, Eleonora; Maiuri, Luigi; D'Andrea, Cosimo; Valentini, Gianluca; Bassi, Andrea

    2016-05-01

    Light sheet fluorescence microscopy has proven to be a powerful tool to image fixed and chemically cleared samples, providing in depth and high resolution reconstructions of intact mouse organs. We applied light sheet microscopy to image the mouse intestine. We found that large portions of the sample can be readily visualized, assessing the organ status and highlighting the presence of regions with impaired morphology. Yet, three-dimensional (3-D) sectioning of the intestine leads to a large dataset that produces unnecessary storage and processing overload. We developed a routine that extracts the relevant information from a large image stack and provides quantitative analysis of the intestine morphology. This result was achieved by a three step procedure consisting of: (1) virtually unfold the 3-D reconstruction of the intestine; (2) observe it layer-by-layer; and (3) identify distinct villi and statistically analyze multiple samples belonging to different intestinal regions. Even if the procedure has been developed for the murine intestine, most of the underlying concepts have a general applicability.

  13. Cholesterol crystal binding of biliary immuno globulin A: visualization by fluorescence light microscopy

    Institute of Scientific and Technical Information of China (English)

    Frank Lammert; Stefan Sudfetd; Norbert Busch; Siegfried Matern

    2001-01-01

    AIM To assess potential contributions of biliary IgA for crystal agglomeration into gallstones, we visualized cholesterol crystal binding of biliary IgA.METHODS Crystal-binding biliary proteins were extracted from human gallbladder bile using lectin affinity chromatography. Biliary IgA was isolated from the bound protein fraction by immunoaffinity chromatography. Pure cholesterol monohydrate crystals were incubated with biliary IgA and fluoresceine isothiocyanate (FITC)-conjugated anti-lgA at 37C. Samples were examined under polarizing and fluorescence light microscopy with digital image processing.RESULTS Binding of biliary IgA to cholesterol monohydrate crystals could be visualized with FITC-conjugated anti-lgA antibodies. Peak fluorescence occurred at crystal edges and dislocations. Controls without biliary IgA or with biliary IgG showed no significant fluorescence.CONCLUSION Fluorescence light microscopy provided evidence for cholesterol crystal binding of biliary IgA. Cholesterol crystalbinding proteins like IgA might be important mediators of crystal agglomeration and growth of cholesterol gallstones by modifying the evolving crystal structures in vivo.

  14. Dynamic light scattering and atomic force microscopy techniques for size determination of polyurethane nanoparticles

    Energy Technology Data Exchange (ETDEWEB)

    Giehl Zanetti-Ramos, Betina [Laboratorio de Bioenergetica e Bioquimica de Macromoleculas, Departamento de Ciencias Farmaceuticas (Brazil)], E-mail: betinagzramos@pq.cnpq.br; Beddin Fritzen-Garcia, Mauricia [Laboratorio de Bioenergetica e Bioquimica de Macromoleculas, Departamento de Ciencias Farmaceuticas (Brazil); Schweitzer de Oliveira, Cristian; Avelino Pasa, Andre [Laboratorio de Filmes Finos e Superficie, Departamento de Fisica (Brazil); Soldi, Valdir [Grupo de Estudos em Materiais Polimericos, Departamento de Quimica, Universidade Federal de Santa Catarina, 88040-900, Florianopolis, SC (Brazil); Borsali, Redouane [Centre de Recherche sur les Macromolecules Vegetales CERMAV/CNRS, 38041 - Grenoble (France); Creczynski-Pasa, Tania Beatriz [Laboratorio de Bioenergetica e Bioquimica de Macromoleculas, Departamento de Ciencias Farmaceuticas (Brazil)

    2009-03-01

    Nanoparticles have applications in various industrial fields principally in drug delivery. Nowadays, there are several processes for manufacturing colloidal polymeric systems and methods of preparation as well as of characterization. In this work, Dynamic Light Scattering and Atomic Force Microscopy techniques were used to characterize polyurethane nanoparticles. The nanoparticles were prepared by miniemulsion technique. The lipophilic monomers, isophorone diisocyanate (IPDI) and natural triol, were emulsified in water containing surfactant. In some formulations the poly(ethylene glycol) was used as co-monomer to obtain the hydrophilic and pegylated nanoparticles. Polyurethane nanoparticles observed by atomic force microscopy (AFM) were spherical with diameter around 209 nm for nanoparticles prepared without PEG. From AFM imaging two populations of nanoparticles were observed in the formulation prepared with PEG (218 and 127 nm) while dynamic light scattering (DLS) measurements showed a monodisperse size distribution around 250 nm of diameters for both formulations. The polydispersity index of the formulations and the experimental procedures could influence the particle size determination with these techniques.

  15. [Ischemic-reperfusion paraplegia in the dog and its light microscopy imaging].

    Science.gov (United States)

    Sulla, I; Marsala, J; Radonák, J

    2004-02-01

    Paraplegia, which develops after operation on aorta, represents a real catastrophe for the patient and for the surgeon. The aim of the present work was to investigate the light microscopy picture of this complication and consequently better understand related processes. Twenty one adult dogs, cross breeds of both sexes, weight 18-25 kg, were divided into four groups: 1. Controls (n = 3); 2.30-min ischemia induced by occlusion of thoracic aorta by a tourniquet, followed for 30 min survival (n = 6); 3.30-min ischemia and 72 h of survival (n = 6); 4) 30-min ischemia and 6 days of survival (n = 6). All these manipulations were made in sterile conditions under general anesthesia. As soon as the planned time of survival passed, the animals were flushed out, in deep pentobarbital anesthesia, with 3,000 ml of sodium chloride and fixed with 3,000 ml of 10% neutral formaldehyde. Sections, 30 microns thick, from L3-S1 medulla segments were processed in the laboratory of Neurobiological Institute by the method of Nauta for light microscopy examination. Neurohistological picture was characterized by a marked damage of the medulla neurons. The changes proved to be irreversible and resulted, in the course of six days of survival, to death of the cells, characterized by their disintegration. The results indicate that the only rational procedure in conditions of threatening ischemic-reperfusion injury of medulla is to prevent it.

  16. Application of micro-PIXE, MRI and light microscopy for research in wood science and dendroecology

    Science.gov (United States)

    Merela, M.; Pelicon, P.; Vavpetič, P.; Regvar, M.; Vogel-Mikuš, K.; Serša, I.; Poličnik, H.; Pokorny, B.; Levanič, T.; Oven, P.

    2009-06-01

    Beech ( Fagus sylvatica L.) branches were topped and after five months the wound response was analyzed by PIXE, 3D-MRI and light microscopy. From freshly cut and deeply frozen sample 30 μm thick longitudinal-radial tissue sections were prepared for anatomical investigations and micro-PIXE analysis. Light microscopy revealed the structural response to wounding, i.e. occurrence of the reaction zone between the exposed and dehydrated dead tissue and healthy sound wood. The reaction zone was characterized by tylosis in vessels and accumulation of colored deposits in parenchyma cells, fibres and vessels. 3D MRI of a parallel sample showed that the moisture content in the reaction zone was three times higher than in normal healthy wood. Micro-PIXE mapping at margins of compromised wood in beech revealed an increased concentration of potassium in the reaction zone. The increase in the calcium concentration was associated with the dehydrated tissue adjacent to reaction zones. In addition, micro-PIXE was used to determine the elemental distribution in annual tree rings. This may be relevant for retrospective assessment of environmental pollution in wood by measuring yearly increments as a biomonitoring tool. The analysis of European larch ( Larix decidua Mill.) wood revealed a high similarity between optical characteristics (i.e. late versus earlywood) and elemental (e.g. Cl, K, Ca, Mn, Zn) distribution.

  17. Application of micro-PIXE, MRI and light microscopy for research in wood science and dendroecology

    Energy Technology Data Exchange (ETDEWEB)

    Merela, M. [University of Ljubljana, BF, Dep. of Wood Science and Technology, Rozna dolina VIII/34, SI-1000 Ljubljana (Slovenia); Pelicon, P. [Jozef Stefan Institute, Jamova 39, SI-1000 Ljubljana (Slovenia)], E-mail: primoz.pelicon@ijs.si; Vavpetic, P. [Jozef Stefan Institute, Jamova 39, SI-1000 Ljubljana (Slovenia); Regvar, M.; Vogel-Mikus, K. [University of Ljubljana, BF, Dep. of Biology, Vecna pot 111, SI-1000 Ljubljana (Slovenia); Sersa, I. [Jozef Stefan Institute, Jamova 39, SI-1000 Ljubljana (Slovenia); Policnik, H. [ERICo Velenje, Ecological Research and Industrial Co-operation, Koroska 58, SI-3320 Velenje (Slovenia); Pokorny, B. [ERICo Velenje, Ecological Research and Industrial Co-operation, Koroska 58, SI-3320 Velenje (Slovenia); Slovenian Forestry Institute, Vecna pot 2, SI-1000 Ljubljana (Slovenia); Levanic, T. [Slovenian Forestry Institute, Vecna pot 2, SI-1000 Ljubljana (Slovenia); Oven, P. [University of Ljubljana, BF, Dep. of Wood Science and Technology, Rozna dolina VIII/34, SI-1000 Ljubljana (Slovenia)

    2009-06-15

    Beech (Fagus sylvatica L.) branches were topped and after five months the wound response was analyzed by PIXE, 3D-MRI and light microscopy. From freshly cut and deeply frozen sample 30 {mu}m thick longitudinal-radial tissue sections were prepared for anatomical investigations and micro-PIXE analysis. Light microscopy revealed the structural response to wounding, i.e. occurrence of the reaction zone between the exposed and dehydrated dead tissue and healthy sound wood. The reaction zone was characterized by tylosis in vessels and accumulation of colored deposits in parenchyma cells, fibres and vessels. 3D MRI of a parallel sample showed that the moisture content in the reaction zone was three times higher than in normal healthy wood. Micro-PIXE mapping at margins of compromised wood in beech revealed an increased concentration of potassium in the reaction zone. The increase in the calcium concentration was associated with the dehydrated tissue adjacent to reaction zones. In addition, micro-PIXE was used to determine the elemental distribution in annual tree rings. This may be relevant for retrospective assessment of environmental pollution in wood by measuring yearly increments as a biomonitoring tool. The analysis of European larch (Larix decidua Mill.) wood revealed a high similarity between optical characteristics (i.e. late versus earlywood) and elemental (e.g. Cl, K, Ca, Mn, Zn) distribution.

  18. Wavefront coding for fast, high-resolution light-sheet microscopy (Conference Presentation)

    Science.gov (United States)

    Olarte, Omar E.; Licea-Rodriguez, Jacob; Loza-Alvarez, Pablo

    2017-02-01

    Some biological experiments demand the observation of dynamics processes in 3D with high spatiotemporal resolution. The use of wavefront coding to extend the depth-of-field (DOF) of the collection arm of a light-sheet microscope is an interesting alternative for fast 3D imaging. Under this scheme, the 3D features of the sample are captured at high volumetric rates while the light sheet is swept rapidly within the extended DOF. The DOF is extended by coding the pupil function of the imaging lens by using a custom-designed phase mask. A posterior restoration step is required to decode the information of the captured images based on the applied phase mask [1]. This hybrid optical-digital approach is known as wavefront coding (WFC). Previously, we have demonstrated this method for performing fast 3D imaging of biological samples at medium resolution [2]. In this work, we present the extension of this approach for high-resolution microscopes. Under these conditions, the effective DOF of a standard high NA objective is of a few micrometers. Here we demonstrate that by the use of WFC, we can extend the DOF more than one order of magnitude keeping the high-resolution imaging. This is demonstrated for two designed phase masks using Zebrafish and C. elegans samples. [1] Olarte, O.E., Andilla, J., Artigas, D., and Loza-Alvarez, P., "Decoupled Illumination-Detection Microscopy. Selected Optics in Year 2105," in Optics and Photonics news 26, p. 41 (2015). [2] Olarte, O.E., Andilla, J., Artigas, D., and Loza-Alvarez, P., "Decoupled illumination detection in light sheet microscopy for fast volumetric imaging," Optica 2(8), 702 (2015).

  19. Optimal experimental design for the detection of light atoms from high-resolution scanning transmission electron microscopy images

    NARCIS (Netherlands)

    Gonnissen, J.; De Backer, A.; Den Dekker, A.J.; Martinez, G.T.; Rosenauer, A.; Sijbers, J.; Van Aert, S.

    2014-01-01

    We report an innovative method to explore the optimal experimental settings to detect light atoms from scanning transmission electron microscopy (STEM) images. Since light elements play a key role in many technologically important materials, such as lithium-battery devices or hydrogen storage

  20. X-ray microscopy as an approach to increasing accuracy and efficiency of serial block-face imaging for correlated light and electron microscopy of biological specimens.

    Science.gov (United States)

    Bushong, Eric A; Johnson, Donald D; Kim, Keun-Young; Terada, Masako; Hatori, Megumi; Peltier, Steven T; Panda, Satchidananda; Merkle, Arno; Ellisman, Mark H

    2015-02-01

    The recently developed three-dimensional electron microscopic (EM) method of serial block-face scanning electron microscopy (SBEM) has rapidly established itself as a powerful imaging approach. Volume EM imaging with this scanning electron microscopy (SEM) method requires intense staining of biological specimens with heavy metals to allow sufficient back-scatter electron signal and also to render specimens sufficiently conductive to control charging artifacts. These more extreme heavy metal staining protocols render specimens light opaque and make it much more difficult to track and identify regions of interest (ROIs) for the SBEM imaging process than for a typical thin section transmission electron microscopy correlative light and electron microscopy study. We present a strategy employing X-ray microscopy (XRM) both for tracking ROIs and for increasing the efficiency of the workflow used for typical projects undertaken with SBEM. XRM was found to reveal an impressive level of detail in tissue heavily stained for SBEM imaging, allowing for the identification of tissue landmarks that can be subsequently used to guide data collection in the SEM. Furthermore, specific labeling of individual cells using diaminobenzidine is detectable in XRM volumes. We demonstrate that tungsten carbide particles or upconverting nanophosphor particles can be used as fiducial markers to further increase the precision and efficiency of SBEM imaging.

  1. Quantifying light scattering with single-mode fiber -optic confocal microscopy

    Directory of Open Access Journals (Sweden)

    Haidekker Mark A

    2009-11-01

    Full Text Available Abstract Background Confocal microscopy has become an important option for examining tissues in vivo as a diagnostic tool and a quality control tool for tissue-engineered constructs. Collagen is one of the primary determinants of biomechanical stability. Since collagen is also the primary scattering element in skin and other soft tissues, we hypothesized that laser-optical imaging methods, particularly confocal scattered-light scanning, would allow us to quantify scattering intensity and determine collagen content in biological layers. Methods We built a fully automated confocal scattered-light scanner to examine how light scatters in Intralipid, a common tissue phantom, and three-dimensional collagen gels. Intralipid with 0.5%, 1.0%, 1.5%, and 2.0% concentration was filled between precisely spaced glass coverslips. Collagen gels at collagen concentrations from 0.30 mg/mL to 3.30 mg/mL were prepared, and all samples underwent A-mode scanning with multiple averaged scans. In Intralipid samples, light reflected from the upper fluid-glass interface was measured. In collagen gels, average scattering intensity inside the actual gel was measured. In both cases, intensity was correlated with concentration. Results By measuring light attenuation at interface reflections of various thicknesses using our device, we were able to determine that the scattering coefficient at 660 nm of Intralipid at increasing concentrations in water to be 39 cm-1 for each percent increase of Intralipid. We were also able to measure the amount of scattering of various concentrations of collagen in gels directly using backscattered light. The results show a highly linear relationship with an increase of 8.2 arbitrary units in backscattering intensity for every 1 mg increase of collagen within a 1 mL gel volume. Conclusion The confocal scattered-light scanner allows to accurately quantify scattering in Intralipid and collagen gels. Furthermore, a linear relationship between

  2. Cellular imaging of deep organ using two-photon Bessel light-sheet nonlinear structured illumination microscopy

    Science.gov (United States)

    Zhao, Ming; Zhang, Han; Li, Yu; Ashok, Amit; Liang, Rongguang; Zhou, Weibin; Peng, Leilei

    2014-01-01

    In vivo fluorescent cellular imaging of deep internal organs is highly challenging, because the excitation needs to penetrate through strong scattering tissue and the emission signal is degraded significantly by photon diffusion induced by tissue-scattering. We report that by combining two-photon Bessel light-sheet microscopy with nonlinear structured illumination microscopy (SIM), live samples up to 600 microns wide can be imaged by light-sheet microscopy with 500 microns penetration depth, and diffused background in deep tissue light-sheet imaging can be reduced to obtain clear images at cellular resolution in depth beyond 200 microns. We demonstrate in vivo two-color imaging of pronephric glomeruli and vasculature of zebrafish kidney, whose cellular structures located at the center of the fish body are revealed in high clarity by two-color two-photon Bessel light-sheet SIM. PMID:24876996

  3. Cellular imaging of deep organ using two-photon Bessel light-sheet nonlinear structured illumination microscopy.

    Science.gov (United States)

    Zhao, Ming; Zhang, Han; Li, Yu; Ashok, Amit; Liang, Rongguang; Zhou, Weibin; Peng, Leilei

    2014-05-01

    In vivo fluorescent cellular imaging of deep internal organs is highly challenging, because the excitation needs to penetrate through strong scattering tissue and the emission signal is degraded significantly by photon diffusion induced by tissue-scattering. We report that by combining two-photon Bessel light-sheet microscopy with nonlinear structured illumination microscopy (SIM), live samples up to 600 microns wide can be imaged by light-sheet microscopy with 500 microns penetration depth, and diffused background in deep tissue light-sheet imaging can be reduced to obtain clear images at cellular resolution in depth beyond 200 microns. We demonstrate in vivo two-color imaging of pronephric glomeruli and vasculature of zebrafish kidney, whose cellular structures located at the center of the fish body are revealed in high clarity by two-color two-photon Bessel light-sheet SIM.

  4. Optimal experimental design for the detection of light atoms from high-resolution scanning transmission electron microscopy images

    OpenAIRE

    Gonnissen, J.; De Backer, A.; Den Dekker, A.J.; Martinez, G. T.; Rosenauer, A.; Sijbers, J.; Van Aert, S.

    2014-01-01

    Abstract: We report an innovative method to explore the optimal experimental settings to detect light atoms from scanning transmission electron microscopy (STEM) images. Since light elements play a key role in many technologically important materials, such as lithium-battery devices or hydrogen storage applications, much effort has been made to optimize the STEM technique in order to detect light elements. Therefore, classical performance criteria, such as contrast or signal-to-noise ratio, a...

  5. Visualizing Cell Architecture and Molecular Location Using Soft X-Ray Tomography and Correlated Cryo-Light Microscopy

    Science.gov (United States)

    McDermott, Gerry; Le Gros, Mark A.; Larabell, Carolyn A.

    2012-05-01

    Living cells are structured to create a range of microenvironments that support specific chemical reactions and processes. Understanding how cells function therefore requires detailed knowledge of both the subcellular architecture and the location of specific molecules within this framework. Here we review the development of two correlated cellular imaging techniques that fulfill this need. Cells are first imaged using cryogenic fluorescence microscopy to determine the location of molecules of interest that have been labeled with fluorescent tags. The same specimen is then imaged using soft X-ray tomography to generate a high-contrast, 3D reconstruction of the cells. Data from the two modalities are then combined to produce a composite, information-rich view of the cell. This correlated imaging approach can be applied across the spectrum of problems encountered in cell biology, from basic research to biotechnological and biomedical applications such as the optimization of biofuels and the development of new pharmaceuticals.

  6. Quantitative mapping of collagen fiber alignment in thick tissue samples using transmission polarized-light microscopy.

    Science.gov (United States)

    Yakovlev, Dmitry D; Shvachkina, Marina E; Sherman, Maria M; Spivak, Andrey V; Pravdin, Alexander B; Yakovlev, Dmitry A

    2016-07-01

    Immersion optical clearing makes it possible to use transmission polarized-light microscopy for characterization of thick (200 to 2000  μm) layers of biological tissues. We discuss polarization properties of thick samples in the context of the problem of characterization of collagen fiber alignment in connective tissues such as sclera and dermis. Optical chirality caused by azimuthal variations of the macroscopic (effective) optic axis of the medium across the sample thickness should be considered in polarization mapping of thick samples of these tissues. We experimentally evaluate to what extent the optical chirality affects the measurement results in typical situations and show under what conditions it can be easily taken into account and does not hinder, but rather helps, in characterization of collagen fiber alignment.

  7. White light scattering spectroscopy and electron microscopy of laser induced melting in single gold nanorods.

    Science.gov (United States)

    Zijlstra, Peter; Chon, James W M; Gu, Min

    2009-07-28

    We present the first measurements of laser induced melting and reshaping of single gold nanorods. Using a combination of white light scattering spectroscopy and electron microscopy we find a melting energy of 260 fJ for nanorods with an average size of 92 x 30 nm. Contrary to previous reports on ensembles of nanorods, this melting energy corresponds well to the theoretical prediction of 225 fJ. We observe a gradual shape change from a long and thin rod to a shorter and wider rod, which eventually collapses into a sphere when enough laser energy is deposited. We also observe that higher aspect ratio particles are thermodynamically less stable, leading to a greater reduction of the aspect ratio at lower laser pulse energy densities.

  8. Light and scanning electron microscopy of the tongue of a degu (Octodon degus).

    Science.gov (United States)

    Cizek, Petr; Hamouzova, Pavla; Jekl, Vladimir; Kvapil, Pavel; Tichy, Frantisek

    2017-09-01

    The tongue of an adult degu was examined by light and scanning electron microscopy. It consists of an apex, corpus, and radix and contains a lingual prominence. The aim of this study was to describe the course of muscle fascicles of the proper lingual muscle, the presence and nature of the lingual salivary glands, and particularly the appearance and distribution of the lingual papillae. Three major types of papillae have been observed: filiform, conical, and vallate. The dorsal surface of the lingual apex extends in caudally bent filiform papillae with two spines. The lingual corpus bears long filiform papillae with a single tip. The lingual radix contains crown-like papillae in the region of the prominence and conical papillae in the remaining areas. Two oval vallate papillae were discovered caudally on the lingual radix. This first description of the lingual structures in a degu could be used for comparative studies or as basic data for differentiation of lingual morphology in this species.

  9. Automatic segmentation and classification of mycobacterium tuberculosis with conventional light microscopy

    Science.gov (United States)

    Xu, Chao; Zhou, Dongxiang; Zhai, Yongping; Liu, Yunhui

    2015-12-01

    This paper realizes the automatic segmentation and classification of Mycobacterium tuberculosis with conventional light microscopy. First, the candidate bacillus objects are segmented by the marker-based watershed transform. The markers are obtained by an adaptive threshold segmentation based on the adaptive scale Gaussian filter. The scale of the Gaussian filter is determined according to the color model of the bacillus objects. Then the candidate objects are extracted integrally after region merging and contaminations elimination. Second, the shape features of the bacillus objects are characterized by the Hu moments, compactness, eccentricity, and roughness, which are used to classify the single, touching and non-bacillus objects. We evaluated the logistic regression, random forest, and intersection kernel support vector machines classifiers in classifying the bacillus objects respectively. Experimental results demonstrate that the proposed method yields to high robustness and accuracy. The logistic regression classifier performs best with an accuracy of 91.68%.

  10. New Aspidoderidae species parasite of Didelphis aurita (Mammalia: Didelphidae): a light and scanning electron microscopy approach.

    Science.gov (United States)

    Chagas-Moutinho, V A; Sant'anna, V; Oliveira-Menezes, A; De Souza, W

    2014-02-01

    Nematodes of the family Aspidoderidae (Nematoda: Heterakoidea) Skrjabin and Schikobalova, 1947, are widely distributed in the Americas. The family Aspidoderidae includes the subfamilies Aspidoderinae Skrjabin and Schikobalova, 1947, and Lauroiinae Skrjabin and Schikobalova, 1951. These two subfamilies are delineated by the presence or absence of cephalic cordons at the anterior region. The nematodes in the subfamily Aspidoderinae, which includes the genus AspidoderaRailliet and Henry, 1912, are represented by nematodes with anterior cephalic cordons at the anterior end. The nematodes of the genus AspidoderaRailliet and Henry, 1912, are found in the cecum and large intestine of mammals of the orders Edentata, Marsupialia and Rodentia. Species within this genus have many morphological similarities. The use of scanning electron microscopy allows the specific characterization of the species within this genus. In the present work, we describe a new species of Aspidodera parasite of the large intestine of Didelphis aurita (Mammalia: Didelphidae) Wied-Neuwied, 1826, collected from Cachoeiras de Macacu, Rio de Janeiro. The combination of light and scanning electron microscopy allowed us a detailed analysis of this nematode.

  11. High-resolution x-ray photoemission electron microscopy at the Advanced Light Source

    Energy Technology Data Exchange (ETDEWEB)

    Stammler, T.; Anders, S.; Padmore, H.A. [Lawrence Berkeley National Lab., CA (United States); Stoehr, J. [IBM Almaden Research Center, San Jose, CA (United States); Scheinfein, M. [Arizona State Univ., Tempe, AZ (United States). Dept. of Physics and Astronomy; Ade, H. [North Carolina State Univ., Raleigh, NC (United States). Dept. of Physics

    1998-12-31

    X-ray Photoemission Electron Microscopy (X-PEEM) is a full-field imaging technique where the sample is illuminated by an x-ray beam and the photoemitted electrons are imaged on a screen by means of an electron optics. It therefore combines two well-established materials analysis techniques--photoemission electron microscopy (PEEM) and x-ray spectroscopy such as near edge x-ray absorption fine structure (NEXAFS) spectroscopy. This combination opens a wide field of new applications in materials research and has proven to be a powerful tool to investigate simultaneously topological, elemental, chemical state, and magnetic properties of surfaces, thin films, and multilayers at high spatial resolution. A new X-PEEM installed at the bend magnet beamline 7.3.1.1 at the Advanced Light Source (ALS) is designed for a spatial resolution of 20 nm and is currently under commissioning. An overview of the ongoing experimental program using X-PEEM in the field of materials research at the ALS is given by elemental and chemical bonding contrast imaging of hard disk coatings and sliders, field emission studies on diamond films as possible candidates for field-emission flat-panel displays, and the study of dewetting and decomposition phenomena of thin polymer blends and bilayers.

  12. Chiral Nematic Structure of Cellulose Nanocrystal Suspensions and Films; Polarized Light and Atomic Force Microscopy

    Directory of Open Access Journals (Sweden)

    Derek G. Gray

    2015-11-01

    Full Text Available Cellulosic liquid crystalline solutions and suspensions form chiral nematic phases that show a rich variety of optical textures in the liquid crystalline state. These ordered structures may be preserved in solid films prepared by evaporation of solvent or suspending medium. Film formation from aqueous suspensions of cellulose nanocrystals (CNC was investigated by polarized light microscopy, optical profilometry and atomic force microscopy (AFM. An attempt is made to interpret qualitatively the observed textures in terms of the orientation of the cellulose nanocrystals in the suspensions and films, and the changes in orientation caused by the evaporative process. Mass transfer within the evaporating droplet resulted in the formation of raised rings whose magnitude depended on the degree of pinning of the receding contact line. AFM of dry films at short length scales showed a radial orientation of the CNC at the free surface of the film, along with a radial height variation with a period of approximately P/2, ascribed to the anisotropic shrinkage of the chiral nematic structure.

  13. Rapid reconstruction of 3D neuronal morphology from light microscopy images with augmented rayburst sampling.

    Science.gov (United States)

    Ming, Xing; Li, Anan; Wu, Jingpeng; Yan, Cheng; Ding, Wenxiang; Gong, Hui; Zeng, Shaoqun; Liu, Qian

    2013-01-01

    Digital reconstruction of three-dimensional (3D) neuronal morphology from light microscopy images provides a powerful technique for analysis of neural circuits. It is time-consuming to manually perform this process. Thus, efficient computer-assisted approaches are preferable. In this paper, we present an innovative method for the tracing and reconstruction of 3D neuronal morphology from light microscopy images. The method uses a prediction and refinement strategy that is based on exploration of local neuron structural features. We extended the rayburst sampling algorithm to a marching fashion, which starts from a single or a few seed points and marches recursively forward along neurite branches to trace and reconstruct the whole tree-like structure. A local radius-related but size-independent hemispherical sampling was used to predict the neurite centerline and detect branches. Iterative rayburst sampling was performed in the orthogonal plane, to refine the centerline location and to estimate the local radius. We implemented the method in a cooperative 3D interactive visualization-assisted system named flNeuronTool. The source code in C++ and the binaries are freely available at http://sourceforge.net/projects/flneurontool/. We validated and evaluated the proposed method using synthetic data and real datasets from the Digital Reconstruction of Axonal and Dendritic Morphology (DIADEM) challenge. Then, flNeuronTool was applied to mouse brain images acquired with the Micro-Optical Sectioning Tomography (MOST) system, to reconstruct single neurons and local neural circuits. The results showed that the system achieves a reasonable balance between fast speed and acceptable accuracy, which is promising for interactive applications in neuronal image analysis.

  14. Rapid reconstruction of 3D neuronal morphology from light microscopy images with augmented rayburst sampling.

    Directory of Open Access Journals (Sweden)

    Xing Ming

    Full Text Available Digital reconstruction of three-dimensional (3D neuronal morphology from light microscopy images provides a powerful technique for analysis of neural circuits. It is time-consuming to manually perform this process. Thus, efficient computer-assisted approaches are preferable. In this paper, we present an innovative method for the tracing and reconstruction of 3D neuronal morphology from light microscopy images. The method uses a prediction and refinement strategy that is based on exploration of local neuron structural features. We extended the rayburst sampling algorithm to a marching fashion, which starts from a single or a few seed points and marches recursively forward along neurite branches to trace and reconstruct the whole tree-like structure. A local radius-related but size-independent hemispherical sampling was used to predict the neurite centerline and detect branches. Iterative rayburst sampling was performed in the orthogonal plane, to refine the centerline location and to estimate the local radius. We implemented the method in a cooperative 3D interactive visualization-assisted system named flNeuronTool. The source code in C++ and the binaries are freely available at http://sourceforge.net/projects/flneurontool/. We validated and evaluated the proposed method using synthetic data and real datasets from the Digital Reconstruction of Axonal and Dendritic Morphology (DIADEM challenge. Then, flNeuronTool was applied to mouse brain images acquired with the Micro-Optical Sectioning Tomography (MOST system, to reconstruct single neurons and local neural circuits. The results showed that the system achieves a reasonable balance between fast speed and acceptable accuracy, which is promising for interactive applications in neuronal image analysis.

  15. Quantitative Connection between Ensemble Thermodynamics and Single-Molecule Kinetics: A Case Study Using Cryogenic Electron Microscopy and Single-Molecule Fluorescence Resonance Energy Transfer Investigations of the Ribosome.

    Science.gov (United States)

    Thompson, Colin D Kinz; Sharma, Ajeet K; Frank, Joachim; Gonzalez, Ruben L; Chowdhury, Debashish

    2015-08-27

    At equilibrium, thermodynamic and kinetic information can be extracted from biomolecular energy landscapes by many techniques. However, while static, ensemble techniques yield thermodynamic data, often only dynamic, single-molecule techniques can yield the kinetic data that describe transition-state energy barriers. Here we present a generalized framework based upon dwell-time distributions that can be used to connect such static, ensemble techniques with dynamic, single-molecule techniques, and thus characterize energy landscapes to greater resolutions. We demonstrate the utility of this framework by applying it to cryogenic electron microscopy (cryo-EM) and single-molecule fluorescence resonance energy transfer (smFRET) studies of the bacterial ribosomal pre-translocation complex. Among other benefits, application of this framework to these data explains why two transient, intermediate conformations of the pre-translocation complex, which are observed in a cryo-EM study, may not be observed in several smFRET studies.

  16. Use of polarized light microscopy is essential in the efficient diagnosis of respiratory amyloidosis and could decrease disease prevalence.

    Science.gov (United States)

    Ma, Dedong; Lu, Hongxiu; Zhang, Cuijuan; Ying, Yangyang; Xiao, Wei

    2015-10-16

    Primary tracheobronchial amyloidosis (TBA) is a rare disease of unknown etiology, with a high misdiagnosis rate. The current gold standard diagnostic criteria require double-positive results for Congo red staining and polarized light microscopy examination. The aim of the present report was to examine the diagnostic value of polarized light microscopy in TBA diagnosis in China. Thirteen cases from the Shandong University Qilu Hospital were reviewed. Polarized light microscopic examination was conducted after searching for cases with positive Congo red staining. Among the 13 patients, eight displayed yellow-green birefringence body with polarized light microscopic examination. This result indicated a false-positive rate of 38.5% with Congo red staining used as the single criteria for TBA diagnosis. After reviewing the Chinese literature and selecting 104 reported cases of TBA in China, we found that diagnosis with the gold standard was confirmed in polarized light microscopy in China is currently limited, and there is a big gap from the international diagnosis standards. There is a need to include polarized light microscopy in routine TBA diagnosis to reduce the misdiagnosis rate, and achieve optimal treatment. © 2015 John Wiley & Sons Ltd.

  17. Stripe artifact elimination based on nonsubsampled contourlet transform for light sheet fluorescence microscopy

    Science.gov (United States)

    Liang, Xiao; Zang, Yali; Dong, Di; Zhang, Liwen; Fang, Mengjie; Yang, Xin; Arranz, Alicia; Ripoll, Jorge; Hui, Hui; Tian, Jie

    2016-10-01

    Stripe artifacts, caused by high-absorption or high-scattering structures in the illumination light path, are a common drawback in both unidirectional and multidirectional light sheet fluorescence microscopy (LSFM), significantly deteriorating image quality. To circumvent this problem, we present an effective multidirectional stripe remover (MDSR) method based on nonsubsampled contourlet transform (NSCT), which can be used for both unidirectional and multidirectional LSFM. In MDSR, a fast Fourier transform (FFT) filter is designed in the NSCT domain to shrink the stripe components and eliminate the noise. Benefiting from the properties of being multiscale and multidirectional, MDSR succeeds in eliminating stripe artifacts in both unidirectional and multidirectional LSFM. To validate the method, MDSR has been tested on images from a custom-made unidirectional LSFM system and a commercial multidirectional LSFM system, clearly demonstrating that MDSR effectively removes most of the stripe artifacts. Moreover, we performed a comparative experiment with the variational stationary noise remover and the wavelet-FFT methods and quantitatively analyzed the results with a peak signal-to-noise ratio, showing an improved noise removal when using the MDSR method.

  18. Molecular orientation sensitive second harmonic microscopy by radially and azimuthally polarized light

    Science.gov (United States)

    Ehmke, Tobias; Nitzsche, Tim Heiko; Knebl, Andreas; Heisterkamp, Alexander

    2014-01-01

    We demonstrate the possibility to switch the z-polarization component of the illumination in the vicinity of the focus of high-NA objective lenses by applying radially and azimuthally polarized incident light. The influence of the field distribution on nonlinear effects was first investigated by the means of simulations. These were performed for high-NA objective lenses commonly used in nonlinear microscopy. Special attention is paid to the influence of the polarization of the incoming field. For linearly, circularly and radially polarized light a considerable polarization component in z-direction is generated by high NA focusing. Azimuthal polarization is an exceptional case: even for strong focusing no z-component arises. Furthermore, the influence of the input polarization on the intensity contributing to the nonlinear signal generation was computed. No distinct difference between comparable input polarization states was found for chosen thresholds of nonlinear signal generation. Differences in signal generation for radially and azimuthally polarized vortex beams were experimentally evaluated in native collagen tissue (porcine cornea). The findings are in good agreement with the theoretical predictions and display the possibility to probe the molecular orientation along the optical axis of samples with known nonlinear properties. The combination of simulations regarding the nonlinear response of materials and experiments with different sample orientations and present or non present z-polarization could help to increase the understanding of nonlinear signal formation in yet unstudied materials. PMID:25071961

  19. The relationship between sperm head retardance using polarized light microscopy and clinical outcomes.

    Science.gov (United States)

    Vermey, Belinda G; Chapman, Michael G; Cooke, Simon; Kilani, Suha

    2015-01-01

    In human sperm head, birefringence can be seen under polarized light resulting from highly ordered structures within the acrosome and nucleus. Selecting sperm with partial head birefringence improves success of clinical pregnancies in patients with severe male factor infertility. The aim of this study was to establish a range of retardance in sperm heads using polarized light microscopy to select an optimum sperm for intracytoplasmic sperm injection (ICSI). Sperm heads of 63 couples undergoing ICSI in women aged 38 years or younger were imaged at the time of ICSI and later analysed for retardance blinded to embryo and cycle outcomes. Sperm head retardance was similar irrespective of whether fertilization occurred. Quality of embryos on day 3 and day 5 were higher when sperm were selected with head retardance ranging from 0.56 nm or greater to 0.91 nm or less. Selection of sperm with head retardance ranging from 0.56 nm or greater to 0.91 nm or less was associated with higher clinical pregnancy rates (OR 3.74 95% CI 1.43 to 9.77). Optimum sperm for selection at the time of ICSI was with head retardance within the range 0.56 nm or greater to 0.91 nm or less. Copyright © 2014 Reproductive Healthcare Ltd. Published by Elsevier Ltd. All rights reserved.

  20. Automatic Evaluation of Collagen Fiber Directions from Polarized Light Microscopy Images.

    Science.gov (United States)

    Novak, Kamil; Polzer, Stanislav; Tichy, Michal; Bursa, Jiri

    2015-08-01

    Mechanical properties of the arterial wall depend largely on orientation and density of collagen fiber bundles. Several methods have been developed for observation of collagen orientation and density; the most frequently applied collagen-specific manual approach is based on polarized light (PL). However, it is very time consuming and the results are operator dependent. We have proposed a new automated method for evaluation of collagen fiber direction from two-dimensional polarized light microscopy images (2D PLM). The algorithm has been verified against artificial images and validated against manual measurements. Finally the collagen content has been estimated. The proposed algorithm was capable of estimating orientation of some 35 k points in 15 min when applied to aortic tissue and over 500 k points in 35 min for Achilles tendon. The average angular disagreement between each operator and the algorithm was -9.3±8.6° and -3.8±8.6° in the case of aortic tissue and -1.6±6.4° and 2.6±7.8° for Achilles tendon. Estimated mean collagen content was 30.3±5.8% and 94.3±2.7% for aortic media and Achilles tendon, respectively. The proposed automated approach is operator independent and several orders faster than manual measurements and therefore has the potential to replace manual measurements of collagen orientation via PLM.

  1. Micron-scale resolution optical tomography of entire mouse brains with confocal light sheet microscopy.

    Science.gov (United States)

    Silvestri, Ludovico; Bria, Alessandro; Costantini, Irene; Sacconi, Leonardo; Peng, Hanchuan; Iannello, Giulio; Pavone, Francesco Saverio

    2013-10-08

    Understanding the architecture of mammalian brain at single-cell resolution is one of the key issues of neuroscience. However, mapping neuronal soma and projections throughout the whole brain is still challenging for imaging and data management technologies. Indeed, macroscopic volumes need to be reconstructed with high resolution and contrast in a reasonable time, producing datasets in the TeraByte range. We recently demonstrated an optical method (confocal light sheet microscopy, CLSM) capable of obtaining micron-scale reconstruction of entire mouse brains labeled with enhanced green fluorescent protein (EGFP). Combining light sheet illumination and confocal detection, CLSM allows deep imaging inside macroscopic cleared specimens with high contrast and speed. Here we describe the complete experimental pipeline to obtain comprehensive and human-readable images of entire mouse brains labeled with fluorescent proteins. The clearing and the mounting procedures are described, together with the steps to perform an optical tomography on its whole volume by acquiring many parallel adjacent stacks. We showed the usage of open-source custom-made software tools enabling stitching of the multiple stacks and multi-resolution data navigation. Finally, we illustrated some example of brain maps: the cerebellum from an L7-GFP transgenic mouse, in which all Purkinje cells are selectively labeled, and the whole brain from a thy1-GFP-M mouse, characterized by a random sparse neuronal labeling.

  2. Dual beam light profile microscopy: a new technique for optical absorption depth profilometry.

    Science.gov (United States)

    Power, J F; Fu, S W

    2004-02-01

    Light profile microscopy (LPM) is a recently developed technique of optical inspection that is used to record micrometer-scale images of thin-film cross-sections on a direct basis. In single beam mode, LPM provides image contrast based on luminescence, elastic, and/or inelastic scatter. However, LPM may also be used to depth profile the optical absorption coefficient of a thin film based on a method of dual beam irradiation presented in this work. The method uses a pair of collimated laser beams to consecutively irradiate a film from two opposing directions along the depth axis. An average profile of the beam's light intensity variation through the material is recovered for each direction and used to compute a depth-dependent differential absorbance profile. This latter quantity is shown from theory to be related to the film's depth-dependent optical absorption coefficient through a simple linear model that may be inverted by standard methods of numerical linear algebra. The inverse problem is relatively well posed, showing good immunity to data errors. This profilometry method is experimentally applied to a set of well-characterized materials with known absorption properties over a scale of tens of micrometers, and the reconstructed absorption profiles were found to be highly consistent with the reference data.

  3. Cerebral vessels segmentation for light-sheet microscopy image using convolutional neural networks

    Science.gov (United States)

    Hu, Chaoen; Hui, Hui; Wang, Shuo; Dong, Di; Liu, Xia; Yang, Xin; Tian, Jie

    2017-03-01

    Cerebral vessel segmentation is an important step in image analysis for brain function and brain disease studies. To extract all the cerebrovascular patterns, including arteries and capillaries, some filter-based methods are used to segment vessels. However, the design of accurate and robust vessel segmentation algorithms is still challenging, due to the variety and complexity of images, especially in cerebral blood vessel segmentation. In this work, we addressed a problem of automatic and robust segmentation of cerebral micro-vessels structures in cerebrovascular images acquired by light-sheet microscope for mouse. To segment micro-vessels in large-scale image data, we proposed a convolutional neural networks (CNNs) architecture trained by 1.58 million pixels with manual label. Three convolutional layers and one fully connected layer were used in the CNNs model. We extracted a patch of size 32x32 pixels in each acquired brain vessel image as training data set to feed into CNNs for classification. This network was trained to output the probability that the center pixel of input patch belongs to vessel structures. To build the CNNs architecture, a series of mouse brain vascular images acquired from a commercial light sheet fluorescence microscopy (LSFM) system were used for training the model. The experimental results demonstrated that our approach is a promising method for effectively segmenting micro-vessels structures in cerebrovascular images with vessel-dense, nonuniform gray-level and long-scale contrast regions.

  4. Lensfree Spectral Light-field Fusion Microscopy for Contrast- and Resolution-enhanced Imaging of Biological Specimens

    CERN Document Server

    Kazemzadeh, Farnoud; Molladavoodi, Sara; Mei, Yu; Emelko, Monica B; Gorbet, Maud B; Wong, Alexander

    2015-01-01

    A lensfree spectral light-field fusion microscopy (LSLFM) system is presented for enabling contrast- and resolution-enhanced imaging of biological specimens. LSLFM consists of a pulsed multispectral lensfree microscope for capturing interferometric light-field encodings at different wavelengths, and Bayesian-based fusion to reconstruct a fused object light-field from the encodings. By fusing unique object detail information captured at different wavelengths, LSLFM can achieve improved resolution, contrast, and signal-to-noise ratio (SNR) over a single-channel lensfree microscopy system. A five-channel LSLFM system was developed and quantitatively evaluated to validate the design. Experimental results demonstrated that the LSLFM system provided SNR improvements of 6.81-16.55 dB, as well as a six-fold improvement in the dispersion index (DI), over that achieved using a single-channel lensfree deconvolution microscopy system at individual wavelengths. Furthermore, the LSLFM system achieved an increase in numeric...

  5. CRYOGENIC MAGNETS

    Science.gov (United States)

    Post, R.F.; Taylor, C.E.

    1963-05-21

    A cryogenic magnet coil is described for generating magnetic fields of the order of 100,000 gauss with a minimum expenditure of energy lost in resistive heating of the coil inductors and energy lost irreversibly in running the coil refrigeration plant. The cryogenic coil comprises a coil conductor for generating a magnetic field upon energization with electrical current, and refrigeration means disposed in heat conductive relation to the coil conductor for cooling to a low temperature. A substantial reduction in the power requirements for generating these magnetic fields is attained by scaling the field generating coil to large size and particular dimensions for a particular conductor, and operating the coil at a particular optimum temperature commensurate with minimum overall power requirements. (AEC)

  6. Cryogenics; Criogenia

    Energy Technology Data Exchange (ETDEWEB)

    Gutierrez R, C.; Jimenez D, J.; Cejudo A, J.; Hernandez M, V. [Instituto Nacional de Investigaciones Nucleares, A.P. 18-1027, 11801 Mexico D.F. (Mexico)

    1997-07-01

    Cryogenics is one of these technologies which contributes to scientific research that supports to the industry in the following benefits: 1. Storage ability and a great quantity of dense gases with cryogenic liquid which is found at high pressure. 2. Production ability at low cost with high purity gases through distillation or condensation. 3. Ability to use low temperatures in the refrigerating materials or alteration of the physical properties. This technology is used for reprocessing of those short and long half life radioactive wastes which always have been required that to be separated with classical methods. In this text we report the radioactive wastes separation by more sophisticated methods but more quickly and reliable. (Author)

  7. [Polarized light microscopy for evaluation of oocytes as a prognostic factor in the evolution of a cycle in assisted reproduction].

    Science.gov (United States)

    González-Ortega, C; Cancino-Villarreal, P; Alonzo-Torres, V E; Martínez-Robles, I; Pérez-Peña, E; Gutiérrez-Gutiérrez, A M

    2016-04-01

    Identification of the best embryos to transfer is a key element for success in assisted reproduction. In the last decade, several morphological criteria of oocytes and embryos were evaluated with regard to their potential for predicting embryo viability. The introduction of polarization light microscopy systems has allowed the visualization of the meiotic spindle and the different layers of the zona pellucida in human oocytes on the basis of birefringence in a non-destructive way. Conflicting results have been reported regarding the predictive value in ICSI cycles. To assess the predictive ability of meiotic spindle and zona pellucida of human oocytes to implant by polarized microscopy in ICSI cycles. Prospective and observational clinical study. 903 oocytes from 94 ICSI cycles were analyzed with polarized microscopy. Meiotic spindle visualization and zona pellucida birefringence values by polarized microscopy were correlated with ICSI cycles results. Meiotic spindle visualization and birefringence values of zona pellucida decreased in a direct basis with increasing age. In patients aged over the 35 years, the percentage of a visible spindle and mean zona pellucida birefringence was lower than in younger patients. Fertilization rate were higher in oocytes with visible meiotic spindle (81.3% vs. 64%; p Polarized light microscopy improves oocyte selection, which significantly impacts in the development of embryos with greater implantation potential. The use of polarized light microscopy with sperm selection methods, blastocyst culture and deferred embryo transfers will contribute to transfer fewer embryos without diminishing rates of live birth and single embryo transfer will be more feasible.

  8. Hierarchical super-structure identified by polarized light microscopy, electron microscopy and nanoindentation: Implications for the limits of biological control over the growth mode of abalone sea shells

    Directory of Open Access Journals (Sweden)

    Schneider Andreas S

    2012-09-01

    Full Text Available Abstract Background Mollusc shells are commonly investigated using high-resolution imaging techniques based on cryo-fixation. Less detailed information is available regarding the light-optical properties. Sea shells of Haliotis pulcherina were embedded for polishing in defined orientations in order to investigate the interface between prismatic calcite and nacreous aragonite by standard materialographic methods. A polished thin section of the interface was prepared with a defined thickness of 60 μm for quantitative birefringence analysis using polarized light and LC-PolScope microscopy. Scanning electron microscopy images were obtained for comparison. In order to study structural-mechanical relationships, nanoindentation experiments were performed. Results Incident light microscopy revealed a super-structure in semi-transparent regions of the polished cross-section under a defined angle. This super-structure is not visible in transmitted birefringence analysis due to the blurred polarization of small nacre platelets and numerous organic interfaces. The relative orientation and homogeneity of calcite prisms was directly identified, some of them with their optical axes exactly normal to the imaging plane. Co-oriented "prism colonies" were identified by polarized light analyses. The nacreous super-structure was also visualized by secondary electron imaging under defined angles. The domains of the super-structure were interpreted to consist of crystallographically aligned platelet stacks. Nanoindentation experiments showed that mechanical properties changed with the same periodicity as the domain size. Conclusions In this study, we have demonstrated that insights into the growth mechanisms of nacre can be obtained by conventional light-optical methods. For example, we observed super-structures formed by co-oriented nacre platelets as previously identified using X-ray Photo-electron Emission Microscopy (X-PEEM [Gilbert et al., Journal of the

  9. Effects of coherence and vector properties of the light on the resolution limit in stimulated emission depletion fluorescence microscopy.

    Science.gov (United States)

    Gao, Wanrong

    2008-06-01

    Stimulated emission depletion (STED) fluorescence microscopy is a diffraction-unlimited microscopy. We report a method of analyzing the intensity distribution in the focal region. The method takes both the coherence and the vector properties of the light into account. By using the Gaussian Schell model to describe the cross-spectral density function of the incident beam, we show that the coherence that exists between the electric field at any two points is one of the factors that limit further increase of the spatial resolution in STED fluorescence microscopy.

  10. Light-sheet microscopy with digital Fourier analysis measures transport properties over large field-of-view.

    Science.gov (United States)

    Wulstein, Devynn M; Regan, Kathryn E; Robertson-Anderson, Rae M; McGorty, Ryan

    2016-09-01

    Using light-sheet microscopy combined with digital Fourier methods we probe the dynamics of colloidal samples and DNA molecules. This combination, referred to as selective-plane illumination differential dynamic microscopy (SPIDDM), has the benefit of optical sectioning to study, with minimal photobleaching, thick samples allowing us to measure the diffusivity of colloidal particles at high volume fractions. Further, SPIDDM exploits the inherent spatially-varying thickness of Gaussian light-sheets. Where the excitation sheet is most focused, we capture high spatial frequency dynamics as the signal-to-background is high. In thicker regions, we capture the slower dynamics as diffusion out of the sheet takes longer.

  11. Summary of 2016 Light Microscopy Module (LMM) Physical Science Experiments on ISS. Update of LMM Science Experiments and Facility Capabilities

    Science.gov (United States)

    Sicker, Ronald J.; Meyer, William V.; Foster, William M.; Fletcher, William A.; Williams, Stuart J.; Lee, Chang-Soo

    2016-01-01

    This presentation will feature a series of short, entertaining, and informative videos that describe the current status and science support for the Light Microscopy Module (LMM) facility on the International Space Station. These interviews will focus on current experiments and provide an overview of future capabilities. The recently completed experiments include nano-particle haloing, 3-D self-assembly with Janus particles and a model system for nano-particle drug delivery. The videos will share perspectives from the scientists, engineers, and managers working with the NASA Light Microscopy program.

  12. Quantitative neuroanatomy of all Purkinje cells with light sheet microscopy and high-throughput image analysis

    Directory of Open Access Journals (Sweden)

    Ludovico eSilvestri

    2015-05-01

    Full Text Available Characterizing the cytoarchitecture of mammalian central nervous system on a brain-wide scale is becoming a compelling need in neuroscience. For example, realistic modeling of brain activity requires the definition of quantitative features of large neuronal populations in the whole brain. Quantitative anatomical maps will also be crucial to classify the cytoarchtitectonic abnormalities associated with neuronal pathologies in a high reproducible and reliable manner. In this paper, we apply recent advances in optical microscopy and image analysis to characterize the spatial distribution of Purkinje cells across the whole cerebellum. Light sheet microscopy was used to image with micron-scale resolution a fixed and cleared cerebellum of an L7-GFP transgenic mouse, in which all Purkinje cells are fluorescently labeled. A fast and scalable algorithm for fully automated cell identification was applied on the image to extract the position of all the fluorescent Purkinje cells. This vectorized representation of the cell population allows a thorough characterization of the complex three-dimensional distribution of the neurons, highlighting the presence of gaps inside the lamellar organization of Purkinje cells, whose density is believed to play a significant role in autism spectrum disorders. Furthermore, clustering analysis of the localized somata permits dividing the whole cerebellum in groups of Purkinje cells with high spatial correlation, suggesting new possibilities of anatomical partition. The quantitative approach presented here can be extended to study the distribution of different types of cell in many brain regions and across the whole encephalon, providing a robust base for building realistic computational models of the brain, and for unbiased morphological tissue screening in presence of pathologies and/or drug treatments.

  13. Comparison of light and electron microscopy in measurement of esophageal intercellular space in children.

    Science.gov (United States)

    Altaf, Muhammad A; Ciecierega, Thomas; Szabo, Sara; Miranda, Adrian; Gorges, Christina; Simpson, Pippa; Sood, Manu R

    2014-08-01

    A good objective marker of esophageal mucosal damage from gastroesophageal reflux disease (GERD) is lacking in children. Increased esophageal epithelial intercellular (EEIC) space measured using electron microscopy (EM) has been proposed as a surrogate of esophageal mucosal damage in adults with GERD. The aim of the present study was to compare EEIC space measured using EM and light microscopy (LM) in children with nonerosive reflux disease (NERD) with asymptomatic controls. Distal esophageal mucosal biopsy was used to measure EEIC space using EM in 35 NERD subjects and 8 controls. In a subset of these patients we used phase contrast LM to measure EEIC space area (26 NERD subjects and 8 controls). The median (range) EEIC space measured using EM in the NERD group was 1.15 (0.74-1.64) μm compared with 0.93 (0.67-1.11) μm in the control group (P = 0.002). The median (range) EEIC space measured using LM was 14.4% (9.6%-26.3%) in the NERD group and 9.6% (8.5%-17.2%) in controls (P = 0.003). Using a cutoff value of 1.02 μm for normal EEIC space measured by EM, we obtained 73% sensitivity and 75% specificity to distinguish the NERD group from the control group, and using a cutoff value of 11.1% for EEIC space measured by LM, we obtained 96% sensitivity and 75% specificity. EEIC space is increased in children with NERD compared with that in controls, suggesting that changes in EEIC space can be a useful marker of esophageal mucosal injury in children with NERD. Our results suggest that the accuracy of EM and LM to evaluate EEIC space changes in NERD is comparable, and LM may be a more cost-effective option.

  14. Diagnostic performance of fluorescent light-emitting diode microscopy for tuberculous lymphadenitis in a high-burden setting.

    Science.gov (United States)

    Abdissa, Ketema; Tadesse, Mulualem; Abdella, Kedir; Bekele, Alemayehu; Bezabih, Mesele; Abebe, Gemeda

    2015-11-01

    Diagnosis of tuberculous lymphadenitis using fine-needle aspiration cytology is a simple and safe but low-specificity method, whereas conventional smear microscopy has variable sensitivity due to low bacterial load. We evaluated the diagnostic performance of fluorescent light-emitting diode (LED) microscopy on routinely collected fine-needle aspirates from tuberculous lymphadenitis presumptive cases. Fine-needle aspirates were collected from patients clinically suspected of having tuberculous lymphadenitis as part of routine diagnosis. Smear preparation was performed from the aspirate and processed for cytology, conventional Ziehl-Neelsen and LED microscopy. The remaining aspirate was processed for culture on Lowenstein-Jensen media. Capilia TB-Neo test was used to differentiate M. tuberculosis complex from non-tuberculous mycobacteria. A total of 144 tuberculous lymphadenitis presumptive cases were included. 66.7% (96/144) were positive for M. tuberculosis complex on culture. Only one isolate was identified as non-tuberculous mycobacteria. The detection rates of Ziehl-Neelsen and LED microscopy were 18.8% (27/144) and 34% (49/144), respectively. As compared to culture, sensitivity was 25.0% [95% CI: 16.3-33.7] for Ziehl-Neelsen microscopy and 45.8% [95% CI: 35.9-55.8] for LED microscopy. The specificity was 93.8% [95% CI: 86.9-100] for Ziehl-Neelsen microscopy and 89.6% [95% CI: 80.9-98.2] for LED microscopy. LED microscopy showed a statistically significant increase in sensitivity and similar specificity compared to Ziehl-Neelsen microscopy. Mean reading time of positive slides was 2.62 min/slide for Ziehl-Neelsen and 1.60 min/slide for LED microscopy. Cytology showed sensitivity of 82.3% and specificity of 54.2%. LED microscopy detected TB bacilli in 33.3% of cases cytologically classified as suppurative abscess. The LED microscopy for tuberculous lymphadenitis had significantly higher sensitivity and shorter screening time than Ziehl-Neelsen microscopy. Use

  15. Three-dimensional reconstruction of specular reflecting technical surfaces using structured light microscopy

    Science.gov (United States)

    Kettel, Johannes; Müller, Claas; Reinecke, Holger

    2014-11-01

    In computer assisted quality control the three-dimensional reconstruction of technical surfaces is playing an ever more important role. Due to the demand on high measurement accuracy and data acquisition rates, structured light optical microscopy has become a valuable solution for the three-dimensional measurement of technical surfaces with high vertical and lateral resolution. However, the three-dimensional reconstruction of specular reflecting technical surfaces with very low surface-roughness and local slopes still remains a challenge to optical measurement principles. Furthermore the high data acquisition rates of current optical measurement systems depend on highly complex and expensive scanning-techniques making them impractical for inline quality control. In this paper we present a novel measurement principle based on a multi-pinhole structured light solution without moving parts which enables the threedimensional reconstruction of specular and diffuse reflecting technical surfaces. This measurement principle is based on multiple and parallel processed point-measurements. These point measurements are realized by spatially locating and analyzing the resulting Point Spread Function (PSF) in parallel for each point measurement. Analysis of the PSF is realized by pattern recognition and model-fitting algorithms accelerated by current Graphics-Processing-Unit (GPU) hardware to reach suitable measurement rates. Using the example of optical surfaces with very low surface-roughness we demonstrate the three-dimensional reconstruction of these surfaces by applying our measurement principle. Thereby we show that the resulting high measurement accuracy enables cost-efficient three-dimensional surface reconstruction suitable for inline quality control.

  16. Atomic Force Microscopy of Red-Light Photoreceptors Using PeakForce Quantitative Nanomechanical Property Mapping

    Science.gov (United States)

    Kroeger, Marie E.; Sorenson, Blaire A.; Thomas, J. Santoro; Stojković, Emina A.; Tsonchev, Stefan; Nicholson, Kenneth T.

    2014-01-01

    Atomic force microscopy (AFM) uses a pyramidal tip attached to a cantilever to probe the force response of a surface. The deflections of the tip can be measured to ~10 pN by a laser and sectored detector, which can be converted to image topography. Amplitude modulation or “tapping mode” AFM involves the probe making intermittent contact with the surface while oscillating at its resonant frequency to produce an image. Used in conjunction with a fluid cell, tapping-mode AFM enables the imaging of biological macromolecules such as proteins in physiologically relevant conditions. Tapping-mode AFM requires manual tuning of the probe and frequent adjustments of a multitude of scanning parameters which can be challenging for inexperienced users. To obtain high-quality images, these adjustments are the most time consuming. PeakForce Quantitative Nanomechanical Property Mapping (PF-QNM) produces an image by measuring a force response curve for every point of contact with the sample. With ScanAsyst software, PF-QNM can be automated. This software adjusts the set-point, drive frequency, scan rate, gains, and other important scanning parameters automatically for a given sample. Not only does this process protect both fragile probes and samples, it significantly reduces the time required to obtain high resolution images. PF-QNM is compatible for AFM imaging in fluid; therefore, it has extensive application for imaging biologically relevant materials. The method presented in this paper describes the application of PF-QNM to obtain images of a bacterial red-light photoreceptor, RpBphP3 (P3), from photosynthetic R. palustris in its light-adapted state. Using this method, individual protein dimers of P3 and aggregates of dimers have been observed on a mica surface in the presence of an imaging buffer. With appropriate adjustments to surface and/or solution concentration, this method may be generally applied to other biologically relevant macromolecules and soft materials. PMID

  17. Optical mapping of conduction in early embryonic quail hearts with light-sheet microscopy (Conference Presentation)

    Science.gov (United States)

    Ma, Pei; Gu, Shi; Wang, Yves T.; Jenkins, Michael W.; Rollins, Andrew M.

    2016-03-01

    Optical mapping (OM) using fluorescent voltage-sensitive dyes (VSD) to measure membrane potential is currently the most effective method for electrophysiology studies in early embryonic hearts due to its noninvasiveness and large field-of-view. Conventional OM acquires bright-field images, collecting signals that are integrated in depth and projected onto a 2D plane, not capturing the 3D structure of the sample. Early embryonic hearts, especially at looping stages, have a complicated, tubular geometry. Therefore, conventional OM cannot provide a full picture of the electrical conduction circumferentially around the heart, and may result in incomplete and inaccurate measurements. Here, we demonstrate OM of Hamburger and Hamilton stage 14 embryonic quail hearts using a new commercially-available VSD, Fluovolt, and depth sectioning using a custom built light-sheet microscopy system. Axial and lateral resolution of the system is 14µm and 8µm respectively. For OM imaging, the field-of-view was set to 900µm×900µm to cover the entire heart. 2D over time OM image sets at multiple cross-sections through the looping-stage heart were recorded. The shapes of both atrial and ventricular action potentials acquired were consistent with previous reports using conventional VSD (di-4-ANNEPS). With Fluovolt, signal-to-noise ratio (SNR) is improved significantly by a factor of 2-10 (compared with di-4-ANNEPS) enabling light-sheet OM, which intrinsically has lower SNR due to smaller sampling volumes. Electrophysiologic parameters are rate dependent. Optical pacing was successfully integrated into the system to ensure heart rate consistency. This will also enable accurately gated reconstruction of full four dimensional conduction maps and 3D conduction velocity measurements.

  18. Estimation of age based on tooth cementum annulations: A comparative study using light, polarized, and phase contrast microscopy.

    Science.gov (United States)

    Kaur, Prabhpreet; Astekar, Madhusudan; Singh, Jappreet; Arora, Karandeep Singh; Bhalla, Gagandeep

    2015-01-01

    The identification of living or deceased persons using unique traits and characteristics of the teeth and jaws is a cornerstone of forensic science. Teeth have been used to estimate age both in the young and old, as well as in the living and dead. Gradual structural changes in teeth throughout life are the basis for age estimation. Tooth cementum annulation (TCA) is a microscopic method for the determination of an individual's age based on the analysis of incremental lines of cementum. To compare ages estimated using incremental lines of cementum as visualized by bright field microscopy, polarized microscopy, and phase contrast microscopy with the actual age of subject and to determine accuracy and feasibility of the method used. Cementum annulations of 60 permanent teeth were analyzed after longitudinal ground sections were made in the mesiodistal plane. The incremental lines were counted manually using a light, polarized and phase contrast microscopy. Ages were estimated and then compared with the actual age of individual. Analysis of variance (ANOVA), Student's t-test, the Pearson product-moment corre (PPMCC) and regression analysis were performed. PPMCC value r = 0.347, 0.542 and 0.989 were obtained using light, polarized and phase contrast microscopy methods respectively. It was concluded that incremental lines of cementum were most clearly visible under a phase contrast microscope, followed by a polarized microscope, and then a light microscope when used for age estimation.

  19. Single objective light-sheet microscopy for high-speed whole-cell 3D super-resolution.

    Science.gov (United States)

    Meddens, Marjolein B M; Liu, Sheng; Finnegan, Patrick S; Edwards, Thayne L; James, Conrad D; Lidke, Keith A

    2016-06-01

    We have developed a method for performing light-sheet microscopy with a single high numerical aperture lens by integrating reflective side walls into a microfluidic chip. These 45° side walls generate light-sheet illumination by reflecting a vertical light-sheet into the focal plane of the objective. Light-sheet illumination of cells loaded in the channels increases image quality in diffraction limited imaging via reduction of out-of-focus background light. Single molecule super-resolution is also improved by the decreased background resulting in better localization precision and decreased photo-bleaching, leading to more accepted localizations overall and higher quality images. Moreover, 2D and 3D single molecule super-resolution data can be acquired faster by taking advantage of the increased illumination intensities as compared to wide field, in the focused light-sheet.

  20. Morphology of the Lingual and Buccal Papillae in Alpaca (Vicugna pacos) - Light and Scanning Electron Microscopy.

    Science.gov (United States)

    Goździewska-Harłajczuk, K; Klećkowska-Nawrot, J; Janeczek, M; Zawadzki, M

    2015-10-01

    The aim of this study was the description of the lingual and buccal papillae in adult alpaca (Vicugna pacos) by light and scanning electron microscopy (SEM). The tongue consisted of apex, body and root. Four types of lingual papillae (filiform, fungiform, conical and circumvallate) in addition to two types of buccal papillae were observed. The filiform papillae, some with secondary papillae, were distributed on both the corpus and apex of the tongue, with stratified epithelium, and layer of keratin coat were recognized. The short (small) cone papillae had pointed top, while bunoform papillae were wide with smooth apex. The much less numerous circumvallate papillae with pseudopapillae on the each rim of the caudal lingual body were present with weak layer of keratin and intra-epithelial taste buds. The small fungiform papillae were found on the dorsal lingual surface, while the large fungiform papillae were situated on the ventral surface of the tongue, especially, in rostral part and were round in shape with numerous gustatory pores and very thin keratin coat. Pseudopapillae were present on the buccal conical 'bunoform' papillae surface, while 'elongate' buccal papillae surface was rather softly folded with thin coat of keratin. Microridges were observed in the less keratinized parts of each type of papillae. The orientation of either lingual or buccal papillae into the throat side facilitates the emptying of oral cavity from nutrient and swallowing of food. In conclusion, the anatomical features of the alpaca tongue are an adaptation to the feeding habits.

  1. Active intracellular transport in metastatic cells studied by spatial light interference microscopy.

    Science.gov (United States)

    Ceballos, Silvia; Kandel, Mikhail; Sridharan, Shamira; Majeed, Hassaan; Monroy, Freddy; Popescu, Gabriel

    2015-01-01

    Spatiotemporal patterns of intracellular transport are very difficult to quantify and, consequently, continue to be insufficiently understood. While it is well documented that mass trafficking inside living cells consists of both random and deterministic motions, quantitative data over broad spatiotemporal scales are lacking. We studied the intracellular transport in live cells using spatial light interference microscopy, a high spatiotemporal resolution quantitative phase imaging tool. The results indicate that in the cytoplasm, the intracellular transport is mainly active (directed, deterministic), while inside the nucleus it is both active and passive (diffusive, random). Furthermore, we studied the behavior of the two-dimensional mass density over 30 h in HeLa cells and focused on the active component. We determined the standard deviation of the velocity distribution at the point of cell division for each cell and compared the standard deviation velocity inside the cytoplasm and the nucleus. We found that the velocity distribution in the cytoplasm is consistently broader than in the nucleus, suggesting mechanisms for faster transport in the cytosol versus the nucleus. Future studies will focus on improving phase measurements by applying a fluorescent tag to understand how particular proteins are transported inside the cell.

  2. The postnatal maturation of efferent tubules in the rat: a light and electron microscopy study.

    Science.gov (United States)

    Francavilla, S; Moscardelli, S; Bruno, B; Barcellona, P S; De Martino, C

    1986-07-01

    The postnatal maturation of the epithelium and tubule wall of efferent tubules in the rat was investigated by light and transmission electron microscopy, from birth to 50 days of age, when sperms were released from the seminiferous tubules and appeared in the genital duct. At the end of the first week of life, an endocytotic apparatus is differentiated in the epithelial cells. During the third week of life, efferent tubules developed specializations for the transport of sperms and fluids, namely the appearance of ciliated elements interspersed among the principal cells of the epithelium, and differentiation of myoid elements in the tubule wall. The appearance of specializations related to endocytosis and fluid transport across the epithelium preceded the canalization of the seminiferous cords which, in fact, is reported to appear at the end of the second week of life in the rat, along with the initial secretion of testicular fluid. This suggested that the maturation of efferent tubules is not triggered by the passage of testicular fluid, as surmised for the postnatal differentiation of caput epididymis. The postnatal maturation of efferent tubules was almost complete 35 days after birth. The appearance of sperms in the genital duct of 50-day-old animals was not associated with any remarkable structural change.

  3. Gradient light interference microscopy (GLIM) for imaging thick specimens (Conference Presentation)

    Science.gov (United States)

    Nguyen, Tan H.; Kandel, Mikhail E.; Popescu, Gabriel

    2016-03-01

    Compared to the Phase Contrast, Differential Interference Contrast (DIC) has been known to give higher depth sectioning as well as a halo-free images when investigating transparent specimens. Thanks to relying on generating two slightly shifted replicas with a small amount of shift, within the coherence area, DIC is able to operate with very low coherence light. More importantly, the method is able to work with very large numerical aperture of the illumination, which offer comparable sectioning capability to bright field microscopy. However, DIC is still a qualitative method, which limits potential applications of the technique. In this paper, we introduce a method that extends the capability of DIC by combining it with a phase shifting module to extract the phase gradient information. A theoretical model of the image formation is developed and the possibility of integrating the gradient function is analyzed.. Our method is benchmarked on imaging embryos during their 7-day development, HeLa cells during mitosis, and control samples.

  4. Structural characterization of the capybara (Hydrochaeris hydrochaeris) tongue by light, scanning, and transmission electron microscopy.

    Science.gov (United States)

    Watanabe, Ii-Sei; Dos Santos Haemmerle, Carlos Alexandre; Dias, Fernando José; Cury, Diego Pulzatto; Da Silva, Marcelo Cavenaghi Pereira; Sosthines, Marcia Consentino Kronka; Dos Santos, Tatiana Carlesco; Guimarães, Juliana Plácido; Miglino, Maria Angélica

    2013-02-01

    Capybara is the largest rodent in the world and displays a seasonally dependent herbivore feeding behavior. Here, we present an anatomical contribution for understand this fact, by light, scanning, and transmission electron microscopy methodologies for tongue tissue analysis. The histological preparations revealed filiform, fungiform, vallate, and foliate papillae on the dorsal mucosa of the capybara tongue. The epithelial layer exhibited a lining of keratinized stratified squamous epithelial cells. The lamina propria was characterized by a dense connective tissue composed of the primary and secondary papillar projections. We also revealed the original aspects of the connective papillae. The shapes of the papillae varied by region of the tongue, and filiform, fungiform, vallate, and foliate papillae and subjacent layers of muscular fibers were observed. Pyriform taste buds occupying the epithelial layer of fungiform, vallate and foliate papillae were identified and the intracellular components of the taste buds and the intracorpuscular amyelinated nerve fibers were observed. The taste buds were characterized by the distribution of granular endoplasmic reticulum throughout the perinuclear area, the Golgi apparatus, and mitochondrial assemblies of various distinct diameters. Mitochondrial accumulation was also observed in the collagen bundle-surrounded amyelinated nerve fibers beside the basal cells. Therefore, these peculiar anatomical descriptions may contribute to understanding the adaptation of the feeding behavior of capybaras in a seasonally changing environment.

  5. Pattern of glomerular diseases in oman: A study based on light microscopy and immunofluorescence

    Directory of Open Access Journals (Sweden)

    Nasar Yousuf Alwahaibi

    2013-01-01

    Full Text Available Light microscopy and immunofluorescence play an important part in the final diagnosis of renal biopsy. The aim of this study was to analyze the pattern of various glomerular diseases in Oman. A total of 424 renal biopsies were retrospectively analyzed at the Sultan Qaboos University Hospital between 1999 and 2010. Focal and segmental glomerulosclerosis (FSGS, minimal change disease (MCD, membranous glomerulopathy (MGN and IgA nephropathy were the most common primary glomerular diseases encountered, accounting for 21.2%, 17%, 12.3% and 8.3%, respectively, of all cases. Lupus nephritis was the most common secondary glomerular disease and was the most prevalent among all biopsies, accounting for 30.4% of all biopsies. Amyloidosis was seen in only two cases. The presence of fluorescein isothiocyanatefibrin in all renal cases was low when compared with IgG, IgA, IgM, C3 and C1q markers. In conclusion, based on the findings of this study, lupus nephritis was the most common of all glomerular diseases and FSGS was the most common primary glomerular disease. The importance of fluorescein isothiocyanate-fibrin in the diagnosis of renal biopsy needs to be further investigated.

  6. Label-Free Imaging of Single Microtubule Dynamics Using Spatial Light Interference Microscopy.

    Science.gov (United States)

    Kandel, Mikhail E; Teng, Kai Wen; Selvin, Paul R; Popescu, Gabriel

    2017-01-24

    Due to their diameter, of only 24 nm, single microtubules are extremely challenging to image without the use of extrinsic contrast agents. As a result, fluorescence tagging is the common method to visualize their motility. However, such investigation is limited by photobleaching and phototoxicity. We experimentally demonstrate the capability of combining label-free spatial light interference microscopy (SLIM) with numerical processing for imaging single microtubules in a gliding assay. SLIM combines four different intensity images to obtain the optical path length map associated with the sample. Because of the use of broadband fields, the sensitivity to path length is better than 1 nm without (temporal) averaging and better than 0.1 nm upon averaging. Our results indicate that SLIM can image the dynamics of microtubules in a full field of view, of 200 × 200 μm(2), over many hours. Modeling the microtubule transport via the diffusion-advection equation, we found that the dispersion relation yields the standard deviation of the velocity distribution, without the need for tracking individual tubes. Interestingly, during a 2 h window, the microtubules begin to decelerate, at 100 pm/s(2) over a 20 min period. Thus, SLIM is likely to serve as a useful tool for understanding molecular motor activity, especially over large time scales, where fluorescence methods are of limited utility.

  7. Microscopical characterization of known postmortem root bands using light and scanning electron microscopy.

    Science.gov (United States)

    Hietpas, Jack; Buscaglia, JoAnn; Richard, Adam H; Shaw, Stephen; Castillo, Hilda S; Donfack, Joseph

    2016-10-01

    A postmortem root band (PMRB) is a distinct microscopic feature that is postulated to occur in hair remaining in the follicle during the postmortem interval [1] (Petraco et al., 1998). The scientific validity of this premise has been highlighted in two recent high-profile criminal cases involving PMRBs [2,3] (State of Florida v. Casey Marie Anthony, 2008; People v. Kogut, 2005). To better understand the fundamental aspects of postmortem root banding, the microscopical properties of known PMRBs(1) were characterized by light microscopy, and scanning electron microscope (SEM) imaging of microtomed sections of hairs showing root banding. The results from this study show that the appearance of the PMRB may be due to the degradation of the chemically labile, non-keratin intermacrofibrillar matrix (IMM) in the pre-keratin/keratogenous region of anagen hairs. In addition, this degradation is confined to the cortex of the hair, with no apparent damage to the layers of the cuticle. These results could provide valuable information for determining the mechanism of band formation, as well as identify a set of microscopic features that could be used to distinguish hairs with known PMRBs from similarly looking environmentally degraded hairs. Published by Elsevier Ireland Ltd.

  8. Isotropic image in structured illumination microscopy patterned with a spatial light modulator.

    Science.gov (United States)

    Chang, Bo-Jui; Chou, Li-Jun; Chang, Yun-Ching; Chiang, Su-Yu

    2009-08-17

    We developed a structured illumination microscopy (SIM) system that uses a spatial light modulator (SLM) to generate interference illumination patterns at four orientations - 0 degrees, 45 degrees, 90 degrees, and 135 degrees, to reconstruct a high-resolution image. The use of a SLM for pattern alterations is rapid and precise, without mechanical calibration; moreover, our design of SLM patterns allows generating the four illumination patterns of high contrast and nearly equivalent periods to achieve a near isotropic enhancement in lateral resolution. We compare the conventional image of 100-nm beads with those reconstructed from two (0 degrees +90 degrees or 45 degrees +135 degrees) and four (0 degrees +45 degrees +90 degrees +135 degrees) pattern orientations to show the differences in resolution and image, with the support of simulations. The reconstructed images of 200-nm beads at various depths and fine structures of actin filaments near the edge of a HeLa cell are presented to demonstrate the intensity distributions in the axial direction and the prospective application to biological systems.

  9. New approach for the quantification of processed animal proteins in feed using light microscopy.

    Science.gov (United States)

    Veys, P; Baeten, V

    2010-07-01

    A revision of European Union's total feed ban on animal proteins in feed will need robust quantification methods, especially for control analyses, if tolerance levels are to be introduced, as for fishmeal in ruminant feed. In 2006, a study conducted by the Community Reference Laboratory for Animal Proteins in feedstuffs (CRL-AP) demonstrated the deficiency of the official quantification method based on light microscopy. The study concluded that the method had to be revised. This paper puts forward an improved quantification method based on three elements: (1) the preparation of permanent slides with an optical adhesive preserving all morphological markers of bones necessary for accurate identification and precision counting; (2) the use of a counting grid eyepiece reticle; and (3) new definitions for correction factors for the estimated portions of animal particles in the sediment. This revised quantification method was tested on feeds adulterated at different levels with bovine meat and bone meal (MBM) and fishmeal, and it proved to be effortless to apply. The results obtained were very close to the expected values of contamination levels for both types of adulteration (MBM or fishmeal). Calculated values were not only replicable, but also reproducible. The advantages of the new approach, including the benefits of the optical adhesive used for permanent slide mounting and the experimental conditions that need to be met to implement the new method correctly, are discussed.

  10. Submicron-resolution photoacoustic microscopy of endogenous light-absorbing biomolecules

    Science.gov (United States)

    Zhang, Chi

    Photoacoustic imaging in biomedicine has the unique advantage of probing endogenous light absorbers at various length scales with a 100% relative sensitivity. Among the several modalities of photoacoustic imaging, optical-resolution photoacoustic microscopy (OR-PAM) can achieve high spatial resolution, on the order of optical wavelength, at detection. We achieved 220 nm lateral resolution in transmission mode, 0.43 microm lateral resolution in reflection mode, 7.6 microm axial resolution in normal tissue, and 5.8 microm axial resolution with silicone oil immersion/injection. The achieved lateral resolution and axial resolution were the finest reported at the time. With high-resolution in 3D, PAM was demonstrated to resolve cellular and subcellular structures in vivo, such as red blood cells and melanosomes in melanoma cells. Compared with previous PAM systems, our high-resolution PAM could resolve capillaries in mouse ears more clearly. As an example application, we demonstrated intracellular temperature imaging, assisted by fluorescence signal detection, with sub-degree temperature resolution and sub-micron lateral resolution. The second part of this dissertation describes the exploration of endogenous light-absorbing biomolecules for PAM. We demonstrated cytochromes and myoglobin as new absorption contrasts for PAM and identified the corresponding optimal wavelengths for imaging. Fixed fibroblasts on slides and mouse ear sections were imaged by PAM at 422 nm and 250 nm wavelengths to reveal cytoplasms and nuclei, respectively, as confirmed by standard hematoxylin and eosin (H&E) histology. By imaging a blood-perfused mouse heart at 532 nm down to 150 microm in depth, we derived the myocardial sheet thickness and the cleavage height from an undehydrated heart for the first time. The findings promote PAM at new wavelengths and open up new possibilities for characterizing biological tissue. Of particular interest, dual-wavelength PAM around 250 nm and 420 nm

  11. Pre-embedding staining of single muscle fibers for light and electron microscopy studies of subcellular organization

    DEFF Research Database (Denmark)

    Ralston, E; Ploug, Thorkil

    1996-01-01

    Skeletal muscle fibers are large, multinucleated cells which pose a challenge to the morphologist. In the course of studies of the distribution of the glucose transporter GLUT4, in muscle, we have compared different preparative procedures, for both light (LM) and electron microscopy (EM) immunocy...

  12. Methods and apparatus of spatially resolved electroluminescence of operating organic light-emitting diodes using conductive atomic force microscopy

    Science.gov (United States)

    Hersam, Mark C. (Inventor); Pingree, Liam S. C. (Inventor)

    2008-01-01

    A conductive atomic force microscopy (cAFM) technique which can concurrently monitor topography, charge transport, and electroluminescence with nanometer spatial resolution. This cAFM approach is particularly well suited for probing the electroluminescent response characteristics of operating organic light-emitting diodes (OLEDs) over short length scales.

  13. Direct Evidence of Lack of Colocalisation of Fluorescently Labelled Gold Labels Used in Correlative Light Electron Microscopy

    Science.gov (United States)

    Miles, Benjamin T.; Greenwood, Alexander B.; Benito-Alifonso, David; Tanner, Hugh; Galan, M. Carmen; Verkade, Paul; Gersen, Henkjan

    2017-01-01

    Fluorescently labelled nanoparticles are routinely used in Correlative Light Electron Microscopy (CLEM) to combine the capabilities of two separate microscope platforms: fluorescent light microscopy (LM) and electron microscopy (EM). The inherent assumption is that the fluorescent label observed under LM colocalises well with the electron dense nanoparticle observed in EM. Herein we show, by combining single molecule fluorescent imaging with optical detection of the scattering from single gold nanoparticles, that for a commercially produced sample of 10 nm gold nanoparticles tagged to Alexa-633 there is in fact no colocalisation between the fluorescent signatures of Alexa-633 and the scattering associated with the gold nanoparticle. This shows that the attached gold nanoparticle quenches the fluorescent signal by ~95%, or less likely that the complex has dissociated. In either scenario, the observed fluorescent signal in fact arises from a large population of untagged fluorophores; rendering these labels potentially ineffective and misleading to the field. PMID:28317888

  14. Grinding and polishing instead of sectioning for the tissue samples with a graft: Implications for light and electron microscopy.

    Science.gov (United States)

    Mukhamadiyarov, Rinat A; Sevostyanova, Victoria V; Shishkova, Daria K; Nokhrin, Andrey V; Sidorova, Olga D; Kutikhin, Anton G

    2016-06-01

    A broad use of the graft replacement requires a detailed investigation of the host-graft interaction, including both histological examination and electron microscopy. A high quality sectioning of the host tissue with a graft seems to be complicated; in addition, it is difficult to examine the same tissue area by both of the mentioned microscopy techniques. To solve these problems, we developed a new technique of epoxy resin embedding with the further grinding, polishing, and staining. Graft-containing tissues prepared by grinding and polishing preserved their structure; however, sectioning frequently required the explantation of the graft and led to tissue disintegration. Moreover, stained samples prepared by grinding and polishing may then be assessed by both light microscopy and backscattered scanning electron microscopy. Therefore, grinding and polishing outperform sectioning when applied to the tissues with a graft.

  15. Small Scroll Pump for Cryogenic Liquids Project

    Data.gov (United States)

    National Aeronautics and Space Administration — The innovation is a compact, reliable, light weight, electrically driven pump capable of pumping cryogenic liquids, based on scroll pump technology. This pump will...

  16. Sperm attributes and morphology on Rusa timorensis: light and scanning electron microscopy.

    Science.gov (United States)

    Mahre, M B; Wahid, H; Rosnina, Y; Jesse, F F A; Azlan, C A; Khumran, A M; Jaji, A Z

    2014-08-01

    This study provides standard information on the attributes of sperm and describes the surface structure of normal and abnormal spermatozoa of Rusa timorensis. Two fertile stags were used as the source of semen collected during the first breeding season commencing from April 5 to July 2, 2012. Another five stags were used as the source of semen collected during the second breeding season commencing from April 1 to June 27, 2013. Semen samples were collected from the stags using an electro-ejaculator. The ejaculate was processed and samples prepared for light and scanning electron microscopy (SEM) according to standard methods. No significant difference (P>0.05) was found between sperm attributes in comparison between different stags and different months of the fertile seasons. The results of this study have also demonstrated that there are no differences in size, shape and surface structure between spermatozoa of the different stags and different months of the fertile seasons. Sperm attributes (volume, pH, sperm concentration, general motility, progressive motility and viability) were 2.2±0.29 ml, 7.2±0.17, 886.3±39.7×10(6) spermatozoa/ml, 78.7±2.01%, 80.8±1.85% and 83.2±0.85%, respectively. Morphological analysis showed low percentage of abnormal spermatozoa 13.9±2.88%. Scanning electron microscopy revealed spermatozoa which consisted of a flat paddle-shaped head, short neck and a tail, which was subdivided into midpiece, principal piece and endpiece. The average spermatozoon was 66.2±0.69 μm in total length. The flat paddle-shaped head was 7.8±0.28 μm long, 4.2±0.15 μm at its widest width, 2.4±0.18 μm basal width and 0.7±0.0 2μm thick. As for the tail, the midpiece length was 13.2±0.14 μm, 0.6±0.04 μm in diameter; the principal piece was 42.6±0.04μm, and 2.8±0.06 μm for the endpiece. Abnormal spermatozoa such as tapered head, microcephalic head, decapitated spermatozoa and bent tails were observed. Results provide standard information

  17. Visualizing viral protein structures in cells using genetic probes for correlated light and electron microscopy.

    Science.gov (United States)

    Ou, Horng D; Deerinck, Thomas J; Bushong, Eric; Ellisman, Mark H; O'Shea, Clodagh C

    2015-11-15

    Structural studies of viral proteins most often use high-resolution techniques such as X-ray crystallography, nuclear magnetic resonance, single particle negative stain, or cryo-electron microscopy (EM) to reveal atomic interactions of soluble, homogeneous viral proteins or viral protein complexes. Once viral proteins or complexes are separated from their host's cellular environment, their natural in situ structure and details of how they interact with other cellular components may be lost. EM has been an invaluable tool in virology since its introduction in the late 1940's and subsequent application to cells in the 1950's. EM studies have expanded our knowledge of viral entry, viral replication, alteration of cellular components, and viral lysis. Most of these early studies were focused on conspicuous morphological cellular changes, because classic EM metal stains were designed to highlight classes of cellular structures rather than specific molecular structures. Much later, to identify viral proteins inducing specific structural configurations at the cellular level, immunostaining with a primary antibody followed by colloidal gold secondary antibody was employed to mark the location of specific viral proteins. This technique can suffer from artifacts in cellular ultrastructure due to compromises required to provide access to the immuno-reagents. Immunolocalization methods also require the generation of highly specific antibodies, which may not be available for every viral protein. Here we discuss new methods to visualize viral proteins and structures at high resolutions in situ using correlated light and electron microscopy (CLEM). We discuss the use of genetically encoded protein fusions that oxidize diaminobenzidine (DAB) into an osmiophilic polymer that can be visualized by EM. Detailed protocols for applying the genetically encoded photo-oxidizing protein MiniSOG to a viral protein, photo-oxidation of the fusion protein to yield DAB polymer staining, and

  18. Laser Light-field Fusion for Wide-field Lensfree On-chip Phase Contrast Microscopy of Nanoparticles

    Science.gov (United States)

    Kazemzadeh, Farnoud; Wong, Alexander

    2016-12-01

    Wide-field lensfree on-chip microscopy, which leverages holography principles to capture interferometric light-field encodings without lenses, is an emerging imaging modality with widespread interest given the large field-of-view compared to lens-based techniques. In this study, we introduce the idea of laser light-field fusion for lensfree on-chip phase contrast microscopy for detecting nanoparticles, where interferometric laser light-field encodings acquired using a lensfree, on-chip setup with laser pulsations at different wavelengths are fused to produce marker-free phase contrast images of particles at the nanometer scale. As a proof of concept, we demonstrate, for the first time, a wide-field lensfree on-chip instrument successfully detecting 300 nm particles across a large field-of-view of ~30 mm2 without any specialized or intricate sample preparation, or the use of synthetic aperture- or shift-based techniques.

  19. Lights Will Guide You : Sample Preparation and Applications for Integrated Laser and Electron Microscopy

    Science.gov (United States)

    Karreman, M. A.

    2013-03-01

    Correlative microscopy is the combined use of two different forms of microscopy in the study of a specimen, allowing for the exploitation of the advantages of both imaging tools. The integrated Laser and Electron Microscope (iLEM), developed at Utrecht University, combines a fluorescence microscope (FM) and a transmission electron microscope (TEM) in a single set-up. The region of interest in the specimen is labeled or tagged with a fluorescent probe and can easily be identified within a large field of view with the FM. Next, this same area is retraced in the TEM and can be studied at high resolution. The iLEM demands samples that can be imaged with both FM and TEM. Biological specimen, typically composed of light elements, generate low image contrast in the TEM. Therefore, these samples are often ‘contrasted’ with heavy metal stains. FM, on the other hand, images fluorescent samples. Sample preparation for correlative microscopy, and iLEM in particular, is complicated by the fact that the heavy metals stains employed for TEM quench the fluorescent signal of the probe that is imaged with FM. The first part of this thesis outlines preparation procedures for biological material yielding specimen that can be imaged with the iLEM. Here, approaches for the contrasting of thin sections of cells and tissue are introduced that do not affect the fluorescence signal of the probe that marks the region of interest. Furthermore, two novel procedures, VIS2FIXH and VIS2FIX­FS are described that allow for the chemical fixation of thin sections of cryo-immobilized material. These procedures greatly expedite the sample preparation process, and open up novel possibilities for the immuno-labeling of difficult antigens, eg. proteins and lipids that are challenging to preserve. The second part of this thesis describes applications of iLEM in research in the field of life and material science. The iLEM was employed in the study of UVC induced apoptosis (programmed cell death) of

  20. Foetal and postnatal equine articular cartilage development: magnetic resonance imaging and polarised light microscopy

    Directory of Open Access Journals (Sweden)

    C Cluzel

    2013-08-01

    Full Text Available Adult articular cartilage (AC has a well described multizonal collagen structure. Knowledge of foetal AC organisation and development may provide a prototype for cartilage repair strategies, and improve understanding of structural changes in developmental diseases such as osteochondrosis (OC. The objective of this study was to describe normal development of the spatial architecture of the collagen network of equine AC using 1.5 T magnetic resonance imaging (MRI and polarised light microscopy (PLM, at sites employed for cartilage repair studies or susceptible to OC. T2-weighted fast-spin echo (FSE sequences and PLM assessment were performed on distal femoral epiphyses of equine foetuses, foals and adults. Both MRI and PLM revealed an early progressive collagen network zonal organisation of the femoral epiphyses, beginning at 4 months of gestation. PLM revealed that the collagen network of equine foetal AC prior to birth was already organised into an evident anisotropic layered structure that included the appearance of a dense tangential zone in the superficial AC in the youngest specimens, with the progressive development of an underlying transitional zone. A third, increasingly birefringent, radial layer developed in the AC from 6 months of gestation. Four laminae were observed on the MR images in the last third of gestation. These included not only the AC but also the superficial growth plate of the epiphysis. These findings provide novel data on normal equine foetal cartilage collagen development, and may serve as a template for cartilage repair studies in this species or a model for developmental studies of OC.

  1. Modeling of fibrin gels based on confocal microscopy and light-scattering data.

    Science.gov (United States)

    Magatti, Davide; Molteni, Matteo; Cardinali, Barbara; Rocco, Mattia; Ferri, Fabio

    2013-03-01

    Fibrin gels are biological networks that play a fundamental role in blood coagulation and other patho/physiological processes, such as thrombosis and cancer. Electron and confocal microscopies show a collection of fibers that are relatively monodisperse in diameter, not uniformly distributed, and connected at nodal points with a branching order of ∼3-4. Although in the confocal images the hydrated fibers appear to be quite straight (mass fractal dimension D(m) = 1), for the overall system 1, joined at randomly distributed nodal points. The resulting 3D network strikingly resembles real fibrin gels and can be sketched as an assembly of densely packed fractal blobs, i.e., regions of size ξ, where the fiber concentration is higher than average. The blobs are placed at a distance ξ0 between their centers of mass so that they are overlapped by a factor η =ξ/ξ0 and have D(m) ∼1.2-1.6. The in silico gels' structure is quantitatively analyzed by its 3D spatial correlation function g(3D)(r) and corresponding power spectrum I(q) = FFT(3D[g3D(r)]), from which ρ, d, D(m), η, and ξ0 can be extracted. In particular, ξ0 provides an excellent estimate of the gel mesh size. The in silico gels' I(q) compares quite well with real gels' elastic light-scattering measurements. We then derived an analytical form factor for accurately fitting the scattering data, which allowed us to directly recover the gels' structural parameters.

  2. A fast image registration approach of neural activities in light-sheet fluorescence microscopy images

    Science.gov (United States)

    Meng, Hui; Hui, Hui; Hu, Chaoen; Yang, Xin; Tian, Jie

    2017-03-01

    The ability of fast and single-neuron resolution imaging of neural activities enables light-sheet fluorescence microscopy (LSFM) as a powerful imaging technique in functional neural connection applications. The state-of-art LSFM imaging system can record the neuronal activities of entire brain for small animal, such as zebrafish or C. elegans at single-neuron resolution. However, the stimulated and spontaneous movements in animal brain result in inconsistent neuron positions during recording process. It is time consuming to register the acquired large-scale images with conventional method. In this work, we address the problem of fast registration of neural positions in stacks of LSFM images. This is necessary to register brain structures and activities. To achieve fast registration of neural activities, we present a rigid registration architecture by implementation of Graphics Processing Unit (GPU). In this approach, the image stacks were preprocessed on GPU by mean stretching to reduce the computation effort. The present image was registered to the previous image stack that considered as reference. A fast Fourier transform (FFT) algorithm was used for calculating the shift of the image stack. The calculations for image registration were performed in different threads while the preparation functionality was refactored and called only once by the master thread. We implemented our registration algorithm on NVIDIA Quadro K4200 GPU under Compute Unified Device Architecture (CUDA) programming environment. The experimental results showed that the registration computation can speed-up to 550ms for a full high-resolution brain image. Our approach also has potential to be used for other dynamic image registrations in biomedical applications.

  3. Inclusion bodies induced by bean rugose mosaic virus seen under light microscopy

    Directory of Open Access Journals (Sweden)

    Carmen Rivera

    2000-12-01

    Full Text Available Two types of inclusion bodies were consistently observed under light microscopy in bean (Phaseolus vulgaris leaf tissue infected with bean rugose mosaic virus (BRMV, a species of the genus Comovirus, family Comoviridae. One type consisted of vacuolated inclusions found mainly in the cytoplasm of epidermal cells. The other type consisted of abundant crystalloid inclusions of different sizes and shapes found consistently in glandular hairs, guard cells, phloem tissue, xylem elements and occasionally in epidermal and mesophyll tissues. The two types of inclusion bodies stained with Azure A and Luxol Brilliant Green Bl-Calcomine Orange 2RS (O-G, and were similar to those seen to be caused by other species of comoviruses.Se observaron dos tipos de inclusiones virales, mediante microscopia de luz, en hojas de plantas de frijol (Phaseolus vulgaris previamente infectadas con el virus del mosaico rugoso del frijol ("bean rugose mosaic comovirus", BRMV, especie del género Comovirus, familia Comoviridae. Se hallaron inclusiones vesiculadas, principalmente en el citoplasma de células de la epidermis, y abundantes inclusiones cristalinas de diferentes formas y tamaños siempre en células guarda, tricomas glandulares, floema, elementos del xilema y ocasionalmente en células epidérmicas y del mesófilo. Ambos tipos de inclusiones tiñeron con Azure A y con la tinción, verde naranja (Luxol Brilliant Green BL-Calcomine Orange 2 RS conocida como OG, y son similares a las inclusiones inducidas por otras especies del género Comovirus.

  4. Micellar aggregates of saponins from Chenopodium quinoa: characterization by dynamic light scattering and transmission electron microscopy.

    Science.gov (United States)

    Verza, S G; de Resende, P E; Kaiser, S; Quirici, L; Teixeira, H F; Gosmann, G; Ferreira, F; Ortega, G G

    2012-04-01

    Entire seeds of Chenopodium quinoa Willd are a rich protein source and are also well-known for their high saponin content. Due to their amphiphily quinoa saponins are able to form intricate micellar aggregates in aqueous media. In this paper we study the aggregates formed by self-association of these compounds from two quinoa saponin fractions (FQ70 and FQ90) as well as several distinctive nanostructures obtained after their complexation with different ratios of cholesterol (CHOL) and phosphatidylcholine (PC). The FQ70 and FQ90 fractions were obtained by reversed-phase preparative chromatography. The structural features of their resulting aggregates were determined by Dynamic Light Scattering (DLS) and Transmission Electron Microscopy (TEM). Novel nanosized spherical vesicles formed by self-association with mean diameter about 100-200 nm were observed in FQ70 aqueous solutions whereas worm-like micelles an approximate width of 20 nm were detected in FQ90 aqueous solutions. Under experimental conditions similar to those reported for the preparation of Quillaja saponaria ISCOM matrices, tubular and ring-like micelles arose from FQ70:CHOL:PC and FQ90:CHOL:PC formulations, respectively. However, under these conditions no cage-like ISCOM matrices were observed. The saponin composition of FQ70 and FQ90 seems to determine the nanosized structures viewed by TEM. Phytolaccagenic acid, predominant in FQ70 and FQ90 fractions, is accountable for the formation of the nanosized vesicles and tubular structures observed by TEM in the aqueous solutions of both samples. Conversely, ring-like micelles observed in FQ90:CHOL:PC complexes can be attributed to the presence of less polar saponins present in FQ90, in particular those derived from oleanolic acid.

  5. In Depth Analyses of LEDs by a Combination of X-ray Computed Tomography (CT) and Light Microscopy (LM) Correlated with Scanning Electron Microscopy (SEM).

    Science.gov (United States)

    Meyer, Jörg; Thomas, Christian; Tappe, Frank; Ogbazghi, Tekie

    2016-06-16

    In failure analysis, device characterization and reverse engineering of light emitting diodes (LEDs), and similar electronic components of micro-characterization, plays an important role. Commonly, different techniques like X-ray computed tomography (CT), light microscopy (LM) and scanning electron microscopy (SEM) are used separately. Similarly, the results have to be treated for each technique independently. Here a comprehensive study is shown which demonstrates the potentials leveraged by linking CT, LM and SEM. In depth characterization is performed on a white emitting LED, which can be operated throughout all characterization steps. Major advantages are: planned preparation of defined cross sections, correlation of optical properties to structural and compositional information, as well as reliable identification of different functional regions. This results from the breadth of information available from identical regions of interest (ROIs): polarization contrast, bright and dark-field LM images, as well as optical images of the LED cross section in operation. This is supplemented by SEM imaging techniques and micro-analysis using energy dispersive X-ray spectroscopy.

  6. Uptake and localization of fluorescent labelled gold nanoparticles in living zebrafish (Danio rerio) using Light Sheet Microscopy

    DEFF Research Database (Denmark)

    Skjolding, Lars Michael; Asmonaite, G.; Jolk, R.

    2015-01-01

    and localization of fluorescent labelled nanoparticles in living whole organisms with minimal sample preparation. Two strains of D. rerio (wildtype AB and transparent Casper) were exposed to 50 nm PEG coated gold nanoparticles (Au NP) synthesized with 1% of a fluorescent probe (FITC). The fish were exposed...... and determine localization on a whole organism level. Furthermore, methods used to identify nanoparticle uptake have been associated with artefacts induced by sample preparation including staining methods for electron microscopy.  This study used Fluorescent Light Sheet Microscopy (FLSM) to determine uptake...... the suitability for whole imaging of living organisms using FLSM....

  7. Virtual-'light-sheet' single-molecule localisation microscopy enables quantitative optical sectioning for super-resolution imaging.

    Science.gov (United States)

    Palayret, Matthieu; Armes, Helen; Basu, Srinjan; Watson, Adam T; Herbert, Alex; Lando, David; Etheridge, Thomas J; Endesfelder, Ulrike; Heilemann, Mike; Laue, Ernest; Carr, Antony M; Klenerman, David; Lee, Steven F

    2015-01-01

    Single-molecule super-resolution microscopy allows imaging of fluorescently-tagged proteins in live cells with a precision well below that of the diffraction limit. Here, we demonstrate 3D sectioning with single-molecule super-resolution microscopy by making use of the fitting information that is usually discarded to reject fluorophores that emit from above or below a virtual-'light-sheet', a thin volume centred on the focal plane of the microscope. We describe an easy-to-use routine (implemented as an open-source ImageJ plug-in) to quickly analyse a calibration sample to define and use such a virtual light-sheet. In addition, the plug-in is easily usable on almost any existing 2D super-resolution instrumentation. This optical sectioning of super-resolution images is achieved by applying well-characterised width and amplitude thresholds to diffraction-limited spots that can be used to tune the thickness of the virtual light-sheet. This allows qualitative and quantitative imaging improvements: by rejecting out-of-focus fluorophores, the super-resolution image gains contrast and local features may be revealed; by retaining only fluorophores close to the focal plane, virtual-'light-sheet' single-molecule localisation microscopy improves the probability that all emitting fluorophores will be detected, fitted and quantitatively evaluated.

  8. Virtual-'light-sheet' single-molecule localisation microscopy enables quantitative optical sectioning for super-resolution imaging.

    Directory of Open Access Journals (Sweden)

    Matthieu Palayret

    Full Text Available Single-molecule super-resolution microscopy allows imaging of fluorescently-tagged proteins in live cells with a precision well below that of the diffraction limit. Here, we demonstrate 3D sectioning with single-molecule super-resolution microscopy by making use of the fitting information that is usually discarded to reject fluorophores that emit from above or below a virtual-'light-sheet', a thin volume centred on the focal plane of the microscope. We describe an easy-to-use routine (implemented as an open-source ImageJ plug-in to quickly analyse a calibration sample to define and use such a virtual light-sheet. In addition, the plug-in is easily usable on almost any existing 2D super-resolution instrumentation. This optical sectioning of super-resolution images is achieved by applying well-characterised width and amplitude thresholds to diffraction-limited spots that can be used to tune the thickness of the virtual light-sheet. This allows qualitative and quantitative imaging improvements: by rejecting out-of-focus fluorophores, the super-resolution image gains contrast and local features may be revealed; by retaining only fluorophores close to the focal plane, virtual-'light-sheet' single-molecule localisation microscopy improves the probability that all emitting fluorophores will be detected, fitted and quantitatively evaluated.

  9. Preservation of protein fluorescence in embedded human dendritic cells for targeted 3D light and electron microscopy.

    Science.gov (United States)

    Höhn, K; Fuchs, J; Fröber, A; Kirmse, R; Glass, B; Anders-Össwein, M; Walther, P; Kräusslich, H-G; Dietrich, C

    2015-08-01

    In this study, we present a correlative microscopy workflow to combine detailed 3D fluorescence light microscopy data with ultrastructural information gained by 3D focused ion beam assisted scanning electron microscopy. The workflow is based on an optimized high pressure freezing/freeze substitution protocol that preserves good ultrastructural detail along with retaining the fluorescence signal in the resin embedded specimens. Consequently, cellular structures of interest can readily be identified and imaged by state of the art 3D confocal fluorescence microscopy and are precisely referenced with respect to an imprinted coordinate system on the surface of the resin block. This allows precise guidance of the focused ion beam assisted scanning electron microscopy and limits the volume to be imaged to the structure of interest. This, in turn, minimizes the total acquisition time necessary to conduct the time consuming ultrastructural scanning electron microscope imaging while eliminating the risk to miss parts of the target structure. We illustrate the value of this workflow for targeting virus compartments, which are formed in HIV-pulsed mature human dendritic cells. © 2015 The Authors Journal of Microscopy © 2015 Royal Microscopical Society.

  10. CRYOGENIC DEWAR

    Science.gov (United States)

    Chamberlain, W.H.; Maseck, H.E.

    1964-01-28

    This patent relates to a dewar for storing cryogenic gase and is of the type having aii inner flask surrounded by a vacuum jacket and having a vent spout through which evaporating gas escapes. Heretofore substantial gas loss has resulted from the radiation of heat towards the flask from the warmer outer elements of the dewar. In this invention, the mask is surrounded by a thermally conducting shield which is disposed in the vacuum space between the flask and the outer elements of the dewar. The shield contacts only the vent spout, which is cooled by the evaporating gas, and thus is maintained at a temperature very close to that of the flask itself. Accordingly, heat radiated toward the flask is intercepted and conducted to the evaporating gas rather than being re-radiated towards the hask. In a liquid helium dewar of typical configniration the mention reduces the boil-off rate by approximately one-half.(AEC)

  11. Lights Will Guide You : Sample Preparation and Applications for Integrated Laser and Electron Microscopy

    NARCIS (Netherlands)

    Karreman, M.A.

    2013-01-01

    Correlative microscopy is the combined use of two different forms of microscopy in the study of a specimen, allowing for the exploitation of the advantages of both imaging tools. The integrated Laser and Electron Microscope (iLEM), developed at Utrecht University, combines a fluorescence microscope

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

    Science.gov (United States)

    Singh, Jasmine N.; Nowlin, Taylor M.; Seedorf, Gregory J.; Abman, Steven H.; Shepherd, Douglas P.

    2017-07-01

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

  13. High-resolution corneal topography and tomography of fish eye using wide-field white light interference microscopy

    Science.gov (United States)

    Srivastava, Vishal; Nandy, Sreyankar; Singh Mehta, Dalip

    2013-04-01

    Topography and tomography of fish cornea is reconstructed using high resolution white light interference microscopy. White light interferograms at different depths were recorded by moving the object axially. For each depth position, five phase shifted interferograms were recorded and analyzed. From the reconstructed phase maps, the corneal topography and hence the refractive index was determined and from amplitude images the cross-sectional image of fish cornea was reconstructed. In the present method, we utilize a nearly common-path interference microscope and wide field illumination and hence do not require any mechanical B-scan. Therefore, the phase stability of the recorded data is improved.

  14. Light depolarization induced by metallic tips in apertureless near-field optical microscopy and tip-enhanced Raman spectroscopy

    Energy Technology Data Exchange (ETDEWEB)

    Gucciardi, P G [CNR-Istituto per i Processi Chimico-Fisici, sezione Messina, Salita Sperone, Contrada Papardo, I-98158 Faro Superiore, Messina (Italy); Lopes, M; Deturche, R; Julien, C; Barchiesi, D; Chapelle, M Lamy de la [Institut Charles Delaunay-CNRS FRE 2848, Laboratoire de Nanotechnologie et d' Instrumentation Optique, Universite de Technologie de Troyes, 12 rue Marie Curie, BP2060, 10010 Troyes (France)

    2008-05-28

    We have investigated the depolarization effects of light scattered by sharp tips used for apertureless near-field optical microscopy. Dielectric and metal coated tips have been investigated and depolarization factors between 5 and 30% have been measured, changing as a function of the incident light polarization and of the tip shape. The experimental results are in good agreement with theoretical calculations performed by the finite element method, giving a near-field depolarization factor close to 10%. The effect of depolarization has been investigated in polarized tip-enhanced Raman spectroscopy (TERS) experiments; the depolarization gives rise to forbidden Raman modes in Si crystals.

  15. Comet assay, cloning assay, and light and electron microscopy on one preselected cell

    Science.gov (United States)

    Koenig, Karsten; Oehring, Hartmut; Halbhuber, Karl-Juergen; Fiedler, Ursula; Bauer, Eckhard; Greulich, Karl-Otto

    1998-01-01

    In order to perform long-term studies up to one week on a preselected single cell after micromanipulation (e.g. UVA and NIR microbeam exposure) in comparison with non-treated neighbor cells (control cells) we applied a variety of single cell diagnostic techniques and developed a special comet assay for single preselected cells. For that purpose adherent cells were grown in low concentrations and maintained in special sterile centimeter-sized glass cell chambers. After preselection, a single cell was marked by means of diamond-produced circles on the outer cell chamber window. During exposure to microbeams, NADH-attributed autofluorescence of the chosen cell was detected by fluorescence imaging and spectroscopy. In addition, cell morphology was video-monitored (formation of pseudopodia, membrane blebbing,...). Maintaining the microchamber in the incubator, the irradiated cell was examined 24 h later for cell division (clone formation) and modifications in autofluorescence and morphology (including daughter cells). In the case that no division occurred the vitality of the light-exposed cell and of the control cells were probed by intranuclear propidium iodide accumulation. After fixation, either electron microscopy or single cell gel electrophoresis (comet assay) was performed. To monitor comet formation indicating photoinduced DNA damage in the preselected single cell in comparison with the non-exposed neighbor cells the chamber was filled with low-melting gel and lysis solution and exposed to an electric field. In contrast to the conventional comet assay, where only randomly chosen cells of a suspension are investigated, the novel optimized electrophoresis technique should enhance the possibilities of DNA damage detection to a true single (preselected) cell level. The single cell techniques applied to UVA microexposed Chinese hamster ovary cells (364 nm, 1 mW, 3.5 W/cm2) revealed significant cell damage for J/cm2 fluences such as modifications of intracellular

  16. Observation platform for colloid science experiments using the ISS Light Microscopy Module

    Science.gov (United States)

    Kurk, Michael Andy; Todd, Paul; Vellinger, John C.

    2012-07-01

    The role of gravity in colloid self-assembly is a long-standing subject of inquiry. The International Space Station Light Microscopy Module (LMM) is a potentially powerful tool for implementing the observation of colloids in a high-quality low-gravity environment. The main requirements for making observations of colloid self-assembly include a small-volume, thermally stabilized environment, the addition and removal of small volumes of fluids (colloidal suspensions or reagents), and on-demand access to electrokinetic and/or magnetophoretic body forces. A modular device has been designed in which a custom electronics module is designed to mate with the existing LMM cold plate and LMM controlling power. All control features, electrical power, microscope illuminator, fluid pumps and valves are components of this module. This module lies under, mates with and serves an experiment module which houses the fluid containers that fit under the LMM objective lenses and fluid transfer tubing. Four versions of the experiment module have been designed: a hollow-slide stopped flow cell, a multiwell quiescent module, a magnetization module and an electrokinetic module. Interfaces can be established in the viewing field using the stopped-flow cell, which is also applicable to living systems such as microbial cultures, suspended blood cells, nematodes, etc. Multi-well modules can be equipped with in-line static mixers that allow the investigator to combine pairs of fluids or to re-homogenize settled samples. The maximum dimension of all modules is 16 cm, so the modules can be transported in large numbers on cargo or manned spacecraft to the ISS. A doubly-contained transfer tool can be used to transfer fluids in and out of the experiment module, which is equipped with a fluid coupling that mates to the transfer tool. Experiment-specific versions of these modules can be prepared for approved experimenters within a 1-year period. The research for these devices is supported by NASA

  17. A Review of Correlative Light and Electron Microscopy (CLEM) Methods, Markers, and Instrument Set Ups to Study Infectious Disease

    Science.gov (United States)

    2016-12-12

    with the oxidation reaction [43], and limited penetration [43, 72]. In fixed and non-fixed cells mitochondria generate reactive oxygen species (ROS... mitochondria during apoptosis. Nat Cell Biol, 2007. 9(9): p. 1057-65. 22. Kong, D. and J. Loncarek, Correlative light and electron microscopy analysis...there is need to study, at nanoscale, objects of interest which are commonly part of rare transient events or afflict a particular cell among a

  18. Analysis of acute brain slices by electron microscopy: a correlative light-electron microscopy workflow based on Tokuyasu cryo-sectioning.

    Science.gov (United States)

    Loussert Fonta, Celine; Leis, Andrew; Mathisen, Cliff; Bouvier, David S; Blanchard, Willy; Volterra, Andrea; Lich, Ben; Humbel, Bruno M

    2015-01-01

    Acute brain slices are slices of brain tissue that are kept vital in vitro for further recordings and analyses. This tool is of major importance in neurobiology and allows the study of brain cells such as microglia, astrocytes, neurons and their inter/intracellular communications via ion channels or transporters. In combination with light/fluorescence microscopies, acute brain slices enable the ex vivo analysis of specific cells or groups of cells inside the slice, e.g. astrocytes. To bridge ex vivo knowledge of a cell with its ultrastructure, we developed a correlative microscopy approach for acute brain slices. The workflow begins with sampling of the tissue and precise trimming of a region of interest, which contains GFP-tagged astrocytes that can be visualised by fluorescence microscopy of ultrathin sections. The astrocytes and their surroundings are then analysed by high resolution scanning transmission electron microscopy (STEM). An important aspect of this workflow is the modification of a commercial cryo-ultramicrotome to observe the fluorescent GFP signal during the trimming process. It ensured that sections contained at least one GFP astrocyte. After cryo-sectioning, a map of the GFP-expressing astrocytes is established and transferred to correlation software installed on a focused ion beam scanning electron microscope equipped with a STEM detector. Next, the areas displaying fluorescence are selected for high resolution STEM imaging. An overview area (e.g. a whole mesh of the grid) is imaged with an automated tiling and stitching process. In the final stitched image, the local organisation of the brain tissue can be surveyed or areas of interest can be magnified to observe fine details, e.g. vesicles or gold labels on specific proteins. The robustness of this workflow is contingent on the quality of sample preparation, based on Tokuyasu's protocol. This method results in a reasonable compromise between preservation of morphology and maintenance of

  19. A new method of evaluation of fracture patterns following microtensile bond strength testing using polarized light microscopy.

    Science.gov (United States)

    Hamama, Hamdi H; Yiu, Cynthia K Y; Burrow, Michael F

    2014-08-01

    This work describes a new method using polarized light microscopy to determine the failure modes of fractured beams following microtensile bond strength testing. The outcomes were validated using SEM and EDX elemental analysis. Resin adhesives and resin composites bonded to caries-free dentin samples as well as disks of adhesive and composite were observed with reflected polarized light microscopy (PLM) to obtain standard images. A set of beams fractured in the microtensile bond test were observed with PLM and compared with the standard images to determine failure mode through PLM color matching with the standard dentin, adhesive, or composite images. Samples were analyzed by EDX under SEM and compared with the PLM outcomes. Reflected PLM images showed that the fractured surfaces covered with resin-based materials (adhesives or composite) appeared pink in color, in contrast to dentin surfaces, which appeared yellow. EDX mapping together with SEM observation confirmed the results obtained by PLM. The results of EDX mapping and SEM observation showed that the use of polarized light microscopy is a simple, viable method for differentiation between the resin-covered dentin surfaces for determining fracture pattern analysis after bond testing.

  20. In situ transmission electron microscopy of light-induced photocatalytic reactions

    DEFF Research Database (Denmark)

    Cavalca, Filippo; Laursen, Anders Bo; Kardynal, Beata

    2012-01-01

    Transmission electron microscopy (TEM) makes it possible to obtain insight into the structure, composition and reactivity of photocatalysts, which are of fundamental interest for sustainable energy research. Such insight can be used for further material optimization. Here, we combine conventional...

  1. Super-Resolution Microscopy: Shedding Light on the Cellular Plasma Membrane.

    Science.gov (United States)

    Stone, Matthew B; Shelby, Sarah A; Veatch, Sarah L

    2017-02-17

    Lipids and the membranes they form are fundamental building blocks of cellular life, and their geometry and chemical properties distinguish membranes from other cellular environments. Collective processes occurring within membranes strongly impact cellular behavior and biochemistry, and understanding these processes presents unique challenges due to the often complex and myriad interactions between membrane components. Super-resolution microscopy offers a significant gain in resolution over traditional optical microscopy, enabling the localization of individual molecules even in densely labeled samples and in cellular and tissue environments. These microscopy techniques have been used to examine the organization and dynamics of plasma membrane components, providing insight into the fundamental interactions that determine membrane functions. Here, we broadly introduce the structure and organization of the mammalian plasma membrane and review recent applications of super-resolution microscopy to the study of membranes. We then highlight some inherent challenges faced when using super-resolution microscopy to study membranes, and we discuss recent technical advancements that promise further improvements to super-resolution microscopy and its application to the plasma membrane.

  2. Scanning superlens microscopy for non-invasive large field-of-view visible light nanoscale imaging

    Science.gov (United States)

    Wang, Feifei; Liu, Lianqing; Yu, Haibo; Wen, Yangdong; Yu, Peng; Liu, Zhu; Wang, Yuechao; Li, Wen Jung

    2016-12-01

    Nanoscale correlation of structural information acquisition with specific-molecule identification provides new insight for studying rare subcellular events. To achieve this correlation, scanning electron microscopy has been combined with super-resolution fluorescent microscopy, despite its destructivity when acquiring biological structure information. Here we propose time-efficient non-invasive microsphere-based scanning superlens microscopy that enables the large-area observation of live-cell morphology or sub-membrane structures with sub-diffraction-limited resolution and is demonstrated by observing biological and non-biological objects. This microscopy operates in both non-invasive and contact modes with ~200 times the acquisition efficiency of atomic force microscopy, which is achieved by replacing the point of an atomic force microscope tip with an imaging area of microspheres and stitching the areas recorded during scanning, enabling sub-diffraction-limited resolution. Our method marks a possible path to non-invasive cell imaging and simultaneous tracking of specific molecules with nanoscale resolution, facilitating the study of subcellular events over a total cell period.

  3. Cryogenic Propulsion Project

    Data.gov (United States)

    National Aeronautics and Space Administration — The storage of cryogenic propellants is challenging because heat leaks into the cryogenic storage tanks no matter how good the insulation, resulting in a necessity...

  4. Cryogenics a textbook

    CERN Document Server

    Thipse, S S

    2013-01-01

    A Textbook covers lucidly various cryogenic applications including cryogenic engines and space and electronic applications. Importance of cryogenic engines in space propulsion, complete thermodynamic analysis of cryogenic systems with special emphasis on cryogenic cycles, Dewar vessels used to store cryogenic fluids and their applications in various industries have also been discussed in detail. Explanation of Superconductivity and its applications with a description of various Cryocoolers used in industry has also been provided with extensive details. Further technical information on cryogens has been specified alongwith the vacuum technology which has been sufficiently described with examples. Science of Cryonics has been elaborated and all aspects of technology related to functioning of cryogenic plants and their construction including valves, pipes has been incorporated in this book.

  5. Energy Efficient Cryogenics

    Science.gov (United States)

    Meneghelli, Barry J.; Notardonato, William; Fesmire, James E.

    2016-01-01

    The Cryogenics Test Laboratory, NASA Kennedy Space Center, works to provide practical solutions to low-temperature problems while focusing on long-term technology targets for the energy-efficient use of cryogenics on Earth and in space.

  6. Integrated single- and two-photon light sheet microscopy using accelerating beams

    DEFF Research Database (Denmark)

    Piksarv, Peeter; Marti, Dominik; Le, Tuan

    2017-01-01

    We demonstrate the first light sheet microscope using propagation invariant, accelerating Airy beams that operates both in single- and two-photon modes. The use of the Airy beam permits us to develop an ultra compact, high resolution light sheet system without beam scanning. In two-photon mode, a...

  7. Comparison of Partec Rapid Malaria Test with Conventional Light Microscopy for Diagnosis of Malaria in Northwest Ethiopia

    Directory of Open Access Journals (Sweden)

    Meseret Birhanie

    2016-01-01

    Full Text Available Background. Laboratory diagnosis of malaria is the key for effective disease management. Diagnosis of malaria infection requires rapid, sensitive, and specific test methods with an affordable cost. This study was aimed to assess the diagnostic performance of Partec rapid malaria test with reference to light microscopy for the diagnosis of malaria in Northwest Ethiopia. Methods. A total of 180 febrile patients were tested for malaria using Giemsa stain microscopy and Partec rapid malaria test from June to July 2013 at Gendewuha health centers, Metema district. Data were analyzed using SPSS version 20 statistical software. Odds ratio with 95% CI was calculated. Result. The sensitivity and specificity of Partec rapid malaria test were 93.8% (95% CI = 87.1%–100% and 87.9% (95% CI = 79.7%–96.1%, respectively, while the positive predictive value and negative predictive value were 6.4% (95% CI = 77.2%–95.5% and 94.6% (95% CI = 88.7%–100%, respectively. There was also an excellent agreement between two tests with Kappa value of 0.811 (95% CI = 0.625–0.996. Conclusion. Partec rapid malaria test showed good sensitivity and specificity with an excellent agreement to the reference light microscopy. Therefore PT can be considered as alternative diagnostic tools in malaria endemic areas.

  8. 3D Visualization of Developmental Toxicity of 2,4,6-Trinitrotoluene in Zebrafish Embryogenesis Using Light-Sheet Microscopy

    Directory of Open Access Journals (Sweden)

    Juneyong Eum

    2016-11-01

    Full Text Available Environmental contamination by trinitrotoluene is of global concern due to its widespread use in military ordnance and commercial explosives. Despite known long-term persistence in groundwater and soil, the toxicological profile of trinitrotoluene and other explosive wastes have not been systematically measured using in vivo biological assays. Zebrafish embryos are ideal model vertebrates for high-throughput toxicity screening and live in vivo imaging due to their small size and transparency during embryogenesis. Here, we used Single Plane Illumination Microscopy (SPIM/light sheet microscopy to assess the developmental toxicity of explosive-contaminated water in zebrafish embryos and report 2,4,6-trinitrotoluene-associated developmental abnormalities, including defects in heart formation and circulation, in 3D. Levels of apoptotic cell death were higher in the actively developing tissues of trinitrotoluene-treated embryos than controls. Live 3D imaging of heart tube development at cellular resolution by light-sheet microscopy revealed trinitrotoluene-associated cardiac toxicity, including hypoplastic heart chamber formation and cardiac looping defects, while the real time PCR (polymerase chain reaction quantitatively measured the molecular changes in the heart and blood development supporting the developmental defects at the molecular level. Identification of cellular toxicity in zebrafish using the state-of-the-art 3D imaging system could form the basis of a sensitive biosensor for environmental contaminants and be further valued by combining it with molecular analysis.

  9. Cryogenic immersion microscope

    Science.gov (United States)

    Le Gros, Mark; Larabell, Carolyn A.

    2010-12-14

    A cryogenic immersion microscope whose objective lens is at least partially in contact with a liquid reservoir of a cryogenic liquid, in which reservoir a sample of interest is immersed is disclosed. When the cryogenic liquid has an index of refraction that reduces refraction at interfaces between the lens and the sample, overall resolution and image quality are improved. A combination of an immersion microscope and x-ray microscope, suitable for imaging at cryogenic temperatures is also disclosed.

  10. Shedding new light on lipid functions with CARS and SRS microscopy.

    Science.gov (United States)

    Yu, Yong; Ramachandran, Prasanna V; Wang, Meng C

    2014-08-01

    Modern optical microscopy has granted biomedical scientists unprecedented access to the inner workings of a cell, and revolutionized our understanding of the molecular mechanisms underlying physiological and disease states. In spite of these advances, however, visualization of certain classes of molecules (e.g. lipids) at the sub-cellular level has remained elusive. Recently developed chemical imaging modalities - Coherent Anti-Stokes Raman Scattering (CARS) microscopy and Stimulated Raman Scattering (SRS) microscopy - have helped bridge this gap. By selectively imaging the vibration of a specific chemical group, these non-invasive techniques allow high-resolution imaging of individual molecules in vivo, and circumvent the need for potentially perturbative extrinsic labels. These tools have already been applied to the study of fat metabolism, helping uncover novel regulators of lipid storage. Here we review the underlying principle of CARS and SRS microscopy, and discuss the advantages and caveats of each technique. We also review recent applications of these tools in the study of lipids as well as other biomolecules, and conclude with a brief guide for interested researchers to build and use CARS/SRS systems for their own research. This article is part of a Special Issue entitled Tools to study lipid functions.

  11. Lasers, lenses and light curves : adaptive optics microscopy and peculiar transiting exoplanets

    NARCIS (Netherlands)

    Werkhoven, Theodorus Isaak Mattheus van

    2014-01-01

    In the first part of this thesis, we present an adaptive optics implementation for multi-photon microscopy correcting sample-induced wavefront aberrations using either direct wavefront sensing to run a close-loop adaptive optics system (Chapter 3), or use a model-based sensorless approach to iterati

  12. Differential localization of SAP102 and PSD-95 is revealed in hippocampal spines using super-resolution light microscopy.

    Science.gov (United States)

    Zheng, Chan-Ying; Wang, Ya-Xia; Kachar, Bechara; Petralia, Ronald S

    2011-01-01

    Synapse-associated protein 102 (SAP102) and postsynaptic density 95 (PSD-95) are two major cytoskeleton proteins in the postsynaptic density (PSD). Both of them belong to the membrane-associated guanylate kinase (MAGUK) family, which clusters and anchors glutamate receptors and other proteins at synapses. In our previous study, we found that SAP102 and PSD-95 have different distributions, using combined light/electron microscopy (LM/EM) methods.1 Here, we double labeled endogenous SAP102 and PSD-95 in mature hippocampal neurons, and then took images by two different kinds of super resolution microscopy-Stimulated Emission Depletion microscopy (STED) and DeltaVision OMX 3D super resolution microscopy. We found that our 2D and 3D super resolution data were consistent with our previous LM/EM data, showing significant differences in the localization of SAP102 and PSD-95 in spines: SAP102 is distributed in both the PSD and cytoplasm of spines, while PSD-95 is concentrated only in the PSD area. These results indicate functional differences between SAP102 and PSD-95 in synaptic organization and plasticity.

  13. Improving signal-to-noise ratio of structured light microscopy based on photon reassignment.

    Science.gov (United States)

    Singh, Vijay Raj; Choi, Heejin; Yew, Elijah Y S; Bhattacharya, Dipanjan; Yuan, Luo; Sheppard, Colin J R; Rajapakse, Jagath C; Barbastathis, George; So, Peter T C

    2012-01-01

    In this paper, we report a method for 3D visualization of a biological specimen utilizing a structured light wide-field microscopic imaging system. This method improves on existing structured light imaging modalities by reassigning fluorescence photons generated from off-focal plane excitation, improving in-focus signal strength. Utilizing a maximum likelihood approach, we identify the most likely fluorophore distribution in 3D that will produce the observed image stacks under structured and uniform illumination using an iterative maximization algorithm. Our results show the optical sectioning capability of tissue specimens while mostly preserving image stack photon count, which is usually not achievable with other existing structured light imaging methods.

  14. Correlative and integrated light and electron microscopy of in-resin GFP fluorescence, used to localise diacylglycerol in mammalian cells

    Energy Technology Data Exchange (ETDEWEB)

    Peddie, Christopher J.; Blight, Ken; Wilson, Emma [Electron Microscopy Unit, London Research Institute, Cancer Research UK, London WC2A 3LY (United Kingdom); Melia, Charlotte [Electron Microscopy Unit, London Research Institute, Cancer Research UK, London WC2A 3LY (United Kingdom); Cell Biophysics Laboratory, London Research Institute, Cancer Research UK, London WC2A 3LY (United Kingdom); Department of Molecular Cell Biology, Leiden University Medical Centre, 2300 RC Leiden (Netherlands); Marrison, Jo [Department of Biology, The University of York, Heslington, York (United Kingdom); Carzaniga, Raffaella [Electron Microscopy Unit, London Research Institute, Cancer Research UK, London WC2A 3LY (United Kingdom); Domart, Marie-Charlotte [Electron Microscopy Unit, London Research Institute, Cancer Research UK, London WC2A 3LY (United Kingdom); Cell Biophysics Laboratory, London Research Institute, Cancer Research UK, London WC2A 3LY (United Kingdom); O' Toole, Peter [Department of Biology, The University of York, Heslington, York (United Kingdom); Larijani, Banafshe [Cell Biophysics Laboratory, London Research Institute, Cancer Research UK, London WC2A 3LY (United Kingdom); Cell Biophysics Laboratory, Unidad de Biofísica (CSIC-UPV/EHU),Sarriena s/n, 48940 Leioa (Spain); IKERBASQUE, Basque Foundation for Science, Bilbao (Spain); Collinson, Lucy M. [Electron Microscopy Unit, London Research Institute, Cancer Research UK, London WC2A 3LY (United Kingdom)

    2014-08-01

    Fluorescence microscopy of GFP-tagged proteins is a fundamental tool in cell biology, but without seeing the structure of the surrounding cellular space, functional information can be lost. Here we present a protocol that preserves GFP and mCherry fluorescence in mammalian cells embedded in resin with electron contrast to reveal cellular ultrastructure. Ultrathin in-resin fluorescence (IRF) sections were imaged simultaneously for fluorescence and electron signals in an integrated light and scanning electron microscope. We show, for the first time, that GFP is stable and active in resin sections in vacuo. We applied our protocol to study the subcellular localisation of diacylglycerol (DAG), a modulator of membrane morphology and membrane dynamics in nuclear envelope assembly. We show that DAG is localised to the nuclear envelope, nucleoplasmic reticulum and curved tips of the Golgi apparatus. With these developments, we demonstrate that integrated imaging is maturing into a powerful tool for accurate molecular localisation to structure. - Highlights: • GFP and mCherry fluorescence are preserved in heavy-metal stained mammalian cells embedded in resin • Fluorophores are stable and intensity is sufficient for detection in ultrathin sections • Overlay of separate LM and EM images from the same ultrathin section improves CLEM protein localisation precision • GFP is stable and active in the vacuum of an integrated light and scanning EM • Integrated light and electron microscopy shows new subcellular locations of the lipid diacylglycerol.

  15. Quantitative analysis with advanced compensated polarized light microscopy on wavelength dependence of linear birefringence of single crystals causing arthritis

    Science.gov (United States)

    Takanabe, Akifumi; Tanaka, Masahito; Taniguchi, Atsuo; Yamanaka, Hisashi; Asahi, Toru

    2014-07-01

    To improve our ability to identify single crystals causing arthritis, we have developed a practical measurement system of polarized light microscopy called advanced compensated polarized light microscopy (A-CPLM). The A-CPLM system is constructed by employing a conventional phase retardation plate, an optical fibre and a charge-coupled device spectrometer in a polarized light microscope. We applied the A-CPLM system to measure linear birefringence (LB) in the visible region, which is an optical anisotropic property, for tiny single crystals causing arthritis, i.e. monosodium urate monohydrate (MSUM) and calcium pyrophosphate dihydrate (CPPD). The A-CPLM system performance was evaluated by comparing the obtained experimental data using the A-CPLM system with (i) literature data for a standard sample, MgF2, and (ii) experimental data obtained using an established optical method, high-accuracy universal polarimeter, for the MSUM. The A-CPLM system was found to be applicable for measuring the LB spectra of the single crystals of MSUM and CPPD, which cause arthritis, in the visible regions. We quantitatively reveal the large difference in LB between MSUM and CPPD crystals. These results demonstrate the usefulness of the A-CPLM system for distinguishing the crystals causing arthritis.

  16. Use of colloidal quantum dots as a digitally switched swept light source for gold nanoparticle based hyperspectral microscopy

    Science.gov (United States)

    Hoshino, Kazunori; Joshi, Pratixa. P.; Bhave, Gauri.; Sokolov, Konstantin V.; Zhang, Xiaojing

    2014-01-01

    We propose a method to utilize colloidal quantum dots (QDs) as a swept light source for hyperspectral microscopy. The use of QD allows for uniform multicolor emission which covers visible-NIR wavelengths. We used 8 colors of CdSe/ZnS and CdTe/ZnS colloidal quantum dots with the peak emission wavelengths from 520 nm to 800 nm. The QDs are packed in a compact enclosure, composing a low-cost, solid-state swept light source that can be easily used in most microscopes. Multicolor emission from the QDs is simply controlled by digitally switching excitation UVLEDs, eliminating the use of mechanically-driven gratings or filters. We used gold nanoparticles as optical markers for hyperspectral microscopy. Due to the effect of localized surface plasmon resonance, gold nanoparticles demonstrate size and shape-dependent absorption spectra. Employed in a standard microscope, the QD light source enabled multispectral absorption imaging of macrophage cells labeled with gold nanorods and nanospheres. PMID:24877018

  17. Optical coherence photoacoustic microscopy (OC-PAM) with an intensity-modulated continuous-wave broadband light source

    Science.gov (United States)

    Liu, Xiaojing; Wen, Rong; Li, Yiwen; Jiao, Shuliang

    2016-06-01

    We developed an optical coherence photoacoustic microscopy system using an intensity-modulated continuous-wave superluminescent diode with a center wavelength of 840 nm. The system can accomplish optical coherence tomography (OCT) and photoacoustic microscopy (PAM) simultaneously. Compared to the system with a pulsed light source, this system is able to achieve OCT imaging with quality as high as conventional spectral-domain OCT. Since both of the OCT and PAM images are generated from the same group of photons, they are intrinsically registered in the lateral directions. The system was tested for multimodal imaging the vasculature of mouse ear in vivo by using gold nanorods as contrast agent for PAM, as well as excised porcine eyes ex vivo. The OCT and PAM images showed complimentary information of the sample.

  18. Maxillary sinus augmentation with Bio-Oss particles: a light, scanning, and transmission electron microscopy study in man.

    Science.gov (United States)

    Orsini, Giovanna; Traini, Tonino; Scarano, Antonio; Degidi, Marco; Perrotti, Vittoria; Piccirilli, Marcello; Piattelli, Adriano

    2005-07-01

    Biological interactions occurring at the bone-biomaterial interface are critical for long-term clinical success. Bio-Oss is a deproteinized, sterilized bovine bone that has been extensively used in bone regeneration procedures. The aim of the present study was a comparative light, scanning, and electron microscopy evaluation of the interface between Bio-Oss and bone in specimens retrieved after sinus augmentation procedures. Under light microscopy, most of the particles were surrounded by newly formed bone, while in a few cases, at the interface of some particles it was possible to observe marrow spaces and biological fluids. Under scanning electron microscopy, in most cases, the particle perimeter appeared lined by bone that was tightly adherent to the biomaterial surface. Transmission electron microscopy showed that the bone tissue around the biomaterial showed all the phases of the bone healing process. In some areas, randomly organized collagen fibers were present, while in other areas, newly formed compact bone was present. In the first bone lamella collagen fibers contacting the Bio-Oss surface were oriented at 243.73 +/- 7.12 degrees (mean +/- SD), while in the rest of the lamella they were oriented at 288.05 +/- 4.86 degrees (mean +/- SD) with a statistically significant difference of 44.32 degrees (p electron-dense layer similar to cement lines. This layer had a variable morphology being, in some areas, a thin line, and in other areas, a thick irregular band. The analyses showed that Bio-Oss particles do not interfere with the normal osseous healing process after sinus lift procedures and promote new bone formation. In conclusion, this study serves as a better understanding of the morphologic characteristics of Bio-Oss and its interaction with the surrounding tissues.

  19. A genetically encoded tag for correlated light and electron microscopy of intact cells, tissues, and organisms.

    Directory of Open Access Journals (Sweden)

    Xiaokun Shu

    2011-04-01

    Full Text Available Electron microscopy (EM achieves the highest spatial resolution in protein localization, but specific protein EM labeling has lacked generally applicable genetically encoded tags for in situ visualization in cells and tissues. Here we introduce "miniSOG" (for mini Singlet Oxygen Generator, a fluorescent flavoprotein engineered from Arabidopsis phototropin 2. MiniSOG contains 106 amino acids, less than half the size of Green Fluorescent Protein. Illumination of miniSOG generates sufficient singlet oxygen to locally catalyze the polymerization of diaminobenzidine into an osmiophilic reaction product resolvable by EM. MiniSOG fusions to many well-characterized proteins localize correctly in mammalian cells, intact nematodes, and rodents, enabling correlated fluorescence and EM from large volumes of tissue after strong aldehyde fixation, without the need for exogenous ligands, probes, or destructive permeabilizing detergents. MiniSOG permits high quality ultrastructural preservation and 3-dimensional protein localization via electron tomography or serial section block face scanning electron microscopy. EM shows that miniSOG-tagged SynCAM1 is presynaptic in cultured cortical neurons, whereas miniSOG-tagged SynCAM2 is postsynaptic in culture and in intact mice. Thus SynCAM1 and SynCAM2 could be heterophilic partners. MiniSOG may do for EM what Green Fluorescent Protein did for fluorescence microscopy.

  20. Combined Use of Electron and Light Microscopy Techniques Reveals False Secondary Shell Units in Megaloolithidae Eggshells.

    Science.gov (United States)

    Moreno-Azanza, Miguel; Bauluz, Blanca; Canudo, José Ignacio; Gasca, José Manuel; Torcida Fernández-Baldor, Fidel

    2016-01-01

    Abnormalities in the histo- and ultrastructure of the amniote eggshell are often related to diverse factors, such as ambient stress during egg formation, pathologies altering the physiology of the egg-laying females, or evolutionarily selected modifications of the eggshell structure that vary the physical properties of the egg, for example increasing its strength so as to avoid fracture during incubation. When dealing with fossil materials, all the above hypotheses are plausible, but a detailed taphonomical study has to be performed to rule out the possibility that secondary processes of recrystallization have occurred during fossilization. Traditional analyses, such as optical microscopy inspection and cathodoluminescence, have proven not to be enough to understand the taphonomic story of some eggshells. Recently, electron backscatter diffraction has been used, in combination with other techniques, to better understand the alteration of fossil eggshells. Here we present a combined study using scanning electron microscopy, optical microscopy, cathodoluminescence and electron backscatter diffraction of eggshell fragments assigned to Megaloolithus cf. siruguei from the Upper Cretaceous outcrops of the Cameros Basin. We focus our study on the presence of secondary shell units that mimic most aspects of the ultrastructure of the eggshell mammillae, but grow far from the inner surface of the eggshell. We call these structures extra-spherulites, describe their crystal structure and demonstrate their secondary origin. Our study has important implications for the interpretation of secondary shell units as biological or pathological structures. Thus, electron backscatter diffraction complements other microscope techniques as a useful tool for understanding taphonomical alterations in fossil eggshells.

  1. Cryogenic heat transfer

    CERN Document Server

    Barron, Randall F

    2016-01-01

    Cryogenic Heat Transfer, Second Edition continues to address specific heat transfer problems that occur in the cryogenic temperature range where there are distinct differences from conventional heat transfer problems. This updated version examines the use of computer-aided design in cryogenic engineering and emphasizes commonly used computer programs to address modern cryogenic heat transfer problems. It introduces additional topics in cryogenic heat transfer that include latent heat expressions; lumped-capacity transient heat transfer; thermal stresses; Laplace transform solutions; oscillating flow heat transfer, and computer-aided heat exchanger design. It also includes new examples and homework problems throughout the book, and provides ample references for further study.

  2. Cryogen Safety Course 8876

    Energy Technology Data Exchange (ETDEWEB)

    Glass, George [Los Alamos National Lab. (LANL), Los Alamos, NM (United States)

    2017-06-13

    Cryogenics (from the Greek word κρvoζ, meaning frost or icy cold) is the study of the behavior of matter at very cold temperatures. The purpose of this course is to provide trainees with an introduction to cryogen use, the hazards and potential accidents related to cryogen systems, cryogen safety components, and the requirements that govern the design and use of cryogen systems at Los Alamos National Laboratory (LANL). The knowledge you gain will help you keep your workplace safe for yourself and your coworkers.

  3. Teager-Kaiser Energy and Higher-Order Operators in White-Light Interference Microscopy for Surface Shape Measurement

    Directory of Open Access Journals (Sweden)

    Abdel-Ouahab Boudraa

    2005-10-01

    Full Text Available In white-light interference microscopy, measurement of surface shape generally requires peak extraction of the fringe function envelope. In this paper the Teager-Kaiser energy and higher-order energy operators are proposed for efficient extraction of the fringe envelope. These energy operators are compared in terms of precision, robustness to noise, and subsampling. Flexible energy operators, depending on order and lag parameters, can be obtained. Results show that smoothing and interpolation of envelope approximation using spline model performs better than Gaussian-based approach.

  4. Discrepancies in quantitative assessment of normal and regenerated peripheral nerve fibers between light and electron microscopy.

    Science.gov (United States)

    Ronchi, Giulia; Jager, Sara Buskbjerg; Vaegter, Christian Bjerggaard; Raimondo, Stefania; Giacobini-Robecchi, Maria Giuseppina; Geuna, Stefano

    2014-09-01

    Quantitative estimation of myelinated nerve fiber number, together with fiber size parameters, is one of the most important tools for nerve regeneration research. In this study we used a design-based stereological method to evaluate the regenerative process in two experimental paradigms: crush injury and autograft repair. Samples were embedded in resin and morphometric counting and measurements were performed using both light and electron microscopes. Results show a significant difference in myelinated fiber number estimation between light and electron microscopes, especially after autograft repair; light microscope significantly underestimates the number of fibers because of the large number of very small axons that can be detected only in electron microscope. The analysis of the size parameters also shows a higher number of small fibers in electron microscopic analysis, especially in regenerated nerves. This comparative study shows that the integration of data obtained in light microscope with those obtained in electron microscope is necessary in revealing very small myelinated fibers that cannot be detected otherwise. Moreover, the difference in the estimation of total number of myelinated fibers between light and electron microscopes must be considered in data analysis to ensure accurate interpretation of the results. © 2014 Peripheral Nerve Society.

  5. Three-dimensional reconstruction of the guinea pig inner ear, comparison of OPFOS and light microscopy, applications of 3D reconstruction

    NARCIS (Netherlands)

    Hofman, R.; Segenhout, J. M.; Wit, H. P.

    2009-01-01

    Three-dimensional (3D) reconstruction of anatomical structures can give additional insight into the morphology and function of these structures. We compare 3D reconstructions of the guinea pig inner ear, using light microscopy and orthogonal plane fluorescence optical sectioning microscopy. Applicat

  6. Lossless Three-Dimensional Parallelization in Digitally Scanned Light-Sheet Fluorescence Microscopy.

    Science.gov (United States)

    Dean, Kevin M; Fiolka, Reto

    2017-08-24

    We introduce a concept that enables parallelized three-dimensional imaging throughout large volumes with isotropic 300-350 nm resolution. By staggering high aspect ratio illumination beams laterally and axially within the depth of focus of a digitally scanned light-sheet fluorescence microscope (LSFM), multiple image planes can be simultaneously imaged with minimal cross-talk and light loss. We present a first demonstration of this concept for parallelized imaging by synthesizing two light-sheets with nonlinear Bessel beams and perform volumetric imaging of fluorescent beads and invasive breast cancer cells. This work demonstrates that in principle any digitally scanned LSFM can be parallelized in a lossless manner, enabling drastically faster volumetric image acquisition rates for a given sample brightness and detector technology.

  7. Programmable aperture microscopy: A computational method for multi-modal phase contrast and light field imaging

    Science.gov (United States)

    Zuo, Chao; Sun, Jiasong; Feng, Shijie; Zhang, Minliang; Chen, Qian

    2016-05-01

    We demonstrate a simple and cost-effective programmable aperture microscope to realize multi-modal computational imaging by integrating a programmable liquid crystal display (LCD) into a conventional wide-field microscope. The LCD selectively modulates the light distribution at the rear aperture of the microscope objective, allowing numerous imaging modalities, such as bright field, dark field, differential phase contrast, quantitative phase imaging, multi-perspective imaging, and full resolution light field imaging to be achieved and switched rapidly in the same setup, without requiring specialized hardwares and any moving parts. We experimentally demonstrate the success of our method by imaging unstained cheek cells, profiling microlens array, and changing perspective views of thick biological specimens. The post-exposure refocusing of a butterfly mouthpart and RFP-labeled dicot stem cross-section is also presented to demonstrate the full resolution light field imaging capability of our system for both translucent and fluorescent specimens.

  8. Light sheet-based fluorescence microscopy (LSFM) reduces phototoxic effects and provides new means for the modern life sciences

    Science.gov (United States)

    Pampaloni, Francesco; Ansari, Nari; Girard, Philippe; Stelzer, Ernst H. K.

    2011-07-01

    Most optical technologies are applied to flat, basically two-dimensional cellular systems. However, physiological meaningful information relies on the morphology, the mechanical properties and the biochemistry of a cell's context. A cell requires the complex three-dimensional relationship to other cells. However, the observation of multi-cellular biological specimens remains a challenge. Specimens scatter and absorb light, thus, the delivery of the probing light and the collection of the signal light become inefficient; many endogenous biochemical compounds also absorb light and suffer degradation of some sort (photo-toxicity), which induces malfunction of a specimen. In conventional and confocal fluorescence microscopy, whenever a single plane, the entire specimen is illuminated. Recording stacks of images along the optical Z-axis thus illuminates the entire specimen once for each plane. Hence, cells are illuminated 10-20 and fish 100-300 times more often than they are observed. This can be avoided by changing the optical arrangement. The basic idea is to use light sheets, which are fed into the specimen from the side and overlap with the focal plane of a wide-field fluorescence microscope. In contrast to an epi-fluorescence arrangement, such an azimuthal fluorescence arrangement uses two independently operated lenses for illumination and detection. Optical sectioning and no photo-toxic damage or photo-bleaching outside a small volume close to the focal plane are intrinsic properties. Light sheet-based fluorescence microscopy (LSFM) takes advantage of modern camera technologies. LSFM can be operated with laser cutters and for fluorescence correlation spectroscopy. During the last few years, LSFM was used to record zebrafish development from the early 32-cell stage until late neurulation with sub-cellular resolution and short sampling periods (60-90 sec/stack). The recording speed was five 4-Megapixel large frames/sec with a dynamic range of 12-14 bit. We followed

  9. Combined Use of Electron and Light Microscopy Techniques Reveals False Secondary Shell Units in Megaloolithidae Eggshells.

    Directory of Open Access Journals (Sweden)

    Miguel Moreno-Azanza

    Full Text Available Abnormalities in the histo- and ultrastructure of the amniote eggshell are often related to diverse factors, such as ambient stress during egg formation, pathologies altering the physiology of the egg-laying females, or evolutionarily selected modifications of the eggshell structure that vary the physical properties of the egg, for example increasing its strength so as to avoid fracture during incubation. When dealing with fossil materials, all the above hypotheses are plausible, but a detailed taphonomical study has to be performed to rule out the possibility that secondary processes of recrystallization have occurred during fossilization. Traditional analyses, such as optical microscopy inspection and cathodoluminescence, have proven not to be enough to understand the taphonomic story of some eggshells. Recently, electron backscatter diffraction has been used, in combination with other techniques, to better understand the alteration of fossil eggshells. Here we present a combined study using scanning electron microscopy, optical microscopy, cathodoluminescence and electron backscatter diffraction of eggshell fragments assigned to Megaloolithus cf. siruguei from the Upper Cretaceous outcrops of the Cameros Basin. We focus our study on the presence of secondary shell units that mimic most aspects of the ultrastructure of the eggshell mammillae, but grow far from the inner surface of the eggshell. We call these structures extra-spherulites, describe their crystal structure and demonstrate their secondary origin. Our study has important implications for the interpretation of secondary shell units as biological or pathological structures. Thus, electron backscatter diffraction complements other microscope techniques as a useful tool for understanding taphonomical alterations in fossil eggshells.

  10. Optical parametric oscillator-based light source for coherent Raman scattering microscopy: practical overview

    Science.gov (United States)

    Brustlein, Sophie; Ferrand, Patrick; Walther, Nico; Brasselet, Sophie; Billaudeau, Cyrille; Marguet, Didier; Rigneault, Hervé

    2011-02-01

    We present the assets and constraints of using optical parametric oscillators (OPOs) to perform point scanning nonlinear microscopy and spectroscopy with special emphasis on coherent Raman spectroscopy. The difterent possible configurations starting with one OPO and two OPOs are described in detail and with comments that are intended to be practically useful for the user. Explicit examples on test samples such as nonlinear organic crystal, polystyrene beads, and fresh mouse tissues are given. Special emphasis is given to background-free coherent Raman anti-Stokes scattering (CARS) imaging, including CARS hyperspectral imaging in a fully automated mode with commercial OPOs.

  11. Self-reconstructing sectioned Bessel beams offer submicron optical sectioning for large fields of view in light-sheet microscopy.

    Science.gov (United States)

    Fahrbach, Florian O; Gurchenkov, Vasily; Alessandri, Kevin; Nassoy, Pierre; Rohrbach, Alexander

    2013-05-06

    One of main challenges in light-sheet microscopy is to design the light-sheet as extended and thin as possible--extended to cover a large field of view, thin to optimize resolution and contrast. However, a decrease of the beam's waist also decreases the illumination beam's depth of field. Here, we introduce a new kind of beam that we call sectioned Bessel beam. These beams can be generated by blocking opposite sections of the beam's angular spectrum. In combination with confocal-line detection the optical sectioning performance of the light-sheet can be decoupled from the depth of field of the illumination beam. By simulations and experiments we demonstrate that these beams exhibit self-reconstruction capabilities and penetration depths into thick scattering media equal to those of conventional Bessel beams. We applied sectioned Bessel beams to illuminate tumor multicellular spheroids and prove the increase in contrast. Sectioned Bessel beams turn out to be highly advantageous for the investigation of large strongly scattering samples in a light-sheet microscope.

  12. Co-Orientation: Quantifying Simultaneous Co-Localization and Orientational Alignment of Filaments in Light Microscopy.

    Directory of Open Access Journals (Sweden)

    Robert P J Nieuwenhuizen

    Full Text Available Co-localization analysis is a widely used tool to seek evidence for functional interactions between molecules in different color channels in microscopic images. Here we extend the basic co-localization analysis by including the orientations of the structures on which the molecules reside. We refer to the combination of co-localization of molecules and orientational alignment of the structures on which they reside as co-orientation. Because the orientation varies with the length scale at which it is evaluated, we consider this scale as a separate informative dimension in the analysis. Additionally we introduce a data driven method for testing the statistical significance of the co-orientation and provide a method for visualizing the local co-orientation strength in images. We demonstrate our methods on simulated localization microscopy data of filamentous structures, as well as experimental images of similar structures acquired with localization microscopy in different color channels. We also show that in cultured primary HUVEC endothelial cells, filaments of the intermediate filament vimentin run close to and parallel with microtubuli. In contrast, no co-orientation was found between keratin and actin filaments. Co-orientation between vimentin and tubulin was also observed in an endothelial cell line, albeit to a lesser extent, but not in 3T3 fibroblasts. These data therefore suggest that microtubuli functionally interact with the vimentin network in a cell-type specific manner.

  13. Co-Orientation: Quantifying Simultaneous Co-Localization and Orientational Alignment of Filaments in Light Microscopy.

    Science.gov (United States)

    Nieuwenhuizen, Robert P J; Nahidiazar, Leila; Manders, Erik M M; Jalink, Kees; Stallinga, Sjoerd; Rieger, Bernd

    2015-01-01

    Co-localization analysis is a widely used tool to seek evidence for functional interactions between molecules in different color channels in microscopic images. Here we extend the basic co-localization analysis by including the orientations of the structures on which the molecules reside. We refer to the combination of co-localization of molecules and orientational alignment of the structures on which they reside as co-orientation. Because the orientation varies with the length scale at which it is evaluated, we consider this scale as a separate informative dimension in the analysis. Additionally we introduce a data driven method for testing the statistical significance of the co-orientation and provide a method for visualizing the local co-orientation strength in images. We demonstrate our methods on simulated localization microscopy data of filamentous structures, as well as experimental images of similar structures acquired with localization microscopy in different color channels. We also show that in cultured primary HUVEC endothelial cells, filaments of the intermediate filament vimentin run close to and parallel with microtubuli. In contrast, no co-orientation was found between keratin and actin filaments. Co-orientation between vimentin and tubulin was also observed in an endothelial cell line, albeit to a lesser extent, but not in 3T3 fibroblasts. These data therefore suggest that microtubuli functionally interact with the vimentin network in a cell-type specific manner.

  14. Noninvasive assessment of articular cartilage surface damage using reflected polarized light microscopy.

    Science.gov (United States)

    Huynh, Ruby N; Nehmetallah, George; Raub, Christopher B

    2017-06-01

    Articular surface damage occurs to cartilage during normal aging, osteoarthritis, and in trauma. A noninvasive assessment of cartilage microstructural alterations is useful for studies involving cartilage explants. This study evaluates polarized reflectance microscopy as a tool to assess surface damage to cartilage explants caused by mechanical scraping and enzymatic degradation. Adult bovine articular cartilage explants were scraped, incubated in collagenase, or underwent scrape and collagenase treatments. In an additional experiment, cartilage explants were subject to scrapes at graduated levels of severity. Polarized reflectance parameters were compared with India ink surface staining, features of histological sections, changes in explant wet weight and thickness, and chondrocyte viability. The polarized reflectance signal was sensitive to surface scrape damage and revealed individual scrape features consistent with India ink marks. Following surface treatments, the reflectance contrast parameter was elevated and correlated with image area fraction of India ink. After extensive scraping, polarized reflectance contrast and chondrocyte viability were lower than that from untreated explants. As part of this work, a mathematical model was developed and confirmed the trend in the reflectance signal due to changes in surface scattering and subsurface birefringence. These results demonstrate the effectiveness of polarized reflectance microscopy to sensitively assess surface microstructural alterations in articular cartilage explants.

  15. New light on ion channel imaging by total internal reflection fluorescence (TIRF microscopy

    Directory of Open Access Journals (Sweden)

    Hisao Yamamura

    2015-05-01

    Full Text Available Ion channels play pivotal roles in a wide variety of cellular functions; therefore, their physiological characteristics, pharmacological responses, and molecular structures have been extensively investigated. However, the mobility of an ion channel itself in the cell membrane has not been examined in as much detail. A total internal reflection fluorescence (TIRF microscope allows fluorophores to be imaged in a restricted region within an evanescent field of less than 200 nm from the interface of the coverslip and plasma membrane in living cells. Thus the TIRF microscope is useful for selectively visualizing the plasmalemmal surface and subplasmalemmal zone. In this review, we focused on a single-molecule analysis of the dynamic movement of ion channels in the plasma membrane using TIRF microscopy. We also described two single-molecule imaging techniques under TIRF microscopy: fluorescence resonance energy transfer (FRET for the identification of molecules that interact with ion channels, and subunit counting for the determination of subunit stoichiometry in a functional channel. TIRF imaging can also be used to analyze spatiotemporal Ca2+ events in the subplasmalemma. Single-molecule analyses of ion channels and localized Ca2+ signals based on TIRF imaging provide beneficial pharmacological and physiological information concerning the functions of ion channels.

  16. A Comparison of Honey Bee-Collected Pollen From Working Agricultural Lands Using Light Microscopy and ITS Metabarcoding.

    Science.gov (United States)

    Smart, M D; Cornman, R S; Iwanowicz, D D; McDermott-Kubeczko, M; Pettis, J S; Spivak, M S; Otto, C R V

    2017-02-01

    Taxonomic identification of pollen has historically been accomplished via light microscopy but requires specialized knowledge and reference collections, particularly when identification to lower taxonomic levels is necessary. Recently, next-generation sequencing technology has been used as a cost-effective alternative for identifying bee-collected pollen; however, this novel approach has not been tested on a spatially or temporally robust number of pollen samples. Here, we compare pollen identification results derived from light microscopy and DNA sequencing techniques with samples collected from honey bee colonies embedded within a gradient of intensive agricultural landscapes in the Northern Great Plains throughout the 2010-2011 growing seasons. We demonstrate that at all taxonomic levels, DNA sequencing was able to discern a greater number of taxa, and was particularly useful for the identification of infrequently detected species. Importantly, substantial phenological overlap did occur for commonly detected taxa using either technique, suggesting that DNA sequencing is an appropriate, and enhancing, substitutive technique for accurately capturing the breadth of bee-collected species of pollen present across agricultural landscapes. We also show that honey bees located in high and low intensity agricultural settings forage on dissimilar plants, though with overlap of the most abundantly collected pollen taxa. We highlight practical applications of utilizing sequencing technology, including addressing ecological issues surrounding land use, climate change, importance of taxa relative to abundance, and evaluating the impact of conservation program habitat enhancement efforts.

  17. Study on validity of a rapid diagnostic test kit versus light microscopy for malaria diagnosis in Ahmedabad city, India.

    Science.gov (United States)

    Vyas, S; Puwar, B; Patel, V; Bhatt, G; Kulkarni, S; Fancy, M

    2014-05-01

    Light microscopy of blood smears for diagnosis of malaria in the field has several limitations, notably delays in diagnosis. This study in Ahmedabad in Gujarat State, India, evaluated the diagnostic performance of a rapid diagnostic test for malaria (SD Bioline Malaria Ag P.f/Pan) versus blood smear examination as the gold standard. All fever cases presenting at 13 urban health centres were subjected to rapid diagnostic testing and thick and thin blood smears. A total of 677 cases with fever were examined; 135 (20.0%) tested positive by rapid diagnostic test and 86 (12.7%) by blood smear. The sensitivity of the rapid diagnostic test for malaria was 98.8%, specificity was 91.5%, positive predictive value 63.0% and negative predictive value 99.8%. For detection of Plasmodium falciparum the sensitivity of rapid diagnostic test was 100% and specificity was 97.3%. The results show the acceptability of the rapid test as an alternative to light microscopy in the field setting.

  18. Hyperspectral reflected light microscopy of plasmonic Au/Ag alloy nanoparticles incubated as multiplex chromatic biomarkers with cancer cells.

    Science.gov (United States)

    Patskovsky, Sergiy; Bergeron, Eric; Rioux, David; Simard, Mikaël; Meunier, Michel

    2014-10-21

    A hyperspectral microscopy system based on a reflected light method for plasmonic nanoparticle (NP) imaging was designed and compared with a conventional darkfield method for spatial localization and spectroscopic identification of single Au, Ag and Au/Ag alloy NPs incubated with fixed human cancer cell preparations. A new synthesis protocol based on co-reduction of Au and Ag salts combined with the seeded growth technique was used for the fabrication of monodispersed alloy NPs with sizes ranging from 30 to 100 nm in diameter. We validated theoretically and experimentally the performance of 60 nm Au, Ag and Au/Ag (50 : 50) NPs as multiplexed biological chromatic markers for biomedical diagnostics and optical biosensing. The advantages of the proposed reflected light microscopy method are presented for NP imaging in a complex and highly diffusing medium such as a cellular environment. The obtained information is essential for the development of a high throughput, selective and efficient strategy for cancer detection and treatment.

  19. An optical investigation of dentinal discoloration due to commonly endodontic sealers, using the transmitted light polarizing microscopy and spectrophotometry.

    Science.gov (United States)

    Suciu, Ioana; Ionescu, Ecaterina; Dimitriu, Bogdan Alexandru; Bartok, Ruxandra Ioana; Moldoveanu, Georgiana Florentina; Gheorghiu, Irina Maria; Suciu, Ileana; Ciocîrdel, Mihai

    2016-01-01

    The aim of this study was to establish the degree of tooth crown staining by commonly used endodontic sealers. Crown discolorations by tooth canal sealers [AH Plus (Dentsply DeTrey Gmbh, Konstanz, Germany); Endofill (Produits Dentaires SA, Vevey, Switzerland); Apexit (Dentsply DeTrey Gmbh, Konstanz, Germany); and MTA Fillapex (Angelus, Londrina, Brazil)] were tested on extracted human premolars. The samples were divided into five groups of five samples each, after root canal sealing. Five teeth were used as control groups. The spectrophotometric method was performed in order to quantify in terms of color change of the coronal part (it was also recorded a track on how the color changes over time). For the microscopic study of the extracted dental specimens subjected to this study, polarized transmitted light microscopy was used. This method involves the development of special microscopic preparations, called "thin sections". In our case, the thin section was performed on 20 prepared and obturated recently extracted teeth. The degree of discoloration was determined after one week and three months using spectrophotometry and polarized light microscopy. All sealers usually cause some degree of discoloration on the cervical aspect of the crowns that increases in time. AH Plus and Endofill caused the greatest discoloration, followed by Apexit and MTA Fillapex.

  20. A LIGHT AND SCANNING ELECTRON MICROSCOPY STUDY OF PLACENTAL VILLI ASSOCIATED WITH OBESITY AND HYPERTENSIÓN

    Directory of Open Access Journals (Sweden)

    Castejón Sandoval OC

    2013-05-01

    Full Text Available The aim of this study was to examine the structure of the placental villi associated with obesity and hypertension using light and scanning electron microscopy.Two placentas at term obtained of woman pregnancy associated to stillborn were taken for microscopical analysis. Woman pregnancy weighed 75 and 85 Kg as body mass index and their hypertension with more of 90/150 mmHg. They had 39 and 41 years old. The placental weights were 600 and 650 gr respectively after of draining all their blood during 30 minutes post delivery. Five small fragments were taken by each placenta and evaluated with light and scanning electron microscopy. Stem villi showed obstructive vessels with damage in their walls and reticular tissue under the syncytio was a permanent finding. Immature intermediate villi were frequent. Severe degenerative changes are noted in peripheric stem villi. Fibrotic villi, very poor developed mature intermediate villi, filiform terminal villi, corangiosis, deposition of fibrinoid and vasodilatation were found in placental villi. No inflammation and low arborization was seen. These results indicate immaturity persistent, low maturation degree and severe degenerative changes affecting the structure of placental villi without inflammation

  1. A comparison of honey bee-collected pollen from working agricultural lands using light microscopy and ITS metabarcoding

    Science.gov (United States)

    Smart, Matthew; Cornman, Robert S.; Iwanowicz, Deborah; McDermott-Kubeczko, Margaret; Pettis, Jeff S; Spivak, Marla S; Otto, Clint R.

    2017-01-01

    Taxonomic identification of pollen has historically been accomplished via light microscopy but requires specialized knowledge and reference collections, particularly when identification to lower taxonomic levels is necessary. Recently, next-generation sequencing technology has been used as a cost-effective alternative for identifying bee-collected pollen; however, this novel approach has not been tested on a spatially or temporally robust number of pollen samples. Here, we compare pollen identification results derived from light microscopy and DNA sequencing techniques with samples collected from honey bee colonies embedded within a gradient of intensive agricultural landscapes in the Northern Great Plains throughout the 2010–2011 growing seasons. We demonstrate that at all taxonomic levels, DNA sequencing was able to discern a greater number of taxa, and was particularly useful for the identification of infrequently detected species. Importantly, substantial phenological overlap did occur for commonly detected taxa using either technique, suggesting that DNA sequencing is an appropriate, and enhancing, substitutive technique for accurately capturing the breadth of bee-collected species of pollen present across agricultural landscapes. We also show that honey bees located in high and low intensity agricultural settings forage on dissimilar plants, though with overlap of the most abundantly collected pollen taxa. We highlight practical applications of utilizing sequencing technology, including addressing ecological issues surrounding land use, climate change, importance of taxa relative to abundance, and evaluating the impact of conservation program habitat enhancement efforts.

  2. Multispectral endoscopy and microscopy imaging system using a spectrally programmable light engine

    Science.gov (United States)

    MacKinnon, N.; Stange, Ulrich; Lane, Pierre M.; MacAulay, Calum E.

    2005-03-01

    We report a spectrally and temporally programmable light engine based on a spatial light modulator that can dynamically create any narrow or broadband spectral profile for hyperspectral, fluorescence, or principal component imaging. Most hyperspectral or multispectral imaging systems use wavelength selection devices such as acousto-optic tunable filters (AOTFs), tunable grating or prism-based monochromators, or filter wheels. While these devices can select wavelengths they cannot create arbitrary spectral profiles. This simple and economical system can be controlled at high speed (up to 5000 illumination profiles per second). Digitally controlled illumination is bit additive with image data providing high dynamic range imaging with monochrome or color imaging devices. This is especially advantageous for endoscopes employing small well CCD or CMOS sensors since the dynamic range now can extend beyond the limits of the sensor itself. In this report we show multispectral images of in vivo tissue and in vitro tissue samples using endoscopes, surgical microscopes and conventional microscopes.

  3. Physically-based in silico light sheet microscopy for visualizing fluorescent brain models.

    Science.gov (United States)

    Abdellah, Marwan; Bilgili, Ahmet; Eilemann, Stefan; Markram, Henry; Schürmann, Felix

    2015-01-01

    We present a physically-based computational model of the light sheet fluorescence microscope (LSFM). Based on Monte Carlo ray tracing and geometric optics, our method simulates the operational aspects and image formation process of the LSFM. This simulated, in silico LSFM creates synthetic images of digital fluorescent specimens that can resemble those generated by a real LSFM, as opposed to established visualization methods producing visually-plausible images. We also propose an accurate fluorescence rendering model which takes into account the intrinsic characteristics of fluorescent dyes to simulate the light interaction with fluorescent biological specimen. We demonstrate first results of our visualization pipeline to a simplified brain tissue model reconstructed from the somatosensory cortex of a young rat. The modeling aspects of the LSFM units are qualitatively analysed, and the results of the fluorescence model were quantitatively validated against the fluorescence brightness equation and characteristic emission spectra of different fluorescent dyes. Modelling and simulation.

  4. Detrapping and retrapping of free carriers in nominally pure single crystal GaP, GaAs and 4H-SiC semiconductors under light illumination at cryogenic temperatures

    CERN Document Server

    Mouneyrac, David; Floch, Jean-Michel Le; Tobar, Michael E; Cros, Dominique; Krupka, Jerzy

    2010-01-01

    We report on extremely sensitive measurements of changes in the microwave properties of high purity non-intentionally-doped single-crystal semiconductor samples of gallium phosphide, gallium arsenide and 4H-silicon carbide when illuminated with light of different wavelengths at cryogenic temperatures. Whispering gallery modes were excited in the semiconductors whilst they were cooled on the coldfinger of a single-stage cryocooler and their frequencies and Q-factors measured under light and dark conditions. With these materials, the whispering gallery mode technique is able to resolve changes of a few parts per million in the permittivity and the microwave losses as compared with those measured in darkness. A phenomenological model is proposed to explain the observed changes, which result not from direct valence to conduction band transitions but from detrapping and retrapping of carriers from impurity/defect sites with ionization energies that lay in the semiconductor band gap. Detrapping and retrapping relax...

  5. Light scattering microscopy measurements of single nuclei compared with GPU-accelerated FDTD simulations

    Science.gov (United States)

    Stark, Julian; Rothe, Thomas; Kieß, Steffen; Simon, Sven; Kienle, Alwin

    2016-04-01

    Single cell nuclei were investigated using two-dimensional angularly and spectrally resolved scattering microscopy. We show that even for a qualitative comparison of experimental and theoretical data, the standard Mie model of a homogeneous sphere proves to be insufficient. Hence, an accelerated finite-difference time-domain method using a graphics processor unit and domain decomposition was implemented to analyze the experimental scattering patterns. The measured cell nuclei were modeled as single spheres with randomly distributed spherical inclusions of different size and refractive index representing the nucleoli and clumps of chromatin. Taking into account the nuclear heterogeneity of a large number of inclusions yields a qualitative agreement between experimental and theoretical spectra and illustrates the impact of the nuclear micro- and nanostructure on the scattering patterns.

  6. Positronium production in cryogenic environments

    Science.gov (United States)

    Cooper, B. S.; Alonso, A. M.; Deller, A.; Liszkay, L.; Cassidy, D. B.

    2016-03-01

    We report measurements of positronium (Ps) formation following positron irradiation of mesoporous SiO2 films and Ge(100) single crystals at temperatures ranging from 12-700 K. As both of these materials generate Ps atoms via nonthermal processes, they are able to function as positron-positronium converters at cryogenic temperatures. Our data show that such Ps formation is possibly provided the targets are not compromised by adsorption of residual gas. In the case of SiO2 films, we observe a strong reduction in the Ps formation efficiency following irradiation with UV laser light (λ =243.01 nm) below 250 K, in accordance with previous observations of radiation-induced surface paramagnetic centers. Conversely, Ps emission from Ge is enhanced by irradiation with visible laser light (λ =532 nm) via a photoemission process that persists at cryogenic temperatures. Both mesoporous SiO2 films and Ge crystals were found to produce Ps efficiently in cryogenic environments. Accordingly, these materials are likely to prove useful in several areas of research, including Ps mediated antihydrogen formation conducted in the cold bore of a superconducting magnet, the production of Rydberg Ps for experiments in which the effects of black-body radiation must be minimized, and the utilization of mesoporous structures that have been modified to produce cold Ps atoms.

  7. Effects of low-intensity polarized visible laser radiation on skin burns: a light microscopy study.

    Science.gov (United States)

    Ribeiro, Martha Simões; Da Silva, Daniela De Fátima Teixeira; De Araújo, Carlos Eugênio Nabuco; De Oliveira, Sérgio Ferreira; Pelegrini, Cleusa Maria Raspantini; Zorn, Telma Maria Tenório; Zezell, Denise Maria

    2004-02-01

    This study was carried out to investigate the influence of low-intensity polarized visible laser radiation on the acceleration of skin wound healing. Low-level laser therapy (LLLT) at adequate wavelength, intensity, and dose can accelerate tissue repair. However, there is still unclear information about light characteristics, such as coherence and polarization. Some studies indicate that linearly polarized light can survive through long propagation distance in biological tissue. Three burns about 6 mm in diameter were created on the back of rats with liquid N(2). Lesion "L(//)" was irradiated by He-Ne laser (lambda = 632.8 nm), D= 1.0 J/cm(2), with linear polarization parallel to the spinal column of the rat. Lesion "L(inverted v)" was irradiated using the same laser and dose, but the light polarization was aligned perpendicularly to the relative orientation. Lesion "C" was not irradiated in order to be considered as control. The animals were sacrificed at day 3-17 after lesion creation. Samples were collected and prepared for histological analysis. Histological analysis showed that the healing of irradiated wounds was faster than that of non-irradiated wounds. Moreover, it was observed that skin wound repair is dependent on polarization orientation with respect to a referential axis as the animal's spinal column. Consequently, "L(//)" was completely healed after 17 days, whereas "L (perpendicular) " showed a moderate degree of healing after the same period. These results indicate that the relative direction of the laser polarization plays an important role in the wound healing process when highly coherent He-Ne laser is used.

  8. A cryogenic test facility

    Science.gov (United States)

    Veenendaal, Ian

    The next generation, space-borne instruments for far infrared spectroscopy will utilize large diameter, cryogenically cooled telescopes in order to achieve unprecedented sensitivities. Low background, ground-based cryogenic facilities are required for the cryogenic testing of materials, components and subsystems. The Test Facility Cryostat (TFC) at the University of Lethbridge is a large volume, closed cycle, 4K cryogenic facility, developed for this purpose. This thesis discusses the design and performance of the facility and associated external instrumentation. An apparatus for measuring the thermal properties of materials is presented, and measurements of the thermal expansion and conductivity of carbon fibre reinforced polymers (CFRPs) at cryogenic temperatures are reported. Finally, I discuss the progress towards the design and fabrication of a demonstrator cryogenic, far infrared Fourier transform spectrometer.

  9. Micromorphology of epicuticular waxes and epistomatal chambers of pine species by electron microscopy and white light scanning interferometry.

    Science.gov (United States)

    Kim, Ki Woo; Lee, In Jung; Kim, Chang Soo; Lee, Don Koo; Park, Eun Woo

    2011-02-01

    High-resolution imaging and quantitative surface analysis of epicuticular waxes and epistomatal chambers of pine species were performed by field emission scanning electron microscopy and white light scanning interferometry. Both juvenile and adult needles were collected from the two-year-old seedlings of Pinus rigida and Pinus densiflora and subjected to surface observations. Epicuticular wax structures developed on the cuticle layer as well as in the epistomatal chambers and appeared to occlude the cavities in the two pine species. The stomata of P. densiflora were characterized by more distinctly raised rings around openings than P. rigida. The most common epicuticular wax structures of the two pine species included tubules with terminal openings and coiled rodlets. Wax platelets were deposited on epistomatal chambers. Either rodlets or tubules seemed to be longer and thicker in P. rigida than those in P. densiflora. White light scanning interferometry revealed quantitative surface profiles, demonstrating more ridged (ca. 4 μm high) stomatal apertures and nearly twofold deeper (ca. 20 μm deep) epistomatal chambers of P. densiflora than those of P. rigida. These results suggest that white light scanning interferometry can be applied to unravel the quantitative surface features of epicuticular sculptures on plant leaves.

  10. New approach to study starch gelatinization applying a combination of hot-stage light microscopy and differential scanning calorimetry.

    Science.gov (United States)

    Li, Qian; Xie, Qin; Yu, Shujuan; Gao, Qunyu

    2013-02-13

    To overcome the difficulty of the original polarizing microscope-based method in monitoring the gelatinization of starch, a new method for dynamically monitoring the gelatinization process, integral optical density (IOD), which was based on the digital image analysis technique, was proposed. Hot-stage light microscopy and differential scanning calorimetry (DSC) techniques were coupled to study the dynamic changes of three types of starches: type A (corn starch), type B (potato starch), and type C (pea starch), during the gelatinization process in an excess water system. A model of response difference change of crystallite could represent the responding intensity of crystallization changes in the process of starch gelatinization. Results demonstrated that three crystalline types of starch underwent a process of swelling, accompanied with gradual disappearing of the crystallite. This difference was mainly associated with the diversity and composition of the starch structure. The IOD method was of advantage compared to the previous traditional methods that are based on a polarization microscope, such as counting the particle number and calculating polarization area methods, because it was the product of two parameters: optical density and area, which would be a response of both light intensity and area of birefringence light. The single peak in DSC corresponded to the combination of crystalline helix-helix dissociation and the reduction of the molecule helix-coil transition, while the gelatinization degree measured by the IOD method mainly corresponded to the helix-helix dissociation. The gelatinization mechanism could be revealed clearer in this study.

  11. Techniques of advanced light microscopy and their applications to morphological analysis of human extra-embryonic membranes.

    Science.gov (United States)

    Ockleford, C D; Mongan, L C; Hubbard, A R

    The science of light microscopy has advanced dramatically in recent years through the introduction of new technology. A brief description of scanning light microscopes, laser illumination, the confocal principle, digital imaging, and image processing reveals a number of theoretical advantages which are particularly useful in improving epifluorescence microscope images. Examples of results from several studies of human extra-embryonic membranes conducted in our laboratory show how the application of these techniques has been used to describe structures such as microtrabeculae and rivets for the first time, to map the microscopic distribution of a wide range of proteins, and to observe the activity of placental villi at the microscopic level in an environmentally controlled microscope stage. High-sensitivity detectors have permitted the "super-resolution" detection of structures smaller than the theoretically calculated limits of light microscope resolution. Rendering images in false colour is demonstrably useful in detecting subtle variations in fluorescence intensity at different intracellular sites and at different sites within tissues of fetal membranes. Processing stacks of digital images using appropriate software allows the 3-D reconstruction of suitably sized extra-embryonic membrane components. These digital images created from optical sections through the tissue are obtained non-destructively, and the relationships in space of the components are well preserved.

  12. Light-sheet microscopy by confocal line scanning of dual-Bessel beams.

    Science.gov (United States)

    Zhang, Pengfei; Phipps, Mary E; Goodwin, Peter M; Werner, James H

    2016-10-01

    We have developed a light-sheet microscope that uses confocal scanning of dual-Bessel beams for illumination. A digital micromirror device (DMD) is placed in the intermediate image plane of the objective used to collect fluorescence and is programmed with two lines of pixels in the “on” state such that the DMD functions as a spatial filter to reject the out-of-focus background generated by the side-lobes of the Bessel beams. The optical sectioning and out-of-focus background rejection capabilities of this microscope were demonstrated by imaging of fluorescently stained actin in human A431 cells. The dual-Bessel beam system enables twice as many photons to be detected per imaging scan, which is useful for low light applications (e.g., single-molecule localization) or imaging at high speed with a superior signal to noise. While demonstrated for two Bessel beams, this approach is scalable to a larger number of beams.

  13. Anatomical and histochemical analysis of Dysphania ambrosioides supported by light and electron microscopy

    Directory of Open Access Journals (Sweden)

    Rafaela D. Sá

    Full Text Available ABSTRACT Dysphania ambrosioides (L. Mosyakin & Clemants (syn: Chenopodium ambrosoides L., Amaranthaceae, popularly known as “mastruz”, is an herb widely used in Brazil as anthelmintic. To contribute to the knowledge about medicinal plants, a microscopic analysis was accomplished to describe the main anatomical characters of root, stem, petiole and leaf blade of D. ambrosioides and histochemical tests were performed on the leaf blade. Cross-sections were obtained, by hand, for microscopic analysis of root, stem, petiole and leaf blade; to the leaf blade were still made paradermal sections, scanning electron microscopy analysis, maceration and histochemical tests. The main characters useful in the identification of the plant were: anomalous secondary thickening in the root and stem; presence of idioblasts containing crystal sand in the root, stem, petiole and leaf blade; in these there are also idioblasts with druses; presence of non-glandular and glandular trichomes in the stem, petiole and leaf blade; stomata on the stem, petiole and leaf blade, identified in these as anomocytic and anisocytic; dorsiventral mesophyll and collateral vascular bundles. Maceration revealed that the vessel elements are helical type. Through the histochemical tests, it was evidenced the presence of lipophilic substances, essential oils, oleoresins, phenolic compounds, starch, lignin and calcium oxalate crystals. This work provides support to the quality control of the species.

  14. Subconjunctival Dirofilaria repens Infestation: A Light and Scanning Electron Microscopy Study.

    Science.gov (United States)

    Melsom, Henrik A; Kurtzhals, Jørgen A L; Qvortrup, Klaus; Bargum, Ralph; Barfod, Toke S; la Cour, Morten; Heegaard, Steffen

    2011-01-01

    To present a case of subconjunctival infestation with Dirofilaria repens which is very rare in Northern Europe. A 61-year-old male presented with a swelling and redness of the left supraorbital region migrating to the eyelid and the left eyeball resulting in conjunctival injection, proptosis and diplopia. The patient underwent incisional extraction of a nine cm long worm, which was analysed histologically. The worm was structureless, greyish-white in colour and measuring nine cm in length and 0.5 mm in diameter. Histopathological examination of the worm showed an outer thick, multi-layered cuticle with longitudinal ridges. Beneath the cuticle, a thick muscle layer was observed and internally the intestine and a single reproductive tube containing spermatozoa were noted. Scanning electron microscopy of the worm showed tapered ends, transverse striations and longitudinal ridges at the anterior end. The tail was relatively short with spirally coiled ridges indicating a male Dirofilaria repens. Humans are an uncommon and accidental host of Dirofilaria repens which is rarely seen in Northern Europe but should be considered as a differential diagnosis to other nematode ocular infections. A travel history is helpful in diagnosing the potential involved organisms. No further treatment is necessary beyond surgical removal since this organism fails to mature and thereby does not cause microfilariaemia in humans.

  15. Hygroscopic Swelling Determination of Cellulose Nanocrystal (CNC) Films by Polarized Light Microscopy Digital Image Correlation.

    Science.gov (United States)

    Shrestha, Shikha; Diaz, Jairo A; Ghanbari, Siavash; Youngblood, Jeffrey P

    2017-05-08

    The coefficient of hygroscopic swelling (CHS) of self-organized and shear-oriented cellulose nanocrystal (CNC) films was determined by capturing hygroscopic strains produced as result of isothermal water vapor intake in equilibrium. Contrast enhanced microscopy digital image correlation enabled the characterization of dimensional changes induced by the hygroscopic swelling of the films. The distinct microstructure and birefringence of CNC films served in exploring the in-plane hygroscopic swelling at relative humidity values ranging from 0% to 97%. Water vapor intake in CNC films was measured using dynamic vapor sorption (DVS) at constant temperature. The obtained experimental moisture sorption and kinetic profiles were analyzed by fitting with Guggenheim, Anderson, and deBoer (GAB) and Parallel Exponential Kinetics (PEK) models, respectively. Self-organized CNC films showed isotropic swelling, CHS ∼0.040 %strain/%C. By contrast, shear-oriented CNC films exhibited an anisotropic swelling, resulting in CHS ∼0.02 and ∼0.30 %strain/%C, parallel and perpendicular to CNC alignment, respectively. Finite element analysis (FEA) further predicted moisture diffusion as the predominant mechanism for swelling of CNC films.

  16. An optical light microscopy study of aggregation of membrane tethered particles

    Science.gov (United States)

    Rädler, Joachim O.; Koltover, Ilya; Safinya, Cyrus R.

    1996-03-01

    We have studied the behavior of polystyrene latex beads tethered to giant phospholipid vesicle membranes via the covalent biotin-streptavidin bond. The vesicles were composed of mixtures of dimyristoyl phosphatidylcholine (DMPC) with small amounts of biotinylated diheptanoyl phosphatidylethanolamin (DHPE-X-biotin). Giant vesicles with attached beads were observed by video-enhanced optical microscopy as a function of local vesicle curvature, membrane tension and temperature. We have found that the beads tend to aggregate at high curvature regions of the membrane above the phospholipid chain-melting temperature (i.e. on a fluid membrane). With increasing number of beads, large bead aggregates form even on spherical vesicles. In the latter case, larger bead aggregates are favored over a number of smaller ones. We compare our findings to recent theories of interactions between particles locally deforming the membranes. footnote Goulian et. al., Europhysics Letters, 1993, v.22, (no.2), 145-50. footnote Dan et. al., J. Phys II, 1994, v.4, (no.10), 1713-25.

  17. Self-organization of the Escherichia coli chemotaxis network imaged with super-resolution light microscopy.

    Directory of Open Access Journals (Sweden)

    Derek Greenfield

    2009-06-01

    Full Text Available The Escherichia coli chemotaxis network is a model system for biological signal processing. In E. coli, transmembrane receptors responsible for signal transduction assemble into large clusters containing several thousand proteins. These sensory clusters have been observed at cell poles and future division sites. Despite extensive study, it remains unclear how chemotaxis clusters form, what controls cluster size and density, and how the cellular location of clusters is robustly maintained in growing and dividing cells. Here, we use photoactivated localization microscopy (PALM to map the cellular locations of three proteins central to bacterial chemotaxis (the Tar receptor, CheY, and CheW with a precision of 15 nm. We find that cluster sizes are approximately exponentially distributed, with no characteristic cluster size. One-third of Tar receptors are part of smaller lateral clusters and not of the large polar clusters. Analysis of the relative cellular locations of 1.1 million individual proteins (from 326 cells suggests that clusters form via stochastic self-assembly. The super-resolution PALM maps of E. coli receptors support the notion that stochastic self-assembly can create and maintain approximately periodic structures in biological membranes, without direct cytoskeletal involvement or active transport.

  18. Introduction to cryogenic engineering

    CERN Document Server

    CERN. Geneva; Vandoni, Giovanna; Niinikoski, Tapio O

    2005-01-01

    Cryogenic engineering is one of the key technologies at CERN. It is widely used in research and has many applications in industry and last but not least in medicine. In research cryogenic engineering and its applications are omnipresent from the smallest laboratories to fusion reactors, hughe detectors and accelerators. With the termination of the LHC, CERN will in fact become the world's largest cryogenic installation. This series of talks intends to introduce the non-cryogenist to the basic principles and challenges of cryogenic engineering and its applications. The course will also provide a basis for practical application as well as for further learning.

  19. Testing a high-power LED based light source for hyperspectral imaging microscopy

    Science.gov (United States)

    Klomkaew, Phiwat; Mayes, Sam A.; Rich, Thomas C.; Leavesley, Silas J.

    2017-02-01

    Our lab has worked to develop high-speed hyperspectral imaging systems that scan the fluorescence excitation spectrum for biomedical imaging applications. Hyperspectral imaging can be used in remote sensing, medical imaging, reaction analysis, and other applications. Here, we describe the development of a hyperspectral imaging system that comprised an inverted Nikon Eclipse microscope, sCMOS camera, and a custom light source that utilized a series of high-power LEDs. LED selection was performed to achieve wavelengths of 350-590 nm. To reduce scattering, LEDs with low viewing angles were selected. LEDs were surface-mount soldered and powered by an RCD. We utilized 3D printed mounting brackets to assemble all circuit components. Spectraradiometric calibration was performed using a spectrometer (QE65000, Ocean Optics) and integrating sphere (FOIS-1, Ocean Optics). Optical output and LED driving current were measured over a range of illumination intensities. A normalization algorithm was used to calibrate and optimize the intensity of the light source. The highest illumination power was at 375 nm (3300 mW/cm2), while the lowest illumination power was at 515, 525, and 590 nm (5200 mW/cm2). Comparing the intensities supplied by each LED to the intensities measured at the microscope stage, we found there was a great loss in power output. Future work will focus on using two of the same LEDs to double the power and finding more LED and/or laser diodes and chips around the range. This custom hyperspectral imaging system could be used for the detection of cancer and the identification of biomolecules.

  20. Optical diffraction tomography microscopy with transport of intensity equation using a light-emitting diode array

    Science.gov (United States)

    Li, Jiaji; Chen, Qian; Zhang, Jialin; Zhang, Zhao; Zhang, Yan; Zuo, Chao

    2017-08-01

    Optical diffraction tomography (ODT) is an effective label-free technique for quantitatively refractive index imaging, which enables long-term monitoring of the internal three-dimensional (3D) structures and molecular composition of biological cells with minimal perturbation. However, existing optical tomographic methods generally rely on interferometric configuration for phase measurement and sophisticated mechanical systems for sample rotation or beam scanning. Thereby, the measurement is suspect to phase error coming from the coherent speckle, environmental vibrations, and mechanical error during data acquisition process. To overcome these limitations, we present a new ODT technique based on non-interferometric phase retrieval and programmable illumination emitting from a light-emitting diode (LED) array. The experimental system is built based on a traditional bright field microscope, with the light source replaced by a programmable LED array, which provides angle-variable quasi-monochromatic illumination with an angular coverage of ±37 degrees in both x and y directions (corresponding to an illumination numerical aperture of ∼0.6). Transport of intensity equation (TIE) is utilized to recover the phase at different illumination angles, and the refractive index distribution is reconstructed based on the ODT framework under first Rytov approximation. The missing-cone problem in ODT is addressed by using the iterative non-negative constraint algorithm, and the misalignment of the LED array is further numerically corrected to improve the accuracy of refractive index quantification. Experiments on polystyrene beads and thick biological specimens show that the proposed approach allows accurate refractive index reconstruction while greatly reduced the system complexity and environmental sensitivity compared to conventional interferometric ODT approaches.

  1. Optomechanical properties of cancer cells revealed by light-induced deformation and quantitative phase microscopy

    Science.gov (United States)

    Kastl, Lena; Budde, Björn; Isbach, Michael; Rommel, Christina; Kemper, Björn; Schnekenburger, Jürgen

    2015-05-01

    There is a growing interest in cell biology and clinical diagnostics in label-free, optical techniques as the interaction with the sample is minimized and substances like dyes or fixatives do not affect the investigated cells. Such techniques include digital holographic microscopy (DHM) and the optical stretching by fiber optical two beam traps. DHM enables quantitative phase contrast imaging and thereby the determination of the cellular refractive index, dry mass and the volume, whereas optical cell stretching reveals the deformability of cells. Since optical stretching strongly depends on the optical properties and the shape of the investigated material we combined the usage of fiber optical stretching and DHM for the characterization of pancreatic tumor cells. The risk of tumors is their potential to metastasize, spread through the bloodstream and build distal tumors/metastases. The grade of dedifferentiation in which the cells lose their cell type specific properties is a measure for this metastatic potential. The less differentiated the cells are, the higher is their risk to metastasize. Our results demonstrate that pancreatic tumor cells, which are from the same tumor but vary in their grade of differentiation, show significant differences in their deformability. The retrieved data show that differentiated cells have a higher stiffness than less differentiated cells of the same tumor. Even cells that differ only in the expression of a single tumor suppressor gene which is responsible for cell-cell adhesions can be distinguished by their mechanical properties. Additionally, results from DHM measurements yield that the refractive index shows only few variations, indicating that it does not significantly influence optical cell stretching. The obtained results show a promising new approach for the phenotyping of different cell types, especially in tumor cell characterization and cancer diagnostics.

  2. Leaf anatomy and histochemistry of three species of Ficus sect. Americanae supported by light and electron microscopy.

    Science.gov (United States)

    Araújo, Nathalia Diniz; Coelho, Victor Peçanha M; Ventrella, Marília Contin; Agra, Maria de Fátima

    2014-02-01

    In this work the leaf anatomy of three species of Ficus section Americanae (Miq.) Miq. from Brazil, whose leaves and latex are used in folk medicine is reported. The work was carried out using light and scanning electron microscopy in order to characterize these species and to evaluate their taxonomic significance, and also contribute to the quality control of their ethnodrugs. The three species (Ficus cyclophylla, Ficus elliotiana, and Ficus caatingae) showed hypostomatic leaves, anomocytic stomata, straight epidermal cell outlines, and a dorsiventral mesophyll. Some micro-morphological characters such as density and distribution of epicuticular waxes, glandular trichomes, the length and width of stomata, as well as the palisade of mesophyll and petiole outlines proved to be the most useful and distinctive characters for the separation of species. These may contribute as additional support for the taxonomy of the section and for the quality control of their ethnodrugs.

  3. 3D exploration of light scattering from live cells in the presence of gold nanomarkers using holographic microscopy

    CERN Document Server

    Joud, Fadwa; Bun, P; Verpillat, Frédéric; Suck, Sarah Y; Tessier, G; Atlan, Michael; Desbiolles, Pierre; Coppey-Moisan, Maite; Abboud, Marie; Gross, Michel

    2011-01-01

    In this paper, we explore the 3D structure of light scattering from dark-field illuminated live 3T3 cells in the presence of 40 nm gold nanomarkers. For this purpose, we use a high resolution holographic microscope combining the off-axis heterodyne geometry and the phase-shifting acquisition of the digital holograms. A comparative study of the 3D reconstructions of the scattered fields allows us to locate the gold markers which yield, contrarily to the cell structures, well defined bright scattering patterns that are not angularly titled and clearly located along the optical axis (z). This characterization is an unambiguous signature of the presence of gold biological nanomarkers, and validates the capability of digital holographic microscopy to discriminate them from background signals in live ce

  4. 5D imaging via light sheet microscopy reveals cell dynamics during the eye-antenna disc primordium formation in Drosophila

    Science.gov (United States)

    Huang, Yu Shan; Ku, Hui Yu; Tsai, Yun Chi; Chang, Chin Hao; Pao, Sih Hua; Sun, Y. Henry; Chiou, Arthur

    2017-03-01

    5D images of engrailed (en) and eye gone (eyg) gene expressions during the course of the eye-antenna disc primordium (EADP) formation of Drosophila embryos from embryonic stages 13 through 16 were recorded via light sheet microscopy and analyzed to reveal the cell dynamics involved in the development of the EADP. Detailed analysis of the time-lapsed images revealed the process of EADP formation and its invagination trajectory, which involved an inversion of the EADP anterior-posterior axis relative to the body. Furthermore, analysis of the en-expression pattern in the EADP provided strong evidence that the EADP is derived from one of the en-expressing head segments.

  5. Proteasome particle-rich structures are widely present in human epithelial neoplasms: correlative light, confocal and electron microscopy study.

    Directory of Open Access Journals (Sweden)

    Vittorio Necchi

    Full Text Available A novel cytoplasmic structure has been recently characterized by confocal and electron microscopy in H. pylori-infected human gastric epithelium, as an accumulation of barrel-like proteasome reactive particles colocalized with polyubiquitinated proteins, H. pylori toxins and the NOD1 receptor. This proteasome particle-rich cytoplasmic structure (PaCS, a sort of focal proteasome hyperplasia, was also detected in dysplastic cells and was found to be enriched in SHP2 and ERK proteins, known to play a role in H. pylori-mediated gastric carcinogenesis. However, no information is available on its occurrence in neoplastic growths. In this study, surgical specimens of gastric cancer and various other human epithelial neoplasms have been investigated for PaCSs by light, confocal and electron microscopy including correlative confocal and electron microscopy (CCEM. PaCSs were detected in gastric cohesive, pulmonary large cell and bronchioloalveolar, thyroid papillary, parotid gland, hepatocellular, ovarian serous papillary, uterine cervix and colon adenocarcinomas, as well as in pancreatic serous microcystic adenoma. H. pylori bodies, their virulence factors (VacA, CagA, urease, and outer membrane proteins and the NOD1 bacterial proteoglycan receptor were selectively concentrated inside gastric cancer PaCSs, but not in PaCSs from other neoplasms which did, however, retain proteasome and polyubiquitinated proteins reactivity. No evidence of actual microbial infection was obtained in most PaCS-positive neoplasms, except for H. pylori in gastric cancer and capsulated bacteria in a colon cancer case. Particle lysis and loss of proteasome distinctive immunoreactivities were seen in some tumour cell PaCSs, possibly ending in sequestosomes or autophagic bodies. It is concluded that PaCSs are widely represented in human neoplasms and that both non-infectious and infectious factors activating the ubiquitin-proteasome system are likely to be involved in their origin

  6. Proteasome particle-rich structures are widely present in human epithelial neoplasms: correlative light, confocal and electron microscopy study.

    Science.gov (United States)

    Necchi, Vittorio; Sommi, Patrizia; Vanoli, Alessandro; Manca, Rachele; Ricci, Vittorio; Solcia, Enrico

    2011-01-01

    A novel cytoplasmic structure has been recently characterized by confocal and electron microscopy in H. pylori-infected human gastric epithelium, as an accumulation of barrel-like proteasome reactive particles colocalized with polyubiquitinated proteins, H. pylori toxins and the NOD1 receptor. This proteasome particle-rich cytoplasmic structure (PaCS), a sort of focal proteasome hyperplasia, was also detected in dysplastic cells and was found to be enriched in SHP2 and ERK proteins, known to play a role in H. pylori-mediated gastric carcinogenesis. However, no information is available on its occurrence in neoplastic growths. In this study, surgical specimens of gastric cancer and various other human epithelial neoplasms have been investigated for PaCSs by light, confocal and electron microscopy including correlative confocal and electron microscopy (CCEM). PaCSs were detected in gastric cohesive, pulmonary large cell and bronchioloalveolar, thyroid papillary, parotid gland, hepatocellular, ovarian serous papillary, uterine cervix and colon adenocarcinomas, as well as in pancreatic serous microcystic adenoma. H. pylori bodies, their virulence factors (VacA, CagA, urease, and outer membrane proteins) and the NOD1 bacterial proteoglycan receptor were selectively concentrated inside gastric cancer PaCSs, but not in PaCSs from other neoplasms which did, however, retain proteasome and polyubiquitinated proteins reactivity. No evidence of actual microbial infection was obtained in most PaCS-positive neoplasms, except for H. pylori in gastric cancer and capsulated bacteria in a colon cancer case. Particle lysis and loss of proteasome distinctive immunoreactivities were seen in some tumour cell PaCSs, possibly ending in sequestosomes or autophagic bodies. It is concluded that PaCSs are widely represented in human neoplasms and that both non-infectious and infectious factors activating the ubiquitin-proteasome system are likely to be involved in their origin. PaCS detection

  7. A Cryogenic Flow Sensor Project

    Data.gov (United States)

    National Aeronautics and Space Administration — Advanced Technologies Group, Inc. proposes the development of a Cryogenic Flow Sensor (CFS) for determining mass flow of cryogens in spacecraft propellant...

  8. [Erythrocyte morphology in urine determined by light microscopy in patients with bladder cancer].

    Science.gov (United States)

    Knezević, Gordana; Parigros, Katarina; Krizaj, Biserka; Anić, Veronika; Pazur, Marina; Jelić-Puskarić, Biljana; Sustercić, Dunja; Kardum-Skelin, Ika

    2011-09-01

    A finding of 80% or more of dysmorphic erythrocytes is assumed to point to kidney glomeruli, and of 80% or more of isomorphic erythrocytes to lower urinary tract as the origin of bleeding. In urine samples without significant origin of bleeding, there were 20%-80% of mixed results with both dysmorphic and isomorphic erythrocytes. The aim of the study was to show the origin of erythrocytes in malignant urine samples. Samples were fresh native urine sediment contrast stained with 0.1% safranin solution and analyzed under light microscope (X40). Out of 72 patients with malignant cells detected in urine, the origin of erythrocytes was identified in 25 patients (nine female and 16 male) through 90 samples (approximately 3-4 samples per patient); 26 (28.9%) samples did not have enough erythrocytes to define their origin, a mixed origin of erythrocytes was identified in 33 (36.7%) samples, dysmorphic erythrocytes were found in 25 (27.9%) samples, and isomorphic erythrocytes in 6 (6.3%) samples. In conclusion, there was no specific connection between malignant cell findings in urine and origin of erythrocytes. However, the high presence of mixed erythrocyte origin in malignant samples may suggest that the existence of a malignant process and renal disease should be taken in consideration.

  9. Spectral Reflectance of Sub-Micron Scale Light Absorbing Impurities Using Hyperspectral Microscopy

    Science.gov (United States)

    Kaspari, S.; Dal Farra, A.; Beach, J.; Schaepman, M. E.; Schwikowski, M.

    2016-12-01

    Light absorbing impurities (LAI) include black carbon, mineral dust and colored organic material. When deposited on highly reflective snow and glacier ice, LAI cause darkening of the surface, resulting in greater absorption of solar energy, heating of the snow/ice, and accelerated snow and glacier melt. Efforts to reduce LAI emissions and deposition have the potential to slow melt in regions where LAI are a substantial driver of snow and/or glacier melt. However, difficulties in characterizing the optical properties of mineral dust and organic LAI impede the assessment of the relative importance of black carbon, dust and organic LAI in driving melt. We developed a new method to optically characterize black carbon, mineral dust and organic matter at the particle scale using a Hyperspectral Microscope (HM, Cytoviva). The HM provides quantitative spectral analysis of nanoscale (128 nm pixel resolution) materials in the visible to near-infrared range (400 nm-1000 nm). We present: 1) an overview of the modifications we made to the HM in order to measure LAI reflectance, 2) reflectance spectra of pure minerals, black carbon, and humic substances measured with the HM at the particle scale, 3) a comparison of the HM measured spectra with bulk measurements made of the same materials using a spectroradiometer, and 4) preliminary results from environmental samples.

  10. Quantification of high-power ultrasound induced damage on potato starch granules using light microscopy.

    Science.gov (United States)

    Zuo, Yue Yue J; Hébraud, Pascal; Hemar, Yacine; Ashokkumar, Muthupandian

    2012-05-01

    A simple light microscopic technique was developed in order to quantify the damage inflicted by high-power low-frequency ultrasound (0-160 W, 20 kHz) treatment on potato starch granules in aqueous dispersions. The surface properties of the starch granules were modified using ethanol and SDS washing methods, which are known to displace proteins and lipids from the surface of the starch granules. The study showed that in the case of normal and ethanol-washed potato starch dispersions, two linear regions were observed. The number of defects first increased linearly with an increase in ultrasound power up to a threshold level. This was then followed by another linear dependence of the number of defects on the ultrasound power. The power threshold where the change-over occurred was higher for the ethanol-washed potato dispersions compared to non-washed potato dispersions. In the case of SDS-washed potato starch, although the increase in defects was linear with the ultrasound power, the power threshold for a second linear region was not observed. These results are discussed in terms of the different possible mechanisms of cavitation induced-damage (hydrodynamic shear stresses and micro-jetting) and by taking into account the hydrophobicity of the starch granule surface.

  11. Light and transmission electron microscopy of Cepedea longa (Opalinidae from Fejervarya limnocharis

    Directory of Open Access Journals (Sweden)

    Li Can

    2017-01-01

    Full Text Available Cepedea longa Bezzenberger, 1904, collected from Fejervarya limnocharis (Amphibia, Anura, Ranidae from Honghu Lake, Hubei Province, China in May–July 2016, is described at both light and transmission electron microscope levels. This is the first electron microscopic study of this species. Cepedea longa possesses a developed fibrillar skeletal system, composed of longitudinal fibrillar bands and transversal fibrils as well as numerous thin microfibrils dispersed in the endoplasm, which may play an important role in morphogenesis and offer some resilience to deformations of the cell. Longitudinal microfibrils are polarizing elements of kineties, bordering the somatic kineties on the left side and possibly responsible for kinetosome alignment. Two types of vesicles exist in the somatic cortex: globular endocytotic vesicles and flattened exocytotic vesicles. As to the nuclei of C. longa, a thick microfibrillar layer was observed to attach to the cytoplasmic face of the nuclear envelope. This fact suggests no necessary connection between the presence of this microfibrillar layer and the number of nuclei. In addition, some unknown tightly-packed microtubular structures in the nucleoplasm were observed for the first time in opalinids; neither their nature nor physiological significance is known. A detailed list of all reported Cepedea species is included.

  12. Quantitative assessment of the age of fibrotic lesions using polarized light microscopy and digital image analysis.

    Science.gov (United States)

    Pickering, J G; Boughner, D R

    1991-05-01

    Reliable histologic methods for gauging the maturity of fibrotic lesions are limited, making interventions in the healing process difficult to assess. As collagen ages there is enhanced birefringence due to increased molecular and fibrillar organization. The purpose of this study was to develop a microscopal technique to quantify this process and to determine its ability to distinguish scars of varying ages. Fibrosis in the rat gracilis muscle was studied 5 to 63 days after superficial injury. Sections were stained with picrosirius red and illuminated with monochromatic, polarized light. The microscope fields were digitized using a computer-video system yielding an image in which noncollagenous material was dark (gray level 0) and collagen was depicted by grey levels 1 to 255. In the fibrosis model used, the collagen area fraction plateaued at 80% by day 21. The median collagen grey level increased progressively as the scar aged. It is concluded that this histologic, nondestructive technique can reliably quantify age-related optical properties of fibrotic collagen and that this could be used to determine the maturity of fibrotic lesions.

  13. LED-FISH: Fluorescence microscopy based on light emitting diodes for the molecular analysis of Her-2/neu oncogene amplification

    Directory of Open Access Journals (Sweden)

    Vollmer Ekkehard

    2008-12-01

    Full Text Available Abstract Light emitting diodes (LED, which are available as small monochromatic light sources with characteristic features such as maximum illumination power combined with minimum energy consumption and extremely long lifespan have already proved as a highly potential low-cost alternative for specific diagnostic applications in clinical medicine such as tuberculosis fluorescence microscopy. Likewise, the most reliable evaluation of Her-2/neu (c-erbB2 gene amplification, which has been established in the last few years for routine diagnosis in clinical pathology as determinant towards Herceptin-based treatment of patients with breast cancer, is based on fluorescence in situ hybridization (FISH and corresponding high priced fluorescence equipment. In order to test the possibility to utilize the advantages of low-cost LED technology on FISH analysis of c-erbB2 gene expression for routine diagnostic purposes, the applicability of a standard bright field Carl Zeiss Axiostar Plus microscope equipped with a Fraen AFTER* LED Fluorescence Microscope Kit for the detection of Her-2/neu gene signals was compared to an advanced Nikon Eclipse 80i fluorescence microscope in combination with a conventional 100W mercury vapor lamp. Both microscopes were fitted with the same Quicam FAST CCD digital camera to unequivocally compare the quality of the captured images. C-erbB2 gene expression was analyzed in 30 different human tissue samples of primary invasive breast cancer, following formalin fixation and subsequent paraffin-embedding. The Her2/neu gene signals (green were identifiable in the tumor cells in all cases and images of equal quality were captured under almost identical conditions by 480 nm (blue LED module equipped standard Axiostar microscope as compared to conventional fluorescence microscopy. In this first attempt, these monochromatic LED elements proved in principle to be suitable for the detection of Her-2/neu gene expression by FISH. Thus, our own

  14. ZERODUR TAILORED for cryogenic application

    Science.gov (United States)

    Jedamzik, R.; Westerhoff, T.

    2014-07-01

    ZERODUR® glass ceramic from SCHOTT is known for its very low thermal expansion coefficient (CTE) at room temperature and its excellent CTE homogeneity. It is widely used for ground-based astronomical mirrors but also for satellite applications. Many reference application demonstrate the excellent and long lasting performance of ZERODUR® components in orbit. For space application a low CTE of the mirror material is required at cryogenic temperatures together with a good match of the thermal expansion to the supporting structure material. It is possible to optimize the coefficient of thermal expansion of ZERODUR® for cryogenic applications. This paper reports on measurements of thermal expansion of ZERODUR® down to cryogenic temperatures of 10 K performed by the PTB (Physikalisch Technische Bundesanstallt, Braunschweig, Germany, the national metrology laboratory). The ZERODUR® TAILORED CRYO presented in this paper has a very low coefficient of thermal expansion down to 70 K. The maximum absolute integrated thermal expansion down to 10 K is only about 20 ppm. Mirror blanks made from ZERODUR® TAILORED CRYO can be light weighted to almost 90% with our modern processing technologies. With ZERODUR® TAILORED CRYO, SCHOTT offers the mirror blank material for the next generation of space telescope applications.

  15. Simultaneous multiview capture and fusion improves spatial resolution in wide-field and light-sheet microscopy

    Science.gov (United States)

    Wu, Yicong; Chandris, Panagiotis; Winter, Peter W.; Kim, Edward Y.; Jaumouillé, Valentin; Kumar, Abhishek; Guo, Min; Leung, Jacqueline M.; Smith, Corey; Rey-Suarez, Ivan; Liu, Huafeng; Waterman, Clare M.; Ramamurthi, Kumaran S.; La Riviere, Patrick J.; Shroff, Hari

    2016-01-01

    Most fluorescence microscopes are inefficient, collecting only a small fraction of the emitted light at any instant. Besides wasting valuable signal, this inefficiency also reduces spatial resolution and causes imaging volumes to exhibit significant resolution anisotropy. We describe microscopic and computational techniques that address these problems by simultaneously capturing and subsequently fusing and deconvolving multiple specimen views. Unlike previous methods that serially capture multiple views, our approach improves spatial resolution without introducing any additional illumination dose or compromising temporal resolution relative to conventional imaging. When applying our methods to single-view wide-field or dual-view light-sheet microscopy, we achieve a twofold improvement in volumetric resolution (~235 nm × 235 nm × 340 nm) as demonstrated on a variety of samples including microtubules in Toxoplasma gondii, SpoVM in sporulating Bacillus subtilis, and multiple protein distributions and organelles in eukaryotic cells. In every case, spatial resolution is improved with no drawback by harnessing previously unused fluorescence. PMID:27761486

  16. Simultaneous localization of six antigens in single sections of transgenic mouse intestine using a combination of light and fluorescence microscopy.

    Science.gov (United States)

    Hermiston, M L; Latham, C B; Gordon, J I; Roth, K A

    1992-09-01

    To study the geographic differentiation of the intestinal epithelium and to understand the complex lineage relationships of its cell populations, it is often necessary to visualize the protein products of multiple genes in sections prepared from different positions along the duodenal-to-colonic and/or crypt-to-villus axes. Multilabel fluorescence or brightfield immunohistochemical techniques have previously been used for this purpose. However, the number of antigens that can be identified on single sections is limited in fluorescence microscopy by the number of fluorophores with non-overlapping absorption and emission characteristics, in brightfield microscopy by the number of visually distinguishable chromogens, and in both methods by the availability of primary antisera raised in multiple species. We have now used a combination of light and fluorescence microscopic techniques to increase the number of antigens that can be detected in a single section to six. Sections were sequentially stained using immunogold with silver intensification, peroxidase-antiperoxidase with diaminobenzidine chromogen, and peroxidase-anti-peroxidase with alpha-naphthol/basic dye as chromogen, followed by simultaneous fluorescent detection with fluorescein, 7-amino-4-methylcoumarin-3-acetic acid, and beta-phycoerythrin. This method enables up to four separate antigens to be visualized within a single cell and two additional antigens to be detected in unrelated cells. The technique is illustrated by examining the cellular patterns of expression of liver fatty acid binding protein/human growth hormone fusion genes in the intestinal epithelium of adult transgenic mice. It should be generally applicable to other experimental systems that require localization of multiple antigens in single tissue sections.

  17. Altered state of primordial follicles in neonatal and early infantile rats due to maternal hypothyroidism: Light and electron microscopy approach.

    Science.gov (United States)

    Danilović Luković, Jelena; Korać, Aleksandra; Milošević, Ivan; Lužajić, Tijana; Puškaš, Nela; Kovačević Filipović, Milica; Radovanović, Anita

    2016-11-01

    Thyroid hormones (TH) are one of the key factors for normal prenatal development in mammals. Previously, we showed that subclinical maternal hypothyroidism leads to premature atresia of ovarian follicles in female rat offspring in the pre-pubertal and pubertal periods. The influence of decreased concentration of TH on primordial follicles pool formation during neonatal and early infantile period of rat pups was not investigated previously. Maternal hypothyroidism during pregnancy has irreversible negative influence on primordial follicles pool formation and population of resting oocytes in female rat offspring. The study was done on neonatal and early infantile control (n-10) and hypothyroid (n-10) female rat pups derived from control (n-6) and propylthiouracil (PTU) treated pregnant dams (n-6), respectively. Ovaries of all pups were removed and processed for light and transmission electron microscopy (TEM). Number of nests, oogonia and oocytes per nest, primordial, primary, secondary and preantral follicles were determined. Screening for overall calcium presence in ovarian tissue was done using Alizarin red staining. Morphology and volume density of nucleus, mitochondria and smooth endoplasmic reticulum (sER) in the oocytes in primordial follicles was also assessed. Caspase-3 and terminal deoxynucleotidyl transferase dUTP nick end labelling (TUNEL), both markers for apoptosis, and proliferating cell nuclear antigen (PCNA) for proliferation were determined in oocytes and granulosa cells in different type of follicles. In neonatal period, ovaries of hypothyroid pups had a decreased number of oogonia, oocytes and nests, an increased number of primordial follicles and a decreased number of primary and secondary follicles, while in early infantile period, increased number of primary, secondary and preantral follicles were found. Alizarin red staining was intense in hypothyroid neonatal rats that also had the highest content of dilated sER. Number of mitochondria with

  18. Quantitative fractography under light microscopy: A digital image processing approach; Quantitative Fraktographie mittels Lichtmikroskopie: Naeherung durch digitale Bildverarbeitung

    Energy Technology Data Exchange (ETDEWEB)

    Horovistiz, A.L.; Ribeiro, L.M.F.; Campos, K.A.; Jesuino, G.A.; Guimaraes, V.A.; Hein, L.R.O. [UNESP, Guaratingueta, SP (Brazil)

    2003-02-01

    This work is an example of the improvement on quantitative fractography by means of digital image processing and light microscopy. Two techniques are presented to investigate the quantitative fracture behavior of Ti-4Al-4V heat-treated alloy specimens, under Charpy impact testing. The first technique is the Minkowski method for fractal dimension measurement from surface profiles, revealing the multifractal character of Ti-4Al-4V fracture. It was not observed a clear positive correlation of fractal values against Charpy energies for Ti-4Al-4V alloy specimens, due to their ductility, microstructural heterogeneities and the dynamic loading characteristics at region near the V-notch. The second technique provides an entire elevation map of fracture surface by extracting in-focus regions for each picture from a stack of images acquired at successive focus positions, then computing the surface roughness. Extended-focus reconstruction has been used to explain the behavior along fracture surface. Since these techniques are based on light microscopy, their inherent low cost is very interesting for failure investigations. (orig.) [German] Diese Arbeit ist ein Beispiel fuer die Verbesserung der quantitativen Fraktographie mittels digitaler Bildverarbeitung und Lichtmikroskopie. Zur Untersuchung des quantitativen Bruchverhaltens von waermebehandelten Ti-4Al-4V-Proben im Charpy-Kerbschlagversuch werden zwei Techniken vorgestellt. Die erste Technik ist die Minkowski-Methode zur Messung der fraktalen Dimensionen aus Oberflaechenprofilen, welche den multifraktalen Charakter des Bruches von Ti-4Al-4V ergibt. Es wurde keine eindeutige positive Korrelation zwischen den fraktalen Werten und den Charpyenergien der Ti-4Al-4V-Proben aufgrund deren Duktilitaet, Gefuegeheterogenitaeten und dynamischen Belastungscharakteristiken im Bereich um den V-Kerb beobachtet. Die zweite Methode bietet eine vollstaendige Erhoehungsabbildung der Bruchoberflaeche durch Extraktion der Fokusierungsbereiche

  19. Polymers at cryogenic temperatures

    CERN Document Server

    Fu, Shao-Yun

    2013-01-01

    Kalia and Fu's novel monograph covers cryogenic treatment, properties and applications of cryo-treated polymer materials. Written by numerous international experts, the twelve chapters in this book offer the reader a comprehensive picture of the latest findings and developments, as well as an outlook on the field. Cryogenic technology has seen remarkable progress in the past few years and especially cryogenic properties of polymers are attracting attention through new breakthroughs in space, superconducting, magnetic and electronic techniques. This book is a valuable resource for researchers, educators, engineers and graduate students in the field and at technical institutions.

  20. SNS Cryogenic Systems Commissioning

    Science.gov (United States)

    Hatfield, D.; Casagrande, F.; Campisi, I.; Gurd, P.; Howell, M.; Stout, D.; Strong, H.; Arenius, D.; Creel, J.; Dixon, K.; Ganni, V.; Knudsen, P.

    2006-04-01

    The Spallation Neutron Source (SNS) is under construction at Oak Ridge National Laboratory. The cold section of the Linac consists of 81 superconducting radio frequency cavities cooled to 2.1K by a 2400 watt cryogenic refrigeration system. The major cryogenic system components include warm helium compressors with associated oil removal and gas management, 4.5K cold box, 7000L liquid helium dewar, 2.1K cold box (consisting of 4 stages of cold compressors), gaseous helium storage, helium purification and gas impurity monitoring system, liquid nitrogen storage and the cryogenic distribution transfer line system. The overall system commissioning and future plans will be presented.

  1. FRIB Cryogenic Plant Status

    Energy Technology Data Exchange (ETDEWEB)

    Dixon, Kelly D. [Thomas Jefferson National Accelerator Facility (TJNAF), Newport News, VA (United States); Ganni, Venkatarao [Thomas Jefferson National Accelerator Facility (TJNAF), Newport News, VA (United States); Knudsen, Peter N. [Thomas Jefferson National Accelerator Facility (TJNAF), Newport News, VA (United States); Casagranda, Fabio [Michigan State Univ., East Lansing, MI (United States)

    2015-12-01

    After practical changes were approved to the initial conceptual design of the cryogenic system for MSU FRIB and an agreement was made with JLab in 2012 to lead the design effort of the cryogenic plant, many activities are in place leading toward a cool-down of the linacs prior to 2018. This is mostly due to using similar equipment used at CHLII for the 12 GeV upgrade at JLab and an aggressive schedule maintained by the MSU Conventional Facilities department. Reported here is an updated status of the cryogenic plant, including the equipment procurement status, plant layout, facility equipment and project schedule.

  2. SNS Cryogenic Systems Commissioning

    Energy Technology Data Exchange (ETDEWEB)

    D. Hatfield; F. Casagrande; I. Campisi; P. Gurd; M. Howell; D. Stout; H. Strong; D. Arenius; J. Creel; K. Dixon; V. Ganni; and P. Knudsen

    2005-08-29

    The Spallation Neutron Source (SNS) is under construction at Oak Ridge National Laboratory. The cold section of the Linac consists of 81 superconducting radio frequency cavities cooled to 2.1K by a 2400 watt cryogenic refrigeration system. The major cryogenic system components include warm helium compressors with associated oil removal and gas management, 4.5K cold box, 7000L liquid helium dewar, 2.1K cold box (consisting of 4 stages of cold compressors), gaseous helium storage, helium purification and gas impurity monitoring system, liquid nitrogen storage and the cryogenic distribution transfer line system. The overall system commissioning and future plans will be presented.

  3. Fundamentals of cryogenic engineering

    CERN Document Server

    Mukhopadhyay, Mamata

    2014-01-01

    The author, with her vast and varied experience in teaching and allied fields, clearly enunciates the behaviour and various properties of common cryogenic fluids, methods of liquefaction, and separation and applications of cryogens with thermodynamic analysis for process selection. This profusely illustrated study with clear-cut diagrams and process charts, should serve not only as a textbook for students but also as an excellent reference for researchers and practising engineers on design of cryogenic refrigeration, and liquefaction and separation process plants for various applications.

  4. Evanescent Light-Scattering Microscopy for Label-Free Interfacial Imaging: From Single Sub-100 nm Vesicles to Live Cells.

    Science.gov (United States)

    Agnarsson, Björn; Lundgren, Anders; Gunnarsson, Anders; Rabe, Michael; Kunze, Angelika; Mapar, Mokhtar; Simonsson, Lisa; Bally, Marta; Zhdanov, Vladimir P; Höök, Fredrik

    2015-12-22

    Advancement in the understanding of biomolecular interactions has benefited greatly from the development of surface-sensitive bioanalytical sensors. To further increase their broad impact, significant efforts are presently being made to enable label-free and specific biomolecule detection with high sensitivity, allowing for quantitative interpretation and general applicability at low cost. In this work, we have addressed this challenge by developing a waveguide chip consisting of a flat silica core embedded in a symmetric organic cladding with a refractive index matching that of water. This is shown to reduce stray light (background) scattering and thereby allow for label-free detection of faint objects, such as individual sub-20 nm gold nanoparticles as well as sub-100 nm lipid vesicles. Measurements and theoretical analysis revealed that light-scattering signals originating from single surface-bound lipid vesicles enable characterization of their sizes without employing fluorescent lipids as labels. The concept is also demonstrated for label-free measurements of protein binding to and enzymatic (phospholipase A2) digestion of individual lipid vesicles, enabling an analysis of the influence on the measured kinetics of the dye-labeling of lipids required in previous assays. Further, diffraction-limited imaging of cells (platelets) binding to a silica surface showed that distinct subcellular features could be visualized and temporally resolved during attachment, activation, and spreading. Taken together, these results underscore the versatility and general applicability of the method, which due to its simplicity and compatibility with conventional microscopy setups may reach a widespread in life science and beyond.

  5. Determining the fibrillar orientation of bast fibres with polarized light microscopy: the modified Herzog test (red plate test) explained.

    Science.gov (United States)

    Haugan, E; Holst, B

    2013-11-01

    The identification of bast fibre samples, in particular, bast fibres used in textiles, is an important issue in archaeology, criminology and other scientific fields. One of the characteristic features of bast fibres is their fibrillar orientation, referred to as Z- or S twist (or alternatively right- and left-handed fibres). An empirical test for determining the fibrillar orientation using polarized light microscopy has been known in the community for many years. It is referred to as the modified Herzog test or red plate test. The test has the reputation for never producing false results, but also for occasionally not working. However, so far, no proper justification has been provided in the literature that the 'no false results' assumption is really correct and it has also not been clear up till now, why the method sometimes does not work. In this paper, we present an analytical model for the modified Herzog test, which explains why the test never gives a false result. We also provide an explanation for why the Herzog test sometimes does not work: According to our model, the Herzog test will not work if none of the three distinct layers in the secondary cell wall is significantly thicker than the others.

  6. Determining the fibrillar orientation of bast fibres with polarized light microscopy: the modified Herzog test (red plate test) explained

    Science.gov (United States)

    HAUGAN, E; HOLST, B

    2013-01-01

    The identification of bast fibre samples, in particular, bast fibres used in textiles, is an important issue in archaeology, criminology and other scientific fields. One of the characteristic features of bast fibres is their fibrillar orientation, referred to as Z- or S twist (or alternatively right- and left-handed fibres). An empirical test for determining the fibrillar orientation using polarized light microscopy has been known in the community for many years. It is referred to as the modified Herzog test or red plate test. The test has the reputation for never producing false results, but also for occasionally not working. However, so far, no proper justification has been provided in the literature that the ‘no false results’ assumption is really correct and it has also not been clear up till now, why the method sometimes does not work. In this paper, we present an analytical model for the modified Herzog test, which explains why the test never gives a false result. We also provide an explanation for why the Herzog test sometimes does not work: According to our model, the Herzog test will not work if none of the three distinct layers in the secondary cell wall is significantly thicker than the others. PMID:24020614

  7. Correlated light and electron microscopy observations of the uterine epithelial cell actin cytoskeleton using fluorescently labeled resin-embedded sections.

    Science.gov (United States)

    Moore, Chad L; Cheng, Delfine; Shami, Gerald J; Murphy, Christopher R

    2016-05-01

    In order to perform correlative light and electron microscopy (CLEM) more precisely, we have modified existing specimen preparation protocols allowing fluorescence retention within embedded and sectioned tissue, facilitating direct observation across length scales. We detail a protocol which provides a precise correlation accuracy using accessible techniques in biological specimen preparation. By combining a pre-embedding uranyl acetate staining step with the progressive lowering of temperature (PLT) technique, a methacrylate embedded tissue specimen is ultrathin sectioned and mounted onto a TEM finder grid for immediate viewing in the confocal and electron microscope. In this study, the protocol is applied to rat uterine epithelial cells in vivo during early pregnancy. Correlative overlay data was used to track changes in filamentous actin that occurs in these cells from fertilization (Day 1) to implantation on Day 6 as part of the plasma membrane transformation, a process essential in the development of uterine receptivity in the rat. CLEM confirmed that the actin cytoskeleton is disrupted as apical microvilli are progressively lost toward implantation, and revealed the thick and continuous terminal web is replaced by a thinner and irregular actin band, with individually distinguishable filaments connecting actin meshworks which correspond with remaining plasma membrane protrusions. Copyright © 2016 Elsevier Ltd. All rights reserved.

  8. Correlative and integrated light and electron microscopy of in-resin GFP fluorescence, used to localise diacylglycerol in mammalian cells.

    Science.gov (United States)

    Peddie, Christopher J; Blight, Ken; Wilson, Emma; Melia, Charlotte; Marrison, Jo; Carzaniga, Raffaella; Domart, Marie-Charlotte; O'Toole, Peter; Larijani, Banafshe; Collinson, Lucy M

    2014-08-01

    Fluorescence microscopy of GFP-tagged proteins is a fundamental tool in cell biology, but without seeing the structure of the surrounding cellular space, functional information can be lost. Here we present a protocol that preserves GFP and mCherry fluorescence in mammalian cells embedded in resin with electron contrast to reveal cellular ultrastructure. Ultrathin in-resin fluorescence (IRF) sections were imaged simultaneously for fluorescence and electron signals in an integrated light and scanning electron microscope. We show, for the first time, that GFP is stable and active in resin sections in vacuo. We applied our protocol to study the subcellular localisation of diacylglycerol (DAG), a modulator of membrane morphology and membrane dynamics in nuclear envelope assembly. We show that DAG is localised to the nuclear envelope, nucleoplasmic reticulum and curved tips of the Golgi apparatus. With these developments, we demonstrate that integrated imaging is maturing into a powerful tool for accurate molecular localisation to structure. Copyright © 2014 The Authors. Published by Elsevier B.V. All rights reserved.

  9. Light and electron microscopy of the European beaver (Castor fiber stomach reveal unique morphological features with possible general biological significance.

    Directory of Open Access Journals (Sweden)

    Natalia Ziółkowska

    Full Text Available Anatomical, histological, and ultrastructural studies of the European beaver stomach revealed several unique morphological features. The prominent attribute of its gross morphology was the cardiogastric gland (CGG, located near the oesophageal entrance. Light microscopy showed that the CGG was formed by invaginations of the mucosa into the submucosa, which contained densely packed proper gastric glands comprised primarily of parietal and chief cells. Mucous neck cells represented <0.1% of cells in the CGG gastric glands and 22-32% of cells in the proper gastric glands of the mucosa lining the stomach lumen. These data suggest that chief cells in the CGG develop from undifferentiated cells that migrate through the gastric gland neck rather than from mucous neck cells. Classical chief cell formation (i.e., arising from mucous neck cells occurred in the mucosa lining the stomach lumen, however. The muscularis around the CGG consisted primarily of skeletal muscle tissue. The cardiac region was rudimentary while the fundus/corpus and pyloric regions were equally developed. Another unusual feature of the beaver stomach was the presence of specific mucus with a thickness up to 950 µm (in frozen, unfixed sections that coated the mucosa. Our observations suggest that the formation of this mucus is complex and includes the secretory granule accumulation in the cytoplasm of pit cells, the granule aggregation inside cells, and the incorporation of degenerating cells into the mucus.

  10. Collagen Architecture of the Posterior Pole: High-Resolution Wide Field of View Visualization and Analysis Using Polarized Light Microscopy

    Science.gov (United States)

    Jan, Ning-Jiun; Lathrop, Kira; Sigal, Ian A.

    2017-01-01

    Purpose The purpose of this study was to leverage polarized light microscopy (PLM) to visualize the collagen fiber architecture of posterior pole and optic nerve head with micrometer-scale resolution and to identify and quantify major organizational components. Methods Eight sheep posterior poles were cryosectioned and imaged using PLM. Collagen fiber orientation was determined by using custom scripts, and the resulting orientation maps were inspected and quantified to identify major structural elements and tested for differences in mean fiber orientation and anisotropy, using linear mixed effect models. Results Images revealed an intricate organization of collagen fibers in the posterior pole. In the lamina cribrosa, interweaving fibers formed large knots and wrapped around nerve fiber pores, with beam insertions into the scleral canal wall that were either narrow and straight or wide. In the peripapillary sclera, three significantly different (P PLM revealed structural aspects of the lamina cribrosa and sclera that may have important biomechanical roles but that were previously unreported or not characterized quantitatively. In the lamina cribrosa, these roles included wide and narrow beam insertions and details of collagen fibers interweaving and wrapping around the pores. In the sclera, we described regions of circumferential, radial, and unaligned “random” fibers. Although there is consensus that circumferential fibers protect neural tissues by resisting canal expansion, the role of the radial fibers remains unclear. PMID:28146238

  11. Modulation of the pupil function of microscope objective lens for multifocal multi-photon microscopy using a spatial light modulator

    Science.gov (United States)

    Matsumoto, Naoya; Okazaki, Shigetoshi; Takamoto, Hisayoshi; Inoue, Takashi; Terakawa, Susumu

    2014-02-01

    We propose a method for high precision modulation of the pupil function of a microscope objective lens to improve the performance of multifocal multi-photon microscopy (MMM). To modulate the pupil function, we adopt a spatial light modulator (SLM) and place it at the conjugate position of the objective lens. The SLM can generate an arbitrary number of spots to excite the multiple fluorescence spots (MFS) at the desired positions and intensities by applying an appropriate computer-generated hologram (CGH). This flexibility allows us to control the MFS according to the photobleaching level of a fluorescent protein and phototoxicity of a specimen. However, when a large number of excitation spots are generated, the intensity distribution of the MFS is significantly different from the one originally designed due to misalignment of the optical setup and characteristics of the SLM. As a result, the image of a specimen obtained using laser scanning for the MFS has block noise segments because the SLM could not generate a uniform MFS. To improve the intensity distribution of the MFS, we adaptively redesigned the CGH based on the observed MFS. We experimentally demonstrate an improvement in the uniformity of a 10 × 10 MFS grid using a dye solution. The simplicity of the proposed method will allow it to be applied for calibration of MMM before observing living tissue. After the MMM calibration, we performed laser scanning with two-photon excitation to observe a real specimen without detecting block noise segments.

  12. Four-wave mixing based light sources for real-world biomedical applications of coherent Raman microscopy

    Science.gov (United States)

    Gottschall, Thomas; Meyer, Tobias; Jauregui, Cesar; Schmitt, Michael; Popp, Jürgen; Limpert, Jens; Tünnermann, Andreas

    2016-03-01

    Stimulated Raman Scattering requires an extremely quiet, widely wavelength tunable laser, which, up to now, is unheard of in fiber lasers. We present a compact and maintenance-free optical parametric oscillator based on degenerate four-wave mixing in a photonic crystal fiber. By employing an all-fiber frequency and repetition rate tunable laser as a seed source, we are able to generate tunable light between 1015 and 1065 nm. After amplification and subsequent conversion in the fiber OPO, signal and idler radiation between 785 and 960 nm and 1177 and 1500 nm may be generated with a repetition rate of 9 MHz. Therefore, we are able to address Raman shifts between 910 and 3030 cm-1. An additional output provides the Stokes radiation at 18 MHz required for the SRS process, which is passively synchronized to the tunable radiation. We measure the relative intensity noise of the Stokes beam at 9 MHz to be -150 dBc enabling high speed SRS imaging with a good signal-to-noise ratio. The combination of FWM based conversion, coupled with all-fiber Yb-based fiber lasers allows for the first turn-key, widely tunable and extremely compact laser systems developed for applications of CRS microscopy in clinics. This source could very well be the missing key instrument that CRS imaging requires for its real world transition.

  13. Light and electron microscopy of the European beaver (Castor fiber) stomach reveal unique morphological features with possible general biological significance.

    Science.gov (United States)

    Ziółkowska, Natalia; Lewczuk, Bogdan; Petryński, Wojciech; Palkowska, Katarzyna; Prusik, Magdalena; Targońska, Krystyna; Giżejewski, Zygmunt; Przybylska-Gornowicz, Barbara

    2014-01-01

    Anatomical, histological, and ultrastructural studies of the European beaver stomach revealed several unique morphological features. The prominent attribute of its gross morphology was the cardiogastric gland (CGG), located near the oesophageal entrance. Light microscopy showed that the CGG was formed by invaginations of the mucosa into the submucosa, which contained densely packed proper gastric glands comprised primarily of parietal and chief cells. Mucous neck cells represented stomach lumen. These data suggest that chief cells in the CGG develop from undifferentiated cells that migrate through the gastric gland neck rather than from mucous neck cells. Classical chief cell formation (i.e., arising from mucous neck cells) occurred in the mucosa lining the stomach lumen, however. The muscularis around the CGG consisted primarily of skeletal muscle tissue. The cardiac region was rudimentary while the fundus/corpus and pyloric regions were equally developed. Another unusual feature of the beaver stomach was the presence of specific mucus with a thickness up to 950 µm (in frozen, unfixed sections) that coated the mucosa. Our observations suggest that the formation of this mucus is complex and includes the secretory granule accumulation in the cytoplasm of pit cells, the granule aggregation inside cells, and the incorporation of degenerating cells into the mucus.

  14. Collagen Architecture of the Posterior Pole: High-Resolution Wide Field of View Visualization and Analysis Using Polarized Light Microscopy.

    Science.gov (United States)

    Jan, Ning-Jiun; Lathrop, Kira; Sigal, Ian A

    2017-02-01

    The purpose of this study was to leverage polarized light microscopy (PLM) to visualize the collagen fiber architecture of posterior pole and optic nerve head with micrometer-scale resolution and to identify and quantify major organizational components. Eight sheep posterior poles were cryosectioned and imaged using PLM. Collagen fiber orientation was determined by using custom scripts, and the resulting orientation maps were inspected and quantified to identify major structural elements and tested for differences in mean fiber orientation and anisotropy, using linear mixed effect models. Images revealed an intricate organization of collagen fibers in the posterior pole. In the lamina cribrosa, interweaving fibers formed large knots and wrapped around nerve fiber pores, with beam insertions into the scleral canal wall that were either narrow and straight or wide. In the peripapillary sclera, three significantly different (P narrow beam insertions and details of collagen fibers interweaving and wrapping around the pores. In the sclera, we described regions of circumferential, radial, and unaligned "random" fibers. Although there is consensus that circumferential fibers protect neural tissues by resisting canal expansion, the role of the radial fibers remains unclear.

  15. High-fidelity functional and structural whole-brain imaging with Bessel-beam light-sheet microscopy (Conference Presentation)

    Science.gov (United States)

    Müllenbroich, Marie Caroline; Silvestri, Ludovico; Turrini, Lapo; Di Giovanna, Antonino Paolo; Alterini, Tommaso; Gheisari, Ali; Ricci, Pietro; Sacconi, Leonardo; Vanzi, Francesco; Pavone, Francesco S.

    2017-02-01

    Light-sheet microscopy (LSM) has proven a useful tool in neuroscience and is particularly well suited to image the entire brain with high frame rates at single cell resolution. On the one hand, LSM is employed in combination with tissue clearing methods like CLARITY which allows for the reconstruction of neuronal or vascular anatomy over cm-sized samples. On the other hand, LSM has been paired with intrinsically transparent samples for real-time recording of neuronal activity with single cell resolution across the entire brain, using calcium indicators like GCaMP6. Despite its intrinsic advantages in terms of high imaging speed and reduced photobleaching, LSM is very sensitive to residual opaque objects present in the sample, which cause dark horizontal stripes in the collected images. In the best case, these artefacts obscure the features of interest in structural imaging; in the worst case, dynamic shadowing introduced by red blood cells significantly alters the fluorescence signal variations related to neuronal activity. We show how the use of Bessel beams in LSM can dramatically reduce such artefacts even in conventional one-sided illumination schemes, thanks to their "self-healing" properties. On the functional side, Bessel-beam LSM allows recording neuronal activity traces without any disturbing flickering caused by the movement of red blood cells. On the structural side, our proposed method is capable of obtaining anatomical information across the entire volume of whole mouse brains allowing tracing blood vessels and neuronal projections also in poorly cleared specimens.

  16. Evaluation of Residual Cellularity and Proliferation on Preoperatively Treated Breast Cancer: A Comparison between Image Analysis and Light Microscopy Analysis

    Directory of Open Access Journals (Sweden)

    Valentina Corletto

    1998-01-01

    Full Text Available Histopathology has been suggested as a reliable method for tumour reduction evaluation of preoperatively treated breast cancer. Immunocytochemistry can be used to enhance the visibility of residual tumour cellularity and in the evaluation of its proliferative activity. We compared Image Analysis (IA with Light Microscopy Analysis (LMA on sections of breast carcinomas treated with preoperative chemo‐ or chemo/radiotherapy in the evaluation of the Neoplastic Cell Density (NCD (69 cases and the Proliferation Index (PI (35 cases. NCD was expressed as the immunoreactive area to cytokeratin over the total original neoplastic area and PI was expressed as the number of immunostained tumoural nuclei with MIB1 MoAb over the total of tumoural nuclei. The intraobserver agreement and that between IA and LMA for both indices were estimated by the common (Kw and the jackknife weighted kappa statistic (K˜w. The extent of agreement of each considered category was also assessed by means of the category‐specific kappa statistics (Kcs. The intraobserver agreement within LMA for NCD and PI and that between IA and LMA for PI were both satisfactory. Upon evaluation of the NCD, the agreement between IA and LMA showed unsatisfactory results, especially when the ratio between the residual tumour cells and the background was critical.

  17. Evaluation of residual cellularity and proliferation on preoperatively treated breast cancer: a comparison between image analysis and light microscopy analysis.

    Science.gov (United States)

    Corletto, V; Verderio, P; Giardini, R; Cipriani, S; Di Palma, S; Rilke, F

    1998-01-01

    Histopathology has been suggested as a reliable method for tumour reduction evaluation of preoperatively treated breast cancer. Immunocytochemistry can be used to enhance the visibility of residual tumour cellularity and in the evaluation of its proliferative activity. We compared Image Analysis (IA) with Light Microscopy Analysis (LMA) on sections of breast carcinomas treated with preoperative chemo- or chemo/radiotherapy in the evaluation of the Neoplastic Cell Density (NCD) (69 cases) and the Proliferation Index (PI) (35 cases). NCD was expressed as the immunoreactive area to cytokeratin over the total original neoplastic area and PI was expressed as the number of immunostained tumoural nuclei with MIB 1 MoAb over the total of tumoural nuclei. The intraobserver agreement and that between IA and LMA for both indices were estimated by the common (kappa(w)) and the jackknife weighted kappa statistic (kappa(w)). The extent of agreement of each considered category was also assessed by means of the category-specific kappa statistics (kappa(cs)). The intraobserver agreement within LMA for NCD and PI and that between IA and LMA for PI were both satisfactory. Upon evaluation of the NCD, the agreement between IA and LMA showed unsatisfactory results, especially when the ratio between the residual tumour cells and the background was critical.

  18. Description of the larva of Amblyomma calcaratum Neumann, 1899 (Acari: Ixodidae) by light and scanning electron microscopy.

    Science.gov (United States)

    Barbieri, Fábio S; Brito, Luciana G; Labruna, Marcelo B; Barros-Battesti, Darci M; Camargo, Luis Marcelo A; Famadas, Kátia M

    2013-12-01

    The larval stage of Amblyomma calcaratum Neumann is described using optical and scanning electron microscopy. Unfed larvae were obtained from a colony of A. calcaratum originating from engorged females collected on Tamandua tetradactyla in the Jaraguá Mountain (23°40'S, 45°44'W), São Paulo County, Brazil. Eleven larvae were prepared and mounted on slides and observed under a light microscope equipped with a drawing tube. Three specimens were prepared for SEM. Several morphological characters are described, including the chaetotaxy of the idiosoma, palpi, and Haller's organ, as well as morphological features of the idiosoma, gnathosoma, and legs of A. calcaratum larvae. In addition, topographical and numerical patterns of integumentary structures on the larval idiosoma are described using a recently proposed nomenclature. On the idiosoma, setaes, lyrifissures, small glands, and large wax glands were found. These structures were observed isolated or associated over the entire idiosoma, except on the scutum, which lacks large wax glands. The topographical and numerical patterns of integumentary structures of the A. calcaratum larva showed only minor differences when compared with patterns of other Amblyomma larvae; however, a few key features can be used to differentiate A. calcaratum from other members of this genus. Copyright © 2013 Elsevier GmbH. All rights reserved.

  19. ON THE IDENTITY OF KARLODINIUM VENEFICUM AND DESCRIPTION OF KARLODINIUM ARMIGER SP. NOV. (DINOPHYCEAE), BASED ON LIGHT AND ELECTRON MICROSCOPY, NUCLEAR-ENCODED LSU RDNA, AND PIGMENT COMPOSITION

    DEFF Research Database (Denmark)

    Bergholtz, Trine; Daugbjerg, Niels; Moestrup, Øjvind

    2006-01-01

    . The culture used by Ballantine to describe Gymnodinium veneficum Ballantine (Plymouth 103) was examined by light and electron microscopy and by partial LSU rDNA. Ultrastructurally, it proved identical to K. micrum (cultures Plymouth 207 and K. Tangen KT-77D, the latter also known as K-0522), and in LSU...

  20. Cryogenics for LHC experiments

    CERN Multimedia

    2001-01-01

    Cryogenic systems will be used by LHC experiments to maximize their performance. Institutes around the world are collaborating with CERN in the construction of these very low temperature systems. The cryogenic test facility in hall 180 for ATLAS magnets. High Energy Physics experiments have frequently adopted cryogenic versions of their apparatus to achieve optimal performance, and those for the LHC will be no exception. The two largest experiments for CERN's new flagship accelerator, ATLAS and CMS, will both use large superconducting magnets operated at 4.5 Kelvin - almost 270 degrees below the freezing point of water. ATLAS also includes calorimeters filled with liquid argon at 87 Kelvin. For the magnets, the choice of a cryogenic version was dictated by a combination economy and transparency to emerging particles. For the calorimeters, liquid argon was selected as the fluid best suited to the experiment's physics requirements. High Energy Physics experiments are the result of worldwide collaborations and...

  1. Advances in Cryogenic Principles

    Science.gov (United States)

    Barron, R. F.

    During the past 50 years, the use of digital computers has significantly influenced the design and analysis of cryogenic systems. At the time when the first Cryogenic Engineering Conference was held, thermodynamic data were presented in graphical or tabular form (the "steam table" format), whereas thermodynamic data for cryogenic system design is computer generated today. The thermal analysis of cryogenic systems in the 1950s involved analytical solutions, graphical solutions, and relatively simple finite-difference approaches. These approaches have been supplanted by finite-element numerical programs which readily solve complicated thermal problems that could not be solved easily using the methods of the 1950s. In distillation column design, the use of the McCabe-Thiele graphical method for determination of the number of theoretical plates has been replaced by numerical methods that allow consideration of several different components in the feed and product streams.

  2. Photomultiplier Tubes at Cryogenic Temperatures

    Science.gov (United States)

    Saunders, Nathan

    2016-09-01

    Liquid noble gas scintillators are widely used in experiments searching for physics beyond the Standard Model. Photomultiplier Tubes (PMTs) working at cryogenic temperatures have been developed as the primary light readout device in those experiments. Three PMTs from Hamamatsu Photonics K.K. (R6041, R11065, and R8520) have been systematically characterized at liquid nitrogen temperature. The high voltage dividing circuits for two of the PMTs were custom-built to make sure there is similar performance at both room and liquid nitrogen temperatures. Their dark count rates at both temperatures were measured. Also measured were their single photoelectron responses at both temperatures using 300, 340, 370, and 420 nm LEDs. The intention is to couple these PMTs directly with inorganic scintillators at liquid nitrogen temperature to achieve high light yeilds for rare-event searches.

  3. Analysis by Light, Scanning, and Transmission Microscopy of the Intima Synovial of the Temporomandibular Joint of Human Fetuses during the Development

    Science.gov (United States)

    Alvez, Carlos Sabu; Carvalho de Moraes, Luis Otavio; Marques, Sergio R.; Tedesco, Roberto C.; Harb, Leandro J. C.; Rodríguez-Vázquez, Jose F.; Mérida-Velasco, Jose R.; Alonso, Luis Garcia

    2014-01-01

    Objective. To characterize morphologically and ultrastructurally using light microscopy, the scanning electron microscopy and transmission electron microscopy the intima synovial of the temporomandibular joint (TMJ) of human fetuses between the 10th and the 38th week of development. Materials and Methods. The TMJ was dissected bilaterally in 37 human fetuses belonging to the Institute of Embryology of the University Complutense of Madrid and of the Federal University of São Paulo. Results. The outcome by light microscopy showed the morphology of the TMJ and that the formation of inferior joint cavity precedes the superior joint cavity and the presence of blood vessels in the synovial. Conclusion. By scanning and transmission electron microscopy we observed the presence of two well-defined cell types in the intima layer of synovial of the TMJ of human fetuses, macrophage-like type A cell and fibroblast-like type B cell, and the presence of the a third cell type, defined by the name of intermediate lining cell in the intima layer of the synovial. PMID:24527214

  4. Visualization of HIV T Cell Virological Synapses and Virus-Containing Compartments by Three-Dimensional Correlative Light and Electron Microscopy

    Science.gov (United States)

    Wang, Lili; Eng, Edward T.; Law, Kenneth; Gordon, Ronald E.; Rice, William J.

    2016-01-01

    ABSTRACT Virological synapses (VS) are adhesive structures that form between infected and uninfected cells to enhance the spread of HIV-1. During T cell VS formation, viral proteins are actively recruited to the site of cell-cell contact where the viral material is efficiently translocated to target cells into heterogeneous, protease-resistant, antibody-inaccessible compartments. Using correlative light and electron microscopy (CLEM), we define the membrane topography of the virus-containing compartments (VCC) where HIV is found following VS-mediated transfer. Focused ion beam scanning electron microscopy (FIB-SEM) and serial sectioning transmission electron microscopy (SS-TEM) were used to better resolve the fluorescent Gag-containing structures within the VCC. We found that small punctate fluorescent signals correlated with single viral particles in enclosed vesicular compartments or surface-localized virus particles and that large fluorescent signals correlated with membranous Gag-containing structures with unknown pathological function. CLEM imaging revealed distinct pools of newly deposited viral proteins within endocytic and nonendocytic compartments in VS target T cells. IMPORTANCE This study directly correlates individual virus-associated objects observed in light microscopy with ultrastructural features seen by electron microscopy in the HIV-1 virological synapse. This approach elucidates which infection-associated ultrastructural features represent bona fide HIV protein complexes. We define the morphology of some HIV cell-to-cell transfer intermediates as true endocytic compartments and resolve unique synapse-associated viral structures created by transfer across virological synapses. PMID:27847357

  5. Observations of grain boundary structures and inclusions in the NEEM ice core by combination of light and scanning electron microscopy

    Science.gov (United States)

    Shigeyama, Wataru; Nagatsuka, Naoko; Homma, Tomoyuki; Takata, Morimasa; Goto-Azuma, Kumiko; Weikusat, Ilka; Drury, Martyn R.; Kuiper, Ernst-Jan N.; Pennock, Gill M.; Mateiu, Ramona V.; Azuma, Nobuhiko; Dahl-Jensen, Dorthe

    2017-04-01

    Dynamics of ice sheets is governed by the flow of the ice and this flow results from the internal deformation of the ice aggregate. The deformation properties of the ice are known to be dependent on several factors, such as microstructure (e.g. crystal grain size and orientation) and impurities. It is well known that ice from glacial periods in ice sheets has a high impurity concentration, and the deformation is reported to be faster than that of non-glacial ice (Faria et al., 2014). However, the mechanisms of the deformation are still not well understood. For a better understanding of ice sheet dynamics, it is a prerequisite to elucidate deformation mechanisms of such impurity-rich ice. The microstructure of a material is a factor that influences mechanical properties and is also an indicator of the dominant deformation mechanisms. The effects of impurities on the deformation and the microstructure depend on chemical compositions, states (viz. insoluble inclusions or soluble ions) and locations of the impurities in the crystal lattice. Therefore, in order to better understand the deformation mechanisms in ice, investigation of relationship between the microstructure and characteristics of the impurities is important. We examined the relationship between grain boundaries and inclusions. Light microscopy (LM) is commonly used to map grain boundary structures on a large area of the ice samples (up to 10 × 10 cm); however, observation of small inclusions NEEM glacial ice (1548 m depth, 19.2 kyr BP). The initial results show inclusions observed by LM formed aggregates of sub-micrometer-sized particles, whose main constituents were silicates. Reference Faria, S. H., I. Weikusat and N. Azuma (2014). The microstructure of polar ice. Part II: State of the art, Journal of Structural Geology 61: 21-49.

  6. Anisotropy of collagen fibre alignment in bovine cartilage: comparison of polarised light microscopy and spatially resolved diffusion-tensor measurements.

    Science.gov (United States)

    de Visser, S K; Bowden, J C; Wentrup-Byrne, E; Rintoul, L; Bostrom, T; Pope, J M; Momot, K I

    2008-06-01

    To compare collagen fibre alignment angles obtained from polarised light microscopy (PLM) and diffusion-tensor imaging (DTI) in bovine articular cartilage. Five samples of bovine articular cartilage from five different animals were studied using magnetic resonance imaging and PLM techniques. T(2)-weighted, diffusion-tensor (DT), and PLM images were acquired for each sample and average depth profiles of the PLM and DTI angles, as well as the banding patterns observed in T(2)-weighted magnetic resonance (MR) images, were compared. Statistical properties of the distributions of the DTI and PLM angles were examined. The samples exhibited a range of alignment morphologies. In the samples with the "conventional" three-zone alignment pattern, a correlation between the PLM and DTI alignment zones and the banding in T(2)-weighted MR images was observed. The shapes of the depth profiles of the PLM and DTI alignment angles were qualitatively similar for each sample. Three samples showed good quantitative correlation between the DT and PLM alignment angles. The correlation between the diffusion and PLM alignment angles was best in the regions of low degree of disorder of fibre alignment. This study provides the first quantitative comparison of DTI of cartilage with the more established PLM techniques. The correlation between alignment angles derived from PLM and DTI data was evident across a wide range of alignment morphologies. The results support the use of DTI for the quantitative measurement of collagen fibre alignment. The microscopic-scale (~10 microm) dispersion of fibre alignment angles appears to be an important factor for understanding the extent of quantitative correlation between PLM and DTI results.

  7. An assessment of the importance ofexposure routes to the uptake and internal localisation of fluorescent nanoparticles in zebrafish (Danio rerio), using light sheet microscopy

    DEFF Research Database (Denmark)

    Skjolding, Lars Michael; Ašmonaitė, G; Jølck, Rasmus Irming

    2017-01-01

    light sheet microscopy (LSM) to define the uptake and localisation of fluorescently labelled nanoparticles in living organisms with minimal sample preparation. Zebrafish (Danio rerio) were exposed to fluorescent gold nanoparticles (Au NPs) and fluorescent polystyrene NPs via aqueous or dietary exposure......A major challenge in nanoecotoxicology is finding suitable methods to determine the uptake and localisation of nanoparticles on a whole-organism level. Some uptake methods have been associated with artefacts induced by sample preparation, including staining for electron microscopy. This study used......, it demonstrates that the localisation of NPs in whole living organisms can be visualised in real-time, using LSM....

  8. Cryogenic Hybrid Magnetic Bearing

    Science.gov (United States)

    Meeks, Crawford R.; Dirusso, Eliseo; Brown, Gerald V.

    1994-01-01

    Cryogenic hybrid magnetic bearing is example of class of magnetic bearings in which permanent magnets and electromagnets used to suspend shafts. Electromagnets provide active control of position of shaft. Bearing operates at temperatures from -320 degrees F (-196 degrees C) to 650 degrees F (343 degrees C); designed for possible use in rocket-engine turbopumps, where effects of cryogenic environment and fluid severely limit lubrication of conventional ball bearings. This and similar bearings also suitable for terrestrial rotating machinery; for example, gas-turbine engines, high-vacuum pumps, canned pumps, precise gimbals that suspend sensors, and pumps that handle corrosive or gritty fluids.

  9. Cryogenic regenerative heat exchangers

    CERN Document Server

    Ackermann, Robert A

    1997-01-01

    An in-depth survey of regenerative heat exchangers, this book chronicles the development and recent commercialization of regenerative devices for cryogenic applications. Chapters cover historical background, concepts, practical applications, design data, and numerical solutions, providing the latest information for engineers to develop advanced cryogenic machines. The discussions include insights into the operation of a regenerator; descriptions of the cyclic and fluid temperature distributions in a regenerator; data for various matrix geometries and materials, including coarse and fine bronze, stainless steel-woven wire mesh screens, and lead spheres; and unique operating features of cryocoolers that produce deviations from ideal regenerator theory.

  10. Identification of Daqingye and Banlangen including crude drugs and decoction dregs from three plant species by normal light and fluorescence microscopy.

    Science.gov (United States)

    Xiaojing, Wan; Liang, Zhitao; Chen, Hu-Biao; Zhao, Zhongzhen; Li, Ping

    2013-08-01

    Daqingye and Banlangen are commonly used Chinese medicinal materials derived from the leaves and roots of Isatis indigotica Fort., respectively, which clinical effects have been confirmed by many studies in recent years. However, many problems have arisen concerning the quality and identity of materials sold in the market under these two names. Thus, the identification of Daqingye and Banlangen has drawn public attention. In this work, transverse sections of Daqingye and Banlangen from I. indigotica Fort. and two easily confused species, namely Baphicacanthus cusia (Nees) Bremek. and Clerodendrum cyrtophyllum Turcz., were investigated with normal light and fluorescence microscopy. The distinguishing features were 7-9 vascular bundles, cystoliths and nonglandular hairs in the leaves of I. indigotica, B. cusia, and C. cyrtophyllum, respectively. The Banlangen could be distinguished according to the characteristics of parenchymous cells, cystoliths, and stone cells. Moreover, the fluorescence features of Daqingye and Banlangen investigated in this study can provide direct points for differentiating those samples. Importantly, whether the crude drugs were decocted could be easily identified by their different fluorescence features, which can ensure their quality in clinical application. This is the first report to distinguish the three species that are commonly found in the market sold as Daqingye and Banlangen by normal light and fluorescence microscopy. This work indicates that the combination of normal light and fluorescence microscopy could be powerful, convenient, and economical for authenticating Daqingye and Banlangen from the three species, including crude drugs and decoction dregs.

  11. Cryogenics Research and Engineering Experience

    Science.gov (United States)

    Toro Medina, Jaime A.

    2013-01-01

    Energy efficient storage, transfer and use of cryogens and cryogenic propellants on Earth and in space have a direct impact on NASA, government and commercial programs. Research and development on thermal insulation, propellant servicing, cryogenic components, material properties and sensing technologies provides industry, government and research institutions with the cross-cutting technologies to manage low-temperature applications. Under the direction of the Cryogenic Testing Lab at Kennedy Space Center, the work experience acquired allowed me to perform research, testing, design and analysis of current and future cryogenic technologies to be applied in several projects.

  12. [Artefacts of confocal microscopy].

    Science.gov (United States)

    Vekshin, N L; Frolov, M S

    2014-01-01

    Typical artefacts caused by using confocal fluorescent microscopy while studying living cells are considered. The role of light scattering, mobility, staining, local concentrations, etc. is discussed.

  13. Seals For Cryogenic Turbomachines

    Science.gov (United States)

    Hendricks, Robert C.; Tam, L. T.; Braun, M. J.; Vlcek, B. L.

    1988-01-01

    Analysis considers effects of seals on stability. Report presents method of calculation of flows of cryogenic fluids through shaft seals. Key to stability is local average velocity in seal. Local average velocity strongly influenced by effects of inlet and outlet and injection of fluid.

  14. High Power Cryogenic Targets

    Energy Technology Data Exchange (ETDEWEB)

    Gregory Smith

    2011-08-01

    The development of high power cryogenic targets for use in parity violating electron scattering has been a crucial ingredient in the success of those experiments. As we chase the precision frontier, the demands and requirements for these targets have grown accordingly. We discuss the state of the art, and describe recent developments and strategies in the design of the next generation of these targets.

  15. Cryogenic current leads

    Energy Technology Data Exchange (ETDEWEB)

    Zizek, F.

    1982-01-01

    Theoretical, technical and design questions are examined of cryogenic current leads for SP of magnetic systems. Simplified mathematical models are presented for the current leads. To illustrate modeling, the calculation is made of the real current leads for 500 A and three variants of current leads for 1500 A for the enterprise ''Shkoda.''

  16. Natural aging behavior of AA7050 Al alloy after cryogenic rolling

    Energy Technology Data Exchange (ETDEWEB)

    Magalhães, Danielle Cristina Camilo, E-mail: danielle_camilo@yahoo.com.br; Hupalo, Marcio Ferreira; Cintho, Osvaldo Mitsuyuki

    2014-01-21

    The effect of cryogenic rolling on the natural aging behavior of a commercial AA7050 aluminum alloy was investigated. Solutionized 10 mm-thick sheets were cryo-rolled to true strains of 0.5, 0.9, 1.1 and 1.4 followed by natural aging (T4 temper) at times ranging from 10 to 1000 h. Light optical microscopy (LOM), scanning electron microscopy (SEM) and differential scanning calorimetry (DSC) were used to follow the microstructural changes upon processing. Mechanical properties were assessed by Vickers hardness measurements and tensile tests. During natural aging the hardness values increased from 100 HV to approximately 145 HV after 100 h. The strength of the undeformed specimen, naturally aged for 100 h, was much higher than that in the as-quenched state. The yield strength (YS) increased from 130 to 375 MPa (188% increase) and the increment of ultimate tensile strength (UTS) was almost 47% (321–470 MPa). A superior combination of mechanical properties was achieved for the specimen cryo-rolled to a true strain of 0.5 followed by natural aging for 1000 h (YS=611 MPa and 15% total elongation). These results suggest that a combination of cryogenic rolling with natural aging is a useful method for achieving optimized mechanical properties for the AA7050 alloy.

  17. Computed tomography of cryogenic cells

    Energy Technology Data Exchange (ETDEWEB)

    Schneider, Gerd; Anderson, E.; Vogt, S.; Knochel, C.; Weiss, D.; LeGros, M.; Larabell, C.

    2001-08-30

    Due to the short wavelengths of X-rays and low numerical aperture of the Fresnel zone plates used as X-ray objectives, the depth of field is several microns. Within the focal depth, imaging a thick specimen is to a good approximation equivalent to projecting the specimen absorption. Therefore, computed tomography based on a tilt series of X-ray microscopic images can be used to reconstruct the local linear absorption coefficient and image the three-dimensional specimen structure. To preserve the structural integrity of biological objects during image acquisition, microscopy is performed at cryogenic temperatures. Tomography based on X-ray microscopic images was applied to study the distribution of male specific lethal 1 (MSL-1), a nuclear protein involved in dosage compensation in Drosophila melanogaster, which ensures that males with single X chromosome have the same amount of most X-linked gene products as females with two X chromosomes. Tomographic reconstructions of X-ray microscopic images were used to compute the local three-dimensional linear absorption coefficient revealing the arrangement of internal structures of Drosophila melanogaster cells. Combined with labelling techniques, nanotomography is a new technique to study the 3D distribution of selected proteins inside whole cells. We want to improve this technique with respect to resolution and specimen preparation. The resolution in the reconstruction can be significantly improved by reducing the angular step size to collect more viewing angles, which requires an automated data acquisition. In addition, fast-freezing with liquid ethane instead of cryogenic He gas will be applied to improve the vitrification of the hydrated samples. We also plan to apply cryo X-ray nanotomography in order to study different types of cells and their nuclear protein distributions.

  18. Minimally invasive ear reshaping with a 1450-nm diode laser using cryogen spray cooling in New Zealand white rabbits.

    Science.gov (United States)

    Holden, Paul K; Chlebicki, Cara; Wong, Brian J F

    2009-01-01

    Otoplasty is the current standard of care for treating prominent ears, a psychologically and sometimes functionally disabling disorder. The technically demanding procedure carries many risks such as poor aesthetic outcome, need for revision surgery, and need for general anesthesia. This study investigates the use of laser irradiation combined with cryogen skin cooling and stenting to reshape cartilage in the ears of New Zealand white rabbits. In this prospective, randomized, internally controlled animal study, the right ears of 9 rabbits were mechanically deformed with a jig and then irradiated with a 1450-nm diode laser combined with cryogen skin cooling (14 J/pulse with cryogen spray for 33 milliseconds per cycle and a 6-mm spot size). The left ear served as the control. The ears were splinted for 1, 3, or 4 weeks. The rabbits were then given a lethal dose of intravenous pentobarbital, and the splints were removed and ears examined and photographed. Light and confocal microscopy were performed on the specimens. Shape change was observed in all 9 treated rabbit ears, while none of the control ears (stenting alone) showed significant change. Qualitatively, reshaped ears were stiffer after 4 weeks of splinting than after 1 or 3 weeks. None of the rabbits showed evidence of skin injury nor did they show signs of postprocedural pain. Findings from histologic analysis in the treated areas showed evidence of an expanded chondrocyte population in the region of laser irradiation, along with some perichondrial thickening and some fibrosis of the deep dermis. Confocal microscopy revealed minimal cellular death at 1 week and none thereafter. Cartilage reshaping using laser energy can be performed safely transcutaneously using cryogen spray cooling in rabbits. This animal model has similarity to human ears with regard to skin and cartilage thickness and is a stepping stone toward developing minimally invasive laser auricle reshaping in humans.

  19. Evaluation of light microscopy and rapid diagnostic test for the detection of malaria under operational field conditions: a household survey in Ethiopia

    Directory of Open Access Journals (Sweden)

    Yohannes Gedeon

    2008-07-01

    Full Text Available Abstract Background In most resource-poor settings, malaria is usually diagnosed based on clinical signs and symptoms and not by detection of parasites in the blood using microscopy or rapid diagnostic tests (RDT. In population-based malaria surveys, accurate diagnosis is important: microscopy provides the gold standard, whilst RDTs allow immediate findings and treatment. The concordance between RDTs and microscopy in low or unstable transmission areas has not been evaluated. Objectives This study aimed to estimate the prevalence of malaria parasites in randomly selected malarious areas of Amhara, Oromia, and Southern Nations, Nationalities and Peoples' (SNNP regions of Ethiopia, using microscopy and RDT, and to investigate the agreement between microscopy and RDT under field conditions. Methods A population-based survey was conducted in 224 randomly selected clusters of 25 households each in Amhara, Oromia and SNNP regions, between December 2006 and February 2007. Fingerpick blood samples from all persons living in even-numbered households were tested using two methods: light microscopy of Giemsa-stained blood slides; and RDT (ParaScreen device for Pan/Pf. Results A total of 13,960 people were eligible for malaria parasite testing of whom 11,504 (82% were included in the analysis. Overall slide positivity rate was 4.1% (95% confidence interval [CI] 3.4–5.0% while ParaScreen RDT was positive in 3.3% (95% CI 2.6–4.1% of those tested. Considering microscopy as the gold standard, ParaScreen RDT exhibited high specificity (98.5%; 95% CI 98.3–98.7 and moderate sensitivity (47.5%; 95% CI 42.8–52.2 with a positive predictive value of 56.8% (95% CI 51.7–61.9 and negative predictive value of 97.6% (95% CI 97.6–98.1% under field conditions. Conclusion Blood slide microscopy remains the preferred option for population-based prevalence surveys of malaria parasitaemia. The level of agreement between microscopy and RDT warrants further

  20. The refrigeration and cryogenic distribution system for the shortpulse x-ray source

    Energy Technology Data Exchange (ETDEWEB)

    Green, Michael A.; Corlett, John N.

    2002-10-20

    This report describes the essential elements of the cryogenic system. The cryogenic distribution system starts at the level of the linac superconducting RF cavities [1] and moves out through the cryogenic piping to the liquid helium refrigeration plant that will be used to cool the RF cavities and the undulator magnets. For this report, the cryogenic distribution system and cryogenic refrigerator includes the following elements: (1) The piping within the linac cryogenic modules will influence the heat transfer through the super-fluid helium from the outer surface of the TESLA niobium cavity and the liquid to gas interface within the horizontal header pipe where the superfluid helium boils. This piping determines the final design of the linac cryogenic module. (2) The acceptable pressure drops determine the supply and return piping dimensions. (3) The helium distribution system is determined by the need to cool down and warm up the various elements in the light source. (4) The size of the cryogenic plant is determined by the heat loads and the probable margin of error on those heat loads. Since the final heat loads are determined by the acceleration gradient in the cavities, a linac with five cryogenic modules will be compared to a linac with only four cryogenic modules. The design assumes that all cryogenic elements in the facility will be cooled using a common cryogenic plant. To minimize vibration effects on the beam lines, this plant is assumed to be located some distance from the synchrotron light beam lines. All of the cryogenic elements in the facility will be attached to the helium refrigeration system through cryogenic transfer lines. The largest single cryogenic load is the main linac, which consists of four or five cryogenic modules depending on the design gradient for the cavities in the linac section. The second largest heat load comes from the cryogenic modules that contain the transverse deflecting RF cavities. The injector linac is the third largest

  1. Light-sheet Bayesian microscopy enables deep-cell super-resolution imaging of heterochromatin in live human embryonic stem cells

    Science.gov (United States)

    Hu, Ying S; Zhu, Quan; Elkins, Keri; Tse, Kevin; Li, Yu; Fitzpatrick, James A J; Verma, Inder M; Cang, Hu

    2016-01-01

    Background Heterochromatin in the nucleus of human embryonic cells plays an important role in the epigenetic regulation of gene expression. The architecture of heterochromatin and its dynamic organization remain elusive because of the lack of fast and high-resolution deep-cell imaging tools. We enable this task by advancing instrumental and algorithmic implementation of the localization-based super-resolution technique. Results We present light-sheet Bayesian super-resolution microscopy (LSBM). We adapt light-sheet illumination for super-resolution imaging by using a novel prism-coupled condenser design to illuminate a thin slice of the nucleus with high signal-to-noise ratio. Coupled with a Bayesian algorithm that resolves overlapping fluorophores from high-density areas, we show, for the first time, nanoscopic features of the heterochromatin structure in both fixed and live human embryonic stem cells. The enhanced temporal resolution allows capturing the dynamic change of heterochromatin with a lateral resolution of 50–60 nm on a time scale of 2.3 s. Conclusion Light-sheet Bayesian microscopy opens up broad new possibilities of probing nanometer-scale nuclear structures and real-time sub-cellular processes and other previously difficult-to-access intracellular regions of living cells at the single-molecule, and single cell level.

  2. Natural enamel caries in polarized light microscopy: differences in histopathological features derived from a qualitative versus a quantitative approach to interpret enamel birefringence.

    Science.gov (United States)

    De Medeiros, R C G; Soares, J D; De Sousa, F B

    2012-05-01

    Lesion area measurement of enamel caries using polarized light microscopy (PLM) is currently performed in a large number of studies, but measurements are based mainly on a mislead qualitative interpretation of enamel birefringence in a single immersion medium. Here, five natural enamel caries lesions are analysed by microradiography and in PLM, and the differences in their histopathological features derived from a qualitative versus a quantitative interpretation of enamel birefringence are described. Enamel birefringence in different immersion media (air, water and quinoline) is interpreted by both qualitative and quantitative approaches, the former leading to an underestimation of the depth of enamel caries mainly when the criterion of validating sound enamel as a negatively birefringent area in immersion in water is used (a current common practice in dental research). Procedures to avoid the shortcomings of a qualitative interpretation of enamel birefringence are presented and discussed. © 2012 The Authors Journal of Microscopy © 2012 Royal Microscopical Society.

  3. Impact of gluten-friendly™ technology on wheat kernel endosperm and gluten protein structure in seeds by light and electron microscopy.

    Science.gov (United States)

    Landriscina, L; D'Agnello, P; Bevilacqua, A; Corbo, M R; Sinigaglia, M; Lamacchia, C

    2017-04-15

    The main aim of this paper was to assess the impact of Gluten-Friendly™ (GF) technology (Italian priority patent n° 102015000084813 filed on 17th December 2015) on wheat kernel endosperm morphology and gluten protein structure, using SEM, light and immunofluorescent microscopy. Microscopy was combined with immunodetection with specific antibodies for gliadins, γ-gliadins, LMW subunits and antigenic epitopes to gain a better understanding of the technology at a molecular level. The results showed significant changes to gluten proteins after GF treatment; cross-reactivity towards the antibodies recognizing almost the entire range of gluten proteins as well as the antigenic epitopes through the sequences QQSF, QQSY, PEQPFPQGC and QQPFP was significantly reduced. The present study confirms the results from our previous work and shows, for the first time, the mechanism by which a chemical-physical treatment abolishes the antigenic capacity of gluten.

  4. Cryogenic treatment of gas

    Science.gov (United States)

    Bravo, Jose Luis [Houston, TX; Harvey, III, Albert Destrehan; Vinegar, Harold J [Bellaire, TX

    2012-04-03

    Systems and methods of treating a gas stream are described. A method of treating a gas stream includes cryogenically separating a first gas stream to form a second gas stream and a third stream. The third stream is cryogenically contacted with a carbon dioxide stream to form a fourth and fifth stream. A majority of the second gas stream includes methane and/or molecular hydrogen. A majority of the third stream includes one or more carbon oxides, hydrocarbons having a carbon number of at least 2, one or more sulfur compounds, or mixtures thereof. A majority of the fourth stream includes one or more of the carbon oxides and hydrocarbons having a carbon number of at least 2. A majority of the fifth stream includes hydrocarbons having a carbon number of at least 3 and one or more of the sulfur compounds.

  5. Cryogenic Control System

    Energy Technology Data Exchange (ETDEWEB)

    Goloborod' ko, S.; /Fermilab

    1989-02-27

    The control system (CS) for the cryogenic arrangement of the DO Liquid Argon Calorimeter consists of a Texas instruments 560/565 Programmable Logical Controller (PLC), two remote bases with Remote Base Controllers and a corresponding set of input/output (I/O) modules, and a PC AST Premium 286 (IBM AT Compatible). The PLC scans a set of inputs and provides a set of outputs based on a ladder logic program and PID control loops. The inputs are logic or analog (current, voltage) signals from equipment status switches or transducers. The outputs are logic or analog (current or voltage) signals for switching solenoids and positioning pneumatic actuators. Programming of the PLC is preformed by using the TISOFT2/560/565 package, which is installed in the PC. The PC communicates to the PLC through a serial RS232 port and provides operator interface to the cryogenic process using Xpresslink software.

  6. Cryogenic treatment of gas

    Energy Technology Data Exchange (ETDEWEB)

    Bravo, Jose Luis [Houston, TX; Harvey, III, Albert Destrehan (Kingwood, TX); Vinegar, Harold J [Bellaire, TX

    2012-04-03

    Systems and methods of treating a gas stream are described. A method of treating a gas stream includes cryogenically separating a first gas stream to form a second gas stream and a third stream. The third stream is cryogenically contacted with a carbon dioxide stream to form a fourth and fifth stream. A majority of the second gas stream includes methane and/or molecular hydrogen. A majority of the third stream includes one or more carbon oxides, hydrocarbons having a carbon number of at least 2, one or more sulfur compounds, or mixtures thereof. A majority of the fourth stream includes one or more of the carbon oxides and hydrocarbons having a carbon number of at least 2. A majority of the fifth stream includes hydrocarbons having a carbon number of at least 3 and one or more of the sulfur compounds.

  7. A correlative study of hydrogen peroxide accumulation after mercury or copper treatment observed in root nodules of Medicago truncatula under light, confocal and electron microscopy.

    Science.gov (United States)

    Górska-Czekaj, Magdalena; Borucki, Wojciech

    2013-01-01

    Heavy metal stress affects both, nodulation and nitrogen fixation of legumes. Mercury triggers disturbances in cellular structure and metabolism but its influence on ROS generation is poorly understood. Copper is redox active metal which in opposition to mercury is an essential micronutrient for plants. Excess of copper is cytotoxic, as it participates in ROS generation via Fenton-type reaction. The present work describes changes in hydrogen peroxide (H₂O₂) accumulation in response to monthly stress caused by mercury (6 mg/L HgCl₂) or copper (60 mg/L CuCl₂) in root nodules. H₂O₂ accumulation viewed with a light microscopy was detected by the use of diaminobenzidine (DAB). 2',7'-Dichlorofluorescein diacetate (H2DCF-DA) was used as a probe for the intracellular localization of H₂O₂ with a confocal laser scanning system. H₂O₂ detection under transmission electron microscopy was performed by the use of cerium method. Histochemical localization and light and confocal microscopy investigations revealed that under Hg or Cu treatments distinct amount of H₂O₂ accumulated mainly in the interzone and nitrogen-fixing zone. Under normal conditions H₂O₂ accumulated predominantly in the interzone. Electron microscopy observations showed H₂O₂ accumulation under Hg or Cu- treatments around peribacteroid membranes of mature symbiosomes located within nitrogen-fixing zone. It should be underlined that under normal conditions H₂O₂ was not detected at the peribacteroid membranes. The main result of our observations is increased accumulation of H₂O₂ in response to mercury and copper treatments at the peribacteroidal membranes, to our knowledge shown for the first time. Therefore, our results revealed that an overproduction of H₂O₂ in response to copper or mercury-treatment may account for lowering of nitrogen fixation rates in heavy-metal affected root nodules. Copyright © 2013 Elsevier Ltd. All rights reserved.

  8. Cryogenic cooling for high power laser amplifiers

    Directory of Open Access Journals (Sweden)

    Perin J.P.

    2013-11-01

    Full Text Available Using DPSSL (Diode Pumped Solid State Lasers as pumping technology, PW-class lasers with enhanced repetition rates are developed. Each of the Yb YAG amplifiers will be diode-pumped at a wavelength of 940 nm. This is a prerequisite for achieving high repetition rates (light amplification duration 1 millisecond and repetition rate 10 Hz. The efficiency of DPSSL is inversely proportional to the temperature, for this reason the slab amplifier have to be cooled at a temperature in the range of 100 K–170 K with a heat flux of 1 MW*m−2. This paper describes the thermo-mechanical analysis for the design of the amplification laser head, presents a preliminary proposal for the required cryogenic cooling system and finally outlines the gain of cryogenic operation for the efficiency of high pulsed laser.

  9. Cryogenic Selective Surfaces

    Science.gov (United States)

    Youngquist, Robert; Nurge, Mark; Gibson, Tracy; Johnson, Wesley

    2017-01-01

    The NASA Innovative Advanced Concept (NIAC) program has been funding work at KSC on a novel coating that should allow cryogenic materials to be stored in deep space. The NIAC Symposium will be the last week of September and it is a requirement that the funded material be presented both orally and at a poster session. This DAA submission is requesting approval to go public with both the presentation and the poster.

  10. Cryogenic Test Technology 1984.

    Science.gov (United States)

    1985-04-01

    aircraft configuration Pathfinder II (Figure 16) made of Vascomax 200, a set of six bodies of revolution (Figure 17) made from 6061 aluminium alloy, a...iron and aluminium alloys appear to be viable candidates. AS loads increase the number of avail- able alloys is severely constrained by toughness...using A-286 screws in four steels and one aluminium alloy. In the absence of loads cryogenic cycling gene- rally produced decreases in breakaway

  11. LEDs for fluorescence microscopy

    NARCIS (Netherlands)

    Young, I.T.; Garini, Y.; Dietrich, H.R.C.; Van Oel, W.; Liqui Lung, G.

    2004-01-01

    Traditional light sources for fluorescence microscopy have been mercury lamps, xenon lamps, and lasers. These sources have been essential in the development of fluorescence microscopy but each can have serious disadvantages: lack of near monochromaticity, heat generation, cost, lifetime of the light

  12. Lighting

    Data.gov (United States)

    Federal Laboratory Consortium — Lighting Systems Test Facilities aid research that improves the energy efficiency of lighting systems. • Gonio-Photometer: Measures illuminance from each portion of...

  13. Study of Collagen Birefringence in Different Grades of Oral Squamous Cell Carcinoma Using Picrosirius Red and Polarized Light Microscopy

    OpenAIRE

    2015-01-01

    Objectives. The present study was done to evaluate birefringence pattern of collagen fibres in different grades of oral squamous cell carcinoma using Picrosirius red stain and polarization microscopy and to determine if there is a change in collagen fibres between different grades of oral squamous cell carcinoma. Materials and Methods. Picrosirius red stained 5 μm thick sections of previously diagnosed different grades of squamous cell carcinoma and normal oral mucosa were studied under polar...

  14. Proteasome Particle-Rich Structures Are Widely Present in Human Epithelial Neoplasms: Correlative Light, Confocal and Electron Microscopy Study

    OpenAIRE

    2011-01-01

    A novel cytoplasmic structure has been recently characterized by confocal and electron microscopy in H. pylori-infected human gastric epithelium, as an accumulation of barrel-like proteasome reactive particles colocalized with polyubiquitinated proteins, H. pylori toxins and the NOD1 receptor. This proteasome particle-rich cytoplasmic structure (PaCS), a sort of focal proteasome hyperplasia, was also detected in dysplastic cells and was found to be enriched in SHP2 and ERK proteins, known to ...

  15. Advances in Helium Cryogenics

    Science.gov (United States)

    Sciver, S. W. Van

    This review provides a survey of major advances that have occurred in recent years in the area of helium cryogenics. Helium-temperature cryogenics is the enabling technology for a substantial and growing number of low-temperature systems from superconducting magnets to space-based experimental facilities. In recent years there have been many advances in the technology of low-temperature helium, driven mostly by new applications. However, to keep the review from being too broad, this presentation focuses mainly on three of the most significant advances. These are: (1) the development of large-scale recuperative refrigeration systems mainly for superconducting magnet applications in accelerators and other research facilities; (2) the use of stored superfluid helium (He II) as a coolant for spacebased astrophysics experiments; and (3) the application of regenerative cryocoolers operating at liquid helium temperatures primarily for cooling superconducting devices. In each case, the reader should observe that critical technologies were developed to facilitate these applications. In addition to these three primary advances, other significant helium cryogenic technologies are briefly reviewed at the end of this chapter, along with some vision for future developments in these areas.

  16. INFLUENCE OF FILM STRUCTURE AND LIGHT ON CHARGE TRAPPING AND DISSIPATION DYNAMICS IN SPUN-CAST ORGANIC THIN-FILM TRANSISTORS MEASURED BY SCANNING KELVIN PROBE MICROSCOPY

    Energy Technology Data Exchange (ETDEWEB)

    Teague, L.; Moth, M.; Anthony, J.

    2012-05-03

    Herein, time-dependent scanning Kelvin probe microscopy of solution processed organic thin film transistors (OTFTs) reveals a correlation between film microstructure and OTFT device performance with the location of trapped charge within the device channel. The accumulation of the observed trapped charge is concurrent with the decrease in I{sub SD} during operation (V{sub G}=-40 V, V{sub SD}= -10 V). We discuss the charge trapping and dissipation dynamics as they relate to the film structure and show that application of light quickly dissipates the observed trapped charge.

  17. Non-invasive red light optogenetic pacing and optical coherence microscopy (OCM) imaging for drosophila melanogaster (Conference Presentation)

    Science.gov (United States)

    Men, Jing; Li, Airong; Jerwick, Jason; Tanzi, Rudolph E.; Zhou, Chao

    2017-02-01

    Cardiac pacing could be a powerful tool for investigating mammalian cardiac electrical conduction systems as well as for treatment of certain cardiac pathologies. However, traditional electrical pacing using pacemaker requires an invasive surgical procedure. Electrical currents from the implanted electrodes can also cause damage to heart tissue, further restricting its utility. Optogenetic pacing has been developed as a promising, non-invasive alternative to electrical stimulation for controlling animal heart rhythms. It induces heart contractions by shining pulsed light on transgene-generated microbial opsins, which in turn activate the light gated ion channels in animal hearts. However, commonly used opsins in optogenetic pacing, such as channelrhodopsin-2 (ChR2), require short light wavelength stimulation (475 nm), which is strongly absorbed and scattered by tissue. Here, we performed optogenetic pacing by expression of recently engineered red-shifted microbial opsins, ReaChR and CsChrimson, in a well-established animal model, Drosophila melanogaster, using the 617 nm stimulation light pulses. The OCM technique enables non-invasive optical imaging of animal hearts with high speed and ultrahigh axial and transverse resolutions. We integrated a customized OCM system with the optical stimulation system to monitor the optogenetic pacing noninvasively. The use of red-sifted opsins enabled deeper penetration of simulating light at lower power, which is promising for applications of optogenetic pacing in mammalian cardiac pathology studies or clinical treatments in the future.

  18. Near-field light detection of a photo induced force by atomic force microscopy with frequency modulation

    Science.gov (United States)

    Satoh, Nobuo; Kobayashi, Kei; Matsushige, Kazumi; Yamada, Hirofumi

    2017-08-01

    We demonstrated near-field light detection using a non contact-mode atomic force microscope (nc-AFM). This system obtains molecular-level resolution by reducing noise in the displacement detection of a Si cantilever. The Si cantilever probe tip was brought close to a glass with a patterned chromium film on a dove prism. The backside of the prism was irradiated by an intensity-modulated laser light to create an evanescent field at the glass surface. We obtained a near-field optical image of the chromium-patterned glass by detecting the amplitude modulation induced by the near-field light while the tip-sample distance was regulated by the frequency modulation method under atmospheric conditions.

  19. Light

    DEFF Research Database (Denmark)

    Prescott, N.B.; Kristensen, Helle Halkjær; Wathes, C.M.

    2004-01-01

    This chapter presents the effect of artificial light environments (light levels, colour, photoperiod and flicker) on the welfare of broilers in terms of vision, behaviour, lameness and mortality......This chapter presents the effect of artificial light environments (light levels, colour, photoperiod and flicker) on the welfare of broilers in terms of vision, behaviour, lameness and mortality...

  20. Direct imaging of light elements by annular dark-field aberration-corrected scanning transmission electron microscopy

    Energy Technology Data Exchange (ETDEWEB)

    Lotnyk, Andriy, E-mail: andriy.lotnyk@iom-leipzig.de; Poppitz, David; Gerlach, Jürgen W.; Rauschenbach, Bernd [Leibniz Institute of Surface Modification, Permoserstr. 15, D-04318 Leipzig (Germany)

    2014-02-17

    In this report, we show that an annular dark-field detector in an aberration-corrected scanning transmission electron microscope allows the direct observation of light element columns in crystalline lattices. At specific imaging conditions, an enhancement of the intensities of light element columns in the presence of heavy element columns is observed. Experimental results are presented for imaging the nitrogen and carbon atomic columns at the GaN-SiC interface and within the GaN and SiC compounds. The crystal polarity of GaN at the interface is identified. The obtained findings are discussed and are well supported by image simulations.

  1. Aerogel Insulation to Support Cryogenic Technologies Project

    Data.gov (United States)

    National Aeronautics and Space Administration — NASA is seeking a high performance thermal insulation material for cryogenic applications in space launch development. Many of the components in cryogenic...

  2. Cryogenic Propellant Storage and Transfer Project

    Data.gov (United States)

    National Aeronautics and Space Administration — The Cryogenic Propellant Storage and Transfer project will demonstrate the capability to safely and efficiently store, transfer and measure cryogenic propellants,...

  3. Direct characterization of ultraviolet-light-induced refractive index structures by scanning near-field optical microscopy

    DEFF Research Database (Denmark)

    Svalgaard, Mikael; Madsen, S.; Hvam, Jørn Märcher;

    1998-01-01

    We have applied a reflection scanning near-field optical microscope to directly probe ultraviolet (UV)-light-induced refractive index structures in planar glass samples. This technique permits direct comparison between topography and refractive index changes (10(-5)-10(-3)) with submicrometer...

  4. A cryogenic optical feedthrough using polarization maintaining fibers.

    Science.gov (United States)

    Nelson, M J; Collins, C J; Speake, C C

    2016-03-01

    Polarization maintaining optical fibers can be used to transmit linearly polarized light over long distances but their use in cryogenic environments has been limited by their sensitivity to temperature changes and associated mechanical stress. We investigate experimentally how thermal stresses affect the polarization maintaining fibers and model the observations with Jones matrices. We describe the design, construction, and testing of a feedthrough and fiber termination assembly that uses polarization maintaining fiber to transmit light from a 633 nm HeNe laser at room temperature to a homodyne polarization-based interferometer in a cryogenic vacuum. We report on the efficiency of the polarization maintaining properties of the feedthrough assembly. We also report that, at cryogenic temperatures, the interferometer can achieve a sensitivity of 8 × 10(-10) rad/√Hz at 0.05 Hz using this feedthrough.

  5. Quantitative phase imaging of biological cells and tissues using singleshot white light interference microscopy and phase subtraction method for extended range of measurement

    Science.gov (United States)

    Mehta, Dalip Singh; Sharma, Anuradha; Dubey, Vishesh; Singh, Veena; Ahmad, Azeem

    2016-03-01

    We present a single-shot white light interference microscopy for the quantitative phase imaging (QPI) of biological cells and tissues. A common path white light interference microscope is developed and colorful white light interferogram is recorded by three-chip color CCD camera. The recorded white light interferogram is decomposed into the red, green and blue color wavelength component interferograms and processed it to find out the RI for different color wavelengths. The decomposed interferograms are analyzed using local model fitting (LMF)" algorithm developed for reconstructing the phase map from single interferogram. LMF is slightly off-axis interferometric QPI method which is a single-shot method that employs only a single image, so it is fast and accurate. The present method is very useful for dynamic process where path-length changes at millisecond level. From the single interferogram a wavelength-dependent quantitative phase imaging of human red blood cells (RBCs) are reconstructed and refractive index is determined. The LMF algorithm is simple to implement and is efficient in computation. The results are compared with the conventional phase shifting interferometry and Hilbert transform techniques.

  6. Visualization of HIV T Cell Virological Synapses and Virus-Containing Compartments by Three-Dimensional Correlative Light and Electron Microscopy.

    Science.gov (United States)

    Wang, Lili; Eng, Edward T; Law, Kenneth; Gordon, Ronald E; Rice, William J; Chen, Benjamin K

    2017-01-15

    Virological synapses (VS) are adhesive structures that form between infected and uninfected cells to enhance the spread of HIV-1. During T cell VS formation, viral proteins are actively recruited to the site of cell-cell contact where the viral material is efficiently translocated to target cells into heterogeneous, protease-resistant, antibody-inaccessible compartments. Using correlative light and electron microscopy (CLEM), we define the membrane topography of the virus-containing compartments (VCC) where HIV is found following VS-mediated transfer. Focused ion beam scanning electron microscopy (FIB-SEM) and serial sectioning transmission electron microscopy (SS-TEM) were used to better resolve the fluorescent Gag-containing structures within the VCC. We found that small punctate fluorescent signals correlated with single viral particles in enclosed vesicular compartments or surface-localized virus particles and that large fluorescent signals correlated with membranous Gag-containing structures with unknown pathological function. CLEM imaging revealed distinct pools of newly deposited viral proteins within endocytic and nonendocytic compartments in VS target T cells.

  7. Accuracy of Mobile Phone and Handheld Light Microscopy for the Diagnosis of Schistosomiasis and Intestinal Protozoa Infections in Côte d'Ivoire.

    Science.gov (United States)

    Coulibaly, Jean T; Ouattara, Mamadou; D'Ambrosio, Michael V; Fletcher, Daniel A; Keiser, Jennifer; Utzinger, Jürg; N'Goran, Eliézer K; Andrews, Jason R; Bogoch, Isaac I

    2016-06-01

    Handheld light microscopy using compact optics and mobile phones may improve the quality of health care in resource-constrained settings by enabling access to prompt and accurate diagnosis. Laboratory technicians were trained to operate two handheld diagnostic devices (Newton Nm1 microscope and a clip-on version of the mobile phone-based CellScope). The accuracy of these devices was compared to conventional light microscopy for the diagnosis of Schistosoma haematobium, S. mansoni, and intestinal protozoa infection in a community-based survey in rural Côte d'Ivoire. One slide of 10 ml filtered urine and a single Kato-Katz thick smear from 226 individuals were subjected to the Newton Nm1 microscope and CellScope for detection of Schistosoma eggs and compared to conventional microscopy. Additionally, 121 sodium acetate-acetic acid-formalin (SAF)-fixed stool samples were examined by the Newton Nm1 microscope and compared to conventional microscopy for the diagnosis of intestinal protozoa. The prevalence of S. haematobium, S. mansoni, Giardia intestinalis, and Entamoeba histolytica/E. dispar, as determined by conventional microscopy, was 39.8%, 5.3%, 20.7%, and 4.9%, respectively. The Newton Nm1 microscope had diagnostic sensitivities for S. mansoni and S. haematobium infection of 91.7% (95% confidence interval (CI) 59.8-99.6%) and 81.1% (95% CI 71.2-88.3%), respectively, and specificities of 99.5% (95% CI 97.0-100%) and 97.1% (95% CI 92.2-99.1%), respectively. The CellScope demonstrated sensitivities for S. mansoni and S. haematobium of 50.0% (95% CI 25.4-74.6%) and 35.6% (95% CI 25.9-46.4%), respectively, and specificities of 99.5% (95% CI 97.0-100%) and 100% (95% CI 86.7-100%), respectively. For G. intestinalis and E. histolytica/E. dispar, the Newton Nm1 microscope had sensitivity of 84.0% (95% CI 63.1-94.7%) and 83.3% (95% CI 36.5-99.1%), respectively, and 100% specificity. Handheld diagnostic devices can be employed in community-based surveys in resource

  8. Accuracy of Mobile Phone and Handheld Light Microscopy for the Diagnosis of Schistosomiasis and Intestinal Protozoa Infections in Cote d'Ivoire.

    Directory of Open Access Journals (Sweden)

    Jean T Coulibaly

    2016-06-01

    Full Text Available Handheld light microscopy using compact optics and mobile phones may improve the quality of health care in resource-constrained settings by enabling access to prompt and accurate diagnosis.Laboratory technicians were trained to operate two handheld diagnostic devices (Newton Nm1 microscope and a clip-on version of the mobile phone-based CellScope. The accuracy of these devices was compared to conventional light microscopy for the diagnosis of Schistosoma haematobium, S. mansoni, and intestinal protozoa infection in a community-based survey in rural Côte d'Ivoire. One slide of 10 ml filtered urine and a single Kato-Katz thick smear from 226 individuals were subjected to the Newton Nm1 microscope and CellScope for detection of Schistosoma eggs and compared to conventional microscopy. Additionally, 121 sodium acetate-acetic acid-formalin (SAF-fixed stool samples were examined by the Newton Nm1 microscope and compared to conventional microscopy for the diagnosis of intestinal protozoa.The prevalence of S. haematobium, S. mansoni, Giardia intestinalis, and Entamoeba histolytica/E. dispar, as determined by conventional microscopy, was 39.8%, 5.3%, 20.7%, and 4.9%, respectively. The Newton Nm1 microscope had diagnostic sensitivities for S. mansoni and S. haematobium infection of 91.7% (95% confidence interval (CI 59.8-99.6% and 81.1% (95% CI 71.2-88.3%, respectively, and specificities of 99.5% (95% CI 97.0-100% and 97.1% (95% CI 92.2-99.1%, respectively. The CellScope demonstrated sensitivities for S. mansoni and S. haematobium of 50.0% (95% CI 25.4-74.6% and 35.6% (95% CI 25.9-46.4%, respectively, and specificities of 99.5% (95% CI 97.0-100% and 100% (95% CI 86.7-100%, respectively. For G. intestinalis and E. histolytica/E. dispar, the Newton Nm1 microscope had sensitivity of 84.0% (95% CI 63.1-94.7% and 83.3% (95% CI 36.5-99.1%, respectively, and 100% specificity.Handheld diagnostic devices can be employed in community-based surveys in resource

  9. Accuracy of Mobile Phone and Handheld Light Microscopy for the Diagnosis of Schistosomiasis and Intestinal Protozoa Infections in Côte d’Ivoire

    Science.gov (United States)

    Coulibaly, Jean T.; Ouattara, Mamadou; D’Ambrosio, Michael V.; Fletcher, Daniel A.; Keiser, Jennifer; Utzinger, Jürg; N’Goran, Eliézer K.

    2016-01-01

    Background Handheld light microscopy using compact optics and mobile phones may improve the quality of health care in resource-constrained settings by enabling access to prompt and accurate diagnosis. Methodology Laboratory technicians were trained to operate two handheld diagnostic devices (Newton Nm1 microscope and a clip-on version of the mobile phone-based CellScope). The accuracy of these devices was compared to conventional light microscopy for the diagnosis of Schistosoma haematobium, S. mansoni, and intestinal protozoa infection in a community-based survey in rural Côte d’Ivoire. One slide of 10 ml filtered urine and a single Kato-Katz thick smear from 226 individuals were subjected to the Newton Nm1 microscope and CellScope for detection of Schistosoma eggs and compared to conventional microscopy. Additionally, 121 sodium acetate-acetic acid-formalin (SAF)-fixed stool samples were examined by the Newton Nm1 microscope and compared to conventional microscopy for the diagnosis of intestinal protozoa. Principal Findings The prevalence of S. haematobium, S. mansoni, Giardia intestinalis, and Entamoeba histolytica/E. dispar, as determined by conventional microscopy, was 39.8%, 5.3%, 20.7%, and 4.9%, respectively. The Newton Nm1 microscope had diagnostic sensitivities for S. mansoni and S. haematobium infection of 91.7% (95% confidence interval (CI) 59.8–99.6%) and 81.1% (95% CI 71.2–88.3%), respectively, and specificities of 99.5% (95% CI 97.0–100%) and 97.1% (95% CI 92.2–99.1%), respectively. The CellScope demonstrated sensitivities for S. mansoni and S. haematobium of 50.0% (95% CI 25.4–74.6%) and 35.6% (95% CI 25.9–46.4%), respectively, and specificities of 99.5% (95% CI 97.0–100%) and 100% (95% CI 86.7–100%), respectively. For G. intestinalis and E. histolytica/E. dispar, the Newton Nm1 microscope had sensitivity of 84.0% (95% CI 63.1–94.7%) and 83.3% (95% CI 36.5–99.1%), respectively, and 100% specificity. Conclusions

  10. Study of Collagen Birefringence in Different Grades of Oral Squamous Cell Carcinoma Using Picrosirius Red and Polarized Light Microscopy

    Directory of Open Access Journals (Sweden)

    Pillai Arun Gopinathan

    2015-01-01

    Full Text Available Objectives. The present study was done to evaluate birefringence pattern of collagen fibres in different grades of oral squamous cell carcinoma using Picrosirius red stain and polarization microscopy and to determine if there is a change in collagen fibres between different grades of oral squamous cell carcinoma. Materials and Methods. Picrosirius red stained 5 μm thick sections of previously diagnosed different grades of squamous cell carcinoma and normal oral mucosa were studied under polarization microscopy for arrangement as well as birefringence of collagen fibres around tumour islands. Results. It was found that thin collagen fibres increased and thick collagen fibres decreased with dedifferentiation of OSCC (P<0.0001 . It was observed that there was change in polarization colours of thick fibres from yellowish orange to greenish yellow with dedifferentiation of OSCC indicating loosely packed fibres (P<0.0001. Conclusion. There was a gradual change of birefringence of collagen from yellowish orange to greenish yellow from well to poorly differentiated squamous cell carcinoma, indicating that there is a change from mature form of collagen to immature form as tumour progresses. Studying collagen fibres with Picrosirius red for stromal changes around tumour islands along with routine staining may help in predicting the prognosis of tumour.

  11. Correlative near-infrared light and cathodoluminescence microscopy using Y2O3:Ln, Yb (Ln = Tm, Er) nanophosphors for multiscale, multicolour bioimaging

    Science.gov (United States)

    Fukushima, S.; Furukawa, T.; Niioka, H.; Ichimiya, M.; Sannomiya, T.; Tanaka, N.; Onoshima, D.; Yukawa, H.; Baba, Y.; Ashida, M.; Miyake, J.; Araki, T.; Hashimoto, M.

    2016-05-01

    This paper presents a new correlative bioimaging technique using Y2O3:Tm, Yb and Y2O3:Er, Yb nanophosphors (NPs) as imaging probes that emit luminescence excited by both near-infrared (NIR) light and an electron beam. Under 980 nm NIR light irradiation, the Y2O3:Tm, Yb and Y2O3:Er, Yb NPs emitted NIR luminescence (NIRL) around 810 nm and 1530 nm, respectively, and cathodoluminescence at 455 nm and 660 nm under excitation of accelerated electrons, respectively. Multimodalities of the NPs were confirmed in correlative NIRL/CL imaging and their locations were visualized at the same observation area in both NIRL and CL images. Using CL microscopy, the NPs were visualized at the single-particle level and with multicolour. Multiscale NIRL/CL bioimaging was demonstrated through in vivo and in vitro NIRL deep-tissue observations, cellular NIRL imaging, and high-spatial resolution CL imaging of the NPs inside cells. The location of a cell sheet transplanted onto the back muscle fascia of a hairy rat was visualized through NIRL imaging of the Y2O3:Er, Yb NPs. Accurate positions of cells through the thickness (1.5 mm) of a tissue phantom were detected by NIRL from the Y2O3:Tm, Yb NPs. Further, locations of the two types of NPs inside cells were observed using CL microscopy.

  12. Combining light microscopy, dielectric spectroscopy, MALDI intact cell mass spectrometry, FTIR spectromicroscopy and multivariate data mining for morphological and physiological bioprocess characterization of filamentous organisms.

    Science.gov (United States)

    Posch, Andreas E; Koch, Cosima; Helmel, Michaela; Marchetti-Deschmann, Martina; Macfelda, Karin; Lendl, Bernhard; Allmaier, Günter; Herwig, Christoph

    2013-02-01

    Along with productivity and physiology, morphological growth behavior is the key parameter in bioprocess design for filamentous fungi. Lacking tools for fast, reliable and efficient analysis however, fungal morphology is still commonly tackled by empirical trial-and-error techniques during strain selection and process development procedures. Bridging the gap, this work presents a comprehensive analytical approach for morphological analysis combining automated high-throughput microscopy, multi-frequency dielectric spectroscopy, MALDI intact cell mass spectrometry and FTIR spectromicroscopy. Industrial fed-batch production processes were investigated in fully instrumented, automated bioreactors using the model system Penicillium chrysogenum. Physiological process characterization was based on the determination of specific conversion rates as scale-independent parameters. Conventional light microscopic morphological analysis was based on holistic determination of time series for more than 30 morphological parameters and their frequency distributions over the respective parameter range by automated high-throughput light microscopy. Characteristic protein patterns enriched in specific morphological and physiological states were further obtained by MALDI intact cell mass spectrometry. Spatial resolution of molecular biomass composition was facilitated by FTIR spectromicroscopy. Real-time in situ monitoring of morphological process behavior was achieved by linking multi-frequency dielectric spectroscopy with above outlined off-line methods. Data integration of complementing orthogonal techniques for morphological and physiological analysis together with multivariate modeling of interdependencies between morphology, physiology and process parameters facilitated complete bioprocess characterization. The suggested approach will thus help understanding morphological and physiological behavior and, in turn, allow to control and optimize those complex processes.

  13. Masked rhodamine dyes of five principal colors revealed by photolysis of a 2-diazo-1-indanone caging group: synthesis, photophysics, and light microscopy applications.

    Science.gov (United States)

    Belov, Vladimir N; Mitronova, Gyuzel Yu; Bossi, Mariano L; Boyarskiy, Vadim P; Hebisch, Elke; Geisler, Claudia; Kolmakov, Kirill; Wurm, Christian A; Willig, Katrin I; Hell, Stefan W

    2014-10-06

    Caged rhodamine dyes (Rhodamines NN) of five basic colors were synthesized and used as "hidden" markers in subdiffractional and conventional light microscopy. These masked fluorophores with a 2-diazo-1-indanone group can be irreversibly photoactivated, either by irradiation with UV- or violet light (one-photon process), or by exposure to intense red light (λ∼750 nm; two-photon mode). All dyes possess a very small 2-diazoketone caging group incorporated into the 2-diazo-1-indanone residue with a quaternary carbon atom (C-3) and a spiro-9H-xanthene fragment. Initially they are non-colored (pale yellow), non-fluorescent, and absorb at λ=330-350 nm (molar extinction coefficient (ε)≈10(4)  M(-1)  cm(-1)) with a band edge that extends to about λ=440 nm. The absorption and emission bands of the uncaged derivatives are tunable over a wide range (λ=511-633 and 525-653 nm, respectively). The unmasked dyes are highly colored and fluorescent (ε=3-8×10(4)  M(-1)  cm(-1) and fluorescence quantum yields (ϕ)=40-85% in the unbound state and in methanol). By stepwise and orthogonal protection of carboxylic and sulfonic acid groups a highly water-soluble caged red-emitting dye with two sulfonic acid residues was prepared. Rhodamines NN were decorated with amino-reactive N-hydroxysuccinimidyl ester groups, applied in aqueous buffers, easily conjugated with proteins, and readily photoactivated (uncaged) with λ=375-420 nm light or intense red light (λ=775 nm). Protein conjugates with optimal degrees of labeling (3-6) were prepared and uncaged with λ=405 nm light in aqueous buffer solutions (ϕ=20-38%). The photochemical cleavage of the masking group generates only molecular nitrogen. Some 10-40% of the non-fluorescent (dark) byproducts are also formed. However, they have low absorbance and do not quench the fluorescence of the uncaged dyes. Photoactivation of the individual molecules of Rhodamines NN (e.g., due to reversible or irreversible

  14. Realization and performance of cryogenic selection mechanisms

    Science.gov (United States)

    Aitink-Kroes, Gabby; Bettonvil, Felix; Kragt, Jan; Elswijk, Eddy; Tromp, Niels

    2014-07-01

    Within Infra-Red large wavelength bandwidth instruments the use of mechanisms for selection of observation modes, filters, dispersing elements, pinholes or slits is inevitable. The cryogenic operating environment poses several challenges to these cryogenic mechanisms; like differential thermal shrinkage, physical property change of materials, limited use of lubrication, high feature density, limited space etc. MATISSE the mid-infrared interferometric spectrograph and imager for ESO's VLT interferometer (VLTI) at Paranal in Chile coherently combines the light from 4 telescopes. Within the Cold Optics Bench (COB) of MATISSE two concepts of selection mechanisms can be distinguished based on the same design principles: linear selection mechanisms (sliders) and rotating selection mechanisms (wheels).Both sliders and wheels are used at a temperature of 38 Kelvin. The selection mechanisms have to provide high accuracy and repeatability. The sliders/wheels have integrated tracks that run on small, accurately located, spring loaded precision bearings. Special indents are used for selection of the slider/wheel position. For maximum accuracy/repeatability the guiding/selection system is separated from the actuation in this case a cryogenic actuator inside the cryostat. The paper discusses the detailed design of the mechanisms and the final realization for the MATISSE COB. Limited lifetime and performance tests determine accuracy, warm and cold and the reliability/wear during life of the instrument. The test results and further improvements to the mechanisms are discussed.

  15. Image formation by linear and nonlinear digital scanned light-sheet fluorescence microscopy with Gaussian and Bessel beam profiles

    Science.gov (United States)

    Olarte, Omar E.; Licea-Rodriguez, Jacob; Palero, Jonathan A.; Gualda, Emilio J.; Artigas, David; Mayer, Jürgen; Swoger, Jim; Sharpe, James; Rocha-Mendoza, Israel; Rangel-Rojo, Raul; Loza-Alvarez, Pablo

    2012-01-01

    We present the implementation of a combined digital scanned light-sheet microscope (DSLM) able to work in the linear and nonlinear regimes under either Gaussian or Bessel beam excitation schemes. A complete characterization of the setup is performed and a comparison of the performance of each DSLM imaging modality is presented using in vivo Caenorhabditis elegans samples. We found that the use of Bessel beam nonlinear excitation results in better image contrast over a wider field of view. PMID:22808423

  16. Quantitative full-colour transmitted light microscopy and dyes for concentration mapping and measurement of diffusion coefficients in microfluidic architectures

    OpenAIRE

    Werts, Martinus H. V.; Raimbault, Vincent; Texier-Picard, Rozenn; Poizat, Rémi; Français, Olivier; Griscom, Laurent; Navarro, Julien R. G.

    2012-01-01

    International audience; A simple and versatile methodology has been developed for the simultaneous measurement of multiple concentration profiles of colourants in transparent microfluidic systems, using a conventional transmitted light microscope, a digital colour (RGB) camera and numerical image processing combined with multicomponent analysis. Rigorous application of the Beer-Lambert law would require monochromatic probe conditions, but in spite of the broad spectral bandwidths of the three...

  17. Glycoproteins and skin-core structure in Nephila clavipes spider silk observed by light and electron microscopy.

    Science.gov (United States)

    Augsten, K; Mühlig, P; Herrmann, C

    2000-01-01

    Microscopical imaging of natural, unstressed draglines or of untreated bulk samples showed two types or threads with diameters of either approximately 1-2 microm or 4-5 microm, which could be identified as products of the minor or major ampullate glands. The threads had a circular profile in serial cross sections and are surrounded by a thin outer layer of a different material within the section. Such fibrillar configurations were also found in untreated threads or in the same serial sections of transmission electron microscopy (TEM) samples by means of the special technique of laser scanning microscopy. In TEM slides, numerous cavities with the same circular profile were detectable, and the length of these cavities is variable from 40-300 nm. The threads are oriented parallel and twisted around themselves to construct a double thread. In the interface between the two single threads, bridge-like structures are prominent. The single untreated thread consists of cylindrical fibers with a diameter of approximately 1-1.5 microm. Apparently more than eight fibers are within a thread and each fiber is composed of a great number of fibrils with a diameter of about 150 nm. The surface of threads is coated with a characteristic layer approximately 150-250 nm thick that contains glycoproteins. These were demonstrated for the first time by labeling with concanavalin A lectin-gold complex and are dependent on the diameter and length of the thread. The same substances could also be detected inside the single thread. The skin can be removed completely or partially by mechanical treatment, or by washing with phosphate-buffered saline or trypsin.

  18. Addressable, large-field second harmonic generation microscopy based on 2D acousto-optical deflector and spatial light modulator.

    Science.gov (United States)

    Shao, Yonghong; Liu, Honghai; Qin, Wan; Qu, Junle; Peng, Xiang; Niu, Hanben; Gao, Bruce Z

    2012-09-01

    We present an addressable, large-field second harmonic generation microscope by combining a 2D acousto-optical deflector with a spatial light modulator. The SLM shapes an incoming mode-locked, near-infrared Ti:Sapphire laser beam into a multifocus array, which can be rapidly scanned by changing the incident angle of the laser beam using a 2D acousto-optical deflector. Compared to the single-beam-scan technique, the multifocus array scan can increase the scanning rate and the field-of-view size with the multi-region imaging ability.

  19. Effects of radiation upon the light-sensing elements of the retina as characterized by scanning electron microscopy

    Science.gov (United States)

    Malachowski, M. J.; Tobias, C. A.; Leith, J. T.

    1977-01-01

    A model system using Necturus maculosus, the common mudpuppy, was established for evaluating effects of radiation upon the light-sensing elements of the retina. Accelerated heavy ions of helium and neon from the Berkeley Bevalac were used. A number of criteria were chosen to characterize radiation damage by observing morphological changes with the scanning electron microscope. The studies indicated retina sensitivity to high-LET (neon) particles at radiation levels below 10 rads (7 particles per visual element) whereas no significant effects were seen from fast helium ions below 50 rads.

  20. Effects of radiation upon the light-sensing elements of the retina as characterized by scanning electron microscopy

    Science.gov (United States)

    Malachowski, M. J.; Tobias, C. A.; Leith, J. T.

    1977-01-01

    A model system using Necturus maculosus, the common mudpuppy, was established for evaluating effects of radiation upon the light-sensing elements of the retina. Accelerated heavy ions of helium and neon from the Berkeley Bevalac were used. A number of criteria were chosen to characterize radiation damage by observing morphological changes with the scanning electron microscope. The studies indicated retina sensitivity to high-LET (neon) particles at radiation levels below 10 rads (7 particles per visual element) whereas no significant effects were seen from fast helium ions below 50 rads.

  1. Cryogenic VPH gratings for the CELT/TMT

    Science.gov (United States)

    Blais-Ouellette, Sebastien; Guzman, Dani; Elgamil, Amal; Rallison, Richard

    2004-09-01

    Characterization of Volume Phase Holographic gratings at cryogenic temperatures have been conducted using a new test facility at Caltech. The new test bench includes a cryostat that allows large angles for incident and diffracted light. Gratings under tests are shielded from thermal background, and precisely and uniformly temperature controlled. Preliminary results are presented and show little temperature dependence of the efficiency function.

  2. Cryogenic Piezoelectric Actuator

    Science.gov (United States)

    Jiang, Xiaoning; Cook, William B.; Hackenberger, Wesley S.

    2009-01-01

    In this paper, PMN-PT single crystal piezoelectric stack actuators and flextensional actuators were designed, prototyped and characterized for space optics applications. Single crystal stack actuators with footprint of 10 mm x10 mm and the height of 50 mm were assembled using 10 mm x10mm x0.15mm PMN-PT plates. These actuators showed stroke > 65 - 85 microns at 150 V at room temperature, and > 30 microns stroke at 77 K. Flextensional actuators with dimension of 10mm x 5 mm x 7.6 mm showed stroke of >50 microns at room temperature at driving voltage of 150 V. A flextensional stack actuator with dimension of 10 mm x 5 mm x 47 mm showed stroke of approx. 285 microns at 150 V at room temperature and > 100 microns at 77K under driving of 150 V should be expected. The large cryogenic stroke and high precision of these actuators are promising for cryogenic optics applications.

  3. Photoswitching of Green mEos2 by Intense 561 nm Light Perturbs Efficient Green-to-Red Photoconversion in Localization Microscopy.

    Science.gov (United States)

    Thédié, Daniel; Berardozzi, Romain; Adam, Virgile; Bourgeois, Dominique

    2017-09-05

    Green-to-red photoconvertible fluorescent proteins (PCFPs) such as mEos2 and its derivatives are widely used in PhotoActivated Localization Microscopy (PALM). However, the complex photophysics of these genetically encoded markers complicates the quantitative analysis of PALM data. Here, we show that intense 561 nm light (∼1 kW/cm(2)) typically used to localize single red molecules considerably affects the green-state photophysics of mEos2 by populating at least two reversible dark states. These dark states retard green-to-red photoconversion through a shelving effect, although one of them is rapidly depopulated by 405 nm light illumination. Multiple mEos2 switching and irreversible photobleaching is thus induced by yellow/green and violet photons before green-to-red photoconversion occurs, contributing to explain the apparent limited signaling efficiency of this PCFP. Our data reveals that the photophysics of PCFPs of anthozoan origin is substantially more complex than previously thought, and suggests that intense 561 nm laser light should be used with care, notably for quantitative or fast PALM approaches.

  4. The internal structure of Early Cambrian fossil embryo Olivooides revealed in the light of Synchrotron X-ray Tomographic Microscopy

    Institute of Scientific and Technical Information of China (English)

    CHEN Fang; DONG XiPing

    2008-01-01

    Countless fossil embryo Olivooides and the hatched larvae, juveniles and adults (the latter two kinds are Punctatus) are recovered by means of acid maceration from the fine-crystalline to medium-crystalline phosphatic limestone and phosphatic micrite of Early Cambrian Kuanchuanpu Formation at the Shizhonggou section, near Kuanchuanpu Village, Ningqiang County, Shaanxi Province, China. Using the technique of Synchrotron X-ray Tomographic Microscopy, the 3D internal structure of Olivooides and Punctatus is reconstructed. The morphological and statistic analyses are also given to the stellae structure of Olivooides and Punctatus, which indicates that this structure is a result of adaptive evolution to a lifestyle of fast-attaching after hatching, probably with the function of mucilage secretion. The internal structure of Punctatus is described and discussed. The ovum-like structure, a common internal feature of Punctatus, is considered as the taphonomic structure, rather than eggs or other biological structure. This structure is thought to be formed after the burial of the animal and before or during the mineralization. The original internal structure of Punctatus is assumed to be tabulae-filled, with soft body grown on them.

  5. EFFECTS OF CHEMICAL PROCESSING AND OXIDE ETHYLENE STERILIZATION ON CORTICAL AND CANCELLOUS RAT BONE: A LIGHT AND ELECTRON SCANNING MICROSCOPY STUDY.

    Science.gov (United States)

    Castiglia, Marcello Teixeira; da Silva, Juliano Voltarelli F; Frezarim Thomazini, José Armendir; Volpon, José Batista

    2009-01-01

    To evaluate, under microscopic examination, the structural changes displayed by the trabecular and cortical bones after being processed chemically and sterilized by ethylene oxide. Samples of cancellous and cortical bones obtained from young female albinus rats (Wistar) were assigned to four groups according to the type of treatment: Group I- drying; Group II- drying and ethylene oxide sterilization; III- chemical treatment; IV- chemical treatment and ethylene oxide sterilization. Half of this material was analyzed under ordinary light microscope and the other half using scanning electron microscopy. In all the samples, regardless the group, there was good preservation of the general morphology. For samples submitted to the chemical processing there was better preservation of the cellular content, whereas there was amalgamation of the fibres when ethylene oxide was used. Treatment with ethylene oxide caused amalgamation of the fibers, possibly because of heating and the chemical treatment contributed to a better cellular preservation of the osseous structure.

  6. Investigating biofilm structure developing on carriers from lab-scale moving bed biofilm reactors based on light microscopy and optical coherence tomography.

    Science.gov (United States)

    Li, Chunyan; Felz, Simon; Wagner, Michael; Lackner, Susanne; Horn, Harald

    2016-01-01

    This study focused on characterizing the structure of biofilms developed on carriers used in lab-scale moving bed biofilm reactors. Both light microscopy (2D) and optical coherence tomography (OCT) were employed to track the biofilm development on carriers of different geometry and under different aeration rates. Biofilm structure was further characterized with respect to average biofilm thickness, biofilm growth velocity, biomass volume, compartment filling degree, surface area, etc. The results showed that carriers with a smaller compartment size stimulated a quick establishment of biofilms. Low aeration rates favored fast development of biofilms. Comparison between the results derived from 2D and 3D images revealed comparable results with respect to average biofilm thickness and compartment filling degree before the carrier compartments were fully willed with biomass. However, 3D imaging with OCT was capable of visualizing and quantifying the heterogeneous structure of biofilms, which cannot be achieved using 2D imaging.

  7. Droplet-based light-sheet fluorescence microscopy for high-throughput sample preparation, 3-D imaging and quantitative analysis on a chip.

    Science.gov (United States)

    Jiang, Hao; Zhu, Tingting; Zhang, Hao; Nie, Jun; Guan, Zeyi; Ho, Chi-Ming; Liu, Sheng; Fei, Peng

    2017-06-27

    We report a novel fusion of droplet microfluidics and light-sheet microscopy, to achieve high-throughput sample compartmentalization, manipulation and three-dimensional imaging on a chip. This optofluidic device characterized by orthogonal plane illumination and rapid liquid handling is compact and cost-effective, and capable of preparing sample droplets with tunable size, frequency and ingredient. Each droplet flowing through the device's imaging region is self-scanned by a laser-sheet, three-dimensionally reconstructed and quantitatively analysed. This simple-and-robust platform combines fast 3-D imaging with efficient sample preparation and eliminates the need of a complicated mechanical scan at the same time. Achieving 500 measurements per second and screening over 30 samples per minute, it shows great potential for various lab-on-a-chip biological studies, such as embryo sorting and cell growth assays.

  8. Grain boundary atomic structures and light-element visualization in ceramics: combination of Cs-corrected scanning transmission electron microscopy and first-principles calculations.

    Science.gov (United States)

    Ikuhara, Yuichi

    2011-01-01

    Grain boundaries and interfaces of crystals have peculiar electronic structures, caused by the disorder in periodicity, providing the functional properties, which cannot be observed in a perfect crystal. In the vicinity of the grain boundaries and interfaces, dopants or impurities are often segregated, and they play a crucial role in deciding the properties of a material. Spherical aberration (Cs)-corrected scanning transmission electron microscopy (STEM), allowing the formation of sub-angstrom-sized electron probes, can directly observe grain boundary-segregated dopants. On the other hand, ceramic materials are composed of light elements, and these light elements also play an important role in the properties of ceramic materials. Recently, annular bright-field (ABF)-STEM imaging has been proposed, which is now known to be a very powerful technique in producing images showing both light- and heavy-element columns simultaneously. In this review, the atomic structure determination of ceramic grain boundaries and direct observation of grain boundary-segregated dopants and light elements in ceramics were shown to combine with the theoretical calculations. Examples are demonstrated for well-defined grain boundaries in rare earth-doped Al(2)O(3) and ZnO ceramics, CeO(2) and SrTiO(3) grain boundary, lithium battery materials and metal hydride, which were characterized by Cs-corrected high-angle annular dark-field and ABF-STEM. It is concluded that the combination of STEM characterization and first-principles calculation is very useful in interpreting the structural information and in understanding the origin of the properties in various ceramics.

  9. Mapping the subcellular distribution of α-synuclein in neurons using genetically encoded probes for correlated light and electron microscopy: implications for Parkinson's disease pathogenesis.

    Science.gov (United States)

    Boassa, Daniela; Berlanga, Monica L; Yang, Mary Ann; Terada, Masako; Hu, Junru; Bushong, Eric A; Hwang, Minju; Masliah, Eliezer; George, Julia M; Ellisman, Mark H

    2013-02-06

    Modifications to the gene encoding human α-synuclein have been linked to the development of Parkinson's disease. The highly conserved structure of α-synuclein suggests a functional interaction with membranes, and several lines of evidence point to a role in vesicle-related processes within nerve terminals. Using recombinant fusions of human α-synuclein, including new genetic tags developed for correlated light microscopy and electron microscopy (the tetracysteine-biarsenical labeling system or the new fluorescent protein for electron microscopy, MiniSOG), we determined the distribution of α-synuclein when overexpressed in primary neurons at supramolecular and cellular scales in three dimensions (3D). We observed specific association of α-synuclein with a large and otherwise poorly characterized membranous organelle system of the presynaptic terminal, as well as with smaller vesicular structures within these boutons. Furthermore, α-synuclein was localized to multiple elements of the protein degradation pathway, including multivesicular bodies in the axons and lysosomes within neuronal cell bodies. Examination of synapses in brains of transgenic mice overexpressing human α-synuclein revealed alterations of the presynaptic endomembrane systems similar to our findings in cell culture. Three-dimensional electron tomographic analysis of enlarged presynaptic terminals in several brain areas revealed that these terminals were filled with membrane-bounded organelles, including tubulovesicular structures similar to what we observed in vitro. We propose that α-synuclein overexpression is associated with hypertrophy of membrane systems of the presynaptic terminal previously shown to have a role in vesicle recycling. Our data support the conclusion that α-synuclein is involved in processes associated with the sorting, channeling, packaging, and transport of synaptic material destined for degradation.

  10. Time-lapse 3-D measurements of a glucose biosensor in multicellular spheroids by light sheet fluorescence microscopy in commercial 96-well plates

    Science.gov (United States)

    Maioli, Vincent; Chennell, George; Sparks, Hugh; Lana, Tobia; Kumar, Sunil; Carling, David; Sardini, Alessandro; Dunsby, Chris

    2016-11-01

    Light sheet fluorescence microscopy has previously been demonstrated on a commercially available inverted fluorescence microscope frame using the method of oblique plane microscopy (OPM). In this paper, OPM is adapted to allow time-lapse 3-D imaging of 3-D biological cultures in commercially available glass-bottomed 96-well plates using a stage-scanning OPM approach (ssOPM). Time-lapse 3-D imaging of multicellular spheroids expressing a glucose Förster resonance energy transfer (FRET) biosensor is demonstrated in 16 fields of view with image acquisition at 10 minute intervals. As a proof-of-principle, the ssOPM system is also used to acquire a dose response curve with the concentration of glucose in the culture medium being varied across 42 wells of a 96-well plate with the whole acquisition taking 9 min. The 3-D image data enable the FRET ratio to be measured as a function of distance from the surface of the spheroid. Overall, the results demonstrate the capability of the OPM system to measure spatio-temporal changes in FRET ratio in 3-D in multicellular spheroids over time in a multi-well plate format.

  11. Qualitative histologic evaluation of the tissue reaction to the polyurethane resin (ricinus communis - based biopolymer implantation assessed by light and scanning electron microscopy

    Directory of Open Access Journals (Sweden)

    Gustavo Campos Belmonte

    2013-01-01

    Full Text Available The tissue reaction of bone tissue accessed by light microscopy and scanning electron microscopy (SEM images after polyurethane resin implantation is presented in this study. Twenty four male rabbits were used, divided into two groups of 12 animals each (experimental group and control group in which full-thickness cranial defect was surgically created. At 30 and 90 days post operation 6 animals of each group were euthanized and bone samples were removed for analysis. The microscopic results indicated no inflammatory foreign body reaction, a perfect union between the polymer and surgical bone bed surface, lack of bone resorption and presence of a thin layer of osteogenic material covering the polymer surface in contact with the surgical bone bed. The SEM images demonstrate the porosity of the resin, with diameters from 120 to 500 µm. This important feature of this polymer is associated with its osteoconductivity, allowing the bone growth inside it, improving the integration between the material and bone tissue. These results confirm that polyurethane resin derived from Ricinuscommunis is an excellent bone substitute for use in repair surgery for great bone losses.

  12. Comparison of Morphometric Aspects of Light and Electron Microscopy of the Hypoglossal Nerve between Young and Aged Male Wistar Rats

    Directory of Open Access Journals (Sweden)

    Mohsen Pourghasem

    2012-01-01

    Full Text Available Objective: Age-related changes occur in many different systems of the body. Many elderlypeople show dysphagia and dysphonia. This research was conducted to evaluatequantitatively the morphometrical changes of the hypoglossal nerve resulting from theaging process in young and aged rats.Materials and Methods: Through an experimental study ten male wistar rats (4 months: 5rats, 24 months: 5 rats were selected randomly from a colony of wistars in the UWC. Aftera fixation process and preparation of samples of the cervical portion of the hypoglossalnerve of these rats, light and electron microscopic imaging were performed. These imageswere evaluated according to the numbers and size of myelinated nerve fibers, nucleoli ofSchwann cells, myelin sheath thickness, axon diameter, and g ratio. All data were analyzedby Mann-Whitney, a non-parametric statistical test.Results: In light microscope, numbers of myelinated nerve fibers, the mean entire nerveperimeters, the mean entire nerve areas and the mean entire nerve diameters in youngand aged rats’ were not significantly different between the two groups.In electron microscope, numbers of myelinated axons, numbers of Schwann cell nucleoliand the mean g ratios of myelinated axon to Schwann cell in young and aged rats werenot significantly different. The myelinated fiber diameters, the myelin sheath thicknesses,myelinated axon diameters and the mean g ratio of axon diameter to myelinated fiberdiameter in young and aged fibers were significantly differentConclusion: The mean g ratio of myelinated nerve fibers of peripheral nerves stabilizes atthe level of 0.6 after maturation and persists without major change during adulthood. Thisratio of axon diameter to fiber diameter (0.6 is optimum for normal conduction velocity ofneural impulses. Our study indicated that the g ratio of myelinated nerve fiber of the hypoglossalnerve decreased prominently in aged rats and can be a cause of impairment innerve function in

  13. Three-dimensional image cytometer based on widefield structured light microscopy and high-speed remote depth scanning.

    Science.gov (United States)

    Choi, Heejin; Wadduwage, Dushan N; Tu, Ting Yuan; Matsudaira, Paul; So, Peter T C

    2015-01-01

    A high throughput 3D image cytometer have been developed that improves imaging speed by an order of magnitude over current technologies. This imaging speed improvement was realized by combining several key components. First, a depth-resolved image can be rapidly generated using a structured light reconstruction algorithm that requires only two wide field images, one with uniform illumination and the other with structured illumination. Second, depth scanning is implemented using the high speed remote depth scanning. Finally, the large field of view, high NA objective lens and the high pixelation, high frame rate sCMOS camera enable high resolution, high sensitivity imaging of a large cell population. This system can image at 800 cell/sec in 3D at submicron resolution corresponding to imaging 1 million cells in 20 min. The statistical accuracy of this instrument is verified by quantitatively measuring rare cell populations with ratio ranging from 1:1 to 1:10(5) . © 2014 International Society for Advancement of Cytometry.

  14. Quantitative full-colour transmitted light microscopy and dyes for concentration mapping and measurement of diffusion coefficients in microfluidic architectures.

    Science.gov (United States)

    Werts, Martinus H V; Raimbault, Vincent; Texier-Picard, Rozenn; Poizat, Rémi; Français, Olivier; Griscom, Laurent; Navarro, Julien R G

    2012-02-21

    A simple and versatile methodology has been developed for the simultaneous measurement of multiple concentration profiles of colourants in transparent microfluidic systems, using a conventional transmitted light microscope, a digital colour (RGB) camera and numerical image processing combined with multicomponent analysis. Rigorous application of the Beer-Lambert law would require monochromatic probe conditions, but in spite of the broad spectral bandwidths of the three colour channels of the camera, a linear relation between the measured optical density and dye concentration is established under certain conditions. An optimised collection of dye solutions for the quantitative optical microscopic characterisation of microfluidic devices is proposed. Using the methodology for optical concentration measurement we then implement and validate a simplified and robust method for the microfluidic measurement of diffusion coefficients using an H-filter architecture. It consists of measuring the ratio of the concentrations of the two output channels of the H-filter. It enables facile determination of the diffusion coefficient, even for non-fluorescent molecules and nanoparticles, and is compatible with non-optical detection of the analyte.

  15. Nanodielectrics for Cryogenic Applications

    Energy Technology Data Exchange (ETDEWEB)

    Tuncer, Enis [ORNL; Sauers, Isidor [ORNL; James, David Randy [ORNL; Ellis, Alvin R [ORNL; Pace, Marshall O [ORNL; More, Karren [Oak Ridge National Laboratory (ORNL); Sathyamurthy, Srivatsan [University of Tennessee, Knoxville (UTK); Woodward, Jonathan [ORNL; Rondinone, Adam Justin [ORNL

    2009-01-01

    In this paper we report the recent advances in nanodielectrics that were developed and tested for cryogenic dielectric applications. The systems studied are composed of nanometer size particles. Particles were produced using either an ex-situ or in-situ technique. It is observed that there are clear differences in the structural properties of materials produced using these two approaches. Either no significant degradation or improvement in the electrical insulation properties were observed for ex-situ nano-particle samples processed with an ultrasonic processor and in-situ nano-particle samples. Nanodielectrics have the potential to be tailored with better thermal and mechanical properties without losing their electrical insulation characteristics.

  16. Cryogenic Cam Butterfly Valve

    Science.gov (United States)

    McCormack, Kenneth J. (Inventor)

    2016-01-01

    A cryogenic cam butterfly valve has a body that includes an axially extending fluid conduit formed there through. A disc lug is connected to a back side of a valve disc and has a circular bore that receives and is larger than a cam of a cam shaft. The valve disc is rotatable for a quarter turn within the body about a lug axis that is offset from the shaft axis. Actuating the cam shaft in the closing rotational direction first causes the camming side of the cam of the cam shaft to rotate the disc lug and the valve disc a quarter turn from the open position to the closed position. Further actuating causes the camming side of the cam shaft to translate the valve disc into sealed contact with the valve seat. Opening rotational direction of the cam shaft reverses these motions.

  17. Cryogenic Tracking Detectors

    CERN Multimedia

    Luukka, P R; Tuominen, E M; Mikuz, M

    2002-01-01

    The recent advances in Si and diamond detector technology give hope of a simple solution to the radiation hardness problem for vertex trackers at the LHC. In particular, we have recently demonstrated that operating a heavily irradiated Si detector at liquid nitrogen (LN$_2$) temperature results in significant recovery of Charge Collection Efficiency (CCE). Among other potential benefits of operation at cryogenic temperatures are the use of large low-resistivity wafers, simple processing, higher and faster electrical signal because of higher mobility and drift velocity of carriers, and lower noise of the readout circuit. A substantial reduction in sensor cost could result The first goal of the approved extension of the RD39 program is to demonstrate that irradiation at low temperature in situ during operation does not affect the results obtained so far by cooling detectors which were irradiated at room temperature. In particular we shall concentrate on processes and materials that could significantly reduce th...

  18. Report on the first VLHC photon stop cryogenic design experiment

    CERN Document Server

    Geynisman, M; Bossert, R; Darve, C; Ewald, K D; Klebaner, A; Limon, P; Martínez, A

    2004-01-01

    As part of Fermilab's study of a Very Large Hadron Collider (VLHC), a water-cooled photon stop was proposed as a device to intercept the synchrotron radiation emitted by the high-energy proton beams in the high-field superconducting magnets with minimal plug-cooling power. Photon stops are radiation absorbers operating at room temperature that protrude into the beam tube at the end of each bending magnet to scrape the synchrotron light emitted by the beam one magnet up- stream. Among the technological challenges regarding photon stops is their cryo-design. The photon stop is water-cooled and operates in a cryogenic environment. A careful cryo-design is therefore essential to enable operation at minimum heat transfer between the room temperature sections and the cryogenic parts. A photon stop cryo- design was developed and a prototype was built. This paper presents the results of the cryogenic experiments conducted on the first VLHC photon-stop prototype.

  19. Light

    CERN Document Server

    Robertson, William C

    2003-01-01

    Why is left right and right left in the mirror? Baffled by the basics of reflection and refraction? Wondering just how the eye works? If you have trouble teaching concepts about light that you don t fully grasp yourself, get help from a book that s both scientifically accurate and entertaining with Light. By combining clear explanations, clever drawings, and activities that use easy-to-find materials, this book covers what science teachers and parents need to know to teach about light with confidence. It uses ray, wave, and particle models of light to explain the basics of reflection and refraction, optical instruments, polarization of light, and interference and diffraction. There s also an entire chapter on how the eye works. Each chapter ends with a Summary and Applications section that reinforces concepts with everyday examples. Whether you need a deeper understanding of how light bends or a good explanation of why the sky is blue, you ll find Light more illuminating and accessible than a college textbook...

  20. Combining gas-phase electrophoretic mobility molecular analysis (GEMMA), light scattering, field flow fractionation and cryo electron microscopy in a multidimensional approach to characterize liposomal carrier vesicles.

    Science.gov (United States)

    Urey, Carlos; Weiss, Victor U; Gondikas, Andreas; von der Kammer, Frank; Hofmann, Thilo; Marchetti-Deschmann, Martina; Allmaier, Günter; Marko-Varga, György; Andersson, Roland

    2016-11-20

    For drug delivery, characterization of liposomes regarding size, particle number concentrations, occurrence of low-sized liposome artefacts and drug encapsulation are of importance to understand their pharmacodynamic properties. In our study, we aimed to demonstrate the applicability of nano Electrospray Gas-Phase Electrophoretic Mobility Molecular Analyser (nES GEMMA) as a suitable technique for analyzing these parameters. We measured number-based particle concentrations, identified differences in size between nominally identical liposomal samples, and detected the presence of low-diameter material which yielded bimodal particle size distributions. Subsequently, we compared these findings to dynamic light scattering (DLS) data and results from light scattering experiments coupled to Asymmetric Flow-Field Flow Fractionation (AF4), the latter improving the detectability of smaller particles in polydisperse samples due to a size separation step prior detection. However, the bimodal size distribution could not be detected due to method inherent limitations. In contrast, cryo transmission electron microscopy corroborated nES GEMMA results. Hence, gas-phase electrophoresis proved to be a versatile tool for liposome characterization as it could analyze both vesicle size and size distribution. Finally, a correlation of nES GEMMA results with cell viability experiments was carried out to demonstrate the importance of liposome batch-to-batch control as low-sized sample components possibly impact cell viability. Copyright © 2016 The Authors. Published by Elsevier B.V. All rights reserved.

  1. Spatially Resolved Imaging on Photocarrier Generations and Band Alignments at Perovskite/PbI2 Heterointerfaces of Perovskite Solar Cells by Light-Modulated Scanning Tunneling Microscopy.

    Science.gov (United States)

    Shih, Min-Chuan; Li, Shao-Sian; Hsieh, Cheng-Hua; Wang, Ying-Chiao; Yang, Hung-Duen; Chiu, Ya-Ping; Chang, Chia-Seng; Chen, Chun-Wei

    2017-02-08

    The presence of the PbI2 passivation layers at perovskite crystal grains has been found to considerably affect the charge carrier transport behaviors and device performance of perovskite solar cells. This work demonstrates the application of a novel light-modulated scanning tunneling microscopy (LM-STM) technique to reveal the interfacial electronic structures at the heterointerfaces between CH3NH3PbI3 perovskite crystals and PbI2 passivation layers of individual perovskite grains under light illumination. Most importantly, this technique enabled the first observation of spatially resolved mapping images of photoinduced interfacial band bending of valence bands and conduction bands and the photogenerated electron and hole carriers at the heterointerfaces of perovskite crystal grains. By systematically exploring the interfacial electronic structures of individual perovskite grains, enhanced charge separation and reduced back recombination were observed when an optimal design of interfacial PbI2 passivation layers consisting of a thickness less than 20 nm at perovskite crystal grains was applied.

  2. A Cryogenic Flow Sensor Project

    Data.gov (United States)

    National Aeronautics and Space Administration — Based on the success of the phase I effort, Advanced Technologies Group, Inc. proposes the development of a Cryogenic Flow Sensor (CFS) for determining mass flow of...

  3. Cryogenic Acoustic Suppression Testing Project

    Data.gov (United States)

    National Aeronautics and Space Administration — The proposed project will explore and test the feasibility and effectiveness of using a cryogenic fluid (liquid nitrogen) to facilitate acoustic suppression in a...

  4. Cryogenic MEMS Pressure Sensor Project

    Data.gov (United States)

    National Aeronautics and Space Administration — A directly immersible cryogenic MEMS pressure sensor will be developed. Each silicon die will contain a vacuum-reference and a tent-like membrane. Offsetting thermal...

  5. Lightweight Inflatable Cryogenic Tank Project

    Data.gov (United States)

    National Aeronautics and Space Administration — This proposal describes the development of an inflatable and lightweight polymer-fabric structured pressure vessel designed for the containment of cryogenic fluids....

  6. Cryogenic Systems and Superconductive Power

    Science.gov (United States)

    The report defines, investigates, and experimentally evaluates the key elements of a representative crogenic turborefrigerator subsystem suitable for providing reliable long-lived cryogenic refrigeration for a superconductive ship propulsion system.

  7. Light microscopy can reveal the consumption of a mixture of psychotropic plant and fungal material in suspicious death.

    Science.gov (United States)

    Wiltshire, Patricia E J; Hawksworth, David L; Edwards, Kevin J

    2015-08-01

    Light microscopical examination of plant and fungal remains in the post mortem gut may be capable of demonstrating the ingestion of unexpected natural psychotropic materials. This is demonstrated here in a case in which a 'shaman' was accused of causing the death of a young man. The deceased had participated in a ceremony which involved the drinking of ayahuasca in order to induce a psychotropic experience. Ayahuasca is an infusion of Banisteriopsis caapi (ayahuasca vine), which produces a monoamine oxidase inhibitor, and one or more additional tropical plants, generally Psychotria viridis (chacruna) which produces dimethyltryptamine (DMT). The monoamine oxidase inhibitor prevents DMT from being broken down in the gut, so enabling its passage into the bloodstream and across the blood/brain barrier. Toxicological tests for DMT demonstrated the presence of this compound in the body. The deceased was reported to be in the habit of using Psilocybe semilanceata (liberty cap). This fungus (popularly called magic mushroom) contains psilocybin which is hydrolysed in the gut to psilocin; this compound mimics a serotonin uptake inhibitor, and also invokes psychotropic experiences. Microscopical examination established that the ileum and colon contained spores of Psilocybe and, in addition, pollen of Cannabis sativa and seeds of Papaver cf. somniferum (opium poppy). Both the plant species yield psychotropic substances. Palynological and mycological analysis of containers from the deceased person's dwelling also yielded abundant trace evidence of pertinent pollen and spores. The police had requested analysis for DMT but there was no screening for other psychotropic substances. Investigators were surprised that a mixture of hallucinogenic materials had been consumed by the deceased. The charge was modified from manslaughter to possession of a 'Class A' drug as the deceased had been consuming psychotropic substances not administered by the 'shaman'. Where death involving drugs

  8. Fluorescence antibunching microscopy

    CERN Document Server

    Schwartz, Osip

    2011-01-01

    Breaking the diffraction limit in microscopy by utilizing quantum properties of light has been the goal of intense research in the recent years. We propose a quantum superresolution technique based on non-classical emission statistics of fluorescent markers, routinely used as contrast labels for bio-imaging. The technique can be readily implemented using standard fluorescence microscopy equipment.

  9. A Piezoelectric Cryogenic Heat Switch

    Science.gov (United States)

    Jahromi, Amir E.; Sullivan, Dan F.

    2014-01-01

    We have measured the thermal conductance of a mechanical heat switch actuated by a piezoelectric positioner, the PZHS (PieZo electric Heat Switch), at cryogenic temperatures. The thermal conductance of the PZHS was measured between 4 K and 10 K, and on/off conductance ratios greater than 100 were achieved when the positioner applied its maximum force of 8 N. We discuss the advantages of using this system in cryogenic applications, and estimate the ultimate performance of an optimized PZHS.

  10. Observations of short-range, high-LET recoil tracks in CR-39 plastic nuclear track detector by visible light microscopy

    Energy Technology Data Exchange (ETDEWEB)

    Benton, E.R., E-mail: eric.benton@okstate.ed [Dept. of Physics, Oklahoma State University, 1110 S. Innovation Way, 100, Stillwater, OK 74078 (United States); Johnson, C.E. [Los Alamos National Laboratory, Los Alamos, NM 87545 (United States); DeWitt, J. [Dept. of Physics, Oklahoma State University, 1110 S. Innovation Way, 100, Stillwater, OK 74078 (United States); Yasuda, N. [National Institute of Radiological Sciences, Chiba (Japan); Benton, E.V. [Dept. of Physics, University of San Francisco, San Francisco, CA 94117 (United States); Moyers, M.H. [Proton Therapy, Inc., Colton, CA 92324 (United States); Frank, A.L. [Dept. of Physics, University of San Francisco, San Francisco, CA 94117 (United States)

    2011-05-15

    Using standard visible light microscopy, we are able to observe particle tracks produced by <10 {mu}m range target fragment recoils in CR-39 plastic nuclear track detector (PNTD) following short chemical etching (bulk etch B {<=}1 {mu}m). In accelerator irradiations, targets of varying composition, including a number of elemental targets of high Z, were exposed in contact with layers of CR-39 PNTD to beams of 60 MeV, 230 MeV, and 1 GeV protons at doses of 10-50 Gy. Chemical etching of CR-39 under standard conditions (50 {sup o}C, 6.25 N NaOH) for 2-4 h (removed layer B = 0.5-1.0 {mu}m) yielded secondary track densities of 10{sup 5}-10{sup 6} cm{sup -2} observable under a standard optical microscope with 500x-800x magnification. Ordinarily such a short duration etch would not be expected to enlarge the tracks sufficiently for them to be resolved by visible light optics. However, due to the short-range of the particles, a longer chemical processing would have over-etched the tracks until they were no longer recognizable. The tracks we observe in CR-39 PNTD irradiated in these experiments are the result of residual heavy recoil fragments returning to equilibrium via evaporation processes following proton-induced knock out of light particles via preequilibrium processes. Because the heavy recoil particles are very near the end of their ranges (i.e. in the Bragg peak), their LET is extremely high and changes rapidly. Consequently, the tracks they produce in CR-39 PNTD often take the form of long tubes rather than the conical etch pits produced by higher energy particles.

  11. Correlative Stochastic Optical Reconstruction Microscopy and Electron Microscopy

    OpenAIRE

    Doory Kim; Deerinck, Thomas J.; Sigal, Yaron M.; Babcock, Hazen P.; Ellisman, Mark H.; Xiaowei Zhuang

    2015-01-01

    Correlative fluorescence light microscopy and electron microscopy allows the imaging of spatial distributions of specific biomolecules in the context of cellular ultrastructure. Recent development of super-resolution fluorescence microscopy allows the location of molecules to be determined with nanometer-scale spatial resolution. However, correlative super-resolution fluorescence microscopy and electron microscopy (EM) still remains challenging because the optimal specimen preparation and ima...

  12. A lens-coupled scintillation counter in cryogenic environment

    CERN Document Server

    Stoykov, A; Amato, A; Bartkowiak, M; Konter, J A; Rodriguez, J; Sedlak, K

    2011-01-01

    In this work we present an elegant solution for a scintillation counter to be integrated into a cryogenic system. Its distinguishing feature is the absence of a continuous light guide coupling the scintillation and the photodetector parts, operating at cryogenic and room temperatures respectively. The prototype detector consists of a plastic scintillator with glued-in wavelength-shifting fiber located inside a cryostat, a Geiger-mode Avalanche Photodiode (G-APD) outside the cryostat, and a lens system guiding the scintillation light re-emitted by the fiber to the G-APD through optical windows in the cryostat shields. With a 0.8mm diameter multiclad fiber and a 1mm active area G-APD the coupling efficiency of the "lens light guide" is about 50%. A reliable performance of the detector down to 3K is demonstrated.

  13. A lens-coupled scintillation counter in cryogenic environment

    Energy Technology Data Exchange (ETDEWEB)

    Stoykov, A; Scheuermann, R; Amato, A; Bartkowiak, M; Konter, J A; Rodriguez, J; Sedlak, K, E-mail: alexey.stoykov@psi.ch [Paul Scherrer Institut, CH-5232 Villigen PSI (Switzerland)

    2011-02-01

    In this work we present an elegant solution for a scintillation counter to be integrated into a cryogenic system. Its distinguishing feature is the absence of a continuous light guide coupling the scintillation and the photodetector parts, operating at cryogenic and room temperatures respectively. The prototype detector consists of a plastic scintillator with glued-in wavelength-shifting fiber located inside a cryostat, a Geiger-mode Avalanche Photodiode (G-APD) outside the cryostat, and a lens system guiding the scintillation light re-emitted by the fiber to the G-APD through optical windows in the cryostat shields. With a 0.8 mm diameter multiclad fiber and a 1 mm active area G-APD the coupling efficiency of the 'lens light guide' is about 50%. A reliable performance of the detector down to 3 K is demonstrated.

  14. Light

    CERN Document Server

    Ditchburn, R W

    2011-01-01

    This classic study, available for the first time in paperback, clearly demonstrates how quantum theory is a natural development of wave theory, and how these two theories, once thought to be irreconcilable, together comprise a single valid theory of light. Aimed at students with an intermediate-level knowledge of physics, the book first offers a historical introduction to the subject, then covers topics such as wave theory, interference, diffraction, Huygens' Principle, Fermat's Principle, and the accuracy of optical measurements. Additional topics include the velocity of light, relativistic o

  15. Cryogenic holographic distortion testing

    Science.gov (United States)

    Michel, David G.

    1994-06-01

    Hughes cryogenic holographic test facility allows for the rapid characterization of optical components and mechanical structures at elevated and reduced temperatures. The facility consists of a 1.6 meter diameter thermal vacuum chamber, vibration isolated experiment test platform, and a holographic camera assembly. Temperatures as low as 12 Kelvin and as high as 350 Kelvin have been demonstrated. Complex aspheric mirrors are tested without the need for auxiliary null lenses and may be tested in either the polished or unpolished state. Structural elements such as optical benches, solar array panels, and spacecraft antennas have been tested. Types of materials tested include beryllium, silicon carbide, aluminum, graphite epoxy, silicon/aluminum matrix material and injection molded plastics. Sizes have ranged from 7 cm X 15 cm to 825 cm X 1125 cm and have weighed as little as 0.2 Kg and as much as 130 Kg. Surface figure changes as little as (lambda) /10 peak-to-valley ((lambda) equals .514 micrometers ) are routinely measured.

  16. Dual Cryogenic Capacitive Density Sensor

    Science.gov (United States)

    Youngquist, Robert; Mata, Carlos; Vokrot, Peter; Cox, Robert

    2009-01-01

    A dual cryogenic capacitive density sensor has been developed. The device contains capacitive sensors that monitor two-phase cryogenic flow density to within 1% accuracy, which, if temperature were known, could be used to determine the ratio of liquid to gas in the line. Two of these density sensors, located a known distance apart, comprise the sensor, providing some information on the velocity of the flow. This sensor was constructed as a proposed mass flowmeter with high data acquisition rates. Without moving parts, this device is capable of detecting the density change within a two-phase cryogenic flow more than 100 times a second. Detection is enabled by a series of two sets of five parallel plates with stainless steel, cryogenically rated tubing. The parallel plates form the two capacitive sensors, which are measured by electrically isolated digital electronics. These capacitors monitor the dielectric of the flow essentially the density of the flow and can be used to determine (along with temperature) the ratio of cryogenic liquid to gas. Combining this information with the velocity of the flow can, with care, be used to approximate the total two-phase mass flow. The sensor can be operated at moderately high pressures and can be lowered into a cryogenic bath. The electronics have been substantially improved over the older sensors, incorporating a better microprocessor, elaborate ground loop protection and noise limiting circuitry, and reduced temperature sensitivity. At the time of this writing, this design has been bench tested at room temperature, but actual cryogenic tests are pending

  17. Image-Guided Cryoablation of the Spine in a Swine Model: Clinical, Radiological, and Pathological Findings with Light and Electron Microscopy

    Energy Technology Data Exchange (ETDEWEB)

    Freitas, Ricardo Miguel Costa de, E-mail: ricardomcfreitas@gmail.com; Andrade, Celi Santos, E-mail: celis.andrade@hotmail.com; Caldas, José Guilherme Mendes Pereira, E-mail: jgmpcaldas@uol.com.br [Faculdade de Medicina da Universidade de São Paulo, Department of Radiology, Interventional Radiology Unit of the Instituto de Radiologia (Brazil); Tsunemi, Miriam Harumi, E-mail: miharumi@gmail.com [Universidade Estadual Paulista Júlio de Mesquita Filho, Department of Biostatistics, Biosciences Institute (Brazil); Ferreira, Lorraine Braga, E-mail: lorraine.braga@gmail.com; Arana-Chavez, Victor Elias, E-mail: vearana@usp.br [Faculdade de Odontologia da Universidade de São Paulo, Department of Oral Pathology (Brazil); Cury, Patrícia Maluf, E-mail: pmcury@hotmail.com [Faculdade de Medicina de São José do Rio Preto, Department of Pathology and Forensic Medicine (Brazil)

    2015-10-15

    PurposeThis study was designed to present the feasibility of an in vivo image-guided percutaneous cryoablation of the porcine vertebral body.MethodsThe institutional animal care committee approved this study. Cone-beam computed tomography (CBCT)-guided vertebral cryoablations (n = 22) were performed in eight pigs with short, 2-min, single or double-freezing protocols. Protective measures to nerves included dioxide carbon (CO{sub 2}) epidural injections and spinal canal temperature monitoring. Clinical, radiological, and pathological data with light (n = 20) or transmission electron (n = 2) microscopic analyses were evaluated after 6 days of clinical follow-up and euthanasia.ResultsCBCT/fluoroscopic-guided transpedicular vertebral body cryoprobe positioning and CO{sub 2} epidural injection were successful in all procedures. No major complications were observed in seven animals (87.5 %, n = 8). A minor complication was observed in one pig (12.5 %, n = 1). Logistic regression model analysis showed the cryoprobe-spinal canal (Cp-Sc) distance as the most efficient parameter to categorize spinal canal temperatures lower than 19 °C (p < 0.004), with a significant Pearson’s correlation test (p < 0.041) between the Cp-Sc distance and the lowest spinal canal temperatures. Ablation zones encompassed pedicles and the posterior wall of the vertebral bodies with an inflammatory rim, although no inflammatory infiltrate was depicted in the surrounding neural structures at light microscopy. Ultrastructural analyses evidenced myelin sheath disruption in some large nerve fibers, although neurological deficits were not observed.ConclusionsCBCT-guided vertebral cryoablation of the porcine spine is feasible under a combination of a short freezing protocol and protective measures to the surrounding nerves. Ultrastructural analyses may be helpful assess the early modifications of the nerve fibers.

  18. Micro-computed tomographic analysis of the radial geometry of intrarenal artery-vein pairs in rats and rabbits: Comparison with light microscopy.

    Science.gov (United States)

    Ngo, Jennifer P; Le, Bianca; Khan, Zohaib; Kett, Michelle M; Gardiner, Bruce S; Smith, David W; Melhem, Mayer M; Maksimenko, Anton; Pearson, James T; Evans, Roger G

    2017-08-10

    We assessed the utility of synchrotron-radiation micro-computed tomography (micro-CT) for quantification of the radial geometry of the renal cortical vasculature. The kidneys of nine rats and six rabbits were perfusion fixed and the renal circulation filled with Microfil. In order to assess shrinkage of Microfil, rat kidneys were imaged at the Australian Synchrotron immediately upon tissue preparation and then post fixed in paraformaldehyde and reimaged 24 hours later. The Microfil shrank only 2-5% over the 24 hour period. All subsequent micro-CT imaging was completed within 24 hours of sample preparation. After micro-CT imaging, the kidneys were processed for histological analysis. In both rat and rabbit kidneys, vascular structures identified in histological sections could be identified in two-dimensional (2D) micro-CT images from the original kidney. Vascular morphology was similar in the two sets of images. Radial geometry quantified by manual analysis of 2D images from micro-CT was consistent with corresponding data generated by light microscopy. However, due to limited spatial resolution when imaging a whole organ using contrast-enhanced micro-CT, only arteries ≥100 and ≥60 μm in diameter, for the rat and rabbit respectively, could be assessed. We conclude that it is feasible and valid to use micro-CT to quantify vascular geometry of the renal cortical circulation in both the rat and rabbit. However, a combination of light microscopic and micro-CT approaches are required to evaluate the spatial relationships between intrarenal arteries and veins over an extensive range of vessel size. © 2017 John Wiley & Sons Australia, Ltd.

  19. Mapping whole-brain activity with cellular resolution by light-sheet microscopy and high-throughput image analysis (Conference Presentation)

    Science.gov (United States)

    Silvestri, Ludovico; Rudinskiy, Nikita; Paciscopi, Marco; Müllenbroich, Marie Caroline; Costantini, Irene; Sacconi, Leonardo; Frasconi, Paolo; Hyman, Bradley T.; Pavone, Francesco S.

    2016-03-01

    Mapping neuronal activity patterns across the whole brain with cellular resolution is a challenging task for state-of-the-art imaging methods. Indeed, despite a number of technological efforts, quantitative cellular-resolution activation maps of the whole brain have not yet been obtained. Many techniques are limited by coarse resolution or by a narrow field of view. High-throughput imaging methods, such as light sheet microscopy, can be used to image large specimens with high resolution and in reasonable times. However, the bottleneck is then moved from image acquisition to image analysis, since many TeraBytes of data have to be processed to extract meaningful information. Here, we present a full experimental pipeline to quantify neuronal activity in the entire mouse brain with cellular resolution, based on a combination of genetics, optics and computer science. We used a transgenic mouse strain (Arc-dVenus mouse) in which neurons which have been active in the last hours before brain fixation are fluorescently labelled. Samples were cleared with CLARITY and imaged with a custom-made confocal light sheet microscope. To perform an automatic localization of fluorescent cells on the large images produced, we used a novel computational approach called semantic deconvolution. The combined approach presented here allows quantifying the amount of Arc-expressing neurons throughout the whole mouse brain. When applied to cohorts of mice subject to different stimuli and/or environmental conditions, this method helps finding correlations in activity between different neuronal populations, opening the possibility to infer a sort of brain-wide 'functional connectivity' with cellular resolution.

  20. Core-shell InGaN/GaN nanowire light emitting diodes analyzed by electron beam induced current microscopy and cathodoluminescence mapping.

    Science.gov (United States)

    Tchernycheva, M; Neplokh, V; Zhang, H; Lavenus, P; Rigutti, L; Bayle, F; Julien, F H; Babichev, A; Jacopin, G; Largeau, L; Ciechonski, R; Vescovi, G; Kryliouk, O

    2015-07-21

    We report on the electron beam induced current (EBIC) microscopy and cathodoluminescence (CL) characterization correlated with compositional analysis of light emitting diodes based on core/shell InGaN/GaN nanowire arrays. The EBIC mapping of cleaved fully operational devices allows to probe the electrical properties of the active region with a nanoscale resolution. In particular, the electrical activity of the p-n junction on the m-planes and on the semi-polar planes of individual nanowires is assessed in top view and cross-sectional geometries. The EBIC maps combined with CL characterization demonstrate the impact of the compositional gradients along the wire axis on the electrical and optical signals: the reduction of the EBIC signal toward the nanowire top is accompanied by an increase of the CL intensity. This effect is interpreted as a consequence of the In and Al gradients in the quantum well and in the electron blocking layer, which influence the carrier extraction efficiency. The interface between the nanowire core and the radially grown layer is shown to produce in some cases a transitory EBIC signal. This observation is explained by the presence of charged traps at this interface, which can be saturated by electron irradiation.

  1. Light and scanning electron microscopy study of in vitro effects of artesunate in newly excysted metacercariae of Echinostoma paraensei (Trematoda: Digenea).

    Science.gov (United States)

    Souza, Joyce G R; Lopes Torres, Eduardo J; Garcia, Juberlan S; Gomes, Ana Paula N; Rodrigues-Silva, Rosangela; Maldonado, Arnaldo; Machado-Silva, José Roberto

    2017-03-01

    Chemotherapy of food-borne trematodes relies on two drugs, praziquantel and tricabendazole, and there is growing interest in finding alternative therapies. Plant oil extracts have long been used in traditional Chinese medicine as sources of bioactive compounds with antiparasitic activity. Species of the genus Echinostoma are used as good models to test effective compounds against food-borne trematodes. This study evaluated the anthelmintic activity of crude artesunate extracts in vitro on newly excysted metacercariae of Echinostoma paraensei by light and scanning electron microscopy (SEM). The flukes were incubated with 1 μg/mL, 10 μg/mL, 25 μg/mL, 50 μg/mL and 100 μg/mL of artesunate for 4, 12, 24, 48 and 72 h. When the exposure time and concentration of artesunate increased, there were changes in motor activity, tegument damage and death. Blebs and swelling were the most common damages quantified on the tegument. The in vitro study reproduced results described for other immature flukes incubated with artemisinin derivatives. Excysted metacercariae of E. paraensei constitute a good model to study in vitro drug effects.

  2. Analysis of the interaction between Bacillus coagulans and Bacillus thuringiensis S-layers and calcium ions by XRD, light microscopy, and FTIR.

    Science.gov (United States)

    Babolmorad, Ghazal; Emtiazi, Giti; Emamzadeh, Rahman

    2014-05-01

    S-layer is a self-assemble regularly crystalline surface that covers major cell wall component of many bacteria and archaea and exhibits a high metal-binding capacity. We have studied the effect of the calcium ions and type of solid support (glass or mica) on the structure of the S-layers from Bacillus coagulans HN-68 and Bacillus thuringiensis MH14 upon simple methods based on light microscopy and AFM. Furthermore, the Fourier transform infrared spectroscopy (FTIR) study is indicated that the calcium-S-layer interaction occurred mainly through the carboxylate groups of the side chains of aspartic acid (Asp) and glutamic acid (Glu) and nitrogen atoms of Lys, Asn, and histidine (His) amino acids and N-H groups of the peptide backbone. Studied FTIR revealed that inner faces of S-layer are mainly negative, and outer faces of S-layer are mainly positive. Probably, calcium ions with positive charges bound to the carboxyl groups of Glu and Asp. Accordingly, calcium ions are anchored in the space between the inner faces of S-layer with negative charge and the surface of mica with negative charge. This leads to regular arrangement of the S-layer subunits.

  3. A Bright Fluorescent Probe for H2S Enables Analyte-Responsive, 3D Imaging in Live Zebrafish Using Light Sheet Fluorescence Microscopy.

    Science.gov (United States)

    Hammers, Matthew D; Taormina, Michael J; Cerda, Matthew M; Montoya, Leticia A; Seidenkranz, Daniel T; Parthasarathy, Raghuveer; Pluth, Michael D

    2015-08-19

    Hydrogen sulfide (H2S) is a critical gaseous signaling molecule emerging at the center of a rich field of chemical and biological research. As our understanding of the complexity of physiological H2S in signaling pathways evolves, advanced chemical and technological investigative tools are required to make sense of this interconnectivity. Toward this goal, we have developed an azide-functionalized O-methylrhodol fluorophore, MeRho-Az, which exhibits a rapid >1000-fold fluorescence response when treated with H2S, is selective for H2S over other biological analytes, and has a detection limit of 86 nM. Additionally, the MeRho-Az scaffold is less susceptible to photoactivation than other commonly used azide-based systems, increasing its potential application in imaging experiments. To demonstrate the efficacy of this probe for H2S detection, we demonstrate the ability of MeRho-Az to detect differences in H2S levels in C6 cells and those treated with AOAA, a common inhibitor of enzymatic H2S synthesis. Expanding the use of MeRho-Az to complex and heterogeneous biological settings, we used MeRho-Az in combination with light sheet fluorescence microscopy (LSFM) to visualize H2S in the intestinal tract of live zebrafish. This application provides the first demonstration of analyte-responsive 3D imaging with LSFM, highlighting the utility of combining new probes and live imaging methods for investigating chemical signaling in complex multicellular systems.

  4. Cryogenics bringing the temperature down, underground

    CERN Multimedia

    2005-01-01

    The first 600m of the LHC cryogenic distribution line (QRL), which will feed the accelerator's superconducting magnets, has passed initial validating tests of its mechanical design at room and cryogenic temperatures.

  5. Cryogenic engineering fifty years of progress

    CERN Document Server

    Reed, Richard

    2007-01-01

    Cryogenic Engineering: Fifty Years of Progress is a benchmark reference work which chronicles the major developments in the field. Starting with an historical background dating to the 1850s, this book reviews the development of data resources now available for cryogenic fields and properties of materials. The advances in cryogenic fundamentals are covered by reviews of cryogenic principles, cryogenic insulation, low-loss storage systems, modern liquefaction processes, helium cryogenics and low-temperature thermometry. Several well-established applications resulting from cryogenic advances include aerospace cryocoolers and refrigerators, use of LTS and HTS systems in electrical applications, and recent changes in cryopreservation. Extensive references are provided for the readers interested in the details of these cryogenic engineering advances.

  6. Cryogenic electron microscopy and single-particle analysis.

    Science.gov (United States)

    Elmlund, Dominika; Elmlund, Hans

    2015-01-01

    About 20 years ago, the first three-dimensional (3D) reconstructions at subnanometer (single-particle analysis. Since then, thousands of structures have been determined to resolutions ranging from 30 Å to near atomic (analysis software have advanced the technology considerably. Near-atomic-resolution reconstructions can now be obtained, not only for megadalton macromolecular complexes or highly symmetrical assemblies but also for proteins of only a few hundred kilodaltons. We discuss the developments that led to this breakthrough in high-resolution structure determination by cryo-EM and point to challenges that lie ahead.

  7. Thermodynamic properties of cryogenic fluids

    CERN Document Server

    Leachman, Jacob; Lemmon, Eric; Penoncello, Steven

    2017-01-01

    This update to a classic reference text provides practising engineers and scientists with accurate thermophysical property data for cryogenic fluids. The equations for fifteen important cryogenic fluids are presented in a basic format, accompanied by pressure-enthalpy and temperature-entropy charts and tables of thermodynamic properties. It begins with a chapter introducing the thermodynamic relations and functional forms for equations of state, and goes on to describe the requirements for thermodynamic property formulations, needed for the complete definition of the thermodynamic properties of a fluid. The core of the book comprises extensive data tables and charts for the most commonly-encountered cryogenic fluids. This new edition sees significant updates to the data presented for air, argon, carbon monoxide, deuterium, ethane, helium, hydrogen, krypton, nitrogen and xenon. The book supports and complements NIST’s REFPROP - an interactive database and tool for the calculation of thermodynamic propertie...

  8. Cryogenic safety organisation at CERN

    CERN Document Server

    CERN. Geneva

    2016-01-01

    With Safety being a top priority of CERN’s general policy, the Organisation defines and implements a Policy that sets out the general principles governing Safety at CERN. To the end of the attainment of said Safety objectives, the organic units (owners/users of the equipment) are assigned the responsibility for the implementation of the CERN Safety Policy at all levels of the organization, whereas the Health and Safety and Environmental Protection Unit (HSE) has the role of providing assistance for the implementation of the Safety Policy, and a monitoring role related to the implementation of continuous improvement of Safety, compliance with the Safety Rules and the handling of emergency situations. This talk will elaborate on the roles, responsibilities and organisational structure of the different stakeholders within the Organization with regards to Safety, and in particular to cryogenic safety. The roles of actors of particular importance such as the Cryogenic Safety Officers (CSOs) and the Cryogenic Sa...

  9. Cryogenic needs for future tokamaks

    Science.gov (United States)

    Katheder, H.

    The ITER tokamak is a machine using superconducting magnets. The windings of these magnets will be subjected to high heat loads resulting from a combination of nuclear energy absorption and AC-losses. It is estimated that about 100 kW at 4.5 K are needed. The total cooling mass flow rate will be around 10 - 15 kg/s. In addition to the large cryogenic power required for the superconducting magnets cryogenic power is also needed for refrigerated radiation shield, various cryopumps, fuel processing and test beds. A general description of the overall layout and the envisaged refrigerator cycle, necessary cold pumps and ancillary equipment is given. The basic cryogenic layout for the ITER tokakmak design, as developed during the conceptual design phase and a short overview about existing tokamak designs using superconducting magnets is given.

  10. Magnetic bearings for cryogenic turbomachines

    Science.gov (United States)

    Iannello, Victor; Sixsmith, Herbert

    1991-01-01

    Magnetic bearings offer a number of advantages over gas bearings for the support of rotors in cryogenic turboexpanders and compressors. Their performance is relatively independent of the temperature or pressure of the process gas for a large range of conditions. Active magnetic bearing systems that use capacitive sensors have been developed for high speed compressors for use in cryogenic refrigerators. Here, the development of a magnetic bearing system for a miniature ultra high speed compressor is discussed. The magnetic bearing has demonstrated stability at rotational speeds exceeding 250,000 rpm. This paper describes the important features of the magnetic bearing and presents test results demonstrating its performance characteristics.

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

    Energy Technology Data Exchange (ETDEWEB)

    Ramanathan, Nathan Muruganathan [ORNL; Darling, Seth B. [Argonne National Laboratory (ANL)

    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.

  12. Photolytic separation of isotopes in cryogenic solution

    Science.gov (United States)

    Freund, S.M.; Maier, W.B. II; Holland, R.F.; Battie, W.H.

    Separation of carbon isotopes by photolysis of CS/sub 2/ in cryogenic solutions of nitrogen, krypton and argon with 206 nm light from an iodine resonance lamp is reported. The spectral distributionn of the ultraviolet absorption depends on solvent. Thus, in liquid nitrogen the photolytic decomposition rate of /sup 13/CS/sub 2/ is greater than that of /sup 12/CS/sub 2/ (because the absorption of 206 nm radiation is greater for /sup 13/CS/sub 2/), whereas in liquid krypton and liquid argon the reverse is true. The shift in ultraviolet spectrum is a general phenomenon readily characterized as a function of solvent polarizability, and exhibits behavior similar to that for vibrational transitions occurring in the infrared.

  13. Electron Microscopy.

    Science.gov (United States)

    Beer, Michael

    1980-01-01

    Reviews technical aspects of structure determination in biological electron microscopy (EM). Discusses low dose EM, low temperature microscopy, electron energy loss spectra, determination of mass or molecular weight, and EM of labeled systems. Cites 34 references. (CS)

  14. Electron Microscopy.

    Science.gov (United States)

    Beer, Michael

    1980-01-01

    Reviews technical aspects of structure determination in biological electron microscopy (EM). Discusses low dose EM, low temperature microscopy, electron energy loss spectra, determination of mass or molecular weight, and EM of labeled systems. Cites 34 references. (CS)

  15. Oocyte polarized light microscopy, assay of specific follicular fluid metabolites, and gene expression in cumulus cells as different approaches to predict fertilization efficiency after ICSI.

    Science.gov (United States)

    Revelli, Alberto; Canosa, Stefano; Bergandi, Loredana; Skorokhod, Oleksii A; Biasoni, Valentina; Carosso, Andrea; Bertagna, Angela; Maule, Milena; Aldieri, Elisabetta; D'Eufemia, Maria Diletta; Evangelista, Francesca; Colacurci, Nicola; Benedetto, Chiara

    2017-06-23

    The complex relationship between oocyte morphology, specific follicular fluid metabolites, gene expression in cumulus granulosa cells, and oocyte competence toward fertilization and embryo development still needs further clarification. Forty-six oocytes retrieved from the largest pre-ovulatory follicle of patients undergoing intra-cytoplasmic sperm injection (ICSI) were considered assessing: (a) oocyte morphological characteristics at polarized light microscopy (PLM), (b) specific follicular fluid (FF) metabolites previously suggested to influence oocyte competence (AMH, markers of redox status and of cytotoxicity), (c) transcription of AMH and AMH type II receptor genes in cumulus cells. Data were analyzed using mono-parametric tests and multivariable logistic analysis in order to correlate morphological and biochemical data with fertilization. Comparing normally fertilized oocytes (n = 29, F group) with unfertilized (n = 17, nF group) we observed that: (a) the meiotic spindle area and major axis were significantly higher in nF group and in fertilized oocytes undergoing an early embryo development arrest; (b) AMH level in FF was comparable in F and nF groups; (c) the FF of nF group contained significantly higher levels of cytotoxicity (lactate dehydrogenase) and oxidative stress (Cu,Zn-superoxide dismutase, catalase, 4-hydroxynonenal-protein conjugates) markers; (d) cumulus cells of nF group showed significantly higher AMH receptor type II gene expression. Taken together, these observations suggest that an excessive cytotoxicity level can alter AMH signal transduction within cumulus cells, in turn leading to partial inhibition of aromatase activity, altered cytoplasmic maturation and increased oxidative stress, factors able to impair oocyte fertilization competence and embryo growth.

  16. Cryogenic technology for tracking detectors

    CERN Document Server

    Granata, V; Watts, S; Borer, K; Janos, S; Pretzl, Klaus P; Dezillie, B; Li, Z; Casagrande, L; Collins, P; Grohmann, S; Heijne, Erik H M; Lourenço, C; Niinikoski, T O; Palmieri, V G; Sonderegger, P; Borchi, E; Bruzzi, Mara; Pirollo, S; Chapuy, S; Dimcovski, Zlatomir; Grigoriev, E; Bell, W; Devine, S R H; O'Shea, V; Ruggiero, G; Smith, K; Berglund, P; de Boer, Wim; Hauler, F; Heising, S; Jungermann, L; Abreu, M C; Rato-Mendes, P; Sousa, P; Cindro, V; Mikuz, M; Zavrtanik, M; Esposito, A P; Konorov, I; Paul, S; Buontempo, S; D'Ambrosio, D; Pagano, S; Eremin, V V; Verbitskaya, E

    2001-01-01

    A low-mass cryogenic cooling technique for silicon sensor modules has been developed in the framework of the RD39 Collaboration at CERN. A prototype low-mass beam tracker cryostat has been designed, constructed and tested for applications in fixed target experiments. We shall report here briefly the main features and results of the system. (2 refs).

  17. USAF Space Sensing Cryogenic Considerations

    Science.gov (United States)

    2010-01-01

    capacitance dilatometer for measuring thermal expansion and magnetostriction Rev. Sci. Instrum. 83, 095102 (2012) Compact radio-frequency resonator...enhancing when the refrigeration system is considered as part of an overall optimization problem. INTRODUCTION The use of cryogenics in space sensing

  18. LHC Cryogenics on the mend

    CERN Multimedia

    2004-01-01

    On 29 September, repairs began on the LHC cryogenic distribution line, or QRL, to replace a faulty part that occurs in the hundreds of elements of the line that are already on-site. The Accelerator Technology Department is designing a work programme to finish the repairs as soon as possible and minimize delays to the rest of the LHC project.

  19. Champagne for the cryogenics teams

    CERN Multimedia

    2005-01-01

    Christmas has come early for the LHC as a complete sector of the cryogenic distribution line has been operating at 10 degrees Kelvin (-263°C) for the past two weeks, just a few degrees above the machine's nominal operating temperature.

  20. Background reduction in cryogenic detectors

    Energy Technology Data Exchange (ETDEWEB)

    Bauer, Daniel A.; /Fermilab

    2005-04-01

    This paper discusses the background reduction and rejection strategy of the Cryogenic Dark Matter Search (CDMS) experiment. Recent measurements of background levels from CDMS II at Soudan are presented, along with estimates for future improvements in sensitivity expected for a proposed SuperCDMS experiment at SNOLAB.