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

Science.gov (United States)

Yamamura, Hisao; Suzuki, Yoshiaki; Imaizumi, Yuji

2015-05-01

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 Ca(2+) events in the subplasmalemma. Single-molecule analyses of ion channels and localized Ca(2+) signals based on TIRF imaging provide beneficial pharmacological and physiological information concerning the functions of ion channels. Copyright © 2015 The Authors. Production and hosting by Elsevier B.V. All rights reserved.

Light reflection models for computer graphics.

Science.gov (United States)

Greenberg, D P

1989-04-14

During the past 20 years, computer graphic techniques for simulating the reflection of light have progressed so that today images of photorealistic quality can be produced. Early algorithms considered direct lighting only, but global illumination phenomena with indirect lighting, surface interreflections, and shadows can now be modeled with ray tracing, radiosity, and Monte Carlo simulations. This article describes the historical development of computer graphic algorithms for light reflection and pictorially illustrates what will be commonly available in the near future.

Variable-angle total internal reflection fluorescence microscopy of intact cells of Arabidopsis thaliana

Directory of Open Access Journals (Sweden)

Kim Myung K

2011-09-01

Full Text Available Abstract Background Total internal reflection fluorescence microscopy (TIRFM is a powerful tool for observing fluorescently labeled molecules on the plasma membrane surface of animal cells. However, the utility of TIRFM in plant cell studies has been limited by the fact that plants have cell walls, thick peripheral layers surrounding the plasma membrane. Recently, a new technique known as variable-angle epifluorescence microscopy (VAEM was developed to circumvent this problem. However, the lack of a detailed analysis of the optical principles underlying VAEM has limited its applications in plant-cell biology. Results Here, we present theoretical and experimental evidence supporting the use of variable-angle TIRFM in observations of intact plant cells. We show that when total internal reflection occurs at the cell wall/cytosol interface with an appropriate angle of incidence, an evanescent wave field of constant depth is produced inside the cytosol. Results of experimental TIRFM observations of the dynamic behaviors of phototropin 1 (a membrane receptor protein and clathrin light chain (a vesicle coat protein support our theoretical analysis. Conclusions These findings demonstrate that variable-angle TIRFM is appropriate for quantitative live imaging of cells in intact tissues of Arabidopsis thaliana.

Biobeam—Multiplexed wave-optical simulations of light-sheet microscopy

Science.gov (United States)

Weigert, Martin; Bundschuh, Sebastian T.

2018-01-01

Sample-induced image-degradation remains an intricate wave-optical problem in light-sheet microscopy. Here we present biobeam, an open-source software package that enables simulation of operational light-sheet microscopes by combining data from 105–106 multiplexed and GPU-accelerated point-spread-function calculations. The wave-optical nature of these simulations leads to the faithful reproduction of spatially varying aberrations, diffraction artifacts, geometric image distortions, adaptive optics, and emergent wave-optical phenomena, and renders image-formation in light-sheet microscopy computationally tractable. PMID:29652879

Light Microscopy at Maximal Precision

Science.gov (United States)

Bierbaum, Matthew; Leahy, Brian D.; Alemi, Alexander A.; Cohen, Itai; Sethna, James P.

2017-10-01

Microscopy is the workhorse of the physical and life sciences, producing crisp images of everything from atoms to cells well beyond the capabilities of the human eye. However, the analysis of these images is frequently little more accurate than manual marking. Here, we revolutionize the analysis of microscopy images, extracting all the useful information theoretically contained in a complex microscope image. Using a generic, methodological approach, we extract the information by fitting experimental images with a detailed optical model of the microscope, a method we call parameter extraction from reconstructing images (PERI). As a proof of principle, we demonstrate this approach with a confocal image of colloidal spheres, improving measurements of particle positions and radii by 10-100 times over current methods and attaining the maximum possible accuracy. With this unprecedented accuracy, we measure nanometer-scale colloidal interactions in dense suspensions solely with light microscopy, a previously impossible feat. Our approach is generic and applicable to imaging methods from brightfield to electron microscopy, where we expect accuracies of 1 nm and 0.1 pm, respectively.

Light Microscopy at Maximal Precision

Directory of Open Access Journals (Sweden)

Matthew Bierbaum

2017-10-01

Full Text Available Microscopy is the workhorse of the physical and life sciences, producing crisp images of everything from atoms to cells well beyond the capabilities of the human eye. However, the analysis of these images is frequently little more accurate than manual marking. Here, we revolutionize the analysis of microscopy images, extracting all the useful information theoretically contained in a complex microscope image. Using a generic, methodological approach, we extract the information by fitting experimental images with a detailed optical model of the microscope, a method we call parameter extraction from reconstructing images (PERI. As a proof of principle, we demonstrate this approach with a confocal image of colloidal spheres, improving measurements of particle positions and radii by 10–100 times over current methods and attaining the maximum possible accuracy. With this unprecedented accuracy, we measure nanometer-scale colloidal interactions in dense suspensions solely with light microscopy, a previously impossible feat. Our approach is generic and applicable to imaging methods from brightfield to electron microscopy, where we expect accuracies of 1 nm and 0.1 pm, respectively.

Light Microscopy Module (LMM)-Emulator

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Levine, Howard G.; Smith, Trent M.; Richards, Stephanie E.

2016-01-01

The Light Microscopy Module (LMM) is a microscope facility developed at Glenn Research Center (GRC) that provides researchers with powerful imaging capability onboard the International Space Station (ISS). LMM has the ability to have its hardware recongured on-orbit to accommodate a wide variety of investigations, with the capability of remotely acquiring and downloading digital images across multiple levels of magnication.

A Simplified, Low-Cost Method for Polarized Light Microscopy

Science.gov (United States)

Maude, Richard J.; Buapetch, Wanchana; Silamut, Kamolrat

2009-01-01

Malaria pigment is an intracellular inclusion body that appears in blood and tissue specimens on microscopic examination and can help in establishing the diagnosis of malaria. In simple light microscopy, it can be difficult to discern from cellular background and artifacts. It has long been known that if polarized light microscopy is used, malaria pigment can be much easier to distinguish. However, this technique is rarely used because of the need for a relatively costly polarization microscope. We describe a simple and economical technique to convert any standard light microscope suitable for examination of malaria films into a polarization microscope. PMID:19861611

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....

Low cost light-sheet microscopy for whole brain imaging

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Kumar, Manish; Nasenbeny, Jordan; Kozorovitskiy, Yevgenia

2018-02-01

Light-sheet microscopy has evolved as an indispensable tool in imaging biological samples. It can image 3D samples at fast speed, with high-resolution optical sectioning, and with reduced photobleaching effects. These properties make light-sheet microscopy ideal for imaging fluorophores in a variety of biological samples and organisms, e.g. zebrafish, drosophila, cleared mouse brains, etc. While most commercial turnkey light-sheet systems are expensive, the existing lower cost implementations, e.g. OpenSPIM, are focused on achieving high-resolution imaging of small samples or organisms like zebrafish. In this work, we substantially reduce the cost of light-sheet microscope system while targeting to image much larger samples, i.e. cleared mouse brains, at single-cell resolution. The expensive components of a lightsheet system - excitation laser, water-immersion objectives, and translation stage - are replaced with an incoherent laser diode, dry objectives, and a custom-built Arduino-controlled translation stage. A low-cost CUBIC protocol is used to clear fixed mouse brain samples. The open-source platforms of μManager and Fiji support image acquisition, processing, and visualization. Our system can easily be extended to multi-color light-sheet microscopy.

Macroscopic self-consistent model for external-reflection near-field microscopy

International Nuclear Information System (INIS)

Berntsen, S.; Bozhevolnaya, E.; Bozhevolnyi, S.

1993-01-01

The self-consistent macroscopic approach based on the Maxwell equations in two-dimensional geometry is developed to describe tip-surface interaction in external-reflection near-field microscopy. The problem is reduced to a single one-dimensional integral equation in terms of the Fourier components of the field at the plane of the sample surface. This equation is extended to take into account a pointlike scatterer placed on the sample surface. The power of light propagating toward the detector as the fiber mode is expressed by using the self-consistent field at the tip surface. Numerical results for trapezium-shaped tips are presented. The authors show that the sharper tip and the more confined fiber mode result in better resolution of the near-field microscope. Moreover, it is found that the tip-surface distance should not be too small so that better resolution is ensured. 14 refs., 10 figs

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...

Validation of Digital Microscopy Compared With Light Microscopy for the Diagnosis of Canine Cutaneous Tumors.

Science.gov (United States)

Bertram, Christof A; Gurtner, Corinne; Dettwiler, Martina; Kershaw, Olivia; Dietert, Kristina; Pieper, Laura; Pischon, Hannah; Gruber, Achim D; Klopfleisch, Robert

2018-07-01

Integration of new technologies, such as digital microscopy, into a highly standardized laboratory routine requires the validation of its performance in terms of reliability, specificity, and sensitivity. However, a validation study of digital microscopy is currently lacking in veterinary pathology. The aim of the current study was to validate the usability of digital microscopy in terms of diagnostic accuracy, speed, and confidence for diagnosing and differentiating common canine cutaneous tumor types and to compare it to classical light microscopy. Therefore, 80 histologic sections including 17 different skin tumor types were examined twice as glass slides and twice as digital whole-slide images by 6 pathologists with different levels of experience at 4 time points. Comparison of both methods found digital microscopy to be noninferior for differentiating individual tumor types within the category epithelial and mesenchymal tumors, but diagnostic concordance was slightly lower for differentiating individual round cell tumor types by digital microscopy. In addition, digital microscopy was associated with significantly shorter diagnostic time, but diagnostic confidence was lower and technical quality was considered inferior for whole-slide images compared with glass slides. Of note, diagnostic performance for whole-slide images scanned at 200× magnification was noninferior in diagnostic performance for slides scanned at 400×. In conclusion, digital microscopy differs only minimally from light microscopy in few aspects of diagnostic performance and overall appears adequate for the diagnosis of individual canine cutaneous tumors with minor limitations for differentiating individual round cell tumor types and grading of mast cell tumors.

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. © 2014 Elsevier Inc. All rights reserved.

Common-path configuration in total internal reflection digital holography microscopy.

Science.gov (United States)

Calabuig, Alejandro; Matrecano, Marcella; Paturzo, Melania; Ferraro, Pietro

2014-04-15

Total Internal Reflection Digital Holographic Microscopy (TIRDHM) is recognized to be a powerful tool for retrieving quantitative phase images of cell-substrate interfaces, adhesions, and tissue structures close to the prism surface. In this Letter, we develop an improved TIRDHM system, taking advantage of a refractive index mismatch between the prism and the sample substrate, to allow phase-shifting DH with just a single-beam interferometric configuration. Instead of the traditional off-axis method, phase-shift method is used to retrieve amplitude and phase images in coherent light and TIR modality. Essentially, the substrate-prism interface acts like a beam splitter generating a reference beam, where the phase-shift dependence on the incident angle is exploited in this common-path configuration. With the aim to demonstrate the technique's validity, some experiments are performed to establish the advantage of this compact and simple configuration, in which the reference arm in the setup is avoided.

Correlation of ''twins'' observed by optical microscopy and transmission electron microscopy in YBa2Cu3O7/sub -//sub x/ superconductors

International Nuclear Information System (INIS)

Hoff, H.A.; Singh, A.K.; Pande, C.S.

1988-01-01

By using transmission electron microscopy and optical microscopy on the same specimens, the patterns of light- and dark-contrast lines seen in reflected polarized light were shown to be an interference pattern due to the variable spacing of suboptical microtwins. These microtwins are mostly [110] reflection twins. The [110] twinning was observed to be cyclic and occasionally pseudotetragonal because of the progressive cycling of contact twin domains. Within a domain, and occasionally in a whole grain, the [110] reflection twins occurred as polysynthetic lamellae. The morphology of the domain structure can be explained from the theory of martensitic transformation

Processing scarce biological samples for light and transmission electron microscopy

Directory of Open Access Journals (Sweden)

P Taupin

2008-06-01

Full Text Available Light microscopy (LM and transmission electron microscopy (TEM aim at understanding the relationship structure-function. With advances in biology, isolation and purification of scarce populations of cells or subcellular structures may not lead to enough biological material, for processing for LM and TEM. A protocol for preparation of scarce biological samples is presented. It is based on pre-embedding the biological samples, suspensions or pellets, in bovine serum albumin (BSA and bis-acrylamide (BA, cross-linked and polymerized. This preparation provides a simple and reproducible technique to process biological materials, present in limited quantities that can not be amplified, for light and transmission electron microscopy.

Field-based dynamic light scattering microscopy: theory and numerical analysis.

Science.gov (United States)

Joo, Chulmin; de Boer, Johannes F

2013-11-01

We present a theoretical framework for field-based dynamic light scattering microscopy based on a spectral-domain optical coherence phase microscopy (SD-OCPM) platform. SD-OCPM is an interferometric microscope capable of quantitative measurement of amplitude and phase of scattered light with high phase stability. Field-based dynamic light scattering (F-DLS) analysis allows for direct evaluation of complex-valued field autocorrelation function and measurement of localized diffusive and directional dynamic properties of biological and material samples with high spatial resolution. In order to gain insight into the information provided by F-DLS microscopy, theoretical and numerical analyses are performed to evaluate the effect of numerical aperture of the imaging optics. We demonstrate that sharp focusing of fields affects the measured diffusive and transport velocity, which leads to smaller values for the dynamic properties in the sample. An approach for accurately determining the dynamic properties of the samples is discussed.

Studies of the Reflection, Refraction and Internal Reflection of Light

Science.gov (United States)

Lanchester, P. C.

2014-01-01

An inexpensive apparatus and associated experiments are described for studying the basic laws of reflection and refraction of light at an air-glass interface, and multiple internal reflections within a glass block. In order to motivate students and encourage their active participation, a novel technique is described for determining the refractive…

The challenge of diagnosing seborrheic keratosis by reflectance confocal microscopy.

Science.gov (United States)

Guo, A; Chen, J; Yang, C; Ding, Y; Zeng, Q; Tan, L

2018-05-24

Seborrheic keratosis (SK) is one of the most common skin tumors seen by dermatologists. It should be differentiated with many diseases, especially skin tumors. Reflectance confocal microscopy (RCM) has been applied for evaluation of SK. There are a few studies that describe the RCM of SK. The aim of the study was to find the challenge of diagnosing seborrheic keratosis by reflectance confocal microscopy. A total of 390 patients with a clinical suspicious diagnosis of seborrheic keratosis were enrolled in this study, and lesions from each patient were imaged with RCM. Thirty-seven of these patients performed a biopsy in order to be given a histological diagnosis. We retrospectively analyzed the outcomes of RCM diagnosis and histological diagnosis, and then found the RCM characteristics of biopsy-proven lesions. According to RCM images, 258 of 390 (66.2%) patients were diagnosed with SK, 97 of 390 (24.9%) patients could not be diagnosed by the dermatologist according to RCM. Of all 37 biopsied lesions, 23 were SK, 6 were actinic keratosis, 2 were basal cell carcinoma, and 2 were squamous cell carcinoma. It is challenge to diagnose seborrheic keratosis by reflectance confocal microscopy. It may due to the variable clinical and RCM appearances of SK, and limited depth of RCM. © 2018 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

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.

Multilayer mounting for long-term light sheet microscopy of zebrafish.

Science.gov (United States)

Weber, Michael; Mickoleit, Michaela; Huisken, Jan

2014-02-27

Light sheet microscopy is the ideal imaging technique to study zebrafish embryonic development. Due to minimal photo-toxicity and bleaching, it is particularly suited for long-term time-lapse imaging over many hours up to several days. However, an appropriate sample mounting strategy is needed that offers both confinement and normal development of the sample. Multilayer mounting, a new embedding technique using low-concentration agarose in optically clear tubes, now overcomes this limitation and unleashes the full potential of light sheet microscopy for real-time developmental biology.

Timing the total reflection of light

International Nuclear Information System (INIS)

Chauvat, Dominique; Bonnet, Christophe; Dunseath, Kevin; Emile, Olivier; Le Floch, Albert

2005-01-01

We have identified for the first time the absolute delay at total reflection, envisioned by Newton. We show that there are in fact two divergent Wigner delays, depending on the polarisation of the incident light. These measurements give a new insight on the passage from total reflection to refraction

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...

Hyperspectral light sheet microscopy

Science.gov (United States)

Jahr, Wiebke; Schmid, Benjamin; Schmied, Christopher; Fahrbach, Florian O.; Huisken, Jan

2015-09-01

To study the development and interactions of cells and tissues, multiple fluorescent markers need to be imaged efficiently in a single living organism. Instead of acquiring individual colours sequentially with filters, we created a platform based on line-scanning light sheet microscopy to record the entire spectrum for each pixel in a three-dimensional volume. We evaluated data sets with varying spectral sampling and determined the optimal channel width to be around 5 nm. With the help of these data sets, we show that our setup outperforms filter-based approaches with regard to image quality and discrimination of fluorophores. By spectral unmixing we resolved overlapping fluorophores with up to nanometre resolution and removed autofluorescence in zebrafish and fruit fly embryos.

Reflectance confocal microscopy: an effective tool for monitoring ultraviolet B phototherapy in psoriasis.

NARCIS (Netherlands)

Wolberink, E.A.W.; Erp, P.E.J. van; Boer-van Huizen, R.T. de; Kerkhof, P.C.M. van de; Gerritsen, M.J.P.

2012-01-01

Background In vivo reflectance confocal microscopy (RCM) is a novel, noninvasive imaging technique which enables imaging of skin at a cellular resolution comparable to conventional microscopy. Objectives We performed a pilot study to evaluate RCM as a noninvasive tool for monitoring ultraviolet (UV)

Measuring Light Reflectance of BGO Crystal Surfaces

Science.gov (United States)

Janecek, Martin; Moses, William W.

2008-10-01

A scintillating crystal's surface reflectance has to be well understood in order to accurately predict and optimize the crystal's light collection through Monte Carlo simulations. In this paper, we measure the inner surface reflectance properties for BGO. The measurements include BGO crystals with a mechanically polished surface, rough-cut surface, and chemically etched surface, and with various reflectors attached, both air-coupled and with coupling compound. The measurements are performed with a laser aimed at the center of a hemispherical shaped BGO crystal. The hemispherical shape eliminates any non-perpendicular angles for light entering and exiting the crystal. The reflected light is collected with an array of photodiodes. The laser can be set at an arbitrary angle, and the photodiode array is rotated to fully cover 2pi of solid angle. The current produced in the photodiodes is readout with a digital multimeter connected through a multiplexer. The two rows of photodiodes achieve 5-degree by 4-degree resolution, and the current measurement has a dynamic range of 105:1. The acquired data was not described by the commonly assumed linear combination of specular and diffuse (Lambertian) distributions, except for a very few surfaces. Surface roughness proved to be the most important parameter when choosing crystal setup. The reflector choice was of less importance and of almost no consequence for rough-cut surfaces. Pure specular reflection distribution for all incidence angles was measured for polished surfaces with VM2000 film, while the most Lambertian distribution for any surface finish was measured for titanium dioxide paint. The distributions acquired in this paper will be used to create more accurate Monte Carlo models for light reflection distribution within BGO crystals.

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.

Reproducibility in light microscopy: Maintenance, standards and SOPs.

Science.gov (United States)

Deagle, Rebecca C; Wee, Tse-Luen Erika; Brown, Claire M

2017-08-01

Light microscopy has grown to be a valuable asset in both the physical and life sciences. It is a highly quantitative method available in individual research laboratories and often centralized in core facilities. However, although quantitative microscopy is becoming a customary tool in research, it is rarely standardized. To achieve accurate quantitative microscopy data and reproducible results, three levels of standardization must be considered: (1) aspects of the microscope, (2) the sample, and (3) the detector. The accuracy of the data is only as reliable as the imaging system itself, thereby imposing the need for routine standard performance testing. Depending on the task some maintenance procedures should be performed once a month, some before each imaging session, while others conducted annually. This text should be implemented as a resource for researchers to integrate with their own standard operating procedures to ensure the highest quality quantitative microscopy data. Copyright © 2017. Published by Elsevier Ltd.

X-ray microscopy resource center at the Advanced Light Source

International Nuclear Information System (INIS)

Meyer-Ilse, W.; Koike, M.; Beguiristain, R.; Maser, J.; Attwood, D.

1992-07-01

An x-ray microscopy resource center for biological x-ray imaging vvill be built at the Advanced Light Source (ALS) in Berkeley. The unique high brightness of the ALS allows short exposure times and high image quality. Two microscopes, an x-ray microscope (XM) and a scanning x-ray microscope (SXM) are planned. These microscopes serve complementary needs. The XM gives images in parallel at comparable short exposure times, and the SXM is optimized for low radiation doses applied to the sample. The microscopes extend visible light microscopy towards significantly higher resolution and permit images of objects in an aqueous medium. High resolution is accomplished by the use of Fresnel zone plates. Design considerations to serve the needs of biological x-ray microscopy are given. Also the preliminary design of the microscopes is presented. Multiple wavelength and multiple view images will provide elemental contrast and some degree of 3D information

Research and application on imaging technology of line structure light based on confocal microscopy

Science.gov (United States)

Han, Wenfeng; Xiao, Zexin; Wang, Xiaofen

2009-11-01

In 2005, the theory of line structure light confocal microscopy was put forward firstly in China by Xingyu Gao and Zexin Xiao in the Institute of Opt-mechatronics of Guilin University of Electronic Technology. Though the lateral resolution of line confocal microscopy can only reach or approach the level of the traditional dot confocal microscopy. But compared with traditional dot confocal microscopy, it has two advantages: first, by substituting line scanning for dot scanning, plane imaging only performs one-dimensional scanning, with imaging velocity greatly improved and scanning mechanism simplified, second, transfer quantity of light is greatly improved by substituting detection hairline for detection pinhole, and low illumination CCD is used directly to collect images instead of photoelectric intensifier. In order to apply the line confocal microscopy to practical system, based on the further research on the theory of the line confocal microscopy, imaging technology of line structure light is put forward on condition of implementation of confocal microscopy. Its validity and reliability are also verified by experiments.

Effects of phase change on reflection in phase-measuring interference microscopy

OpenAIRE

Dubois , Arnaud

2004-01-01

International audience; We show by analytical and numerical calculations that the phase change on reflection that occurs in interference microscopy is almost independent of the numerical aperture of the objective. The shift of the microscope interferogram response due to the phase change on reflection, however, increases with the numerical aperture. Measurements of the interferogram shift are made with a Linnik interference microscope equipped with various numerical-aperture objectives and ar...

Bidirectional reflectance distribution function of Spectralon white reflectance standard illuminated by incoherent unpolarized and plane-polarized light.

Science.gov (United States)

Bhandari, Anak; Hamre, Børge; Frette, Øvynd; Zhao, Lu; Stamnes, Jakob J; Kildemo, Morten

2011-06-01

A Lambert surface would appear equally bright from all observation directions regardless of the illumination direction. However, the reflection from a randomly scattering object generally has directional variation, which can be described in terms of the bidirectional reflectance distribution function (BRDF). We measured the BRDF of a Spectralon white reflectance standard for incoherent illumination at 405 and 680 nm with unpolarized and plane-polarized light from different directions of incidence. Our measurements show deviations of the BRDF for the Spectralon white reflectance standard from that of a Lambertian reflector that depend both on the angle of incidence and the polarization states of the incident light and detected light. The non-Lambertian reflection characteristics were found to increase more toward the direction of specular reflection as the angle of incidence gets larger.

Photometric measurements of red blood cell aggregation: light transmission versus light reflectance

NARCIS (Netherlands)

Baskurt, O.K.; Uyuklu, M.; Hardeman, M.R.; Meiselman, H.J.

2009-01-01

Red blood cell (RBC) aggregation is the reversible and regular clumping in the presence of certain macromolecules. This is a clinically important phenomenon, being significantly enhanced in the presence of acute phase reactants (e. g., fibrinogen). Both light reflection (LR) and light transmission

Photometric measurements of red blood cell aggregation: light transmission versus light reflectance

NARCIS (Netherlands)

Baskurt, Oguz K.; Uyuklu, Mehmet; Hardeman, Max R.; Meiselman, Herbert J.

2009-01-01

Red blood cell (RBC) aggregation is the reversible and regular clumping in the presence of certain macromolecules. This is a clinically important phenomenon, being significantly enhanced in the presence of acute phase reactants (e.g., fibrinogen). Both light reflection (LR) and light transmission

Reflected Light Curves of Extrasolar Planets

Science.gov (United States)

Green, D.; Matthews, J.; Kuschnig, R.; Seager, S.

The planned launches of ultra-precise photometric satellites such as MOST, COROT and MONS should provide the first opportunity to study the reflected light curves from extrasolar planets. To predict the capabilities of these missions, we have constructed a series of models of such light curves, improving upon the Monte Carlo simulations by Seager et al. (2000). These models include more realistic features such limb darkening of the star and broad band photometry. For specific models, the resulting planet light curves exhibit unique behavior with the variation of radius, inclination and presence or absence of clouds.

Quantum Dot Immunocytochemical Localization of Somatostatin in Somatostatinoma by Widefield Epifluorescence, Super-resolution Light, and Immunoelectron Microscopy

Science.gov (United States)

Lai, Ken; Wu, Xiaojuan; Yong, Jim L. C.; Lee, C. Soon

2012-01-01

Quantum dot nanocrystal probes (QDs) have been used for detection of somatostatin hormone in secretory granules of somatostatinoma tumor cells by immunofluorescence light microscopy, super-resolution light microscopy, and immunoelectron microscopy. Immunostaining for all modalities was done using sections taken from an epoxy resin-embedded tissue specimen and a similar labeling protocol. This approach allowed assessment of labeling at light microscopy level before examination at super-resolution and electron microscopy level and was a significant aid in interpretation. Etching of ultrathin sections with saturated sodium metaperiodate was a critical step presumably able to retrieve some tissue antigenicity masked by processing in epoxy resin. Immunofluorescence microscopy of QD-immunolabeled sections showed somatostatin hormone localization in cytoplasmic granules. Some variable staining of tumor gland-like structures appeared related to granule maturity and dispersal of granule contents within the tumor cell cytoplasm. Super-resolution light microscopy demonstrated localization of somatostatin within individual secretory granules to be heterogeneous, and this staining pattern was confirmed by immunoelectron microscopy. PMID:22899862

Quantum dot immunocytochemical localization of somatostatin in somatostatinoma by Widefield Epifluorescence, super-resolution light, and immunoelectron microscopy.

Science.gov (United States)

Killingsworth, Murray C; Lai, Ken; Wu, Xiaojuan; Yong, Jim L C; Lee, C Soon

2012-11-01

Quantum dot nanocrystal probes (QDs) have been used for detection of somatostatin hormone in secretory granules of somatostatinoma tumor cells by immunofluorescence light microscopy, super-resolution light microscopy, and immunoelectron microscopy. Immunostaining for all modalities was done using sections taken from an epoxy resin-embedded tissue specimen and a similar labeling protocol. This approach allowed assessment of labeling at light microscopy level before examination at super-resolution and electron microscopy level and was a significant aid in interpretation. Etching of ultrathin sections with saturated sodium metaperiodate was a critical step presumably able to retrieve some tissue antigenicity masked by processing in epoxy resin. Immunofluorescence microscopy of QD-immunolabeled sections showed somatostatin hormone localization in cytoplasmic granules. Some variable staining of tumor gland-like structures appeared related to granule maturity and dispersal of granule contents within the tumor cell cytoplasm. Super-resolution light microscopy demonstrated localization of somatostatin within individual secretory granules to be heterogeneous, and this staining pattern was confirmed by immunoelectron microscopy.

Localization of fluorescently labeled structures in frozen-hydrated samples using integrated light electron microscopy.

Science.gov (United States)

Faas, F G A; Bárcena, M; Agronskaia, A V; Gerritsen, H C; Moscicka, K B; Diebolder, C A; van Driel, L F; Limpens, R W A L; Bos, E; Ravelli, R B G; Koning, R I; Koster, A J

2013-03-01

Correlative light and electron microscopy is an increasingly popular technique to study complex biological systems at various levels of resolution. Fluorescence microscopy can be employed to scan large areas to localize regions of interest which are then analyzed by electron microscopy to obtain morphological and structural information from a selected field of view at nm-scale resolution. Previously, an integrated approach to room temperature correlative microscopy was described. Combined use of light and electron microscopy within one instrument greatly simplifies sample handling, avoids cumbersome experimental overheads, simplifies navigation between the two modalities, and improves the success rate of image correlation. Here, an integrated approach for correlative microscopy under cryogenic conditions is presented. Its advantages over the room temperature approach include safeguarding the native hydrated state of the biological specimen, preservation of the fluorescence signal without risk of quenching due to heavy atom stains, and reduced photo bleaching. The potential of cryo integrated light and electron microscopy is demonstrated for the detection of viable bacteria, the study of in vitro polymerized microtubules, the localization of mitochondria in mouse embryonic fibroblasts, and for a search into virus-induced intracellular membrane modifications within mammalian cells. Copyright © 2012 Elsevier Inc. All rights reserved.

An active one-particle microrheometer: incorporating magnetic tweezers to total internal reflection microscopy.

Science.gov (United States)

Gong, Xiangjun; Hua, Li; Wu, Chi; Ngai, To

2013-03-01

We present a novel microrheometer by incorporating magnetic tweezers in the total internal reflection microscopy (TIRM) that enables measuring of viscoelastic properties of materials near solid surface. An evanescent wave generated by a solid∕liquid interface in the TIRM is used as the incident light source in the microrheometer. When a probe particle (of a few micrometers diameter) moves near the interface, it can interact with the evanescent field and reflect its position with respect to the interface by the scattered light intensity. The exponential distance dependence of the evanescent field, on the one hand, makes this technique extremely sensitive to small changes from z-fluctuations of the probe (with a resolution of several nanometers), and on the other, it does not require imaging of the probe with high lateral resolution. Another distinct advantage is the high sensitivity in determining the z position of the probe in the absence of any labeling. The incorporated magnetic tweezers enable us to effectively manipulate the distance of the embedded particle from the interface either by a constant or an oscillatory force. The force ramp is easy to implement through a coil current ramp. In this way, the local viscous and elastic properties of a given system under different confinements can therefore be measured by resolving the near-surface particle motion. To test the feasibility of applying this microrheology to soft materials, we measured the viscoelastic properties of sucrose and poly(ethylene glycol) solutions and compared the results to bulk rheometry. In addition, we applied this technique in monitoring the structure and properties of deformable microgel particles near the flat surface.

Reflection of circularly polarized light and the effect of particle distribution on circular dichroism in evaporation induced self-assembled cellulose nanocrystal thin films

Directory of Open Access Journals (Sweden)

D. Hewson

2017-06-01

Full Text Available Evaporation induced self-assembled (EISA thin films of cellulose nanocrystals (CNCs have shown great potential for displaying structural colour across the visible spectrum. They are believed primarily to reflect left handed circularly polarised (LCP light due to their natural tendency to form structures comprising left handed chirality. Accordingly the fabrication of homogenously coloured CNC thin films is challenging. Deposition of solid material towards the edge of a dried droplet, via the coffee-stain effect, is one such difficulty in achieving homogenous colour across CNC films. These effects are most easily observed in films prepared from droplets where observable reflection of visible light is localised around the edge of the dry film. We report here, the observation of both left and right hand circularly polarised (LCP/RCP light in reflection from distinct separate regions of CNC EISA thin films and we elucidate how these reflections are dependent on the distribution of CNC material within the EISA thin film. Optical models of reflection are presented which are based on structures revealed using high resolution transmission electron microscopy (TEM images of film cross sections. We have also employed spectroscopic characterisation techniques to evaluate the distribution of solid CNC material within a selection of CNC EISA thin films and we have correlated this distribution with polarised light spectra collected from each film. We conclude that film regions from which RCP light was reflected were associated with lower CNC concentrations and thicker film regions.

Generating structured light with phase helix and intensity helix using reflection-enhanced plasmonic metasurface at 2 μm

Science.gov (United States)

Zhao, Yifan; Du, Jing; Zhang, Jinrun; Shen, Li; Wang, Jian

2018-04-01

Mid-infrared (2-20 μm) light has been attracting great attention in many areas of science and technology. Beyond the extended wavelength range from visible and near-infrared to mid-infrared, shaping spatial structures may add opportunities to grooming applications of mid-infrared photonics. Here, we design and fabricate a reflection-enhanced plasmonic metasurface and demonstrate efficient generation of structured light with the phase helix and intensity helix at 2 μm. This work includes two distinct aspects. First, structured light (phase helix, intensity helix) generation at 2 μm, which is far beyond the ability of conventional spatial light modulators, is enabled by the metasurface with sub-wavelength engineered structures. Second, the self-referenced intensity helix against environmental noise is generated without using a spatially separated light. The demonstrations may open up advanced perspectives to structured light applications at 2 μm, such as phase helix for communications and non-communications (imaging, sensing) and intensity helix for enhanced microscopy and advanced metrology.

Rapid diagnosis of malaria by fluorescent microscopy with light microscope and interface filter

International Nuclear Information System (INIS)

Hussain, I.; Tayyib, M.; Farooq, M.; Ahmed, N.

2008-01-01

The present study is planned to compare acridine orange (A.O) staining with Giemsa staining by using light microscopy with IF and also with fluorescent microscopy for detection of parasites in peripheral blood of patients suffering from clinically suspected cases of malaria. 200 patients with fever and shivering were included. General investigations like Hb, TLC and platelets were done by sysmex K-1000. Thin and thick blood films were made and stained according to protocol given i.e. by Giemsa and AO stains and slides were examined by different microscopes i.e. light microscope, light microscope with IFS and fluorescent microscope. Out of 200 subjects, 170 (85%) patients showed positive parasitaemia and 30 (15%) subjects were negative for malaria parasites. fib, TLC and platelets were reduced when comparing with MP negative cases. IFS microscope with acridine orange staining showed early detection of malaria parasites by counting fewer fields as compared to light microscopy with Giemsa stains. Time consumed for detection of parasites was also significantly reduced in IFS microscope by using AO stains. (author)

Localization of fluorescently labeled structures in frozen-hydrated samples using integrated light electron microscopy

NARCIS (Netherlands)

Faas, F.G.A.; Bárcena, M.A.; Agronskaia, A.V.; Gerritsen, H.C.; Moscicka, K.B.; Diebolder, C.A.; Driel, L.F.; Limpens, R.W.A.L.; Bos, E.; Ravelli, R.B.G.; Koning, R.I.; Koster, A.J.

2013-01-01

Correlative light and electron microscopy is an increasingly popular technique to study complex biological systems at various levels of resolution. Fluorescence microscopy can be employed to scan large areas to localize regions of interest which are then analyzed by electron microscopy to obtain

Emulation and design of terahertz reflection-mode confocal scanning microscopy based on virtual pinhole

Science.gov (United States)

Yang, Yong-fa; Li, Qi

2014-12-01

In the practical application of terahertz reflection-mode confocal scanning microscopy, the size of detector pinhole is an important factor that determines the performance of spatial resolution characteristic of the microscopic system. However, the use of physical pinhole brings some inconvenience to the experiment and the adjustment error has a great influence on the experiment result. Through reasonably selecting the parameter of matrix detector virtual pinhole (VPH), it can efficiently approximate the physical pinhole. By using this approach, the difficulty of experimental calibration is reduced significantly. In this article, an imaging scheme of terahertz reflection-mode confocal scanning microscopy that is based on the matrix detector VPH is put forward. The influence of detector pinhole size on the axial resolution of confocal scanning microscopy is emulated and analyzed. Then, the parameter of VPH is emulated when the best axial imaging performance is reached.

Nonimaging light concentration using total internal reflection films.

Science.gov (United States)

Ouellette, G; Waltham, C E; Drees, R M; Poon, A; Schubank, R; Whitehead, L A

1992-05-01

We present a method of fabricating nonimaging light concentrators from total internal reflection film. A prototype has been made and tested and found to operate in agreement with predictions of ray-tracing codes. The performance of the prototype is comparable with that of concentrators made from specular reflecting materials.

Reflection of a polarized light cone

Science.gov (United States)

Brody, Jed; Weiss, Daniel; Berland, Keith

2013-01-01

We introduce a visually appealing experimental demonstration of Fresnel reflection. In this simple optical experiment, a polarized light beam travels through a high numerical-aperture microscope objective, reflects off a glass slide, and travels back through the same objective lens. The return beam is sampled with a polarizing beam splitter and produces a surprising geometric pattern on an observation screen. Understanding the origin of this pattern requires careful attention to geometry and an understanding of the Fresnel coefficients for S and P polarized light. We demonstrate that in addition to a relatively simple experimental implementation, the shape of the observed pattern can be computed both analytically and by using optical modeling software. The experience of working through complex mathematical computations and demonstrating their agreement with a surprising experimental observation makes this a highly educational experiment for undergraduate optics or advanced-lab courses. It also provides a straightforward yet non-trivial system for teaching students how to use optical modeling software.

Live cell refractometry using Hilbert phase microscopy and confocal reflectance microscopy.

Science.gov (United States)

Lue, Niyom; Choi, Wonshik; Popescu, Gabriel; Yaqoob, Zahid; Badizadegan, Kamran; Dasari, Ramachandra R; Feld, Michael S

2009-11-26

Quantitative chemical analysis has served as a useful tool for understanding cellular metabolisms in biology. Among many physical properties used in chemical analysis, refractive index in particular has provided molecular concentration that is an important indicator for biological activities. In this report, we present a method of extracting full-field refractive index maps of live cells in their native states. We first record full-field optical thickness maps of living cells by Hilbert phase microscopy and then acquire physical thickness maps of the same cells using a custom-built confocal reflectance microscope. Full-field and axially averaged refractive index maps are acquired from the ratio of optical thickness to physical thickness. The accuracy of the axially averaged index measurement is 0.002. This approach can provide novel biological assays of label-free living cells in situ.

Light Reflectance Spectroscopy to Detect Positive Surgical Margins on Prostate Cancer Specimens.

Science.gov (United States)

Morgan, Monica S C; Lay, Aaron H; Wang, Xinlong; Kapur, Payal; Ozayar, Asim; Sayah, Maryam; Zeng, Li; Liu, Hanli; Roehrborn, Claus G; Cadeddu, Jeffrey A

2016-02-01

Intraoperative frozen section analysis is not routinely performed to determine positive surgical margins at radical prostatectomy due to time requirements and unproven clinical usefulness. Light reflectance spectroscopy, which measures light intensity reflected or backscattered from tissues, can be applied to differentiate malignant from benign tissue. We used a novel light reflectance spectroscopy probe to evaluate positive surgical margins on ex vivo radical prostatectomy specimens and correlate its findings with pathological examination. Patients with intermediate to high risk disease undergoing radical prostatectomy were enrolled. Light reflectance spectroscopy was performed on suspected malignant and benign prostate capsule immediately following organ extraction. Each light reflectance spectroscopy at 530 to 830 nm was analyzed and correlated with pathological results. A regression model and forward sequential selection algorithm were developed for optimal feature selection. Eighty percent of light reflectance spectroscopy data were selected to train a logistic regression model, which was evaluated by the remaining 20% data. This was repeated 5 times to calculate averaged sensitivity, specificity and accuracy. Light reflectance spectroscopy analysis was performed on 17 ex vivo prostate specimens, on which a total of 11 histologically positive and 22 negative surgical margins were measured. Two select features from 700 to 830 nm were identified as unique to malignant tissue. Cross-validation when performing the predictive model showed that the optical probe predicted positive surgical margins with 85% sensitivity, 86% specificity, 86% accuracy and an AUC of 0.95. Light reflectance spectroscopy can identify positive surgical margins accurately in fresh ex vivo radical prostatectomy specimens. Further study is required to determine whether such analysis may be used in real time to improve surgical decision making and decrease positive surgical margin rates

Attenuated total internal reflection infrared microscopy of multilayer plastic packaging foils.

Science.gov (United States)

van Dalen, Gerard; Heussen, Patricia C M; den Adel, Ruud; Hoeve, Robert B J

2007-06-01

Multilayer plastic foils are important packaging materials that are used to extend the shelf life of food products and drinks. Fourier transform infrared (FT-IR) spectroscopic imaging using attenuated total internal reflection (ATR) can be used for the identification and localization of different layers in multilayer foils. A new type of ATR crystal was used in combination with a linear array detector through which large sample areas (400 x 400 microm(2)) could be imaged with a pixel size of 1.6 microm. The method was tested on laminated plastic packing materials containing 5 to 12 layers. The results of the identification of the different materials using ATR-FT-IR were compared with differential scanning calorimetry (DSC) and the layer thickness of the individual layers measured by ATR-FT-IR was compared with polarized light microscopy (LM) and scanning electron microscopy (SEM). It has been demonstrated that individual layers with a thickness of about 3 microm could be identified in multilayer foils with a total thickness ranging from 100 to 150 microm. The results show a spatial resolution of about 4 microm (measured at wavenumbers ranging from 1000 to 1730 cm(-1)), which is about a factor of two better than can be obtained using transmission FT-IR imaging. An additional advantage of ATR is the ease of sample preparation. A good correspondence was found between visible and FT-IR images. The results of ATR-FT-IR imaging were in agreement with those obtained by LM, SEM, and DSC. ATR-FT-IR is superior to the combination of these techniques because it delivers both spatial and chemical information.

In vivo Diagnosis of Basal Cell Carcinoma Subtype by Reflectance Confocal Microscopy

NARCIS (Netherlands)

Peppelman, M.; Wolberink, E.A.W.; Blokx, W.A.M.; Kerkhof, P.C.M. van de; Erp, P.E.J. van; Gerritsen, M.J.P.

2013-01-01

Background: Reflectance confocal microscopy (RCM) is a noninvasive imaging technique. Currently, RCM is mainly used for the diagnosis of melanoma and nonmelanoma skin cancer including basal cell carcinoma (BCC). Until now, it has not been possible to distinguish between subtypes of BCC using RCM.

Imaging a seizure model in zebrafish with structured illumination light sheet microscopy

Science.gov (United States)

Liu, Yang; Dale, Savannah; Ball, Rebecca; VanLeuven, Ariel J.; Baraban, Scott; Sornborger, Andrew; Lauderdale, James D.; Kner, Peter

2018-02-01

Zebrafish are a promising vertebrate model for elucidating how neural circuits generate behavior under normal and pathological conditions. The Baraban group first demonstrated that zebrafish larvae are valuable for investigating seizure events and can be used as a model for epilepsy in humans. Because of their small size and transparency, zebrafish embryos are ideal for imaging seizure activity using calcium indicators. Light-sheet microscopy is well suited to capturing neural activity in zebrafish because it is capable of optical sectioning, high frame rates, and low excitation intensities. We describe work in our lab to use light-sheet microscopy for high-speed long-time imaging of neural activity in wildtype and mutant zebrafish to better understand the connectivity and activity of inhibitory neural networks when GABAergic signaling is altered in vivo. We show that, with light-sheet microscopy, neural activity can be recorded at 23 frames per second in twocolors for over 10 minutes allowing us to capture rare seizure events in mutants. We have further implemented structured illumination to increase resolution and contrast in the vertical and axial directions during high-speed imaging at an effective frame rate of over 7 frames per second.

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.

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......, an increase in the field of view over the use of a standard Gaussian beam by a factor of six is demonstrated. This implementation for light sheet microscopy opens up new possibilities across a wide range of biomedical applications, especially for the study of neuronal processes....

Spectrum of reflected light by self-focusing of light in a laser plasma

International Nuclear Information System (INIS)

Gorbunov, L.M.

1983-01-01

The spectrum of the radiation reflected by a laser-produced plasma is considered. In this situation, self-focusing occurs and a region of low density (caviton) is formed. It is shown that the process leads to a considerable broadening of the spectrum on the ''red'' side, and to the appearance of a line structure in the spectrum. The results can explain data for the reflected light spectrum [L. M. Gorbunov et al., FIAN Preprint No. 126 (1979)] as being due to the nonstationary self-focusing of light in a laser-produced plasma that has recently been observed [V. L. Artsimovich et al., FIAN Preprint No. 252 (1981); Sov. Phys. Doklady 27, 618 (1982)

A four-phase strategy for the implementation of reflectance confocal microscopy in dermatology

NARCIS (Netherlands)

Hoogedoorn, L.; Gerritsen, M.J.P.; Wolberink, E.A.W.; Peppelman, M.; Kerkhof, P.C.M. van de; Erp, P.E.J. van

2016-01-01

BACKGROUND: Reflectance confocal microscopy (RCM) is gradually implemented in dermatology. Strategies for further implementation and practical 'hands on' guidelines are lacking. OBJECTIVE: The primary outcome was to conduct a general strategy for further implementation of RCM. The secondary outcome

Do cephalopods communicate using polarized light reflections from their skin?

Science.gov (United States)

Mäthger, Lydia M; Shashar, Nadav; Hanlon, Roger T

2009-07-01

Cephalopods (squid, cuttlefish and octopus) are probably best known for their ability to change color and pattern for camouflage and communication. This is made possible by their complex skin, which contains pigmented chromatophore organs and structural light reflectors (iridophores and leucophores). Iridophores create colorful and linearly polarized reflective patterns. Equally interesting, the photoreceptors of cephalopod eyes are arranged in a way to give these animals the ability to detect the linear polarization of incoming light. The capacity to detect polarized light may have a variety of functions, such as prey detection, navigation, orientation and contrast enhancement. Because the skin of cephalopods can produce polarized reflective patterns, it has been postulated that cephalopods could communicate intraspecifically through this visual system. The term 'hidden' or 'private' communication channel has been given to this concept because many cephalopod predators may not be able to see their polarized reflective patterns. We review the evidence for polarization vision as well as polarization signaling in some cephalopod species and provide examples that tend to support the notion--currently unproven--that some cephalopods communicate using polarized light signals.

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.

Contributed Review: Review of integrated correlative light and electron microscopy

International Nuclear Information System (INIS)

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

Reflection and refraction of light from a moving block of glass

International Nuclear Information System (INIS)

Lerche, I.

1975-01-01

For a block of glass moving with speed βc we present calculations which give the changes in the laws of reflection and refraction of light from the laws which hold when β = 0. In particular we show (i) that changes in the critical internal reflection condition, although formally of order β 2 , are obtainable by measuring the refraction angle to O(β); (ii) the reflection coefficient depends not only on the polarization plane of the incident light but also on whether the plane of incidence does, or does not, include the direction of motion of the glass block. For instance, for light incident in a plane perpendicular to the direction of motion there is no Brewster angle for either plane of polarization, while for light incident in a plane containing the direction of motion at least one (and sometimes both) plane of polarization possess a Brewster angle. We have done these calculations in order to demonstrate that the shearing of one medium relative to another can have profound effects on the reflection and refraction of light. A major point here is that even when it might appear a priori that O(β 2 ) phenomena are unobservable in the laboratory, the present calculations show that this is not always so

An overview of the legislation and light microscopy for detection of processed animal proteins in feeds.

Science.gov (United States)

Liu, Xian; Han, Lujia; Veys, Pascal; Baeten, Vincent; Jiang, Xunpeng; Dardenne, Pierre

2011-08-01

From the first cases of bovine spongiform encephalopathy (BSE) among cattle in the United Kingdom in 1986, the route of infection of BSE is generally believed by means of feeds containing low level of processed animal proteins (PAPs). Therefore, many feed bans and alternative and complementary techniques were resulted for the BSE safeguards in the world. Now the feed bans are expected to develop into a "species to species" ban, which requires the corresponding species-specific identification methods. Currently, banned PAPs can be detected by various methods as light microscopy, polymerase chain reaction, enzyme-linked immunosorbent assay, near infrared spectroscopy, and near infrared microscopy. Light microscopy as described in the recent Commission Regulation EC/152/2009 is the only official method for the detection and characterization of PAPs in feed in the European Union. It is able to detect the presence of constituents of animal origin in feed at the level of 1 g/kg with hardly any false negative. Nevertheless, light microscopy has the limitation of lack of species specificity. This article presents a review of legislations on the use of PAPs in feedstuff, the detection details of animal proteins by light microscopy, and also presents and discusses the analysis procedure and expected development of the technique. Copyright © 2010 Wiley-Liss, Inc.

Developement of a Light Attenuator Based on Glassy Reflections ...

African Journals Online (AJOL)

The light intensity control of a luminous source is a very important operation in many optical applications. Several types of light attenuator exploiting different optical phenomena like diffraction, absorption, and reflection exist and they differ principally in the maximum attenuation rate, the control range, the sensitivity and the ...

Method of Detecting Coliform Bacteria from Reflected Light

Science.gov (United States)

Vincent, Robert K. (Inventor)

2014-01-01

The present invention relates to a method of detecting coliform bacteria in water from reflected light, and also includes devices for the measurement, calculation and transmission of data relating to that method.

Method of Detecting Coliform Bacteria and Escherichia Coli Bacteria from Reflected Light

Science.gov (United States)

Vincent, Robert (Inventor)

2013-01-01

The present invention relates to a method of detecting coliform bacteria in water from reflected light and a method of detecting Eschericha Coli bacteria in water from reflected light, and also includes devices for the measurement, calculation and transmission of data relating to that method.

Noise analysis of a white-light supercontinuum light source for multiple wavelength confocal laser scanning fluorescence microscopy

Energy Technology Data Exchange (ETDEWEB)

McConnell, Gail [Centre for Biophotonics, Strathclyde Institute for Biomedical Sciences, University of Strathclyde, 27 Taylor Street, Glasgow, G4 0NR (United Kingdom)

2005-08-07

Intensity correlations of a Ti : sapphire, Kr/Ar and a white-light supercontinuum were performed to quantify the typical signal amplitude fluctuations and hence ascertain the comparative output stability of the white-light supercontinuum source for confocal laser scanning microscopy (CLSM). Intensity correlations across a two-pixel sample (n = 1000) of up to 98%, 95% and 94% were measured for the Ti : sapphire, Kr/Ar and white-light supercontinuum source, respectively. The white-light supercontinuum noise level is therefore acceptable for CLSM, with the added advantage of wider wavelength flexibility over traditional CLSM excitation sources. The relatively low-noise white-light supercontinuum was then used to perform multiple wavelength sequential CLSM of guinea pig detrusor to confirm the reliability of the system and to demonstrate system flexibility.

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

Science.gov (United States)

Gualda, Emilio J.; Simão, Daniel; Pinto, Catarina; Alves, Paula M.; Brito, Catarina

2014-01-01

The development of three dimensional (3D) 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 3D matrix, highly resembling physiological conditions. Light sheet fluorescence microscopy (LSFM) is becoming an excellent tool for fast imaging of such 3D 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. PMID:25161607

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.

Towards comprehensive cell lineage reconstructions in complex organisms using light-sheet microscopy.

Science.gov (United States)

Amat, Fernando; Keller, Philipp J

2013-05-01

Understanding the development of complex multicellular organisms as a function of the underlying cell behavior is one of the most fundamental goals of developmental biology. The ability to quantitatively follow cell dynamics in entire developing embryos is an indispensable step towards such a system-level understanding. In recent years, light-sheet fluorescence microscopy has emerged as a particularly promising strategy for recording the in vivo data required to realize this goal. Using light-sheet fluorescence microscopy, entire complex organisms can be rapidly imaged in three dimensions at sub-cellular resolution, achieving high temporal sampling and excellent signal-to-noise ratio without damaging the living specimen or bleaching fluorescent markers. The resulting datasets allow following individual cells in vertebrate and higher invertebrate embryos over up to several days of development. However, the complexity and size of these multi-terabyte recordings typically preclude comprehensive manual analyses. Thus, new computational approaches are required to automatically segment cell morphologies, accurately track cell identities and systematically analyze cell behavior throughout embryonic development. We review current efforts in light-sheet microscopy and bioimage informatics towards this goal, and argue that comprehensive cell lineage reconstructions are finally within reach for many key model organisms, including fruit fly, zebrafish and mouse. © 2013 The Authors Development, Growth & Differentiation © 2013 Japanese Society of Developmental Biologists.

Reflectance confocal microscopy features of thin versus thick melanomas.

Science.gov (United States)

Kardynal, Agnieszka; Olszewska, Małgorzata; de Carvalho, Nathalie; Walecka, Irena; Pellacani, Giovanni; Rudnicka, Lidia

2018-01-24

In vivo reflectance confocal microscopy (RCM) plays an increasingly important role in differential diagnosis of melanoma. The aim of the study was to assess typical confocal features of thin (≤1mm according to Breslow index) versus thick (>1mm) melanomas. 30 patients with histopathologically confirmed cutaneous melanoma were included in the study. Reflectance confocal microscopy was performed with Vivascope equipment prior to excision. Fifteen melanomas were thin (Breslow thickness ≤ 1mm) and 15 were thick melanomas (Breslow thickness >1mm). In the RCM examination, the following features were more frequently observed in thin compared to thick melanomas: edged papillae (26.7% vs 0%, p=0.032) and areas with honeycomb or cobblestone pattern (33.3% vs 6.7%, p=0.068). Both features are present in benign melanocytic lesions, so in melanoma are good prognostic factors. The group of thick melanomas compared to the group of thin melanomas in the RCM images presented with greater frequency of roundish cells (100% vs 40%, p=0.001), non-edged papillae (100% vs 60%, p=0.006), numerous pagetoid cells (73.3% vs 33.3%, p=0.028), numerous atypical cells at dermal-epidermal junction (53.3% vs 20%, p=0.058) and epidermal disarray (93.3% vs 66.7%, p=0.068). Non-invasive imaging methods helps in deepening of knowledge about the evolution and biology of melanoma. The most characteristic features for thin melanomas in confocal examination are: fragments of cobblestone or honeycomb pattern and edged papillae (as good prognostic factors). The features of thick melanomas in RCM examination are: roundish cells, non-edged papillae, numerous pagetoid cells at dermal-epidermal junction and epidermal disarray.

The development of optical microscopy techniques for the advancement of single-particle studies

Science.gov (United States)

Marchuk, Kyle

Single particle orientation and rotational tracking (SPORT) has recently become a powerful optical microscopy tool that can expose many molecular motions. Unfortunately, there is not yet a single microscopy technique that can decipher all particle motions in all environmental conditions, thus there are limitations to current technologies. Within, the two powerful microscopy tools of total internal reflection and interferometry are advanced to determine the position, orientation, and optical properties of metallic nanoparticles in a variety of environments. Total internal reflection is an optical phenomenon that has been applied to microscopy to produce either fluorescent or scattered light. The non-invasive far-field imaging technique is coupled with a near-field illumination scheme that allows for better axial resolution than confocal microscopy and epi-fluorescence microscopy. By controlling the incident illumination angle using total internal reflection fluorescence (TIRF) microscopy, a new type of imaging probe called "non-blinking" quantum dots (NBQDs) were super-localized in the axial direction to sub-10-nm precision. These particles were also used to study the rotational motion of microtubules being propelled by the motor protein kinesin across the substrate surface. The same instrument was modified to function under total internal reflection scattering (TIRS) microscopy to study metallic anisotropic nanoparticles and their dynamic interactions with synthetic lipid bilayers. Utilizing two illumination lasers with opposite polarization directions at wavelengths corresponding to the short and long axis surface plasmon resonance (SPR) of the nanoparticles, both the in-plane and out-of-plane movements of many particles could be tracked simultaneously. When combined with Gaussian point spread function (PSF) fitting for particle super-localization, the binding status and rotational movement could be resolved without degeneracy. TIRS microscopy was also used to

The development of optical microscopy techniques for the advancement of single-particle studies

Energy Technology Data Exchange (ETDEWEB)

Marchuk, Kyle [Iowa State Univ., Ames, IA (United States)

2013-05-15

Single particle orientation and rotational tracking (SPORT) has recently become a powerful optical microscopy tool that can expose many molecular motions. Unfortunately, there is not yet a single microscopy technique that can decipher all particle motions in all environmental conditions, thus there are limitations to current technologies. Within, the two powerful microscopy tools of total internal reflection and interferometry are advanced to determine the position, orientation, and optical properties of metallic nanoparticles in a variety of environments. Total internal reflection is an optical phenomenon that has been applied to microscopy to produce either fluorescent or scattered light. The non-invasive far-field imaging technique is coupled with a near-field illumination scheme that allows for better axial resolution than confocal microscopy and epi-fluorescence microscopy. By controlling the incident illumination angle using total internal reflection fluorescence (TIRF) microscopy, a new type of imaging probe called “non-blinking” quantum dots (NBQDs) were super-localized in the axial direction to sub-10-nm precision. These particles were also used to study the rotational motion of microtubules being propelled by the motor protein kinesin across the substrate surface. The same instrument was modified to function under total internal reflection scattering (TIRS) microscopy to study metallic anisotropic nanoparticles and their dynamic interactions with synthetic lipid bilayers. Utilizing two illumination lasers with opposite polarization directions at wavelengths corresponding to the short and long axis surface plasmon resonance (SPR) of the nanoparticles, both the in-plane and out-of-plane movements of many particles could be tracked simultaneously. When combined with Gaussian point spread function (PSF) fitting for particle super-localization, the binding status and rotational movement could be resolved without degeneracy. TIRS microscopy was also used to

Monte Carlo simulation of light reflection from cosmetic powders on the skin

Science.gov (United States)

Okamoto, Takashi; Motoda, Masafumi; Igarashi, Takanori; Nakao, Keisuke

2011-07-01

The reflection and scattering properties of light incident on skin covered with powder particles have been investigated. A three-layer skin structure with a spot is modeled, and the propagation of light in the skin and the scattering of light by particles on the skin surface are simulated by means of a Monte Carlo method. Under the condition in which only single scattering of light occurs in the powder layer, the reflection spectra of light from the skin change dramatically with the size of powder particles. The color difference between normal skin and spots is found to diminish more when powder particles smaller than the wavelength of light are used. It is shown that particle polydispersity suppresses substantially the extreme spectral change caused by monodisperse particles with a size comparable to the light wavelength.

Microplate-compatible total internal reflection fluorescence microscopy for receptor pharmacology

Science.gov (United States)

Chen, Minghan; Zaytseva, Natalya V.; Wu, Qi; Li, Min; Fang, Ye

2013-05-01

We report the use of total internal reflection fluorescence (TIRF) microscopy for analyzing receptor pharmacology and the development of a microplate-compatible TIRF imaging system. Using stably expressed green fluorescence protein tagged β2-adrenergic receptor as the reporter, we found that the activation of different receptors results in distinct kinetic signatures of the TIRF intensity of cells. These TIRF signatures closely resemble the characteristics of their respective label-free dynamic mass redistribution signals in the same cells. This suggests that TIRF in microplate can be used for profiling and screening drugs.

Total Internal Reflection Fluorescence Microscopy Imaging-Guided Confocal Single-Molecule Fluorescence Spectroscopy

OpenAIRE

Zheng, Desheng; Kaldaras, Leonora; Lu, H. Peter

2013-01-01

We have developed an integrated spectroscopy system combining total internal reflection fluorescence microscopy imaging with confocal single-molecule fluorescence spectroscopy for two-dimensional interfaces. This spectroscopy approach is capable of both multiple molecules simultaneously sampling and in situ confocal fluorescence dynamics analyses of individual molecules of interest. We have demonstrated the calibration with fluorescent microspheres, and carried out single-molecule spectroscop...

Further improvement in the light output power of InGaN-based light emitting diodes by reflective current blocking design

International Nuclear Information System (INIS)

Tsai, Chun-Fu; Su, Yan-Kuin; Lin, Chun-Liang

2011-01-01

In this study, the fabrication and characterization of InGaN/GaN multiple-quantum-well light-emitting diodes (LEDs) with further improvement by the design of a reflective current blocking layer (CBL) were described, and these are demonstrated to be an inexpensive and feasible way for improving the performance of LEDs. With the reflective CBL, not only was the injected current forced to spread outside instead of flowing directly downward under a p-pad, but the light generated from the active region could also be extracted outside of the LED by reflection under the p-pad. At 20 mA, as compared to the conventional LED, the light output power of the LEDs with the normal and reflective CBL can be increased by 15.7% and 25.8%, respectively. We found that the forward voltages of the LEDs with CBL structure were both about 3.7 V at 20 mA, which was slightly higher than that of the conventional LED (3.6 V). In our experiment, the further increase in the light output power of the reflective CBL LED could be attributed to more current injection into the light-emitting active region outside of the p-pad by the CBL and a reduction in optical absorption at the p-pad with more extraction by the reflective design

Proximal design for a multimodality endoscope with multiphoton microscopy, optical coherence microscopy and visual modalities

Science.gov (United States)

Kiekens, Kelli C.; Talarico, Olivia; Barton, Jennifer K.

2018-02-01

A multimodality endoscope system has been designed for early detection of ovarian cancer. Multiple illumination and detection systems must be integrated in a compact, stable, transportable configuration to meet the requirements of a clinical setting. The proximal configuration presented here supports visible light navigation with a large field of view and low resolution, high resolution multiphoton microscopy (MPM), and high resolution optical coherence microscopy (OCM). All modalities are integrated into a single optical system in the endoscope. The system requires two light sources: a green laser for visible light navigation and a compact fiber based femtosecond laser for MPM and OCM. Using an inline wavelength division multiplexer, the two sources are combined into a single mode fiber. To accomplish OCM, a fiber coupler is used to separate the femtosecond laser into a reference arm and signal arm. The reflected reference arm and the signal from the sample are interfered and wavelength separated by a reflection grating and detected using a linear array. The MPM signal is collimated and goes through a series of filters to separate the 2nd and 3rd harmonics as well as twophoton excitation florescence (2PEF) and 3PEF. Each signal is independently detected on a photo multiplier tube and amplified. The visible light is collected by multiple high numerical aperture fibers at the endoscope tip which are bundled into one SMA adapter at the proximal end and connected to a photodetector. This integrated system design is compact, efficient and meets both optical and mechanical requirements for clinical applications.

On the reflection point where light reflects to a known destination on quadratic surfaces.

Science.gov (United States)

Gonçalves, Nuno

2010-01-15

We address the problem of determining the reflection point on a specular surface where a light ray that travels from a source to a target is reflected. The specular surfaces considered are those expressed by a quadratic equation. So far, there is no closed form explicit equation for the general solution of this determination of the reflection point, and the usual approach is to use the Snell law or the Fermat principle whose equations are derived in multidimensional nonlinear minimizations. We prove in this Letter that one can impose a set of three restrictions to the reflection point that can impose a set of three restrictions that culminates in a very elegant formalism of searching the reflection point in a unidimensional curve in space. This curve is the intersection of two quadratic equations. Some applications of this framework are also discussed.

Three-dimensional particle tracking in concave structures made by ultraviolet nanoimprint via total internal reflection fluorescence microscopy and refractive-index-matching method

Science.gov (United States)

Fujinami, Taku; Kigami, Hiroshi; Unno, Noriyuki; Taniguchi, Jun; Satake, Shin-ichi

2018-03-01

Total internal reflection fluorescence microscopy (TIRFM) is a promising method for measuring fluid flow close to a wall with nanoscale resolution in a process that is termed "multilayer nanoparticle image velocimetry" (MnPIV). TIRFM uses evanescent light that is generated on a substrate (typically a glass slide) by total internal reflection of light. Many researchers have previously studied x-y-z (3D) flows of water close to flat glass slides using MnPIV. On the other hand, a fluid flow close to a structured surface is also important. To measure flows of water near micro-patterns, we previously developed an MnPIV technique that uses a refractive-index-matching method. In previous study, the micropattern is made of a thermoplastic material with a refractive index that closely matches that of water. In this study, ultraviolet nanoimprint lithography was used for fabricating the appropriate micro-patterns because this technique can fabricate a pattern with a high resolution. As a result, we succeeded in performing MnPIV in water with a circular hole array pattern made by ultraviolet nanoimprint using a refractive-index-matching method. We believe that this technique will be helpful in elucidating fluid flows around microstructures.

Imaging rat esophagus using combination of reflectance confocal and multiphoton microscopy

International Nuclear Information System (INIS)

Zhuo, S M; Chen, J X; Jiang, X S; Lu, K C; Xie, S S

2008-01-01

We combine reflectance confocal microscopy (RCM) with multiphoton microscopy (MPM) to image rat esophagus. The two imaging modalities allow detection of layered–resolved complementary information from esophagus. In the keratinizing layer, the keratinocytes boundaries can be characterized by RCM, while the keratinocytes cytoplasm (keratin) can be further imaged by multiphoton autofluorescence signal. In the epithelium, the epithelial cellular boundaries and nucleus can be detected by RCM, and MPM can be used for imaging epithelial cell cytoplasm and monitoring metabolic state of epithelium. In the stroma, multiphoton autofluorescence signal is used to image elastin and second harmonic generation signal is utilized to detect collagen, while RCM is used to determine the optical property of stroma. Overall, these results suggest that the combination of RCM and MPM has potential to provide more important and comprehensive information for early diagnosis of esophageal cancer

Multispectral digital lensless holographic microscopy: from femtosecond laser to white light LED

International Nuclear Information System (INIS)

Garcia-Sucerquia, J

2015-01-01

The use of femtosecond laser radiation and super bright white LED in digital lensless holographic microscopy is presented. For the ultrafast laser radiation two different configurations of operation of the microscope are presented and the dissimilar performance of each one analyzed. The microscope operating with a super bright white light LED in combination with optical filters shows very competitive performance as it is compared with more expensive optical sources. The broadband emission of both radiation sources allows the multispectral imaging of biological samples to obtain spectral responses and/or full color images of the microscopic specimens; sections of the head of a Drosophila melanogaster fly are imaged in this contribution. The simple, solid, compact, lightweight, and reliable architecture of digital lensless holographic microscopy operating with broadband light sources to image biological specimens exhibiting micrometer-sized details is evaluated in the present contribution. (paper)

The light-sheet microscopy revolution

Science.gov (United States)

Girkin, J. M.; Carvalho, M. T.

2018-05-01

This paper reviews the rapid advances that have been made in one form of optical biological imaging in the last decade, namely that of light sheet microscopy. Although the concept was originally presented over one hundred years ago, at the time it was a methodology that lacked the technology to really make it a viable tool for practical everyday imaging in the biologist’s laboratory. However, since its re-discovery, it has started to transform in vivo and increasingly intact organ imaging in a number of areas of biology. This review looks back at the beginning of the method and then the crucial role that modern optical technology, frequently developed for other fields, has played in advancing the instrumentation. This paper will also look at the OpenSPIM route that was developed whereby, through the purchase of a few optical components, researchers have been able to develop their own bespoke instruments and we consider if this may be a route forward for the rapid development of other technological breakthroughs.

Electro-optic study of PZT ferroelectric ceramics using modulation of reflected light

Science.gov (United States)

Kniazkov, A. V.

2016-04-01

Electro-optic coefficients of variations in the refractive index of PZT and PLZT ceramic materials induced by ac electric field are estimated using modulation of reflected light. The electro-optic coefficients of PLZT ceramics measured with the aid of conventional birefringence using the phase shift of transmitted radiation and the proposed method of birefringence using the modulation of reflected light are compared.

Graphical Approach to Fresnel's Equations for Reflection and Refraction of Light.

Science.gov (United States)

Doyle, William T.

1980-01-01

Develops a coordinate-free approach to Fresnel's equations for the reflection and refraction of light at a plane interface. Describes a graphical construction for finding the vector amplitudes of the reflected and transmitted waves. (Author/CS)

Structure of ordered polyelectrolyte films from atomic-force microscopy and X-ray reflectivity data

International Nuclear Information System (INIS)

Belyaev, V.V.; Tolstikhina, A.L.; Stepina, N.D.; Kayushina, R.L.

1998-01-01

The possible application of atomic-force microscopy and X-ray reflectometry methods to structural studies of polyelectrolyte films obtained due to alternating adsorption of oppositely charged polyanion [sodium polysterenesulfonate (PSS)] and polycation [poly(allylamine) hydrochloride (PAA)] layers on solid substrates has been considered. The atomic-force microscopy study has revealed the characteristic features of the surface topography of samples consisting of different numbers of polyelectrolyte layers deposited from solutions characterized by different ionic strength values. It is shown that the shape of the reflectivity curves obtained from thin polyelectrolyte films depends on their surface structure

Setting up and running an advanced light microscopy and imaging facility.

Science.gov (United States)

Sánchez, Carlos; Muñoz, Ma Ángeles; Villalba, Maite; Labrador, Verónica; Díez-Guerra, F Javier

2011-07-01

During the last twenty years, interest in light microscopy and imaging techniques has grown in various fields, such as molecular and cellular biology, developmental biology, and neurobiology. In addition, the number of scientific articles and journals using these techniques is rapidly increasing. Nowadays, most research institutions require sophisticated microscopy systems to cover their investigation demands. In general, such instruments are too expensive and complex to be purchased and managed by a single laboratory or research group, so they have to be shared with other groups and supervised by specialized personnel. This is the reason why microscopy and imaging facilities are becoming so important at research institutions nowadays. In this unit, we have gathered and presented a number of issues and considerations from our own experience that we hope will be helpful when planning or setting up a new facility.

Microcavity-coupled fiber Bragg grating with tunable reflection spectra and speed of light.

Science.gov (United States)

Chen, Lei; Han, Ya; Liu, Qian; Liu, Yan-Ge; Zhang, Weigang; Chou, Keng C

2018-04-15

After a fiber Bragg grating (FBG) is fabricated, the reflection spectrum of the FBG is generally not tunable without mechanical deformation or temperature adjustment. Here we present a microcavity-coupled FBG with both a tunable reflection lineshape and dispersion using electromagnetically induced transparency. The Fano interference of light in the FBG and the microcavity allows for dramatic modification of the reflection spectrum. The phase of the reflected spectrum is continuously tunable between 0 and 2π to produce various Fano lineshapes. The dispersion of the output light is adjustable from normal dispersion to abnormal dispersion, consequently providing an adjustable speed of light. Additionally, it allows the FBG to switch from a notch filter to a bandpass filter at the resonant wavelength, which is not possible in a conventional uniform FBG.

Comparing phototoxicity during the development of a zebrafish craniofacial bone using confocal and light sheet fluorescence microscopy techniques.

Science.gov (United States)

Jemielita, Matthew; Taormina, Michael J; Delaurier, April; Kimmel, Charles B; Parthasarathy, Raghuveer

2013-12-01

The combination of genetically encoded fluorescent proteins and three-dimensional imaging enables cell-type-specific studies of embryogenesis. Light sheet microscopy, in which fluorescence excitation is provided by a plane of laser light, is an appealing approach to live imaging due to its high speed and efficient use of photons. While the advantages of rapid imaging are apparent from recent work, the importance of low light levels to studies of development is not well established. We examine the zebrafish opercle, a craniofacial bone that exhibits pronounced shape changes at early developmental stages, using both spinning disk confocal and light sheet microscopies of fluorescent osteoblast cells. We find normal and aberrant opercle morphologies for specimens imaged with short time intervals using light sheet and spinning disk confocal microscopies, respectively, under equivalent exposure conditions over developmentally-relevant time scales. Quantification of shapes reveals that the differently imaged specimens travel along distinct trajectories in morphological space. Copyright © 2013 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Single-molecule fluorescence microscopy review: shedding new light on old problems.

Science.gov (United States)

Shashkova, Sviatlana; Leake, Mark C

2017-08-31

Fluorescence microscopy is an invaluable tool in the biosciences, a genuine workhorse technique offering exceptional contrast in conjunction with high specificity of labelling with relatively minimal perturbation to biological samples compared with many competing biophysical techniques. Improvements in detector and dye technologies coupled to advances in image analysis methods have fuelled recent development towards single-molecule fluorescence microscopy, which can utilize light microscopy tools to enable the faithful detection and analysis of single fluorescent molecules used as reporter tags in biological samples. For example, the discovery of GFP, initiating the so-called 'green revolution', has pushed experimental tools in the biosciences to a completely new level of functional imaging of living samples, culminating in single fluorescent protein molecule detection. Today, fluorescence microscopy is an indispensable tool in single-molecule investigations, providing a high signal-to-noise ratio for visualization while still retaining the key features in the physiological context of native biological systems. In this review, we discuss some of the recent discoveries in the life sciences which have been enabled using single-molecule fluorescence microscopy, paying particular attention to the so-called 'super-resolution' fluorescence microscopy techniques in live cells, which are at the cutting-edge of these methods. In particular, how these tools can reveal new insights into long-standing puzzles in biology: old problems, which have been impossible to tackle using other more traditional tools until the emergence of new single-molecule fluorescence microscopy techniques. © 2017 The Author(s).

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.

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.

Ultraviolet reflecting photonic microstructures in the King Penguin beak.

Science.gov (United States)

Dresp, Birgitta; Jouventin, Pierre; Langley, Keith

2005-09-22

King and emperor penguins (Aptenodytes patagonicus and Aptenodytes forsteri) are the only species of marine birds so far known to reflect ultraviolet (UV) light from their beaks. Unlike humans, most birds perceive UV light and several species communicate using the near UV spectrum. Indeed, UV reflectance in addition to the colour of songbird feathers has been recognized as an important signal when choosing a mate. The king penguin is endowed with several highly coloured ornaments, notably its beak horn and breast and auricular plumage, but only its beak reflects UV, a property considered to influence its sexual attraction. Because no avian UV-reflecting pigments have yet been identified, the origin of such reflections is probably structural. In an attempt to identify the structures that give rise to UV reflectance, we combined reflectance spectrophotometry and morphological analysis by both light and electron microscopy, after experimental removal of surface layers of the beak horn. Here, we characterize for the first time a multilayer reflector photonic microstructure that produces the UV reflections in the king penguin beak.

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.

Signal improvement in multiphoton microscopy by reflection with simple mirrors near the sample

Science.gov (United States)

Rehberg, Markus; Krombach, Fritz; Pohl, Ulrich; Dietzel, Steffen

2010-03-01

In conventional fluorescence or confocal microscopy, emitted light is generated not only in the focal plane but also above and below. The situation is different in multiphoton-induced fluorescence and multiphoton-induced higher harmonic generation. Here, restriction of signal generation to a single focal point permits that all emitted photons can contribute to image formation if collected, regardless of their path through the specimen. Often, the intensity of the emitted light is rather low in biological specimens. We present a method to significantly increase the fraction of photons collected by an epi (backward) detector by placing a simple mirror, an aluminum-coated coverslip, directly under the sample. Samples investigated include fluorescent test slides, collagen gels, and thin-layered, intact mouse skeletal muscles. Quantitative analysis revealed an intensity increase of second- and third-harmonic generated signal in skeletal muscle of nine- and sevenfold respectively, and of fluorescent signal in test slides of up to twofold. Our approach thus allows significant signal improvement also for situations were a forward detection is impossible, e.g., due to the anatomy of animals in intravital microscopy.

Optimizing low-light microscopy with back-illuminated electron multiplying charge-coupled device: enhanced sensitivity, speed, and resolution.

Science.gov (United States)

Coates, Colin G; Denvir, Donal J; McHale, Noel G; Thornbury, Keith D; Hollywood, Mark A

2004-01-01

The back-illuminated electron multiplying charge-coupled device (EMCCD) camera is having a profound influence on the field of low-light dynamic cellular microscopy, combining highest possible photon collection efficiency with the ability to virtually eliminate the readout noise detection limit. We report here the use of this camera, in 512 x 512 frame-transfer chip format at 10-MHz pixel readout speed, in optimizing a demanding ultra-low-light intracellular calcium flux microscopy setup. The arrangement employed includes a spinning confocal Nipkow disk, which, while facilitating the need to both generate images at very rapid frame rates and minimize background photons, yields very weak signals. The challenge for the camera lies not just in detecting as many of these scarce photons as possible, but also in operating at a frame rate that meets the temporal resolution requirements of many low-light microscopy approaches, a particular demand of smooth muscle calcium flux microscopy. Results presented illustrate both the significant sensitivity improvement offered by this technology over the previous standard in ultra-low-light CCD detection, the GenIII+intensified charge-coupled device (ICCD), and also portray the advanced temporal and spatial resolution capabilities of the EMCCD. Copyright 2004 Society of Photo-Optical Instrumentation Engineers.

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.

Invited Review Article: Advanced light microscopy for biological space research

International Nuclear Information System (INIS)

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

2014-01-01

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

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.

Cellular features of psoriatic skin: imaging and quantification using in vivo reflectance confocal microscopy

NARCIS (Netherlands)

Wolberink, E.A.W.; Erp, P.E.J. van; Teussink, M.M.; Kerkhof, P.C.M. van de; Gerritsen, M.J.P.

2011-01-01

BACKGROUND: In vivo reflectance confocal microscopy (RCM) is a novel, exciting imaging technique. It provides images of cell-and tissue structures and dynamics in situ, in real time, without the need for ex vivo tissue samples. RCM visualizes the superficial part of human skin up to a depth of 250

ELiXIR—Solid-State Luminaire With Enhanced Light Extraction by Internal Reflection

Science.gov (United States)

Allen, Steven C.; Steckl, Andrew J.

2007-06-01

A phosphor-converted light-emitting diode (pcLED) luminaire featuring enhanced light extraction by internal reflection (ELiXIR) with efficacy of 60 lm/W producing 18 lumens of yellowish green light at 100 mA is presented. The luminaire consists of a commercial blue high power LED, a polymer hemispherical shell lens with interior phosphor coating, and planar aluminized reflector. High extraction efficiency of the phosphor-converted light is achieved by separating the phosphor from the LED and using internal reflection to steer the light away from lossy reflectors and the LED package and out of the device. At 10 and 500 mA, the luminaire produces 2.1 and 66 lumens with efficacies of 80 and 37 lm/W, respectively. Technological improvements over existing commercial LEDs, such as more efficient pcLED packages or, alternatively, higher efficiency green or yellow for color mixing, will be essential to achieving 150 200 lm/W solid-state lighting. Advances in both areas are demonstrated.

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.

Correlative light and immuno-electron microscopy of retinal tissue cryostat sections

Science.gov (United States)

Burgoyne, Thomas; Lane, Amelia; Laughlin, William E.; Cheetham, Michael E.

2018-01-01

Correlative light-electron microscopy (CLEM) is a powerful technique allowing localisation of specific macromolecules within fluorescence microscopy (FM) images to be mapped onto corresponding high-resolution electron microscopy (EM) images. Existing methods are applicable to limited sample types and are technically challenging. Here we describe novel methods to perform CLEM and immuno-electron microscopy (iEM) on cryostat sections utilising the popular FM embedding solution, optimal cutting temperature (OCT) compound. Utilising these approaches, we have (i) identified the same phagosomes by FM and EM in the retinal pigment epithelium (RPE) of retinal tissue (ii) shown the correct localisation of rhodopsin on photoreceptor outer segment disc like-structures in iPSC derived optic cups and (iii) identified a novel interaction between peroxisomes and melanosomes as well as phagosomes in the RPE. These data show that cryostat sections allow easy characterisation of target macromolecule localisation within tissue samples, thus providing a substantial improvement over many conventional methods that are limited to cultured cells. As OCT embedding is routinely used for FM this provides an easily accessible and robust method for further analysis of existing samples by high resolution EM. PMID:29315318

'En face' ex vivo reflectance confocal microscopy to help the surgery of basal cell carcinoma of the eyelid.

Science.gov (United States)

Espinasse, Marine; Cinotti, Elisa; Grivet, Damien; Labeille, Bruno; Prade, Virginie; Douchet, Catherine; Cambazard, Frédéric; Thuret, Gilles; Gain, Philippe; Perrot, Jean Luc

2017-07-01

Ex vivo confocal microscopy is a recent imaging technique for the perioperative control of skin tumour margins. Up to date, it has been used in the fluorescence mode and with vertical sections of the specimen margins. The aim of this study was to evaluate its use in the reflectance mode and with a horizontal ('en face') scanning of the surgical specimen in a series of basal cell carcinoma of the eyelid. Prospective consecutive cohort study was performed at the University Hospital of Saint-Etienne, France. Forty-one patients with 42 basal cell carcinoma of the eyelid participated in this study. Basal cell carcinomas were excised with a 2-mm-wide clinically safe margin. The surgical specimens were analysed under ex vivo confocal microscopy in the reflectance mode and with an en face scanning in order to control at a microscopic level if the margins were free from tumour invasion. Histopathogical examination was later performed in order to compare the results. Sensitivity and specificity of ex vivo confocal microscopy for the presence of tumour-free margins. Ex vivo confocal microscopy results were consistent with histopathology in all cases (tumour-free margins in 40 out of 42 samples; sensitivity and specificity of 100%). Ex vivo confocal microscopy in the reflectance mode with an 'en face' scanning can control tumour margins of eyelid basal cell carcinomas and optimize their surgical management. This procedure has the advantage on the fluorescent mode of not needing any contrast agent to examine the samples. © 2016 Royal Australian and New Zealand College of Ophthalmologists.

Studies on silica deposition in sugarcane (Saccharum spp. ) using scanning electron microscopy, energy-dispersive X-ray analysis, neutron activation analysis, and light microscopy

Energy Technology Data Exchange (ETDEWEB)

Kaufman, P B; Takeoka, Y; Carlson, T J; Bigelow, W C; Jones, J D; Moore, P H; Ghosheh, N S [Michigan Univ., Ann Arbor (USA)

1979-06-01

Marked differences in silicon content in internodes of two sugarcane cultivars as revealed by neutron activation analysis, were closely correlated with number of silica cells per unit area in the epidermal system of the internodes of the two cultivars, as indicated by scanning electron microscopy and X-ray analysis. Light microscopy of epidermal peels showed that silica cells are capable of transmitting significantly more light through themselves than do other types of adjacent epidermal cells. This could be of great significance to total amount of carbon fixed by photosynthesizing mesophyll cells in leaves and cortical cells in internodes below the epidermis, especially in sugarcane cultivars with high densities of silica cells in their shoot epidermal systems. This has led to propose a window hypothesis, which indicates that silica cells in sugarcane, and in other grasses, act like windows in the epidermal system, allowing more light to be transmitted to photosynthetic tissue below than would occur if silica cells were absent.

Properties of light reflected from road signs in active imaging for driving safety

Science.gov (United States)

Halstuch, Aviran; Yitzhaky, Yitzhak

2007-10-01

Night-vision systems in vehicles are a new emerging technology. A crucial problem in active (illumination-based) systems is distortion of images by saturation and blooming, due to strong retro-reflections from road signs. In this work we quantified this phenomenon. We measured the Mueller matrices and the polarization state of the reflected light from three different types of road signs commonly used. Measurements of the reflected intensity were taken also with respect to the angle of reflection. We found that different types of signs have different reflection properties. It is concluded from our measurements that the optimal solution for attenuating the retro-reflected intensity is using a linear horizontal polarized light source and a linear vertical polarizer. Unfortunately, while the performance of this solution is good for two types of road signs, it is less efficient for the third sign type.

Effects of snow-reflected light levels on human visual comfort.

Science.gov (United States)

Yilmaz, Hasan; Demircioglu Yildiz, Nalan; Yilmaz, Sevgi

2008-09-01

The intensity of the sunlight reflected by the snow-covered surfaces is so high that it may disturb humans many times. This study aims to determine the reflected sunlight intensities from snow covered areas at points near (at a distance of 2 m) and under an individual tree and among trees (in the forest area) by accepting the open area as control; the reducing effects of the plant materials on reflected sunlight in percentage by comparing with the values of the open (control) area; and critical reflected sunlight threshold values for human visual comfort. The study was carried out over 22 clear and calm, i.e. sky was cloudless and wind was calm, days between the 1st and 31st days of January 2004, at 8:30 in the morning, at 12:30 at noon and at 14:30 in the afternoon in Erzurum. In order to determine the discomforting light intensity levels, 25 females and 26 male (totally 51) student subjects whose mean age was 20 and who had no visual disorders were selected. Considering the open area as control, mean reflected sunlight reducing effects were found to be 19.0, 66.0 and 82.7% for the 2 m near a tree, under a tree, and forest area, respectively. According to the responses of 51 subjects in the study, visually "very comfortable" range is between 5,000 and 8,000 lx; "comfortable" range is between 11,000 and 75,000 lx (mostly at 12,000 lx); "uncomfortable" condition is above the light intensity value of 43,000 lx and "very uncomfortable" condition is above the intensity of 80,000 lx. Great majority of the subjects (91%) found the value of 103,000 lx to be "very uncomfortable." As it is not an applicable way to use the great and dense tree masses in the cities, at least individual trees should be used along the main pedestrian axels in the cities having the same features with Erzurum to prevent the natural light pollution and discomforting effects of the snow-reflected sunlight.

Hyaline articular cartilage dissected by papain: light and scanning electron microscopy and micromechanical studies.

OpenAIRE

O'Connor, P; Brereton, J D; Gardner, D L

1984-01-01

Papain was used to digest the hyaline femoral condylar cartilages of 30 adult Wistar rats. Matrix proteoglycan degradation was assessed by the light microscopy of paraffin sections stained with toluidine blue. The extent of surface structural change was estimated by scanning electron microscopy, and the structural integrity of the hyaline cartilage tested by the controlled impact of a sharp pin. The results demonstrated an early loss of cartilage metachromasia, increasing with time of papain ...

Robust reflective ghost imaging against different partially polarized thermal light

Science.gov (United States)

Li, Hong-Guo; Wang, Yan; Zhang, Rui-Xue; Zhang, De-Jian; Liu, Hong-Chao; Li, Zong-Guo; Xiong, Jun

2018-03-01

We theoretically study the influence of degree of polarization (DOP) of thermal light on the contrast-to-noise ratio (CNR) of the reflective ghost imaging (RGI), which is a novel and indirect imaging modality. An expression for the CNR of RGI with partially polarized thermal light is carefully derived, which suggests a weak dependence of CNR on the DOP, especially when the ratio of the object size to the speckle size of thermal light has a large value. Different from conventional imaging approaches, our work reveals that RGI is much more robust against the DOP of the light source, which thereby has advantages in practical applications, such as remote sensing.

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.

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.

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

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

Decoupled illumination detection in light sheet microscopy for fast volumetric imaging

OpenAIRE

Olarte, Omar; Andilla, Jordi; Artigas García, David; Loza-Alvarez, Pablo

2015-01-01

Current microscopy demands the visualization of large three-dimensional samples with increased sensitivity, higher resolution, and faster speed. Several imaging techniques based on widefield, point-scanning, and light-sheet strategies have been designed to tackle some of these demands. Although successful, all these require the illuminated volumes to be tightly coupled with the detection optics to accomplish efficient optical sectioning. Here, we break this paradigm and produce optical sectio...

Evaluation of mobile digital light-emitting diode fluorescence microscopy in Hanoi, Viet Nam.

Science.gov (United States)

Chaisson, L H; Reber, C; Phan, H; Switz, N; Nilsson, L M; Myers, F; Nhung, N V; Luu, L; Pham, T; Vu, C; Nguyen, H; Nguyen, A; Dinh, T; Nahid, P; Fletcher, D A; Cattamanchi, A

2015-09-01

Hanoi Lung Hospital, Hanoi, Viet Nam. To compare the accuracy of CellScopeTB, a manually operated mobile digital fluorescence microscope, with conventional microscopy techniques. Patients referred for sputum smear microscopy to the Hanoi Lung Hospital from May to September 2013 were included. Ziehl-Neelsen (ZN) smear microscopy, conventional light-emitting diode (LED) fluorescence microscopy (FM), CellScopeTB-based LED FM and Xpert(®) MTB/RIF were performed on sputum samples. The sensitivity and specificity of microscopy techniques were determined in reference to Xpert results, and differences were compared using McNemar's paired test of proportions. Of 326 patients enrolled, 93 (28.5%) were Xpert-positive for TB. The sensitivity of ZN microscopy, conventional LED FM, and CellScopeTB-based LED FM was respectively 37.6% (95%CI 27.8-48.3), 41.9% (95%CI 31.8-52.6), and 35.5% (95%CI 25.8-46.1). The sensitivity of CellScopeTB was similar to that of conventional LED FM (difference -6.5%, 95%CI -18.2 to 5.3, P = 0.33) and ZN microscopy (difference -2.2%, 95%CI -9.2 to 4.9, P = 0.73). The specificity was >99% for all three techniques. CellScopeTB performed similarly to conventional microscopy techniques in the hands of experienced TB microscopists. However, the sensitivity of all sputum microscopy techniques was low. Options enabled by digital microscopy, such as automated imaging with real-time computerized analysis, should be explored to increase sensitivity.

Improving your four-dimensional image: traveling through a decade of light-sheet-based fluorescence microscopy research.

Science.gov (United States)

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

2017-06-01

Light-sheet-based fluorescence microscopy features optical sectioning in the excitation process. This reduces phototoxicity and photobleaching by up to four orders of magnitude compared with that caused by confocal fluorescence microscopy, simplifies segmentation and quantification for three-dimensional cell biology, and supports the transition from on-demand to systematic data acquisition in developmental biology applications.

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

OpenAIRE

Ayman F.A. Foad, MD

2017-01-01

Virtual microscopy has an established role in medical practice and education across all medical disciplines. It provides economical and pedagogical advantages, albeit with some shortcomings. 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 bette...

Application of Reflectance Confocal Microscopy in Dermatology Practice

Directory of Open Access Journals (Sweden)

Ayşe Esra Koku Aksu

2015-03-01

Full Text Available In vivo reflectance confocal microscopy (RCM is a non-invasive method, imaging cellular structures in living skin at a level close to the histological resolution. It is easier to diagnose melanocytic and non-melanocytic skin tumors especially in difficult cases when RCM features have been identified. Determination of the cellular features, presence of cellular and structural atypia with RCM allows the discrimination of benign and malignant lesions. Preoperative differential diagnosis of malignant lesions, determining preoperative lesion borders in complicated cases, identification of local recurrence after excision of malignant lesions, monitoring the treatment efficacy in patients using topical treatment and who can not be operated, are the main areas of RCM in tumoral lesions. Besides, RCM is helpful in the establishing the diagnosis of inflammatory disease like psoriasis, contact dermatitis, lichen planus and in evaluation of therapeutic efficacy, detecting of infestation like tinea, skabiyes, demodicosis and determining the level of bullae in bullous disease. Due to being noninvasive, RCM is preferred in cosmetology, in clinical research and practice for the evaluation of the effectiveness of cosmetic products and cosmetic procedures.

Reflection and transmission of light at periodic layered metamaterial films

Science.gov (United States)

Paul, Thomas; Menzel, Christoph; Śmigaj, Wojciech; Rockstuhl, Carsten; Lalanne, Philippe; Lederer, Falk

2011-09-01

The appropriate description of light scattering (transmission/reflection) at a bulky artificial medium, consisting of a sequence of functional metamaterial and natural material films, represents a major challenge in current theoretical nano-optics. Because in many relevant cases, in particular, in the optical domain, a metamaterial must not be described by an effective permittivity and permeability the usual Fresnel formalism cannot be applied. A reliable alternative consists in using a Bloch mode formalism known, e.g., from the theory of photonic crystals. It permits to split this complex issue into two more elementary ones, namely the study of light propagation in an infinitely extended metamaterial and the analysis of light scattering at interfaces between adjacent meta and natural materials. The first problem is routinely solved by calculating the relevant Bloch modes and their dispersion relations. The second task is more involved and represents the subject of the present study. It consists in using the general Bloch mode orthogonality to derive rigorous expressions for the reflection and transmission coefficients at an interface between two three-dimensional absorptive periodic media for arbitrary incidence. A considerable simplification can be achieved if only the fundamental Bloch modes of both media govern the scattering properties at the interface. If this approximation is valid, which depends on the longitudinal metamaterial period, the periodic metamaterial may be termed homogeneous. Only in this case the disentanglement of the fundamental modes of both media can be performed and the reflection/transmission coefficients can be expressed in terms of two impedances, each depending solely on the properties of the fundamental mode of the respective medium. In order to complement the picture, we apply the present formalism to the quite general problem of reflection/transmission at a metamaterial film sandwiched between a dissimilar metamaterial. This

Quantifying the Assembly of Multicomponent Molecular Machines by Single-Molecule Total Internal Reflection Fluorescence Microscopy.

Science.gov (United States)

Boehm, E M; Subramanyam, S; Ghoneim, M; Washington, M Todd; Spies, M

2016-01-01

Large, dynamic macromolecular complexes play essential roles in many cellular processes. Knowing how the components of these complexes associate with one another and undergo structural rearrangements is critical to understanding how they function. Single-molecule total internal reflection fluorescence (TIRF) microscopy is a powerful approach for addressing these fundamental issues. In this article, we first discuss single-molecule TIRF microscopes and strategies to immobilize and fluorescently label macromolecules. We then review the use of single-molecule TIRF microscopy to study the formation of binary macromolecular complexes using one-color imaging and inhibitors. We conclude with a discussion of the use of TIRF microscopy to examine the formation of higher-order (i.e., ternary) complexes using multicolor setups. The focus throughout this article is on experimental design, controls, data acquisition, and data analysis. We hope that single-molecule TIRF microscopy, which has largely been the province of specialists, will soon become as common in the tool box of biophysicists and biochemists as structural approaches have become today. © 2016 Elsevier Inc. All rights reserved.

The 'grey area' between small cell and non-small cell lung carcinomas. Light and electron microscopy versus clinical data in 14 cases

NARCIS (Netherlands)

Mooi, W. J.; van Zandwijk, N.; Dingemans, K. P.; Koolen, M. G.; Wagenvoort, C. A.

1986-01-01

We studied 14 lung tumours which on light microscopy had posed difficulties on classification as either small cell or non-small cell carcinomas. The light and electron microscopical features were compared with patient follow-up data. Electron microscopy showed neuroendocrine granules in 12 cases,

Reflectance dependence of polytetrafluoroethylene on thickness for xenon scintillation light

Energy Technology Data Exchange (ETDEWEB)

Haefner, J.; Neff, A.; Arthurs, M.; Batista, E.; Morton, D.; Okunawo, M.; Pushkin, K.; Sander, A. [Randall Laboratory of Physics, University of Michigan, Ann Arbor, MI 48109-1040 (United States); Stephenson, S. [Randall Laboratory of Physics, University of Michigan, Ann Arbor, MI 48109-1040 (United States); University of California Davis, Department of Physics, One Shields Ave., Davis, CA 95616 (United States); Wang, Y. [Randall Laboratory of Physics, University of Michigan, Ann Arbor, MI 48109-1040 (United States); Lorenzon, W., E-mail: lorenzon@umich.edu [Randall Laboratory of Physics, University of Michigan, Ann Arbor, MI 48109-1040 (United States)

2017-06-01

Many rare event searches including dark matter direct detection and neutrinoless double beta decay experiments take advantage of the high VUV reflective surfaces made from polytetrafluoroethylene (PTFE) reflector materials to achieve high light collection efficiency in their detectors. As the detectors have grown in size over the past decade, there has also been an increased need for ever thinner detector walls without significant loss in reflectance to reduce dead volumes around active noble liquids, outgassing, and potential backgrounds. We report on the experimental results to measure the dependence of the reflectance on thickness of two PTFE samples at wavelengths near 178 nm. No change in reflectance was observed as the wall thickness of a cylindrically shaped PTFE vessel immersed in liquid xenon was varied between 1 mm to 9.5 mm.

Inducing fluorescence of uranyl acetate as a dual-purpose contrast agent for correlative light-electron microscopy with nanometre precision.

Science.gov (United States)

Tuijtel, Maarten W; Mulder, Aat A; Posthuma, Clara C; van der Hoeven, Barbara; Koster, Abraham J; Bárcena, Montserrat; Faas, Frank G A; Sharp, Thomas H

2017-09-05

Correlative light-electron microscopy (CLEM) combines the high spatial resolution of transmission electron microscopy (TEM) with the capability of fluorescence light microscopy (FLM) to locate rare or transient cellular events within a large field of view. CLEM is therefore a powerful technique to study cellular processes. Aligning images derived from both imaging modalities is a prerequisite to correlate the two microscopy data sets, and poor alignment can limit interpretability of the data. Here, we describe how uranyl acetate, a commonly-used contrast agent for TEM, can be induced to fluoresce brightly at cryogenic temperatures (-195 °C) and imaged by cryoFLM using standard filter sets. This dual-purpose contrast agent can be used as a general tool for CLEM, whereby the equivalent staining allows direct correlation between fluorescence and TEM images. We demonstrate the potential of this approach by performing multi-colour CLEM of cells containing equine arteritis virus proteins tagged with either green- or red-fluorescent protein, and achieve high-precision localization of virus-induced intracellular membrane modifications. Using uranyl acetate as a dual-purpose contrast agent, we achieve an image alignment precision of ~30 nm, twice as accurate as when using fiducial beads, which will be essential for combining TEM with the evolving field of super-resolution light microscopy.

Hybrid fluorescence and electron cryo-microscopy for simultaneous electron and photon imaging.

Science.gov (United States)

Iijima, Hirofumi; Fukuda, Yoshiyuki; Arai, Yoshihiro; Terakawa, Susumu; Yamamoto, Naoki; Nagayama, Kuniaki

2014-01-01

Integration of fluorescence light and transmission electron microscopy into the same device would represent an important advance in correlative microscopy, which traditionally involves two separate microscopes for imaging. To achieve such integration, the primary technical challenge that must be solved regards how to arrange two objective lenses used for light and electron microscopy in such a manner that they can properly focus on a single specimen. To address this issue, both lateral displacement of the specimen between two lenses and specimen rotation have been proposed. Such movement of the specimen allows sequential collection of two kinds of microscopic images of a single target, but prevents simultaneous imaging. This shortcoming has been made up by using a simple optical device, a reflection mirror. Here, we present an approach toward the versatile integration of fluorescence and electron microscopy for simultaneous imaging. The potential of simultaneous hybrid microscopy was demonstrated by fluorescence and electron sequential imaging of a fluorescent protein expressed in cells and cathodoluminescence imaging of fluorescent beads. Copyright © 2013 Elsevier Inc. All rights reserved.

Reflection and transmission of polarized light by planetary atmospheres

International Nuclear Information System (INIS)

Rooij, W.A. de.

1985-01-01

In this thesis the reflection and transmission of sunlight by planetary atmospheres is studied, taking full account of the polarization of light. The atmospheres are treated as being locally plane-parallel, and are assumed to consist of a number of homogeneous layers, the lowest one being either a ground surface or a semi-infinite homogeneous layer. (Auth.)

Detection of UV-induced pigmentary and epidermal changes over time using in vivo reflectance confocal microscopy

NARCIS (Netherlands)

Middelkamp-Hup, Maritza A.; Park, H.-Y.; Lee, Jin; Gilchrest, Barbara A.; Gonzalez, Salvador

2006-01-01

In vivo reflectance confocal microscopy (RCM) provides high-resolution optical sections of the skin in its native state, without needing to fix or section the tissue. Melanin provides an excellent contrast for RCM, giving a bright signal in the confocal images. The pigmented guinea-pig is a common

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

Single-Molecule Light-Sheet Imaging of Suspended T Cells.

Science.gov (United States)

Ponjavic, Aleks; McColl, James; Carr, Alexander R; Santos, Ana Mafalda; Kulenkampff, Klara; Lippert, Anna; Davis, Simon J; Klenerman, David; Lee, Steven F

2018-05-08

Adaptive immune responses are initiated by triggering of the T cell receptor. Single-molecule imaging based on total internal reflection fluorescence microscopy at coverslip/basal cell interfaces is commonly used to study this process. These experiments have suggested, unexpectedly, that the diffusional behavior and organization of signaling proteins and receptors may be constrained before activation. However, it is unclear to what extent the molecular behavior and cell state is affected by the imaging conditions, i.e., by the presence of a supporting surface. In this study, we implemented single-molecule light-sheet microscopy, which enables single receptors to be directly visualized at any plane in a cell to study protein dynamics and organization in live, resting T cells. The light sheet enabled the acquisition of high-quality single-molecule fluorescence images that were comparable to those of total internal reflection fluorescence microscopy. By comparing the apical and basal surfaces of surface-contacting T cells using single-molecule light-sheet microscopy, we found that most coated-glass surfaces and supported lipid bilayers profoundly affected the diffusion of membrane proteins (T cell receptor and CD45) and that all the surfaces induced calcium influx to various degrees. Our results suggest that, when studying resting T cells, surfaces are best avoided, which we achieve here by suspending cells in agarose. Copyright © 2018. Published by Elsevier Inc.

Label free imaging of cell-substrate contacts by holographic total internal reflection microscopy.

Science.gov (United States)

Mandracchia, Biagio; Gennari, Oriella; Marchesano, Valentina; Paturzo, Melania; Ferraro, Pietro

2017-09-01

The study of cell adhesion contacts is pivotal to understand cell mechanics and interaction at substrates or chemical and physical stimuli. We designed and built a HoloTIR microscope for label-free quantitative phase imaging of total internal reflection. Here we show for the first time that HoloTIR is a good choice for label-free study of focal contacts and of cell/substrate interaction as its sensitivity is enhanced in comparison with standard TIR microscopy. Finally, the simplicity of implementation and relative low cost, due to the requirement of less optical components, make HoloTIR a reasonable alternative, or even an addition, to TIRF microscopy for mapping cell/substratum topography. As a proof of concept, we studied the formation of focal contacts of fibroblasts on three substrates with different levels of affinity for cell adhesion. © 2017 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim.

Combining total internal reflection sum frequency spectroscopy spectral imaging and confocal fluorescence microscopy.

Science.gov (United States)

Allgeyer, Edward S; Sterling, Sarah M; Gunewardene, Mudalige S; Hess, Samuel T; Neivandt, David J; Mason, Michael D

2015-01-27

Understanding surface and interfacial lateral organization in material and biological systems is critical in nearly every field of science. The continued development of tools and techniques viable for elucidation of interfacial and surface information is therefore necessary to address new questions and further current investigations. Sum frequency spectroscopy (SFS) is a label-free, nonlinear optical technique with inherent surface specificity that can yield critical organizational information on interfacial species. Unfortunately, SFS provides no spatial information on a surface; small scale heterogeneities that may exist are averaged over the large areas typically probed. Over the past decade, this has begun to be addressed with the advent of SFS microscopy. Here we detail the construction and function of a total internal reflection (TIR) SFS spectral and confocal fluorescence imaging microscope directly amenable to surface investigations. This instrument combines, for the first time, sample scanning TIR-SFS imaging with confocal fluorescence microscopy.

Light reflection by the cuticle of C. aurigans scarabs: a biological broadband reflector of left handed circularly polarized light

Science.gov (United States)

Libby, E.; Azofeifa, D. E.; Hernández-Jiménez, M.; Barboza-Aguilar, C.; Solís, A.; García-Aguilar, I.; Arce-Marenco, L.; Hernández, A.; Vargas, W. E.

2014-08-01

Measured reflection spectra from elytra of Chrysina aurigans scarabs are reported. They show a broad reflection band for wavelengths from 0.525 to 1.0 μm with a sequence of maxima and minima reflection values superimposed on a mean value of around 40% for the high reflection band. Different mechanisms contributing to the reflection spectra have been considered, with the dominant effect, reflection of left handed circularly polarized light, being produced by a laminated left handed twisted structure whose pitch changes with depth through the procuticle in a more complex way than that characterizing broad band circular polarizers based on cholesteric liquid crystals.

Light reflection by the cuticle of C. aurigans scarabs: a biological broadband reflector of left handed circularly polarized light

International Nuclear Information System (INIS)

Libby, E; Azofeifa, D E; Hernández-Jiménez, M; García-Aguilar, I; Arce-Marenco, L; Hernández, A; Vargas, W E; Barboza-Aguilar, C; Solís, A

2014-01-01

Measured reflection spectra from elytra of Chrysina aurigans scarabs are reported. They show a broad reflection band for wavelengths from 0.525 to 1.0 μm with a sequence of maxima and minima reflection values superimposed on a mean value of around 40% for the high reflection band. Different mechanisms contributing to the reflection spectra have been considered, with the dominant effect, reflection of left handed circularly polarized light, being produced by a laminated left handed twisted structure whose pitch changes with depth through the procuticle in a more complex way than that characterizing broad band circular polarizers based on cholesteric liquid crystals. (fast track communication)

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. © 2014 Blackwell Verlag GmbH.

Scanning light-sheet microscopy in the whole mouse brain with HiLo background rejection

Science.gov (United States)

Mertz, Jerome; Kim, Jinhyun

2010-01-01

It is well known that light-sheet illumination can enable optically sectioned wide-field imaging of macroscopic samples. However, the optical sectioning capacity of a light-sheet macroscope is undermined by sample-induced scattering or aberrations that broaden the thickness of the sheet illumination. We present a technique to enhance the optical sectioning capacity of a scanning light-sheet microscope by out-of-focus background rejection. The technique, called HiLo microscopy, makes use of two images sequentially acquired with uniform and structured sheet illumination. An optically sectioned image is then synthesized by fusing high and low spatial frequency information from both images. The benefits of combining light-sheet macroscopy and HiLo background rejection are demonstrated in optically cleared whole mouse brain samples, using both green fluorescent protein (GFP)-fluorescence and dark-field scattered light contrast.

Exploring the brain on multiple scales with correlative two-photon and light sheet microscopy

Science.gov (United States)

Silvestri, Ludovico; Allegra Mascaro, Anna Letizia; Costantini, Irene; Sacconi, Leonardo; Pavone, Francesco S.

2014-02-01

One of the unique features of the brain is that its activity cannot be framed in a single spatio-temporal scale, but rather spans many orders of magnitude both in space and time. A single imaging technique can reveal only a small part of this complex machinery. To obtain a more comprehensive view of brain functionality, complementary approaches should be combined into a correlative framework. Here, we describe a method to integrate data from in vivo two-photon fluorescence imaging and ex vivo light sheet microscopy, taking advantage of blood vessels as reference chart. We show how the apical dendritic arbor of a single cortical pyramidal neuron imaged in living thy1-GFP-M mice can be found in the large-scale brain reconstruction obtained with light sheet microscopy. Starting from the apical portion, the whole pyramidal neuron can then be segmented. The correlative approach presented here allows contextualizing within a three-dimensional anatomic framework the neurons whose dynamics have been observed with high detail in vivo.

SVBRDF-Invariant Shape and Reflectance Estimation from a Light-Field Camera.

Science.gov (United States)

Wang, Ting-Chun; Chandraker, Manmohan; Efros, Alexei A; Ramamoorthi, Ravi

2018-03-01

Light-field cameras have recently emerged as a powerful tool for one-shot passive 3D shape capture. However, obtaining the shape of glossy objects like metals or plastics remains challenging, since standard Lambertian cues like photo-consistency cannot be easily applied. In this paper, we derive a spatially-varying (SV)BRDF-invariant theory for recovering 3D shape and reflectance from light-field cameras. Our key theoretical insight is a novel analysis of diffuse plus single-lobe SVBRDFs under a light-field setup. We show that, although direct shape recovery is not possible, an equation relating depths and normals can still be derived. Using this equation, we then propose using a polynomial (quadratic) shape prior to resolve the shape ambiguity. Once shape is estimated, we also recover the reflectance. We present extensive synthetic data on the entire MERL BRDF dataset, as well as a number of real examples to validate the theory, where we simultaneously recover shape and BRDFs from a single image taken with a Lytro Illum camera.

Observation Platform for Dynamic Biomedical and Biotechnology Experiments using the ISS Light Microscopy Module, Phase I

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...

3D single-molecule super-resolution microscopy with a tilted light sheet.

Science.gov (United States)

Gustavsson, Anna-Karin; Petrov, Petar N; Lee, Maurice Y; Shechtman, Yoav; Moerner, W E

2018-01-09

Tilted light sheet microscopy with 3D point spread functions (TILT3D) combines a novel, tilted light sheet illumination strategy with long axial range point spread functions (PSFs) for low-background, 3D super-localization of single molecules as well as 3D super-resolution imaging in thick cells. Because the axial positions of the single emitters are encoded in the shape of each single-molecule image rather than in the position or thickness of the light sheet, the light sheet need not be extremely thin. TILT3D is built upon a standard inverted microscope and has minimal custom parts. The result is simple and flexible 3D super-resolution imaging with tens of nm localization precision throughout thick mammalian cells. We validate TILT3D for 3D super-resolution imaging in mammalian cells by imaging mitochondria and the full nuclear lamina using the double-helix PSF for single-molecule detection and the recently developed tetrapod PSFs for fiducial bead tracking and live axial drift correction.

Association between dermoscopic and reflectance confocal microscopy features of cutaneous melanoma with BRAF mutational status.

Science.gov (United States)

Bombonato, C; Ribero, S; Pozzobon, F C; Puig-Butille, J A; Badenas, C; Carrera, C; Malvehy, J; Moscarella, E; Lallas, A; Piana, S; Puig, S; Argenziano, G; Longo, C

2017-04-01

Melanomas harbouring common genetic mutations might share certain morphological features detectable with dermoscopy and reflectance confocal microscopy. BRAF mutational status is crucial for the management of metastatic melanoma. To correlate the dermoscopic characteristics of primary cutaneous melanomas with BRAF mutational status. Furthermore, a subset of tumours has also been analysed for the presence of possible confocal features that might be linked with BRAF status. Retrospectively acquired dermoscopic and confocal images of patients with melanoma in tertiary referral academic centres: Skin Cancer Unit in Reggio Emilia and at the Melanoma Unit in Barcelona. Kruskal-Wallis test, logistic regressions, univariate and multivariate analyses have been performed to find dermoscopic and confocal features significantly correlated with BRAF mutational status. Dermoscopically, the presence of irregular peripheral streaks and ulceration were positive predictors of BRAF-mutated melanomas with a statistically significance value, while dotted vessels were more represented in wild-type melanomas. None of the evaluated reflectance confocal microscopy features were correlated with genetic profiling. Ulceration and irregular peripheral streaks represent dermoscopic feature indicative for BRAF-mutated melanoma, while dotted vessels are suggestive for wild-type melanoma. © 2016 European Academy of Dermatology and Venereology.

[Correction of light refraction and reflection in medical transmission optical tomography].

Science.gov (United States)

Tereshchenko, S A; Potapov, D A

2002-01-01

The effects of light refraction and reflection on the quality of image reconstruction in medical transmission optical tomography of high-scattering media are considered. It has been first noted that light refraction not only distorts the geometric scheme of measurements, but may lead to the appearance of object areas that cannot be scanned. Some ways of decreasing the effect of refraction on the reconstruction of spatial distribution of the extinction coefficient are stated.

Correlative Light-Electron Microscopy of Lipid-Encapsulated Fluorescent Nanodiamonds for Nanometric Localization of Cell Surface Antigens.

Science.gov (United States)

Hsieh, Feng-Jen; Chen, Yen-Wei; Huang, Yao-Kuan; Lee, Hsien-Ming; Lin, Chun-Hung; Chang, Huan-Cheng

2018-02-06

Containing an ensemble of nitrogen-vacancy centers in crystal matrices, fluorescent nanodiamonds (FNDs) are a new type of photostable markers that have found wide applications in light microscopy. The nanomaterial also has a dense carbon core, making it visible to electron microscopy. Here, we show that FNDs encapsulated in biotinylated lipids (bLs) are useful for subdiffraction imaging of antigens on cell surface with correlative light-electron microscopy (CLEM). The lipid encapsulation enables not only good dispersion of the particles in biological buffers but also high specific labeling of live cells. By employing the bL-encapsulated FNDs to target CD44 on HeLa cell surface through biotin-mediated immunostaining, we obtained the spatial distribution of these antigens by CLEM with a localization accuracy of ∼50 nm in routine operations. A comparative study with dual-color imaging, in which CD44 was labeled with FND and MICA/MICB was labeled with Alexa Fluor 488, demonstrated the superior performance of FNDs as fluorescent fiducial markers for CLEM of cell surface antigens.

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.

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.

High-Throughput Light Sheet Microscopy for the Automated Live Imaging of Larval Zebrafish

Science.gov (United States)

Baker, Ryan; Logan, Savannah; Dudley, Christopher; Parthasarathy, Raghuveer

The zebrafish is a model organism with a variety of useful properties; it is small and optically transparent, it reproduces quickly, it is a vertebrate, and there are a large variety of transgenic animals available. Because of these properties, the zebrafish is well suited to study using a variety of optical technologies including light sheet fluorescence microscopy (LSFM), which provides high-resolution three-dimensional imaging over large fields of view. Research progress, however, is often not limited by optical techniques but instead by the number of samples one can examine over the course of an experiment, which in the case of light sheet imaging has so far been severely limited. Here we present an integrated fluidic circuit and microscope which provides rapid, automated imaging of zebrafish using several imaging modes, including LSFM, Hyperspectral Imaging, and Differential Interference Contrast Microscopy. Using this system, we show that we can increase our imaging throughput by a factor of 10 compared to previous techniques. We also show preliminary results visualizing zebrafish immune response, which is sensitive to gut microbiota composition, and which shows a strong variability between individuals that highlights the utility of high throughput imaging. National Science Foundation, Award No. DBI-1427957.

X-ray holographic microscopy experiments at the Brookhaven synchrotron light source

International Nuclear Information System (INIS)

Howells, M.R.; Iarocci, M.; Kenney, J.; Kirz, J.; Rarback, H.

1983-01-01

Soft x-ray holographic microscopy is discussed from an experimental point of view. Three series of measurements have been carried out using the Brookhaven 750 MeV storage ring as an x-ray source. Young slits fringes, Gabor (in line) holograms and various data pertaining to the soft x-ray performance of photographic plates are reported. The measurements are discussed in terms of the technique for recording them and the experimental limitations in effect. Some discussion is also given of the issues involved in reconstruction using visible light

Production of a diffuse very high reflectivity material for light collection in nuclear detectors

CERN Document Server

Pichler, B J; Mirzoyan, R; Weiss, L; Ziegler, S I

2000-01-01

A diffuse very high reflectivity material, based on polytetrafluorethylene (PTFE) for optimization of light-collection efficiency has been developed. PTFE powder was used to produce reflector block material. The powder was pressed with 525 kPa in a form and sintered at 375 deg. C. The reflectivity was above 98% within the spectral range from 350 to 1000 nm. The blocks of this material are machinable with saws, drilling and milling machines. The reflector is used as a housing for scintillating crystals in a nuclear medicine application (small animal positron emission tomograph). It is also used as a light collector in very high-energy gamma-ray astrophysicas experiments, HEGRA and MAGIC. The application of this inexpensive, easy to make diffuse reflector may allow the optimization of light collection in a wide range of low-level light-detector configurations.

Fluorescence microscopy.

Science.gov (United States)

Sanderson, Michael J; Smith, Ian; Parker, Ian; Bootman, Martin D

2014-10-01

Fluorescence microscopy is a major tool with which to monitor cell physiology. Although the concepts of fluorescence and its optical separation using filters remain similar, microscope design varies with the aim of increasing image contrast and spatial resolution. The basics of wide-field microscopy are outlined to emphasize the selection, advantages, and correct use of laser scanning confocal microscopy, two-photon microscopy, scanning disk confocal microscopy, total internal reflection, and super-resolution microscopy. In addition, the principles of how these microscopes form images are reviewed to appreciate their capabilities, limitations, and constraints for operation. © 2014 Cold Spring Harbor Laboratory Press.

Geometrically distributed one-dimensional photonic crystals for light-reflection in all angles.

Science.gov (United States)

Alagappan, G; Wu, P

2009-07-06

We demonstrate that a series of one-dimensional photonic crystals made of any dielectric materials, with the periods are distributed in a geometrical progression of a common ratio, r rc (theta,P), where rc is a structural parameter that depends on the angle of incidence, theta, and polarization, P, is capable of blocking light of any spectral range. If an omni-directional reflection is desired for all polarizations and for all incident angles smaller than thetao, then r rc (theta(o),p), where p is the polarization with the electric field parallel to the plane of incidence. We present simple and formula like expressions for rc, width of the bandgap, and minimum number of photonic crystals to achieve a perfect light reflection.

Ex Vivo (Fluorescence) Confocal Microscopy in Surgical Pathology: State of the Art.

Science.gov (United States)

Ragazzi, Moira; Longo, Caterina; Piana, Simonetta

2016-05-01

First developed in 1957, confocal microscopy is a powerful imaging tool that can be used to obtain near real-time reflected light images of untreated human tissue with nearly histologic resolution. Besides its research applications, in the last decades, confocal microscopy technology has been proposed as a useful device to improve clinical diagnosis, especially in ophthalmology, dermatology, and endomicroscopy settings, thanks to advances in instrument development. Compared with the wider use of the in vivo tissue assessment, ex vivo applications of confocal microscopy are not fully explored. A comprehensive review of the current literature was performed here, focusing on the reliable applications of ex vivo confocal microscopy in surgical pathology and on some potential evolutions of this new technique from pathologists' viewpoint.

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.

Microscopy and Image Analysis.

Science.gov (United States)

McNamara, George; Difilippantonio, Michael; Ried, Thomas; Bieber, Frederick R

2017-07-11

This unit provides an overview of light microscopy, including objectives, light sources, filters, film, and color photography for fluorescence microscopy and fluorescence in situ hybridization (FISH). We believe there are excellent opportunities for cytogeneticists, pathologists, and other biomedical readers, to take advantage of specimen optical clearing techniques and expansion microscopy-we briefly point to these new opportunities. © 2017 by John Wiley & Sons, Inc. Copyright © 2017 John Wiley & Sons, Inc.

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

International Nuclear Information System (INIS)

Samson, Edward Carlo; Blanca, Carlo Mar

2007-01-01

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

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

Light propagation with phase discontinuities: generalized laws of reflection and refraction.

Science.gov (United States)

Yu, Nanfang; Genevet, Patrice; Kats, Mikhail A; Aieta, Francesco; Tetienne, Jean-Philippe; Capasso, Federico; Gaburro, Zeno

2011-10-21

Conventional optical components rely on gradual phase shifts accumulated during light propagation to shape light beams. New degrees of freedom are attained by introducing abrupt phase changes over the scale of the wavelength. A two-dimensional array of optical resonators with spatially varying phase response and subwavelength separation can imprint such phase discontinuities on propagating light as it traverses the interface between two media. Anomalous reflection and refraction phenomena are observed in this regime in optically thin arrays of metallic antennas on silicon with a linear phase variation along the interface, which are in excellent agreement with generalized laws derived from Fermat's principle. Phase discontinuities provide great flexibility in the design of light beams, as illustrated by the generation of optical vortices through use of planar designer metallic interfaces.

Beam line optics technologies series (7). Orthopedic treatment of sharp of light (reflecting mirror)

International Nuclear Information System (INIS)

Uruga, Tomoya; Nomura, Masaharu

2006-01-01

A reflecting mirror (mirror) is the most popular light device for orthopedic treatment of the shape of light. The paper explains the kinds of mirror for hard X-ray field and its applications in order to think the objects of mirror and how to adjust it when user experiment on the beam-line. The basic knowledge of reflection of X-ray, a use of mirror, the kinds of condenser mirror, the influence factors on the condenser size, arrangement of mirror in the hard X-ray beam-line, what kinds of mirror are necessary, evaluation of performance of mirror and adjustment, and troubles and measures are described. Layout in optics hutch at BL01B1 at SPring-8, refraction and total reflection of X-rays at surface, reflectivity of Rh and Pt with ideal surface as a function of photon energy, effects of surface roughness on reflectivity of Rh, calculated beam sizes for typical SPring-8 mirror as a function of magnification, schematic drawing of mirror, standard mirror system for vertical deflection in bending magnet beam-line, and observed and calculated reflectivity of Rh double mirror at BL01B1 at SPring-8 are illustrated. (S.Y)

Nanograting-based plasmon enhancement for total internal reflection fluorescence microscopy of live cells

International Nuclear Information System (INIS)

Kim, Kyujung; Cho, Eun-Jin; Suh, Jin-Suck; Huh, Yong-Min; Kim, Donghyun; Kim, Dong Jun

2009-01-01

We investigated evanescent field enhancement based on subwavelength nanogratings for improved sensitivity in total internal reflection microscopy of live cells. The field enhancement is associated with subwavelength-grating-coupled plasmon excitation. An optimum sample employed a silver grating on a silver film and an SF10 glass substrate. Field intensity was enhanced by approximately 90% when measured by fluorescent excitation of microbeads relative to that on a bare prism as a control, which is in good agreement with numerical results. The subwavelength-grating-mediated field enhancement was also applied to live cell imaging of quantum dots, which confirmed the sensitivity enhancement qualitatively.

Scanless multitarget-matching multiphoton excitation fluorescence microscopy

Directory of Open Access Journals (Sweden)

Junpeng Qiu

2018-03-01

Full Text Available Using the combination of a reflective blazed grating and a reflective phase-only diffractive spatial light modulator (SLM, scanless multitarget-matching multiphoton excitation fluorescence microscopy (SMTM-MPM was achieved. The SLM shaped an incoming mode-locked, near-infrared Ti:sapphire laser beam into an excitation pattern with addressable shapes and sizes that matched the samples of interest in the field of view. Temporal and spatial focusing were simultaneously realized by combining an objective lens and a blazed grating. The fluorescence signal from illuminated areas was recorded by a two-dimensional sCMOS camera. Compared with a conventional temporal focusing multiphoton microscope, our microscope achieved effective use of the laser power and decreased photodamage with higher axial resolution.

Charting Monosynaptic Connectivity Maps by Two-Color Light-Sheet Fluorescence Microscopy

Directory of Open Access Journals (Sweden)

Christian J. Niedworok

2012-11-01

Full Text Available Cellular resolution three-dimensional (3D visualization of defined, fluorescently labeled long-range neuronal networks in the uncut adult mouse brain has been elusive. Here, a virus-based strategy is described that allowed fluorescent labeling of centrifugally projecting neuronal populations in the ventral forebrain and their directly, monosynaptically connected bulbar interneurons upon a single stereotaxic injection into select neuronal populations. Implementation of improved tissue clearing combined with light-sheet fluorescence microscopy permitted imaging of the resulting connectivity maps in a single whole-brain scan. Subsequent 3D reconstructions revealed the exact distribution of the diverse neuronal ensembles monosynaptically connected with distinct bulbar interneuron populations. Moreover, rehydratation of brains after light-sheet fluorescence imaging enabled the immunohistochemical identification of synaptically connected neurons. Thus, this study describes a method for identifying monosynaptic connectivity maps from distinct, virally labeled neuronal populations that helps in better understanding of information flow in neural systems.

Full optical model of micro-endoscope with optical coherence microscopy, multiphoton microscopy and visible capabilities

Science.gov (United States)

Vega, David; Kiekens, Kelli C.; Syson, Nikolas C.; Romano, Gabriella; Baker, Tressa; Barton, Jennifer K.

2018-02-01

While Optical Coherence Microscopy (OCM), Multiphoton Microscopy (MPM), and narrowband imaging are powerful imaging techniques that can be used to detect cancer, each imaging technique has limitations when used by itself. Combining them into an endoscope to work in synergy can help achieve high sensitivity and specificity for diagnosis at the point of care. Such complex endoscopes have an elevated risk of failure, and performing proper modelling ensures functionality and minimizes risk. We present full 2D and 3D models of a multimodality optical micro-endoscope to provide real-time detection of carcinomas, called a salpingoscope. The models evaluate the endoscope illumination and light collection capabilities of various modalities. The design features two optical paths with different numerical apertures (NA) through a single lens system with a scanning optical fiber. The dual path is achieved using dichroic coatings embedded in a triplet. A high NA optical path is designed to perform OCM and MPM while a low NA optical path is designed for the visible spectrum to navigate the endoscope to areas of interest and narrowband imaging. Different tests such as the reflectance profile of homogeneous epithelial tissue were performed to adjust the models properly. Light collection models for the different modalities were created and tested for efficiency. While it is challenging to evaluate the efficiency of multimodality endoscopes, the models ensure that the system is design for the expected light collection levels to provide detectable signal to work for the intended imaging.

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.

Diagnostics of intestinal parasites in light microscopy among the population of children in eastern Afghanistan

Directory of Open Access Journals (Sweden)

Krzysztof Korzeniewski

2016-09-01

The variety of detected intestinal pathogens in examined children’s population has required the use of combination of multiple diagnostic methods in light microscopy, and finally improved the detection rates of intestinal parasites and helped eliminate infections with nematodes, cestodes, trematodes, and protozoa using appropriate treatment in the study population.

Monte Carlo simulation of reflection spectra of random multilayer media strongly scattering and absorbing light

International Nuclear Information System (INIS)

Meglinskii, I V

2001-01-01

The reflection spectra of a multilayer random medium - the human skin - strongly scattering and absorbing light are numerically simulated. The propagation of light in the medium and the absorption spectra are simulated by the stochastic Monte Carlo method, which combines schemes for calculations of real photon trajectories and the statistical weight method. The model takes into account the inhomogeneous spatial distribution of blood vessels, water, and melanin, the degree of blood oxygenation, and the hematocrit index. The attenuation of the incident radiation caused by reflection and refraction at Fresnel boundaries of layers inside the medium is also considered. The simulated reflection spectra are compared with the experimental reflection spectra of the human skin. It is shown that a set of parameters that was used to describe the optical properties of skin layers and their possible variations, despite being far from complete, is nevertheless sufficient for the simulation of the reflection spectra of the human skin and their quantitative analysis. (laser applications and other topics in quantum electronics)

Spectral and spatial properties of polarized light reflections from the arms of squid (Loligo pealeii) and cuttlefish (Sepia officinalis L.).

Science.gov (United States)

Chiou, Tsyr-Huei; Mäthger, Lydia M; Hanlon, Roger T; Cronin, Thomas W

2007-10-01

On every arm of cuttlefish and squid there is a stripe of high-reflectance iridophores that reflects highly polarized light. Since cephalopods possess polarization vision, it has been hypothesized that these polarized stripes could serve an intraspecific communication function. We determined how polarization changes when these boneless arms move. By measuring the spectral and polarizing properties of the reflected light from samples at various angles of tilt and rotation, we found that the actual posture of the arm has little or no effect on partial polarization or the e-vector angle of the reflected light. However, when the illumination angle changed, the partial polarization of the reflected light also changed. The spectral reflections of the signals were also affected by the angle of illumination but not by the orientation of the sample. Electron microscope samples showed that these stripes are composed of several groups of multilayer platelets within the iridophores. The surface normal to each group is oriented at a different angle, which produces essentially constant reflection of polarized light over a range of viewing angles. These results demonstrate that cuttlefish and squid could send out reliable polarization signals to a receiver regardless of arm orientation.

Determination of lead in clay enameled by X-ray fluorescence technique in Total reflection and by Scanning Electron Microscopy

International Nuclear Information System (INIS)

Zarazua O, G.; Carapia M, L.

2000-01-01

This work has the objective of determining lead free in the glazed commercial stewing pans using the X-ray fluorescence technique in Total reflection (FRX) and the observation and semiquantitative determination of lead by Analytical Scanning Electron Microscopy (ASEM). (Author)

Live Cell Refractometry Using Hilbert Phase Microscopy and Confocal Reflectance Microscopy†

Science.gov (United States)

Lue, Niyom; Choi, Wonshik; Popescu, Gabriel; Yaqoob, Zahid; Badizadegan, Kamran; Dasari, Ramachandra R.; Feld, Michael S.

2010-01-01

Quantitative chemical analysis has served as a useful tool for understanding cellular metabolisms in biology. Among many physical properties used in chemical analysis, refractive index in particular has provided molecular concentration that is an important indicator for biological activities. In this report, we present a method of extracting full-field refractive index maps of live cells in their native states. We first record full-field optical thickness maps of living cells by Hilbert phase microscopy and then acquire physical thickness maps of the same cells using a custom-built confocal reflectance microscope. Full-field and axially averaged refractive index maps are acquired from the ratio of optical thickness to physical thickness. The accuracy of the axially averaged index measurement is 0.002. This approach can provide novel biological assays of label-free living cells in situ. PMID:19803506

Identification of a Multicomponent Traditional Herbal Medicine by HPLC-MS and Electron and Light Microscopy.

Science.gov (United States)

Liu, Ju-Han; Cheng, Yung-Yi; Hsieh, Chen-Hsi; Tsai, Tung-Hu

2017-12-15

Commercial pharmaceutical herbal products have enabled people to take traditional Chinese medicine (TCM) in a convenient and accessible form. However, the quantity and quality should be additionally inspected. To address the issue, a combination of chemical and physical inspection methods were developed to evaluate the amount of an herbal formula, Xiang-Sha-Liu-Jun-Zi-Tang (XSLJZT), in clinical TCM practice. A high-performance liquid chromatography-tandem mass spectrometry (HPLC-MS) method with electrospray ionization was developed to measure the herbal biomarkers of guanosine, atractylenolide III, glycyrrhizic acid, dehydrocostus lactone, hesperidin, and oleanolic acid from XSLJZT. Scanning electron microscopy (SEM) photographs and light microscopy photographs with Congo red and iodine-KI staining were used to identify the cellulose fibers and starch content. Furthermore, solubility analysis, swelling power test, and crude fiber analysis were contributed to measure the starch additive in pharmaceutical products. The results demonstrated large variations in the chemical components of different pharmaceutical brands. The SEM photographs revealed that the starch was oval, smooth, and granular, and that the raw herbal powder appears stripy, stretched, and filiform. The stained light microscopy photographs of all of the pharmaceutical products showed added starch and raw herbal powder as extenders. The developed chemical and physical methods provide a standard operating procedure for the quantity control of the herbal pharmaceutical products of XSLJZT.

Sample Preparation and Mounting of Drosophila Embryos for Multiview Light Sheet Microscopy.

Science.gov (United States)

Schmied, Christopher; Tomancak, Pavel

2016-01-01

Light sheet fluorescent microscopy (LSFM), and in particular its most widespread flavor Selective Plane Illumination Microscopy (SPIM), promises to provide unprecedented insights into developmental dynamics of entire living systems. By combining minimal photo-damage with high imaging speed and sample mounting tailored toward the needs of the specimen, it enables in toto imaging of embryogenesis with high spatial and temporal resolution. Drosophila embryos are particularly well suited for SPIM imaging because the volume of the embryo does not change from the single cell embryo to the hatching larva. SPIM microscopes can therefore image Drosophila embryos embedded in rigid media, such as agarose, from multiple angles every few minutes from the blastoderm stage until hatching. Here, we describe sample mounting strategies to achieve such a recording. We also provide detailed protocols to realize multiview, long-term, time-lapse recording of Drosophila embryos expressing fluorescent markers on the commercially available Zeiss Lightsheet Z.1 microscope and the OpenSPIM.

Optical spectroscopy and microscopy of radiation-induced light-emitting point defects in lithium fluoride crystals and films

Science.gov (United States)

Montereali, R. M.; Bonfigli, F.; Menchini, F.; Vincenti, M. A.

2012-08-01

Broad-band light-emitting radiation-induced F2 and F3+ electronic point defects, which are stable and laser-active at room temperature in lithium fluoride crystals and films, are used in dosimeters, tuneable color-center lasers, broad-band miniaturized light sources and novel radiation imaging detectors. A brief review of their photoemission properties is presented, and their behavior at liquid nitrogen temperatures is discussed. Some experimental data from optical spectroscopy and fluorescence microscopy of these radiation-induced point defects in LiF crystals and thin films are used to obtain information about the coloration curves, the efficiency of point defect formation, the effects of photo-bleaching processes, etc. Control of the local formation, stabilization, and transformation of radiation-induced light-emitting defect centers is crucial for the development of optically active micro-components and nanostructures. Some of the advantages of low temperature measurements for novel confocal laser scanning fluorescence microscopy techniques, widely used for spatial mapping of these point defects through the optical reading of their visible photoluminescence, are highlighted.

Single cell adhesion strength assessed with variable-angle total internal reflection fluorescence microscopy

Directory of Open Access Journals (Sweden)

Marcelina Cardoso Dos Santos

2017-06-01

Full Text Available We propose a new strategy to evaluate adhesion strength at the single cell level. This approach involves variable-angle total internal reflection fluorescence microscopy to monitor in real time the topography of cell membranes, i.e. a map of the membrane/substrate separation distance. According to the Boltzmann distribution, both potential energy profile and dissociation energy related to the interactions between the cell membrane and the substrate were determined from the membrane topography. We have highlighted on glass substrates coated with poly-L-lysine and fibronectin, that the dissociation energy is a reliable parameter to quantify the adhesion strength of MDA-MB-231 motile cells.

Correlative Stochastic Optical Reconstruction Microscopy and Electron Microscopy

Science.gov (United States)

Kim, Doory; Deerinck, Thomas J.; Sigal, Yaron M.; Babcock, Hazen P.; Ellisman, Mark H.; Zhuang, Xiaowei

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 imaging conditions for super-resolution fluorescence microscopy and EM are often not compatible. Here, we have developed several experiment protocols for correlative stochastic optical reconstruction microscopy (STORM) and EM methods, both for un-embedded samples by applying EM-specific sample preparations after STORM imaging and for embedded and sectioned samples by optimizing the fluorescence under EM fixation, staining and embedding conditions. We demonstrated these methods using a variety of cellular targets. PMID:25874453

Waveguide evanescent field fluorescence microscopy & its application in cell biology

Science.gov (United States)

Hassanzadeh, Abdollah

There are many powerful microscopy technologies available for the investigation of bulk materials as well as for thin film samples. Nevertheless, for imaging an interface, especially live cells on a substrate and ultra thin-films, only Total Internal Reflection Fluorescence (TIRF) microscopy is available. This TIRF microscopy allows imaging without interference of the bulk. Various approaches are employed in fluorescence microscopy applications to restrict the excitation and detection of fluorophores to a thin region of the specimen. Elimination of background fluorescence from outside the focal plane can dramatically improve the signal-to-noise ratio, and consequently, the spatial resolution of the features or events of interest. TIRF microscopy is an evanescent field based microscopy. In this method, fluorescent dyes are only excited within an evanescent field: roughly within 100 nm above a glass coverslip. This will allow imaging surface and interfacial issues of the glass coverslip and an adjacent material. Waveguide evanescent field fluorescence (WEFF) microscopy is a new development for imaging cell-substrate interactions in real time and in vitro. It is an alternative to TIRF microscopy. In this method the light is coupled into a waveguide via an optical grating. The coupled light propagates as a waveguide mode and exhibits an evanescent field on top of the waveguide. This can be used as a surface-bound illumination source to excite fluorophores. This evanescent field serves as an extremely powerful tool for quality control of thin films, to study cell-substrate contacts, and investigating the effect of external agents and drugs on the cell-substrate interaction in real time and in vitro. This new method has been established and optimized to minimize non-uniformity, scattering and photo bleaching issues. Visualizing and quantifying of the cell-substrates and solid thin films have been carried out by WEFF microscopy. The images of the cell-substrate interface

Characterizing Cool Giant Planets in Reflected Light

Science.gov (United States)

Marley, Mark

2016-01-01

While the James Webb Space Telescope will detect and characterize extrasolar planets by transit and direct imaging, a new generation of telescopes will be required to detect and characterize extrasolar planets by reflected light imaging. NASA's WFIRST space telescope, now in development, will image dozens of cool giant planets at optical wavelengths and will obtain spectra for several of the best and brightest targets. This mission will pave the way for the detection and characterization of terrestrial planets by the planned LUVOIR or HabEx space telescopes. In my presentation I will discuss the challenges that arise in the interpretation of direct imaging data and present the results of our group's effort to develop methods for maximizing the science yield from these planned missions.

REFLECTED LIGHT CURVES, SPHERICAL AND BOND ALBEDOS OF JUPITER- AND SATURN-LIKE EXOPLANETS

Energy Technology Data Exchange (ETDEWEB)

Dyudina, Ulyana; Kopparla, Pushkar; Ingersoll, Andrew P.; Yung, Yuk L. [Division of Geological and Planetary Sciences, 150-21 California Institute of Technology, Pasadena, CA 91125 (United States); Zhang, Xi [University of California Santa Cruz 1156 High Street, Santa Cruz, CA 95064 (United States); Li, Liming [Department of Physics, University of Houston, Houston, TX 77204 (United States); Dones, Luke [Southwest Research Institute, 1050 Walnut Street, Suite 300, Boulder CO 80302 (United States); Verbiscer, Anne, E-mail: ulyana@gps.caltech.edu [Department of Astronomy, University of Virginia, Charlottesville, VA 22904-4325 (United States)

2016-05-10

Reflected light curves observed for exoplanets indicate that a few of them host bright clouds. We estimate how the light curve and total stellar heating of a planet depends on forward and backward scattering in the clouds based on Pioneer and Cassini spacecraft images of Jupiter and Saturn. We fit analytical functions to the local reflected brightnesses of Jupiter and Saturn depending on the planet’s phase. These observations cover broadbands at 0.59–0.72 and 0.39–0.5 μ m, and narrowbands at 0.938 (atmospheric window), 0.889 (CH4 absorption band), and 0.24–0.28 μ m. We simulate the images of the planets with a ray-tracing model, and disk-integrate them to produce the full-orbit light curves. For Jupiter, we also fit the modeled light curves to the observed full-disk brightness. We derive spherical albedos for Jupiter and Saturn, and for planets with Lambertian and Rayleigh-scattering atmospheres. Jupiter-like atmospheres can produce light curves that are a factor of two fainter at half-phase than the Lambertian planet, given the same geometric albedo at transit. The spherical albedo is typically lower than for a Lambertian planet by up to a factor of ∼1.5. The Lambertian assumption will underestimate the absorption of the stellar light and the equilibrium temperature of the planetary atmosphere. We also compare our light curves with the light curves of solid bodies: the moons Enceladus and Callisto. Their strong backscattering peak within a few degrees of opposition (secondary eclipse) can lead to an even stronger underestimate of the stellar heating.

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......-walled multinucleate spores in the fish stomach as a response to the low pH. The hyphae then penetrate the digestive tract and rupture when they reach a blood vessel (neutral pH), whereby uni- and binucleate bodies and/or amoeboid bodies are released. The small cells are transported in the blood vessels and spread...

Comparative morphology of zebra (Dreissena polymorpha) and quagga (Dreissena bugensis) mussel sperm: Light and electron microscopy

Science.gov (United States)

Walker, G.K.; Black, M.G.; Edwards, C.A.

1996-01-01

Adult zebra (Dreissena polymorpha) and quagga (Dreissena bugensis) mussels were induced to release large quantities of live spermatozoa by the administration of 5-hydroxytryptamine (serotonin). Sperm were photographed alive using phase-contrast microscopy and were fixed subsequently with glutaraldehyde followed by osmium tetroxide for eventual examination by transmission or scanning electron microscopy. The sperm of both genera are of the ect-aquasperm type. Their overall dimensions and shape allow for easy discrimination at the light and scanning electron microscopy level. Transmission electron microscopy of the cells reveals a barrel-shaped nucleus in zebra mussel sperm and an elongated nucleus in quagga mussel sperm. In both species, an acrosome is cradled in a nuclear fossa. The ultrastructure of the acrosome and axial body, however, is distinctive for each species. The structures of the midpiece are shown, including a unique mitochondrial "skirt" that includes densely packed parallel cristae and extends in a narrow sheet from the mitochondria.

Light Microscopy Module: International Space Station Premier Automated Microscope

Science.gov (United States)

Sicker, Ronald J.; Foster, William M.; Motil, Brian J.; Meyer, William V.; Chiaramonte, Francis P.; Abbott-Hearn, Amber; Atherton, Arthur; Beltram, Alexander; Bodzioney, Christopher; Brinkman, John;

[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 vs. 39%; p=0.01). Fertilization rate was higher in oocytes with positive values of birefringence (77.5 % vs. 68.5% p=0.005) with similar embryo quality. Conception cycles showed oocytes with higher mean value of zona birefringence and visible spindle vs. no-conception cycles (pPolarized 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.

Polarization contrast in reflection near-field optical microscopy with uncoated fibre tips

DEFF Research Database (Denmark)

Bozhevolnyi, Sergey I.; Langbein, Wolfgang; Hvam, Jørn Märcher

1999-01-01

Using cross-hatched, patterned semiconductor surfaces and round 20-nm-thick gold pads on semiconductor wafers, we investigate the imaging characteristics of a reflection near-field optical microscope with an uncoated fibre tip for different polarization configurations and light wavelengths....... Is is shown that cross-polarized detection allows one to effectively suppress far-field components in the detected signal and to realise imaging of optical contrast on the sub-wavelength scale. The sensitivity window of our microscope, i.e. the scale on which near-field optical images represent mainly optical...

Wavefronts, light rays and caustic of a circular wave reflected by an arbitrary smooth curve

International Nuclear Information System (INIS)

Marciano-Melchor, Magdalena; Silva-Ortigoza, Ramón; Montiel-Piña, Enrique; Román-Hernández, Edwin; Santiago-Santiago, José Guadalupe; Silva-Ortigoza, Gilberto; Rosado, Alfonso; Suárez-Xique, Román

2011-01-01

The aim of the present work is to obtain expressions for both the wavefront train and the caustic associated with the light rays reflected by an arbitrary smooth curve after being emitted by a point light source located at an arbitrary position in the two-dimensional free space. To this end, we obtain an expression for the k-function associated with the general integral of Stavroudis to the eikonal equation that describes the evolution of the reflected light rays. The caustic is computed by using the definitions of the critical and caustic sets of the two-dimensional map that describes the evolution of an arbitrary wavefront associated with the general integral. The general results are applied to circular and parabolic mirrors. The main motivation to carry out this research is to establish, in future work, the caustic touching theorem in a two-dimensional optical medium and to study the diffraction problem by using the k-function concept. Both problems are important in the computation of the image of an arbitrary object under reflection and refraction

Method and apparatus for detecting phycocyanin-pigmented algae and bacteria from reflected light

Science.gov (United States)

Vincent, Robert (Inventor)

2013-01-01

The present invention relates to a method of detecting phycocyanin algae or bacteria in water from reflected light, and also includes devices for the measurement, calculation and transmission of data relating to that method.

Widefield and total internal reflection fluorescent structured illumination microscopy with scanning galvo mirrors

Science.gov (United States)

Chen, Youhua; Cao, Ruizhi; Liu, Wenjie; Zhu, Dazhao; Zhang, Zhiming; Kuang, Cuifang; Liu, Xu

2018-04-01

We present an alternative approach to realize structured illumination microscopy (SIM), which is capable for live cell imaging. The prototype utilizes two sets of scanning galvo mirrors, a polarization converter and a piezo-platform to generate a fast shifted, s-polarization interfered and periodic variable illumination patterns. By changing the angle of the scanning galvanometer, we can change the position of the spots at the pupil plane of the objective lens arbitrarily, making it easy to switch between widefield and total internal reflection fluorescent-SIM mode and adapting the penetration depth in the sample. Also, a twofold resolution improvement is achieved in our experiments. The prototype offers more flexibility of pattern period and illumination orientation changing than previous systems.

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.

Imaging the dorsal hippocampus: light reflectance relationships to electroencephalographic patterns during sleep

DEFF Research Database (Denmark)

Rector, D M; Poe, G R; Kristensen, Morten Pilgaard

1995-01-01

We assessed the correspondence of 660 nm light reflectance changes from the dorsal hippocampus with slow wave electroencephalographic (EEG) activity during quiet sleep (QS) and rapid eye movement (REM) sleep in four cats. An optic probe, attached to a charge-coupled-device (CCD) video camera...... as EEG changes. Dividing the image into 10 subregions revealed that reflectance changes at the rhythmical slow wave activity band (RSA, 4-6 Hz) persisted in localized regions during QS and REM sleep, but regional changes showed considerable wave-by-wave independence between areas and from slow wave...

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.

Bismuth titanate nanorods and their visible light photocatalytic properties

International Nuclear Information System (INIS)

Pei, L.Z.; Liu, H.D.; Lin, N.; Yu, H.Y.

2015-01-01

Highlights: • Bismuth titanate nanorods have been synthesized by a simple hydrothermal process. • The size of bismuth titanate nanorods can be controlled by growth conditions. • Bismuth titanate nanorods show good photocatalytic activities of methylene blue and Rhodamine B. - Abstract: Bismuth titanate nanorods have been prepared using a facile hydrothermal process without additives. The bismuth titanate products were characterized by X-ray diffraction (XRD), scanning electron microscopy (SEM), transmission electron microscopy (TEM), high-resolution TEM (HRTEM) and UV-vis diffusion reflectance spectrum. XRD pattern shows that the bismuth titanate nanorods are composed of cubic Bi 2 Ti 2 O 7 phase. Electron microscopy images show that the length and diameter of the bismuth titanate nanorods are 50-200 nm and 2 μm, respectively. Hydrothermal temperature and reaction time play important roles on the formation and size of the bismuth titanate nanorods. UV-vis diffusion reflectance spectrum indicates that bismuth titanate nanorods have a band gap of 2.58 eV. The bismuth titanate nanorods exhibit good photocatalytic activities in the photocatalytic degradation of methylene blue (MB) and Rhodamine B (RB) under visible light irradiation. The bismuth titanate nanorods with cubic Bi 2 Ti 2 O 7 phase are a promising candidate as a visible light photocatalyst

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.

Sinusoidal obstruction syndrome (SOS): A light and electron microscopy study in human liver.

Science.gov (United States)

Vreuls, C P H; Driessen, A; Olde Damink, S W M; Koek, G H; Duimel, H; van den Broek, M A J; Dejong, C H C; Braet, F; Wisse, E

2016-05-01

Oxaliplatin is an important chemotherapeutic agent, used in the treatment of hepatic colorectal metastases, and known to induce the sinusoidal obstruction syndrome (SOS). Pathophysiological knowledge concerning SOS is based on a rat model. Therefore, the aim was to perform a comprehensive study of the features of human SOS, using both light microscopy (LM) and electron microscopy (EM). Included were all patients of whom wedge liver biopsies were collected during a partial hepatectomy for colorectal liver metastases, in a 4-year period. The wedge biopsy were perfusion fixated and processed for LM and EM. The SOS lesions were selected by LM and details were studied using EM. Material was available of 30 patients, of whom 28 patients received neo-adjuvant oxaliplatin. Eighteen (64%) of the 28 patients showed SOS lesions, based on microscopy. The lesions consisted of sinusoidal endothelial cell detachment from the space of Disse on EM. In the enlarged space of Disse a variable amount of erythrocytes were located. Sinusoidal endothelial cell detachment was present in human SOS, accompanied by enlargement of the space of Disse and erythrocytes in this area. These findings, originally described in a rat model, were now for the first time confirmed in human livers under clinically relevant settings. Copyright © 2016 Elsevier Ltd. All rights reserved.

Development of a new light collection and detection system optimized for ion beam induced fluorescence microscopy

International Nuclear Information System (INIS)

Vanga, Sudheer Kumar; Mi, Zhaohong; Koh, Long Cheng; Tao, Ye; Bettiol, Andrew A.; Watt, Frank

2015-01-01

Ion beam induced fluorescence microscopy is a new imaging technique which has the potential to achieve sub-50 nm spatial resolution fluorescence images. Currently the resolution of the technique has been limited to around 150 nm mainly because of inefficient collection and detection of emitted photons from the sample. To overcome this limitation, a new light collection system based on a custom made parabolic mirror is employed to enhance the fluorescence collection. The custom made mirror is designed so as to obtain both structural (scanning transmission ion microscopy) and ion beam induced fluorescence imaging simultaneously. The design and characterization of the parabolic mirror is discussed in detail

Fluorescence photooxidation with eosin: a method for high resolution immunolocalization and in situ hybridization detection for light and electron microscopy

Science.gov (United States)

1994-01-01

A simple method is described for high-resolution light and electron microscopic immunolocalization of proteins in cells and tissues by immunofluorescence and subsequent photooxidation of diaminobenzidine tetrahydrochloride into an insoluble osmiophilic polymer. By using eosin as the fluorescent marker, a substantial improvement in sensitivity is achieved in the photooxidation process over other conventional fluorescent compounds. The technique allows for precise correlative immunolocalization studies on the same sample using fluorescence, transmitted light and electron microscopy. Furthermore, because eosin is smaller in size than other conventional markers, this method results in improved penetration of labeling reagents compared to gold or enzyme based procedures. The improved penetration allows for three-dimensional immunolocalization using high voltage electron microscopy. Fluorescence photooxidation can also be used for high resolution light and electron microscopic localization of specific nucleic acid sequences by in situ hybridization utilizing biotinylated probes followed by an eosin-streptavidin conjugate. PMID:7519623

Electron microscopy of surfaces

International Nuclear Information System (INIS)

Venables, J.A.

1981-01-01

Electron beam techniques used to study clean surfaces and surface processes on a microscopic scale are reviewed. Recent experimental examples and possible future developments are discussed. Special emphasis is given to (i) transmission diffraction and microscopy techniques, including atomic imaging; (ii) Auger microscopy on bulk and thin film samples; (iii) secondary electron microscopy, especially low energy secondaries for work-function imaging and photoelectron imaging; and (iv) reflection electron microscopy and diffraction. (orig.)

In vivo pump-probe microscopy of melanoma and pigmented lesions

Science.gov (United States)

Wilson, Jesse W.; Degan, Simone; Mitropoulos, Tanya; Selim, M. Angelica; Zhang, Jennifer Y.; Warren, Warren S.

2012-03-01

A growing number of dermatologists and pathologists are concerned that the rapidly rising incidence of melanoma reflects not a true 'epidemic' but an increasing tendency to overdiagnose pigmented lesions. Addressing this problem requires both a better understanding of early-stage melanoma and new diagnostic criteria based on more than just cellular morphology and architecture. Here we present a method for in-vivo optical microscopy that utilizes pump-probe spectroscopy to image the distribution of the two forms of melanin in skin: eumelanin and pheomelanin. Images are acquired in a scanning microscope with a sensitive modulation transfer technique by analyzing back-scattered probe light with a lock-in amplifier. Early-stage melanoma is studied in a human skin xenografted mouse model. Individual melanocytes have been observed, in addition to pigmented keratinocytes. Combining the pump-probe images simultaneously with other noninvasive laser microscopy methods (confocal reflectance, multiphoton autofluorescence, and second harmonic generation) allows visualization of the skin architecture, framing the functional pump-probe image in the context of the surrounding tissue morphology. It is found that pump-probe images of melanin can be acquired with low peak intensities, enabling wide field-of-view pigmentation surveys. Finally, we investigate the diagnostic potential of the additional chemical information available from pump-probe microscopy.

Position Detection Based on Intensities of Reflected Infrared Light

DEFF Research Database (Denmark)

Christensen, Henrik Vie

measurements of reflected light intensities, and includes easy calibration. The method for reconstructing 3D positions has been implemented in a prototype of a “non-Touch Screen” for a computer, so that the user can control a cursor in three dimensions by moving his/hers hand in front of the computer screen....... The 2D position reconstruction method is mplemented in a prototype of a human-machine interface (HMI) for an electrically powered wheelchair, such that the wheelchair user can control the movement of the wheelchair by head movements. Both “non-Touch Screen” prototype and wheelchair HMI has been tested...

Image analysis using reflected light: an underutilized tool for interpreting magnetic fabrics

Science.gov (United States)

Waters-Tormey, C. L.; Liner, T.; Miller, B.; Kelso, P. R.

2010-12-01

Grain shape fabric analysis is one of the most common tools used to compare magnetic fabric and handsample scale rock fabric. Usually, this image analysis uses photomicrographs taken under plane or polarized light, which may be problematic if there are several dominant magnetic carriers (e.g., magnetite and pyrrhotite). The method developed for this study uses reflected light photomicrographs, and is effective in assessing the relative contribution of different phases to the opaque mineral shape-preferred orientation (SPO). Mosaics of high-resolution photomicrographs are first assembled and processed in Adobe Photoshop®. The Adobe Illustrator® “Live Trace” tool, whose settings can be optimized for reflected light images, completes initial automatic grain tracing and phase separation. Checking and re-classification of phases using reflected light properties and trace editing occurs manually. Phase identification is confirmed by microprobe or quantitative EDS, after which grain traces are easily reclassified as needed. Traces are imported into SPO2003 (Launeau and Robin, 2005) for SPO analysis. The combination of image resolution and magnification used here includes grains down to 10 microns. This work is part of an ongoing study examining fabric development across strain gradients in the granulite facies Capricorn ridge shear zone exposed in the Mt. Hay block of central Australia (Waters-Tormey et al., 2009). Strain marker shape fabrics, mesoscale structures, and strain localization adjacent to major lithologic boundaries all indicate that the deformation involved flattening, but that components of the deformation have been partitioned into different lithological domains. Thin sections were taken from the two gabbroic map units which volumetrically dominate the shear zone (northern and southern) using samples with similar outcrop fabric intensity. Prior thermomagnetic analyses indicate these units contain magnetite ± titanomagnetite ± ilmenite ± pyrrhotite

Identification of crystals in Hanford nuclear waste using polarized light microscopy

International Nuclear Information System (INIS)

Herting, D.L.

1984-09-01

The use of polarized light microscopy for identifying crystals encountered in Rockwell Hanford Operations chemical studies is described. Identifying characteristics and full-color photographs are presented for crystals commonly found in Hanford Site nuclear waste, including sodium nitrate, sodium nitrite, sodium aluminate, sodium phosphate, sodium fluoride, ammonium heptafluorozirconate, sodium sulfate, sodium carbonate, and ammonium nitrate. These characteristics are described in terms of birefringence, extinction position, interference figure, sign of elongation, optic sign, and crystal morphology. Background information on crystal optics is presented so that these traits can be understood by the nonmicroscopist. Detailed operational instructions are given so that the novice microscope user can make the proper adjustments of the instrument to search for and observe the identifying features of the crystals

Live Imaging of Cellular Internalization of Single Colloidal Particle by Combined Label-Free and Fluorescence Total Internal Reflection Microscopy.

Science.gov (United States)

Byrne, Gerard D; Vllasaliu, Driton; Falcone, Franco H; Somekh, Michael G; Stolnik, Snjezana

2015-11-02

In this work we utilize the combination of label-free total internal reflection microscopy and total internal reflectance fluorescence (TIRM/TIRF) microscopy to achieve a simultaneous, live imaging of single, label-free colloidal particle endocytosis by individual cells. The TIRM arm of the microscope enables label free imaging of the colloid and cell membrane features, while the TIRF arm images the dynamics of fluorescent-labeled clathrin (protein involved in endocytosis via clathrin pathway), expressed in transfected 3T3 fibroblasts cells. Using a model polymeric colloid and cells with a fluorescently tagged clathrin endocytosis pathway, we demonstrate that wide field TIRM/TIRF coimaging enables live visualization of the process of colloidal particle interaction with the labeled cell structure, which is valuable for discerning the membrane events and route of colloid internalization by the cell. We further show that 500 nm in diameter model polystyrene colloid associates with clathrin, prior to and during its cellular internalization. This association is not apparent with larger, 1 μm in diameter colloids, indicating an upper particle size limit for clathrin-mediated endocytosis.

Visible light optical coherence microscopy imaging of the mouse cortex with femtoliter volume resolution

Science.gov (United States)

Merkle, Conrad W.; Chong, Shau Poh; Kho, Aaron M.; Zhu, Jun; Kholiqov, Oybek; Dubra, Alfredo; Srinivasan, Vivek J.

2018-02-01

Most flying-spot Optical Coherence Tomography (OCT) and Optical Coherence Microscopy (OCM) systems use a symmetric confocal geometry, where the detection path retraces the illumination path starting from and ending with the spatial mode of a single mode optical fiber. Here, we describe a visible light OCM instrument that breaks this symmetry to improve transverse resolution without sacrificing collection efficiency in scattering tissue. This was achieved by overfilling a 0.3 numerical aperture (NA) water immersion objective on the illumination path, while maintaining a conventional Gaussian mode detection path (1/e2 intensity diameter 0.82 Airy disks), enabling 1.1 μm full-width at half-maximum (FWHM) transverse resolution. At the same time, a 0.9 μm FWHM axial resolution in tissue, achieved by a broadband visible light source, enabled femtoliter volume resolution. We characterized this instrument according to paraxial coherent microscopy theory, and then used it to image the meningeal layers, intravascular red blood cell-free layer, and myelinated axons in the mouse neocortex in vivo through the thinned skull. Finally, by introducing a 0.8 NA water immersion objective, we improved the lateral resolution to 0.44 μm FWHM, which provided a volumetric resolution of 0.2 fL, revealing cell bodies in cortical layer I of the mouse brain with OCM for the first time.

Structured illumination microscopy and its new developments

Directory of Open Access Journals (Sweden)

Jianling Chen

2016-05-01

Full Text Available Optical microscopy allows us to observe the biological structures and processes within living cells. However, the spatial resolution of the optical microscopy is limited to about half of the wavelength by the light diffraction. Structured illumination microscopy (SIM, a type of new emerging super-resolution microscopy, doubles the spatial resolution by illuminating the specimen with a patterned light, and the sample and light source requirements of SIM are not as strict as the other super-resolution microscopy. In addition, SIM is easier to combine with the other imaging techniques to improve their imaging resolution, leading to the developments of diverse types of SIM. SIM has great potential to meet the various requirements of living cells imaging. Here, we review the recent developments of SIM and its combination with other imaging techniques.

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...

Correlative Super-Resolution Microscopy: New Dimensions and New Opportunities.

Science.gov (United States)

Hauser, Meghan; Wojcik, Michal; Kim, Doory; Mahmoudi, Morteza; Li, Wan; Xu, Ke

2017-06-14

Correlative microscopy, the integration of two or more microscopy techniques performed on the same sample, produces results that emphasize the strengths of each technique while offsetting their individual weaknesses. Light microscopy has historically been a central method in correlative microscopy due to its widespread availability, compatibility with hydrated and live biological samples, and excellent molecular specificity through fluorescence labeling. However, conventional light microscopy can only achieve a resolution of ∼300 nm, undercutting its advantages in correlations with higher-resolution methods. The rise of super-resolution microscopy (SRM) over the past decade has drastically improved the resolution of light microscopy to ∼10 nm, thus creating exciting new opportunities and challenges for correlative microscopy. Here we review how these challenges are addressed to effectively correlate SRM with other microscopy techniques, including light microscopy, electron microscopy, cryomicroscopy, atomic force microscopy, and various forms of spectroscopy. Though we emphasize biological studies, we also discuss the application of correlative SRM to materials characterization and single-molecule reactions. Finally, we point out current limitations and discuss possible future improvements and advances. We thus demonstrate how a correlative approach adds new dimensions of information and provides new opportunities in the fast-growing field of SRM.

Fluorescence (Multiwave) Confocal Microscopy.

Science.gov (United States)

Welzel, J; Kästle, Raphaela; Sattler, Elke C

2016-10-01

In addition to reflectance confocal microscopy, multiwave confocal microscopes with different laser wavelengths in combination with exogenous fluorophores allow fluorescence mode confocal microscopy in vivo and ex vivo. Fluorescence mode confocal microscopy improves the contrast between the epithelium and the surrounding soft tissue and allows the depiction of certain structures, like epithelial tumors, nerves, and glands. Copyright © 2016 Elsevier Inc. All rights reserved.

Refractive Index Imaging of Cells with Variable-Angle Near-Total Internal Reflection (TIR) Microscopy.

Science.gov (United States)

Bohannon, Kevin P; Holz, Ronald W; Axelrod, Daniel

2017-10-01

The refractive index in the interior of single cells affects the evanescent field depth in quantitative studies using total internal reflection (TIR) fluorescence, but often that index is not well known. We here present method to measure and spatially map the absolute index of refraction in a microscopic sample, by imaging a collimated light beam reflected from the substrate/buffer/cell interference at variable angles of incidence. Above the TIR critical angle (which is a strong function of refractive index), the reflection is 100%, but in the immediate sub-critical angle zone, the reflection intensity is a very strong ascending function of incidence angle. By analyzing the angular position of that edge at each location in the field of view, the local refractive index can be estimated. In addition, by analyzing the steepness of the edge, the distance-to-substrate can be determined. We apply the technique to liquid calibration samples, silica beads, cultured Chinese hamster ovary cells, and primary culture chromaffin cells. The optical technique suffers from decremented lateral resolution, scattering, and interference artifacts. However, it still provides reasonable results for both refractive index (~1.38) and for distance-to-substrate (~150 nm) for the cells, as well as a lateral resolution to about 1 µm.

Light-free magnetic resonance force microscopy for studies of electron spin polarized systems

International Nuclear Information System (INIS)

Pelekhov, Denis V.; Selcu, Camelia; Banerjee, Palash; Chung Fong, Kin; Chris Hammel, P.; Bhaskaran, Harish; Schwab, Keith

2005-01-01

Magnetic resonance force microscopy is a scanned probe technique capable of three-dimensional magnetic resonance imaging. Its excellent sensitivity opens the possibility for magnetic resonance studies of spin accumulation resulting from the injection of spin polarized currents into a para-magnetic collector. The method is based on mechanical detection of magnetic resonance which requires low noise detection of cantilever displacement; so far, this has been accomplished using optical interferometry. This is undesirable for experiments on doped silicon, where the presence of light is known to enhance spin relaxation rates. We report a non-optical displacement detection scheme based on sensitive microwave capacitive readout

LightLeaves: computer controlled kinetic reflection hologram installation and a brief discussion of earlier work

International Nuclear Information System (INIS)

Connors Chen, Betsy

2013-01-01

LightLeaves is an installation combining leaf shaped, white light reflection holograms of landscape images with a special kinetic lighting device that houses a lamp and moving leaf shaped masks. The masks are controlled by an Arduino microcontroller and servomotors that position the masks in front of the illumination source of the holograms. The work is the most recent in a long series of landscapes that combine multi-hologram installations with computer controlled devices that play with the motion of the holograms, the light, sound or other elements in the work. LightLeaves was first exhibited at the Peabody Essex Museum in Salem, Massachusetts in a show titled E ye Spy: Playing with Perception .

LightLeaves: computer controlled kinetic reflection hologram installation and a brief discussion of earlier work

Energy Technology Data Exchange (ETDEWEB)

Connors Chen, Betsy, E-mail: acmeholo@gmail.com [ACME Holography, Somerville, Massachusetts USA 02144 (United States)

2013-02-22

LightLeaves is an installation combining leaf shaped, white light reflection holograms of landscape images with a special kinetic lighting device that houses a lamp and moving leaf shaped masks. The masks are controlled by an Arduino microcontroller and servomotors that position the masks in front of the illumination source of the holograms. The work is the most recent in a long series of landscapes that combine multi-hologram installations with computer controlled devices that play with the motion of the holograms, the light, sound or other elements in the work. LightLeaves was first exhibited at the Peabody Essex Museum in Salem, Massachusetts in a show titled {sup E}ye Spy: Playing with Perception{sup .}.

Real-time visualization of melanin granules in normal human skin using combined multiphoton and reflectance confocal microscopy.

Science.gov (United States)

Majdzadeh, Ali; Lee, Anthony M D; Wang, Hequn; Lui, Harvey; McLean, David I; Crawford, Richard I; Zloty, David; Zeng, Haishan

2015-05-01

Recent advances in biomedical optics have enabled dermal and epidermal components to be visualized at subcellular resolution and assessed noninvasively. Multiphoton microscopy (MPM) and reflectance confocal microscopy (RCM) are noninvasive imaging modalities that have demonstrated promising results in imaging skin micromorphology, and which provide complementary information regarding skin components. This study assesses whether combined MPM/RCM can visualize intracellular and extracellular melanin granules in the epidermis and dermis of normal human skin. We perform MPM and RCM imaging of in vivo and ex vivo skin in the infrared domain. The inherent three-dimensional optical sectioning capability of MPM/RCM is used to image high-contrast granular features across skin depths ranging from 50 to 90 μm. The optical images thus obtained were correlated with conventional histologic examination including melanin-specific staining of ex vivo specimens. MPM revealed highly fluorescent granular structures below the dermal-epidermal junction (DEJ) region. Histochemical staining also demonstrated melanin-containing granules that correlate well in size and location with the granular fluorescent structures observed in MPM. Furthermore, the MPM fluorescence excitation wavelength and RCM reflectance of cell culture-derived melanin were equivalent to those of the granules. This study suggests that MPM can noninvasively visualize and quantify subepidermal melanin in situ. © 2015 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

Intraocular light scatter, reflections, fluorescence and absorption: what we see in the slit lamp.

Science.gov (United States)

van den Berg, Thomas J T P

2018-01-01

Much knowledge has been collected over the past 20 years about light scattering in the eye- in particular in the eye lens- and its visual effect, called straylight. It is the purpose of this review to discuss how these insights can be applied to understanding the slit lamp image. The slit lamp image mainly results from back scattering, whereas the effects on vision result mainly from forward scatter. Forward scatter originates from particles of about wavelength size distributed throughout the lens. Most of the slit lamp image originates from small particle scatter (Rayleigh scatter). For a population of middle aged lenses it will be shown that both these scatter components remove around 10% of the light from the direct beam. For slit lamp observation close to the reflection angles, zones of discontinuity (Wasserspalten) at anterior and posterior parts of the lens show up as rough surface reflections. All these light scatter effects increase with age, but the correlations with age, and also between the different components, are weak. For retro-illumination imaging it will be argued that the density or opacity seen in areas of cortical or posterior subcapsular cataract show up because of light scattering, not because of light loss. NOTES: (1) Light scatter must not be confused with aberrations. Light penetrating the eye is divided into two parts: a relatively small part is scattered, and removed from the direct beam. Most of the light is not scattered, but continues as the direct beam. This non-scattered part is the basis for functional imaging, but its quality is under the control of aberrations. Aberrations deflect light mainly over small angles (light scatter is important because of the straylight effects over large angles (>1°), causing problems like glare and hazy vision. (2) The slit lamp image in older lenses and nuclear cataract is strongly influenced by absorption. However, this effect is greatly exaggerated by the light path lengths concerned. This

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.

Reduction of parasitic interferences in digital holographic microscopy by numerically decreased coherence length

Science.gov (United States)

Kosmeier, S.; Langehanenberg, P.; von Bally, G.; Kemper, B.

2012-01-01

Due to the large coherence length of laser light, optical path length (OPL) resolution in laser based digital holographic microscopy suffers from parasitic interferences caused by multiple reflections within the experimental setup. Use of partially coherent light reduces this drawback but requires precise and stable matching of object and reference arm's OPLs and limits the spatial frequency of the interference pattern in off-axis holography. Here, we investigate if the noise properties of spectrally broadened light sources can be generated numerically. Therefore, holograms are coherently captured at different laser wavelengths and the corresponding reconstructed wave fields are numerically superimposed utilizing variable weightings. Gaussian and rectangular spectral shapes of the so synthesized field are analyzed with respect to the resulting noise level, which is quantified in OPL distributions of a reflective test target. Utilizing a Gaussian weighting, the noise level is found to be similar to the one obtained with the partially coherent light of a superluminescent diode. With a rectangular shaped synthesized spectrum, noise is reduced more efficient than with a Gaussian one. The applicability of the method in label-free cell analysis is demonstrated by quantitative phase contrast images obtained from living cancer cells.

Two dimensional crystals of LH2 light-harvesting complexes from Ectothiorhodospira sp. and Rhodobacter capsulatus investigated by electron microscopy

NARCIS (Netherlands)

Oling, Frank; Boekema, EJ; deZarate, IO; Visschers, R; vanGrondelle, R; Keegstra, W; Brisson, A; Picorel, R

1996-01-01

Two-dimensional crystals of LH2 (B800-850) light-harvesting complexes from Ectothiorhodospira sp, and Rhodobacter capsulatus were obtained by reconstitution of purified protein into phospholipid vesicles and characterized by electron microscopy. The size of the crystals was up to several

Two-dimensional crystals of LH2 light-harvesting complexes from Ectothiorhodospira sp. and Rhodobacter capsulatus investigated by electron microscopy.

NARCIS (Netherlands)

Oling, F.; Boekema, E.J.; Ortiz de Zarate, I.; Visschers, R.W.; van Grondelle, R.; Keegstra, W.; Brisson, A.; Picorel, R.

1996-01-01

Two-dimensional crystals of LH2 (B800-850) light-harvesting complexes from Ectothiorhodospira sp. and Rhodobacter capsulatus were obtained by reconstitution of purified protein into phospholipid vesicles and characterized by electron microscopy. The size of the crystals was up to several

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. Copyright © 2016 Elsevier Ltd. All rights reserved.

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.

Light polarization management via reflection from arrays of sub-wavelength metallic twisted bands

Science.gov (United States)

Nawrot, M.; Haberko, J.; Zinkiewicz, Ł.; Wasylczyk, P.

2017-12-01

With constant progress of nano- and microfabrication technologies, photolithography in particular, a number of sub-wavelength metallic structures have been demonstrated that can be used to manipulate light polarization. Numerical simulations of light propagation hint that helical twisted bands can have interesting polarization properties. We use three-dimensional two-photon photolithography (direct laser writing) to fabricate a few-micrometer-thick arrays of twisted bands and coat them uniformly with metal. We demonstrate that circular polarization can be generated from linear polarization upon reflection from such structures over a broad range of frequencies in the mid infrared.

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

International Nuclear Information System (INIS)

Merela, M.; Pelicon, P.; Vavpetic, P.; Regvar, M.; Vogel-Mikus, K.; Sersa, I.; Policnik, H.; Pokorny, B.; Levanic, T.; Oven, P.

2009-01-01

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.

Active Appearance Segmentation for Intensity Inhomogeneity in Light Sheet Fluorescence Microscopy

DEFF Research Database (Denmark)

Jensen, Casper Bo; Lyksborg, Mark; Hecksher-Sørensen, J.

2016-01-01

inhomogeneities which are often seen in Light Sheet Fluorescence Microscopy (LSFM) images. This robustness is achieved by modelling the appearance of an image as a regularized Normalized Gradient Field (rNGF). We perform two experiments to challenge the model. First it is tested using a repeated leave......Active Appearance Models (AAM) are used for annotating or segmenting shapes in biomedical images. Performance relies heavily on the image data used to train the AAM. In this paper we improve the generalization properties of the model by making it robust to slowly varying spatial intensity......-one-out approach on images with minimal imperfections where the left out images are corrupted by a simulated bias field and segmented using the AAM. Secondly we test the model on LSFM images with common acquisition problems. In both experiments the proposed approach outperforms the often used AAM implementation...

X-ray microscopy in Aarhus

International Nuclear Information System (INIS)

Uggerhoej, Erik; Abraham-Peskir, Joanna V.

2000-01-01

The Aarhus imaging soft X-ray microscope is now a busy multi-user facility. The optical set-up will be described and project highlights discussed. a) Metal-induced structural changes in whole cells in solution. The effects of aluminum, copper, nickel and zinc on protozoa investigated by using a combination of light microscopy, confocal scanning laser microscopy and X-ray microscopy. b) Botanical studies by X-ray microscopy used to compliment electron microscopy studies. c) Sludge morphology and iron precipitation in Danish freshwater plants by combining X-ray, scanning electron and transmission electron microscopy

Automated Diatom Analysis Applied to Traditional Light Microscopy: A Proof-of-Concept Study

Science.gov (United States)

Little, Z. H. L.; Bishop, I.; Spaulding, S. A.; Nelson, H.; Mahoney, C.

2017-12-01

Diatom identification and enumeration by high resolution light microscopy is required for many areas of research and water quality assessment. Such analyses, however, are both expertise and labor-intensive. These challenges motivate the need for an automated process to efficiently and accurately identify and enumerate diatoms. Improvements in particle analysis software have increased the likelihood that diatom enumeration can be automated. VisualSpreadsheet software provides a possible solution for automated particle analysis of high-resolution light microscope diatom images. We applied the software, independent of its complementary FlowCam hardware, to automated analysis of light microscope images containing diatoms. Through numerous trials, we arrived at threshold settings to correctly segment 67% of the total possible diatom valves and fragments from broad fields of view. (183 light microscope images were examined containing 255 diatom particles. Of the 255 diatom particles present, 216 diatoms valves and fragments of valves were processed, with 170 properly analyzed and focused upon by the software). Manual analysis of the images yielded 255 particles in 400 seconds, whereas the software yielded a total of 216 particles in 68 seconds, thus highlighting that the software has an approximate five-fold efficiency advantage in particle analysis time. As in past efforts, incomplete or incorrect recognition was found for images with multiple valves in contact or valves with little contrast. The software has potential to be an effective tool in assisting taxonomists with diatom enumeration by completing a large portion of analyses. Benefits and limitations of the approach are presented to allow for development of future work in image analysis and automated enumeration of traditional light microscope images containing diatoms.

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.

The value of in vivo reflectance confocal microscopy in the diagnosis and monitoring of inflammatory and infectious skin diseases: a systematic review

NARCIS (Netherlands)

Hoogedoorn, L.; Peppelman, M.; Kerkhof, P.C.M. van de; Erp, P.E.J. van; Gerritsen, M.J.P.

2015-01-01

In vivo examination of the skin by reflectance confocal microscopy (RCM) has been performed for about 20 years, leading to a broad spectrum of imaged infectious and inflammatory skin diseases (ISD) with many described RCM features. We systematically reviewed all available literature concerning ISD

Corneal confocal microscopy alterations in Sjögren's syndrome dry eye.

Science.gov (United States)

Lanza, Michele; Iaccarino, Stefania; Varricchi, Gilda; D'Errico, Tito; Gironi Carnevale, Ugo Antonello; Bifani, Mario

2017-08-01

To evaluate light backscattering (LB) in corneal layers in patients with primary Sjögren's syndrome dry eye (SSDE) utilizing in vivo corneal confocal microscopy (IVCM) and to determine the eventual association with the lacrimal functional test values. A complete ophthalmic evaluation, Schirmer test with and without stimulation, break-up time (BUT) and IVCM were performed on 55 patients affected by SSDE and in an age- and sex-matched cohort of healthy participants (HP). Light backscattering, measures as light reflectivity unit (LRU), detected by IVCM at Bowman's membrane (BM) at 50 μm, at 100 μm and at 200 μm deeper than BM was compared in the two groups. The correlations between LB values and lacrimal function results were evaluated. In patients affected by SSDE, LB was significantly higher (p lacrimal function. Light backscattering (LB) could be very useful for clinical diagnosis and management of SSDE. © 2016 Acta Ophthalmologica Scandinavica Foundation. Published by John Wiley & Sons Ltd.

Online analysis of protein inclusion bodies produced in E. coli by monitoring alterations in scattered and reflected light.

Science.gov (United States)

Ude, Christian; Ben-Dov, Nadav; Jochums, André; Li, Zhaopeng; Segal, Ester; Scheper, Thomas; Beutel, Sascha

2016-05-01

The online monitoring of recombinant protein aggregate inclusion bodies during microbial cultivation is an immense challenge. Measurement of scattered and reflected light offers a versatile and non-invasive measurement technique. Therefore, we investigated two methods to detect the formation of inclusion bodies and monitor their production: (1) online 180° scattered light measurement (λ = 625 nm) using a sensor platform during cultivation in shake flask and (2) online measurement of the light reflective interference using a porous Si-based optical biosensor (SiPA). It could be shown that 180° scattered light measurement allows monitoring of alterations in the optical properties of Escherichia coli BL21 cells, associated with the formation of inclusion bodies during cultivation. A reproducible linear correlation between the inclusion body concentration of the non-fluorescent protein human leukemia inhibitory factor (hLIF) carrying a thioredoxin tag and the shift ("Δamp") in scattered light signal intensity was observed. This was also observed for the glutathione-S-transferase-tagged green fluorescent protein (GFP-GST). Continuous online monitoring of reflective interference spectra reveals a significant increase in the bacterium refractive index during hLIF production in comparison to a non-induced reference that coincide with the formation of inclusion bodies. These online monitoring techniques could be applied for fast and cost-effective screening of different protein expression systems.

Label-free and live cell imaging by interferometric scattering microscopy.

Science.gov (United States)

Park, Jin-Sung; Lee, Il-Buem; Moon, Hyeon-Min; Joo, Jong-Hyeon; Kim, Kyoung-Hoon; Hong, Seok-Cheol; Cho, Minhaeng

2018-03-14

Despite recent remarkable advances in microscopic techniques, it still remains very challenging to directly observe the complex structure of cytoplasmic organelles in live cells without a fluorescent label. Here we report label-free and live-cell imaging of mammalian cell, Escherischia coli , and yeast, using interferometric scattering microscopy, which reveals the underlying structures of a variety of cytoplasmic organelles as well as the underside structure of the cells. The contact areas of the cells attached onto a glass substrate, e.g. , focal adhesions and filopodia, are clearly discernible. We also found a variety of fringe-like features in the cytoplasmic area, which may reflect the folded structures of cytoplasmic organelles. We thus anticipate that the label-free interferometric scattering microscopy can be used as a powerful tool to shed interferometric light on in vivo structures and dynamics of various intracellular phenomena.

Catheter based mid-infrared reflectance and reflectance generated absorption spectroscopy

Science.gov (United States)

Holman, Hoi-Ying N

2013-10-29

A method of characterizing conditions in a tissue, by (a) providing a catheter that has a light source that emits light in selected wavenumbers within the range of mid-IR spectrum; (b) directing the light from the catheter to an area of tissue at a location inside a blood vessel of a subject; (c) collecting light reflected from the location and generating a reflectance spectra; and (d) comparing the reflectance spectra to a reference spectra of normal tissue, whereby a location having an increased number of absorbance peaks at said selected wavenumbers indicates a tissue inside the blood vessel containing a physiological marker for atherosclerosis.

Adhesion of living cells revealed by variable-angle total internal reflection fluorescence microscopy (Conference Presentation)

Science.gov (United States)

Cardoso Dos Santos, Marcelina; Vézy, Cyrille; Jaffiol, Rodolphe

2016-02-01

Total Internal Reflection Fluorescence Microscopy (TIRFM) is a widespread technique to study cellular process occurring near the contact region with the glass substrate. In this field, determination of the accurate distance from the surface to the plasma membrane constitutes a crucial issue to investigate the physical basis of cellular adhesion process. However, quantitative interpretation of TIRF pictures regarding the distance z between a labeled membrane and the substrate is not trivial. Indeed, the contrast of TIRF images depends on several parameters more and less well known (local concentration of dyes, absorption cross section, angular emission pattern…). The strategy to get around this problem is to exploit a series of TIRF pictures recorded at different incident angles in evanescent regime. This technique called variable-angle TIRF microscopy (vaTIRFM), allowing to map the membrane-substrate separation distance with a nanometric resolution (10-20 nm). vaTIRFM was developed by Burmeister, Truskey and Reichert in the early 1990s with a prism-based TIRF setup [Journal of Microscopy 173, 39-51 (1994)]. We propose a more convenient prismless setup, which uses only a rotatable mirror to adjust precisely the laser beam on the back focal plane of the oil immersion objective (no azimuthal scanning is needed). The series of TIRF images permit us to calculate accurately membrane-surface distances in each pixel. We demonstrate that vaTIRFM are useful to quantify the adhesion of living cells for specific and unspecific membrane-surface interactions, achieved on various functionalized substrates with polymers (BSA, poly-L-lysin) or extracellular matrix proteins (collagen and fibronectin).

Degrees of polarization of reflected light eliciting polarotaxis in dragonflies (Odonata), mayflies (Ephemeroptera) and tabanid flies (Tabanidae).

Science.gov (United States)

Kriska, György; Bernáth, Balázs; Farkas, Róbert; Horváth, Gábor

2009-12-01

With few exceptions insects whose larvae develop in freshwater possess positive polarotaxis, i.e., are attracted to sources of horizontally polarized light, because they detect water by means of the horizontal polarization of light reflected from the water surface. These insects can be deceived by artificial surfaces (e.g. oil lakes, asphalt roads, black plastic sheets, dark-coloured cars, black gravestones, dark glass surfaces, solar panels) reflecting highly and horizontally polarized light. Apart from the surface characteristics, the extent of such a 'polarized light pollution' depends on the illumination conditions, direction of view, and the threshold p* of polarization sensitivity of a given aquatic insect species. p* means the minimum degree of linear polarization p of reflected light that can elicit positive polarotaxis from a given insect species. Earlier there were no quantitative data on p* in aquatic insects. The aim of this work is to provide such data. Using imaging polarimetry in the red, green and blue parts of the spectrum, in multiple-choice field experiments we measured the threshold p* of ventral polarization sensitivity in mayflies, dragonflies and tabanid flies, the positive polarotaxis of which has been shown earlier. In the blue (450nm) spectral range, for example, we obtained the following thresholds: dragonflies: Enallagma cyathigerum (0%

A New Method for Automated Identification and Morphometry of Myelinated Fibers Through Light Microscopy Image Analysis

OpenAIRE

Novas, Romulo Bourget; Fazan, Valeria Paula Sassoli; Felipe, Joaquim Cezar

2015-01-01

Nerve morphometry is known to produce relevant information for the evaluation of several phenomena, such as nerve repair, regeneration, implant, transplant, aging, and different human neuropathies. Manual morphometry is laborious, tedious, time consuming, and subject to many sources of error. Therefore, in this paper, we propose a new method for the automated morphometry of myelinated fibers in cross-section light microscopy images. Images from the recurrent laryngeal nerve of adult rats and ...

Measurement of refractive indices of tunicates' tunics: light reflection of the transparent integuments in an ascidian Rhopalaea sp. and a salp Thetys vagina.

Science.gov (United States)

Kakiuchida, Hiroshi; Sakai, Daisuke; Nishikawa, Jun; Hirose, Euichi

2017-01-01

Tunic is a cellulosic, integumentary matrix found in tunicates (Subphylum Tunicata or Urochordata). The tunics of some ascidian species and pelagic tunicates, such as salps, are nearly transparent, which is useful in predator avoidance. Transparent materials can be detected visually using light reflected from their surfaces, with the different refractive indices between two media, i.e., tunic and seawater, being the measure of reflectance. A larger difference in refractive indices thus provides a larger measure of reflectance. We measured the refractive indices of the transparent tunic of Thetys vagina (salp: Thaliacea) and Rhopalae a sp. (ascidian: Ascidiacea) using an Abbe refractometer and an ellipsometer to estimate the light reflection at the tunic surface and evaluate the anti-reflection effect of the nipple array structure on the tunic surface of T. vagina . At D-line light (λ = 589 nm), the refractive indices of the tunics were 0.002-0.004 greater than seawater in the measurements by Abbe refractometer, and 0.02-0.03 greater than seawater in the measurements by ellipsometer. The refractive indices of tunics were slightly higher than that of seawater. According to the simulation of light reflection based on rigorous coupled wave analysis (RCWA), light at a large angle of incidence will be completely reflected from a surface when its refractive indices are smaller than seawater. Therefore, the refractive index of integument is important for enabling transparent organisms to remain invisible in the water column. In order to minimize reflectance, the refractive index should be similar to, but never smaller than, that of the surrounding seawater. The simulation also indicated that the presence or absence of a nipple array does not cause significant difference in reflectance on the surface. The nipple array on the tunic of the diurnal salp may have another function, such as bubble repellence, other than anti-reflection.

Topography and refractometry of sperm cells using spatial light interference microscopy.

Science.gov (United States)

Liu, Lina; Kandel, Mikhail E; Rubessa, Marcello; Schreiber, Sierra; Wheeler, Mathew B; Popescu, Gabriel

2018-02-01

Characterization of spermatozoon viability is a common test in treating infertility. Recently, it has been shown that label-free, phase-sensitive imaging can provide a valuable alternative for this type of assay. We employ spatial light interference microscopy (SLIM) to perform high-accuracy single-cell phase imaging and decouple the average thickness and refractive index information for the population. This procedure was enabled by quantitative-phase imaging cells on media of two different refractive indices and using a numerical tool to remove the curvature from the cell tails. This way, we achieved ensemble averaging of topography and refractometry of 100 cells in each of the two groups. The results show that the thickness profile of the cell tail goes down to 150 nm and the refractive index can reach values of 1.6 close to the head. (2018) COPYRIGHT Society of Photo-Optical Instrumentation Engineers (SPIE).

The Photosensitivity of Rhodopsin Bleaching and Light-Induced Increases of Fundus Reflectance in Mice Measured In Vivo With Scanning Laser Ophthalmoscopy

Science.gov (United States)

Zhang, Pengfei; Goswami, Mayank; Zawadzki, Robert J.; Pugh, Edward N.

2016-01-01

Purpose To quantify bleaching-induced changes in fundus reflectance in the mouse retina. Methods Light reflected from the fundus of albino (Balb/c) and pigmented (C57Bl/6J) mice was measured with a multichannel scanning laser ophthalmoscopy optical coherence tomography (SLO-OCT) optical system. Serial scanning of small retinal regions was used for bleaching rhodopsin and measuring reflectance changes. Results Serial scanning generated a saturating reflectance increase centered at 501 nm with a photosensitivity of 1.4 × 10−8 per molecule μm2 in both strains, 2-fold higher than expected were irradiance at the rod outer segment base equal to that at the retinal surface. The action spectrum of the reflectance increase corresponds to the absorption spectrum of mouse rhodopsin in situ. Spectra obtained before and after bleaching were fitted with a model of fundus reflectance, quantifying contributions from loss of rhodopsin absorption with bleaching, absorption by oxygenated hemoglobin (HbO2) in the choroid (Balb/c), and absorption by melanin (C57Bl/6J). Both mouse strains exhibited light-induced broadband reflectance changes explained as bleaching-induced reflectivity increases at photoreceptor inner segment/outer segment (IS/OS) junctions and OS tips. Conclusions The elevated photosensitivity of rhodopsin bleaching in vivo is explained by waveguide condensing of light in propagation from rod inner segment (RIS) to rod outer segment (ROS). The similar photosensitivity of rhodopsin in the two strains reveals that little light backscattered from the sclera can enter the ROS. The bleaching-induced increases in reflectance at the IS/OS junctions and OS tips resemble results previously reported in human cones, but are ascribed to rods due to their 30/1 predominance over cones in mice and to the relatively minor amount of cone M-opsin in the regions scanned. PMID:27403994

Visual Understanding of Light Absorption and Waveguiding in Standing Nanowires with 3D Fluorescence Confocal Microscopy.

Science.gov (United States)

Frederiksen, Rune; Tutuncuoglu, Gozde; Matteini, Federico; Martinez, Karen L; Fontcuberta I Morral, Anna; Alarcon-Llado, Esther

2017-09-20

Semiconductor nanowires are promising building blocks for next-generation photonics. Indirect proofs of large absorption cross sections have been reported in nanostructures with subwavelength diameters, an effect that is even more prominent in vertically standing nanowires. In this work we provide a three-dimensional map of the light around vertical GaAs nanowires standing on a substrate by using fluorescence confocal microscopy, where the strong long-range disruption of the light path along the nanowire is illustrated. We find that the actual long-distance perturbation is much larger in size than calculated extinction cross sections. While the size of the perturbation remains similar, the intensity of the interaction changes dramatically over the visible spectrum. Numerical simulations allow us to distinguish the effects of scattering and absorption in the nanowire leading to these phenomena. This work provides a visual understanding of light absorption in semiconductor nanowire structures, which is of high interest for solar energy conversion applications.

Dictionary of Microscopy

Science.gov (United States)

Heath, Julian

2005-10-01

The past decade has seen huge advances in the application of microscopy in all areas of science. This welcome development in microscopy has been paralleled by an expansion of the vocabulary of technical terms used in microscopy: terms have been coined for new instruments and techniques and, as microscopes reach even higher resolution, the use of terms that relate to the optical and physical principles underpinning microscopy is now commonplace. The Dictionary of Microscopy was compiled to meet this challenge and provides concise definitions of over 2,500 terms used in the fields of light microscopy, electron microscopy, scanning probe microscopy, x-ray microscopy and related techniques. Written by Dr Julian P. Heath, Editor of Microscopy and Analysis, the dictionary is intended to provide easy navigation through the microscopy terminology and to be a first point of reference for definitions of new and established terms. The Dictionary of Microscopy is an essential, accessible resource for: students who are new to the field and are learning about microscopes equipment purchasers who want an explanation of the terms used in manufacturers' literature scientists who are considering using a new microscopical technique experienced microscopists as an aide mémoire or quick source of reference librarians, the press and marketing personnel who require definitions for technical reports.

Food for thought: Conditions for discourse reflection in the light of environmental assessment

International Nuclear Information System (INIS)

Runhaar, Hens; Runhaar, Piety R.; Oegema, Tammo

2010-01-01

People tend to take notice of what is happening around them selectively. Discourses-frames through which actors give meaning to aspects of the world-act as built-in filters that distinguish relevant from irrelevant data. Use of knowledge generated by environmental assessments (EAs) in decision-making may be understood from this perspective. Environmental knowledge that is inconsistent with dominant discourses runs the risk of being ignored. Discourses on the value of EA as a tool for decision-making may have a similar effect. Stimulating decision-makers and stakeholders to critically reflect on and reconsider their discourses in the light of EAs-also known as frame reflection or policy learning-may enhance the probability that these assessments and the knowledge that they generate impact upon decision-making. Up to now little has been written about how discourse reflection in the context of EA can be promoted. Valuable inputs are fragmented over different bodies of literature. In this paper we draw from these bodies to identify favourable conditions for discourse reflection.

Glycogen in the Nervous System. I; Methods for Light and Electron Microscopy

Science.gov (United States)

Estable, Rosita F. De; Estable-Puig, J. F.; Miquel, J.

1964-01-01

'l'he relative value of different methods for combined light and electron microscopical studies of glycogen in the nervous tissue was investigated. Picroalcoholic fixatives preserve glycogen in a considerable amount but give an inadequate morphological image of glycogen distribution and are unsuitable for ultrastructural studies. Fixation by perfusion, with Dalton's chromeosmic fluid seems adequate for ultrastructural cytochemistry of glycogen. Furthermore it permits routine paraffin embedding of brain slices adjacent to those used for electron microscopy. Dimedone blocking is a necessary step for a selective staining of glycogen with PAS after osmic fixation. Enzymatic removal of glycogen in osmic fixed nervous tissue can be done In paraffin-embedded tissue. It can also be performed in glycolmethacrylate-embedded tissue without removal of the embedding medium. Paraphenylenediamine stains glycogen following periodic acid oxidation.

Dose determination with nitro blue tetrazolium containing radiochromic dye films by measuring absorbed and reflected light

DEFF Research Database (Denmark)

Kovács, A.; Baranyai, M.; Wojnárovits, L.

2000-01-01

determination in a wide dose range both by absorbance and reflectance measurements. The concept of measuring reflected light from dose labels has been discussed earlier and emerged recently due to the requirement of introducing semiquantitative label dose indicators for quarantine control. The usefulness...... of the method was studied using the newly developed radiochromic dye films as well as already existing ones. (C) 2000 Elsevier Science Ltd. All rights reserved....

Cell reactions with biomaterials: the microscopies

Directory of Open Access Journals (Sweden)

Curtis A. S.G.

2001-01-01

Full Text Available The methods and results of optical microscopy that can be used to observe cell reactions to biomaterials are Interference Reflection Microscopy (IRM, Total Internal Reflection Fluorescence Microscopy (TIRFM, Surface Plasmon Resonance Microscopy (SPRM and Forster Resonance Energy Transfer Microscopy (FRETM and Standing Wave Fluorescence Microscopy. The last three are new developments, which have not yet been fully perfected. TIRFM and SPRM are evanescent wave methods. The physics of these methods depend upon optical phenomena at interfaces. All these methods give information on the dimensions of the gap between cell and the substratum to which it is adhering and thus are especially suited to work with biomaterials. IRM and FRETM can be used on opaque surfaces though image interpretation is especially difficult for IRM on a reflecting opaque surface. These methods are compared with several electron microscopical methods for studying cell adhesion to substrata. These methods all yield fairly consistent results and show that the cell to substratum distance on many materials is in the range 5 to 30 nm. The area of contact relative to the total projected area of the cell may vary from a few per cent to close to 100% depending on the cell type and substratum. These methods show that those discrete contact areas well known as focal contacts are frequently present. The results of FRETM suggest that the separation from the substratum even in a focal contact is about 5 nm.

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

Science.gov (United States)

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

2017-06-14

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.

Imaging Early Steps of Sindbis Virus Infection by Total Internal Reflection Fluorescence Microscopy

Directory of Open Access Journals (Sweden)

Youling Gu

2011-01-01

Full Text Available Sindbis virus (SINV is an alphavirus that has a broad host range and has been widely used as a vector for recombinant gene transduction, DNA-based vaccine production, and oncolytic cancer therapy. The mechanism of SINV entry into host cells has yet to be fully understood. In this paper, we used single virus tracking under total internal reflection fluorescence microscopy (TIRFM to investigate SINV attachment to cell surface. Biotinylated viral particles were labeled with quantum dots, which retained viral viability and infectivity. By time-lapse imaging, we showed that the SINV exhibited a heterogeneous dynamics on the surface of the host cells. Analysis of SINV motility demonstrated a two-step attachment reaction. Moreover, dual color TIRFM of GFP-Rab5 and SINV suggested that the virus was targeted to the early endosomes after endocytosis. These findings demonstrate the utility of quantum dot labeling in studying the early steps and behavior of SINV infection.

Classifying distinct basal cell carcinoma subtype by means of dermatoscopy and reflectance confocal microscopy.

Science.gov (United States)

Longo, Caterina; Lallas, Aimilios; Kyrgidis, Athanassios; Rabinovitz, Harold; Moscarella, Elvira; Ciardo, Silvana; Zalaudek, Iris; Oliviero, Margaret; Losi, Amanda; Gonzalez, Salvador; Guitera, Pascale; Piana, Simonetta; Argenziano, Giuseppe; Pellacani, Giovanni

2014-10-01

The current guidelines for the management of basal cell carcinoma (BCC) suggest a different therapeutic approach according to histopathologic subtype. Although dermatoscopic and confocal criteria of BCC have been investigated, no specific studies were performed to evaluate the distinct reflectance confocal microscopy (RCM) aspects of BCC subtypes. To define the specific dermatoscopic and confocal criteria for delineating different BCC subtypes. Dermatoscopic and confocal images of histopathologically confirmed BCCs were retrospectively evaluated for the presence of predefined criteria. Frequencies of dermatoscopic and confocal parameters are provided. Univariate and adjusted odds ratios were calculated. Discriminant analyses were performed to define the independent confocal criteria for distinct BCC subtypes. Eighty-eight BCCs were included. Dermatoscopically, superficial BCCs (n=44) were primarily typified by the presence of fine telangiectasia, multiple erosions, leaf-like structures, and revealed cords connected to the epidermis and epidermal streaming upon RCM. Nodular BCCs (n=22) featured the classic dermatoscopic features and well outlined large basaloid islands upon RCM. Infiltrative BCCs (n=22) featured structureless, shiny red areas, fine telangiectasia, and arborizing vessels on dermatoscopy and dark silhouettes upon RCM. The retrospective design. Dermatoscopy and confocal microscopy can reliably classify different BCC subtypes. Copyright © 2014 American Academy of Dermatology, Inc. Published by Elsevier Inc. All rights reserved.

CR-39 (PADC) Reflection and Transmission of Light in the Ultraviolet-Near-Infrared (UV-NIR) Range.

Science.gov (United States)

Traynor, Nathan B J; McLauchlin, Christopher; Dodge, Kenneth; McGarrah, James E; Padalino, Stephen J; McCluskey, Michelle; Sangster, T C; McLean, James G

2018-04-01

The spectral reflection (specular and diffuse) and transmission of Columbia Resin 39 (CR-39) were measured for incoherent light with wavelengths in the range of 200-2500 nm. These results will be of use for the optical characterization of CR-39, as well as in investigations of the chemical modifications of the polymer caused by ultraviolet (UV) exposure. A Varian Cary 5000 was used to perform spectroscopy on several different thicknesses of CR-39. With proper analysis for the interdependence of reflectance and transmittance, results are consistent across all samples. The reflectivity from each CR-39-air boundary reveals an increase in the index of refraction in the near-UV. Absorption observations are consistent with the Beer-Lambert law. Strong absorption of UV light of wavelength shorter than 350 nm suggests an optical band gap of 3.5 eV, although the standard analysis is not conclusive. Absorption features observed in the near infrared are assigned to molecular vibrations, including some that are new to the literature.

Super-resolution and super-localization microscopy: A novel tool for imaging chemical and biological processes

Energy Technology Data Exchange (ETDEWEB)

Dong, Bin [Iowa State Univ., Ames, IA (United States)

2015-01-01

Optical microscopy imaging of single molecules and single particles is an essential method for studying fundamental biological and chemical processes at the molecular and nanometer scale. The best spatial resolution (~ λ/2) achievable in traditional optical microscopy is governed by the diffraction of light. However, single molecule-based super-localization and super-resolution microscopy imaging techniques have emerged in the past decade. Individual molecules can be localized with nanometer scale accuracy and precision for studying of biological and chemical processes.This work uncovered the heterogeneous properties of the pore structures. In this dissertation, the coupling of molecular transport and catalytic reaction at the single molecule and single particle level in multilayer mesoporous nanocatalysts was elucidated. Most previous studies dealt with these two important phenomena separately. A fluorogenic oxidation reaction of non-fluorescent amplex red to highly fluorescent resorufin was tested. The diffusion behavior of single resorufin molecules in aligned nanopores was studied using total internal reflection fluorescence microscopy (TIRFM).

Shedding light on endocytosis with optimized super-resolution microscopy

NARCIS (Netherlands)

Leyton Puig, D.M.

2017-01-01

Super-resolution microscopy is a relatively new microscopy technique that is still under optimization. In this thesis we focus on the improvement of the quality of super-resolution images, to apply them to the study of the processes of cell signaling and endocytosis. First, we show that the use of a

Multi-modality photoacoustic tomography, ultrasound, and light sheet microscopy for volumetric tumor margin detection

Science.gov (United States)

Sangha, Gurneet S.; Hu, Bihe; Bolus, Daniel; Wang, Mei; Skidmore, Shelby J.; Sholl, Andrew B.; Brown, J. Quincy; Goergen, Craig J.

2018-02-01

Current methods for breast tumor margin detection are invasive, time consuming, and typically result in a reoperative rate of over 25%. This marks a clear clinical need to develop improved tools to intraoperatively differentiate negative versus positive tumor margins. Here, we utilize photoacoustic tomography (PAT), ultrasound (US), and inverted Selective Plane Illumination Microscopy (iSPIM) to assess breast tumor margins in eight human breast biopsies. Our PAT/US system consists of a tunable Nd:YAG laser (NT 300, EKSPLA) coupled with a 40MHz central frequency US probe (Vevo2100, FUJIFILM Visual Sonics). This system allows for the delivery of 10Hz, 5ns pulses with fluence of 40mJ/cm2 to the tissue with PAT and US axial resolutions of 125μm and 40μm, respectively. For this study, we used a linear stepper motor to acquire volumetric PAT/US images of the breast biopsies using 1100nm light to identify bloodrich "tumor" regions and 1210nm light to identify lipid-rich "healthy" regions. iSPIM (Applied Scientific Instrumentation) is an advanced microscopy technique with lateral resolution of 1.5μm and axial resolution of 7μm. We used 488nm laser excitation and acridine orange as a general comprehensive histology stain. Our results show that PAT/US can be used to identify lipid-rich regions, dense areas of arterioles and arteries, and other internal structures such as ducts. iSPIM images correlate well with histopathology slides and can verify nuclear features, cell type and density, stromal features, and microcalcifications. Together, this multimodality approach has the potential to improve tumor margin detection with a high degree of sensitivity and specificity.

An Entertaining Method of Teaching Concepts of Linear Light Propagation, Reflection and Refraction Using a Simple Optical Mechanism

Science.gov (United States)

Yurumezoglu, K.

2009-01-01

An activity has been designed for the purpose of teaching how light is dispersed in a straight line and about the interaction between matter and light as well as the related concepts of shadows, partial shadows, reflection, refraction, primary colours and complementary (secondary) colours, and differentiating the relationship between colours, all…

Soft X-Ray Microscopy and Spectroscopy at the Molecular Environmental Science Beamline at the Advanced Light Source

Energy Technology Data Exchange (ETDEWEB)

Bluhm, Hendrik; Andersson, Klas J.; Araki, Tohru; Benzerara, Karim; Brown, Gordon E.; Dynes, Jay J.; Ghosal, Sutapa; Gilles, Mary K.; Hansen, Hans C.; Hemminger, J. C.; Hitchcock, Adam P.; Ketteler, Guido; Kilcoyne, Arthur L.; Kneedler, Eric M.; Lawrence, John R.; Leppard, Gary G.; Majzlam, Juraj; Mun, B. S.; Myneni, Satish C.; Nilsson, Anders R.; Ogasawara, Hirohito; Ogletree, D. F.; Pecher, Klaus H.; Salmeron, Miquel B.; Shuh, David K.; Tonner, Brian; Tyliszczak, Tolek; Warwick, Tony; Yoon, T. H.

2006-02-01

We present examples of the application of synchrotron-based spectroscopies and microscopies to environmentally-relevant samples. The experiments were performed at the Molecular Environmental Science beamline (11.0.2) at the Advanced Light Source, Lawrence Berkeley National Laboratory. Examples range from the study of water monolayers on Pt(111) single crystal surfaces using X-ray emission spectroscopy and the examination of alkali halide solution/water vapor interfaces using ambient pressure photoemission spectroscopy, to the investigation of actinides, river-water biofilms, Al-containing colloids and mineral-bacteria suspensions using scanning transmission X-ray spectromicroscopy. The results of our experiments show that spectroscopy and microscopy in the soft X-ray energy range are excellent tools for the investigation of environmentally relevant samples under realistic conditions, i.e. with water or water vapor present at ambient temperature.

Exploiting total internal reflection geometry for efficient optical modulation of terahertz light

Directory of Open Access Journals (Sweden)

Xudong Liu

2016-10-01

Full Text Available Efficient methods to modulate terahertz (THz light are essential for realizing rapid THz imaging and communication applications. Here we report a novel THz modulator which utilizes the evanescent wave in a total internal reflection setup coupled with a conductive interface to enhance the attenuation efficiency of THz light. This approach makes it possible to achieve close to 100% modulation with a small interface conductivity of 12 mS. The frequency dependence of this technique is linked to the optical properties of the materials: a material with close to frequency independent conductivity that is also controllable will result in an achromatic modulation response, and the device performance can be optimized further by tuning the internal reflection angle. In this work, we focus on applying the technique in the terahertz frequency range. Using an LED array with a pump intensity of 475 mW/cm2 to produce carriers in a silicon wafer, we have achieved a modulation depth of up to 99.9% in a broad frequency range of 0.1 THz–0.8 THz. The required pumping power for the generation of the required free carriers is low because the sheet conductivity needed is far less than required for traditional transmission techniques. Consequently, the device can be modulated by an LED making it a very practical, low cost, and scalable solution for THz modulation.

Comparison of mouse mammary gland imaging techniques and applications: Reflectance confocal microscopy, GFP Imaging, and ultrasound

International Nuclear Information System (INIS)

Tilli, Maddalena T; Parrish, Angela R; Cotarla, Ion; Jones, Laundette P; Johnson, Michael D; Furth, Priscilla A

2008-01-01

Genetically engineered mouse models of mammary gland cancer enable the in vivo study of molecular mechanisms and signaling during development and cancer pathophysiology. However, traditional whole mount and histological imaging modalities are only applicable to non-viable tissue. We evaluated three techniques that can be quickly applied to living tissue for imaging normal and cancerous mammary gland: reflectance confocal microscopy, green fluorescent protein imaging, and ultrasound imaging. In the current study, reflectance confocal imaging offered the highest resolution and was used to optically section mammary ductal structures in the whole mammary gland. Glands remained viable in mammary gland whole organ culture when 1% acetic acid was used as a contrast agent. Our application of using green fluorescent protein expressing transgenic mice in our study allowed for whole mammary gland ductal structures imaging and enabled straightforward serial imaging of mammary gland ducts in whole organ culture to visualize the growth and differentiation process. Ultrasound imaging showed the lowest resolution. However, ultrasound was able to detect mammary preneoplastic lesions 0.2 mm in size and was used to follow cancer growth with serial imaging in living mice. In conclusion, each technique enabled serial imaging of living mammary tissue and visualization of growth and development, quickly and with minimal tissue preparation. The use of the higher resolution reflectance confocal and green fluorescent protein imaging techniques and lower resolution ultrasound were complementary

Comparative morphology of the snake spectacle using light and transmission electron microscopy

DEFF Research Database (Denmark)

Da Silva, Mari-Ann O; Bertelsen, Mads F; Wang, Tobias

2016-01-01

OBJECTIVE: To determine the interspecific variation in the morphology of the snake spectacle. ANIMALS STUDIED: About 43 snakes of 14 different species, belonging to three different families: Boidae, Colubridae, and Pythonidae. PROCEDURE: The spectacles were examined by light and transmission...... electron microscopy. The thickness of the stromal layer was measured and the location of the blood vessels was noted. The shape of the transition zone located at the rim of the spectacle and the presence of pigment herein were also recorded. RESULTS: The spectacles of all species examined consisted...... of three layers. The outer epithelium was made of basal cells with overlaying keratin layers, the stroma comprised layers of organized collagen fibrils, and the inner epithelium was a layer of squamous cells with microvilli. Blood vessels were found in the stroma of all spectacles: in boas and pythons...

Platinum replica electron microscopy: Imaging the cytoskeleton globally and locally.

Science.gov (United States)

Svitkina, Tatyana M

2017-05-01

Structural studies reveal how smaller components of a system work together as a whole. However, combining high resolution of details with full coverage of the whole is challenging. In cell biology, light microscopy can image many cells in their entirety, but at a lower resolution, whereas electron microscopy affords very high resolution, but usually at the expense of the sample size and coverage. Structural analyses of the cytoskeleton are especially demanding, because cytoskeletal networks are unresolvable by light microscopy due to their density and intricacy, whereas their proper preservation is a challenge for electron microscopy. Platinum replica electron microscopy can uniquely bridge the gap between the "comfort zones" of light and electron microscopy by allowing high resolution imaging of the cytoskeleton throughout the entire cell and in many cells in the population. This review describes the principles and applications of platinum replica electron microscopy for studies of the cytoskeleton. Copyright © 2017 Elsevier Ltd. All rights reserved.

Observation of two regions of selective light reflection from a thin film of a cholesteric liquid crystal

International Nuclear Information System (INIS)

Alaverdyan, R B; Dadalyan, T K; Chilingaryan, Yurii S

2013-01-01

Two regions of selective light reflection (in the short- and long- wavelength parts of the visible spectrum) from a thin film of a cholesteric liquid crystal (CLC), consisting of the mixture of two CLCs with opposite chirality and a nematic liquid crystal, are experimentally found for the first time. The spectral position of the reflection regions and the separation between them varies depending on the CLC composition and the temperature. The long-wavelength region of reflection corresponds to the region of Bragg reflection from the CLC helix, while the short-wavelength region is probably due to the defects in the structure of the CLC film. (letters)

Electrocatalytic activity mapping of model fuel cell catalyst films using scanning electrochemical microscopy

International Nuclear Information System (INIS)

Nicholson, P.G.; Zhou, S.; Hinds, G.; Wain, A.J.; Turnbull, A.

2009-01-01

Scanning electrochemical microscopy has been employed to spatially map the electrocatalytic activity of model proton exchange membrane fuel cell (PEMFC) catalyst films towards the hydrogen oxidation reaction (the PEMFC anode reaction). The catalyst films were composed of platinum-loaded carbon nanoparticles, similar to those typically used in PEMFCs. The electrochemical characterisation was correlated with a detailed physical characterisation using dynamic light scattering, transmission electron microscopy and field-emission scanning electron microscopy. The nanoparticles were found to be reasonably mono-dispersed, with a tendency to agglomerate into porous bead-type structures when spun-cast. The number of carbon nanoparticles with little or no platinum was surprisingly higher than would be expected based on the platinum-carbon mass ratio. Furthermore, the platinum-rich carbon particles tended to agglomerate and the clusters formed were non-uniformly distributed. This morphology was reflected in a high degree of heterogeneity in the film activity towards the hydrogen oxidation reaction.

Zinc vanadate nanorods and their visible light photocatalytic activity

International Nuclear Information System (INIS)

Pei, L.Z.; Lin, N.; Wei, T.; Liu, H.D.; Yu, H.Y.

2015-01-01

Highlights: • Zinc vanadate nanorods have been synthesized by a facile hydrothermal process. • The size of zinc vanadate nanorods can be controlled by growth conditions. • Zinc vanadate nanorods show good photocatalytic activities of methylene blue under solar light. - Abstract: Zinc vanadate nanorods have been synthesized by a simple hydrothermal process using zinc acetate and sodium vanadate as the raw materials. The zinc vanadate nanorods have been characterized by X-ray diffraction (XRD), scanning electron microscopy (SEM), transmission electron microscopy (TEM), high-resolution TEM (HRTEM) and solid UV–vis diffuse reflectance spectrum. XRD pattern and HRTEM image show that the zinc vanadate nanorods are composed of single crystalline monoclinic Zn 2 V 2 O 7 phase. SEM and TEM observations show that the diameter and length of the zinc vanadate nanorods are 50–100 nm and about 5 μm, respectively. Sodium dodecyl sulfonate (SDS) has an essential role in the formation of zinc vanadate nanorods. The SDS-assisted nucleation and growth process have been proposed to explain the formation and growth of the zinc vanadate nanorods. Solid UV–vis diffuse reflectance spectrum shows that the zinc vanadate nanorods have a band gap of 2.76 eV. The photocatalytic activities of the zinc vanadate nanorods have been evaluated by the photocatalytic degradation of methylene blue (MB) under solar light irradiation. The MB with the concentration of 10 mg L −1 can be degraded totally under the solar light irradiation for 4 h. It is suggested that the zinc vanadate nanorods exhibit promising application potential for the degradation of organic pollutants under solar light irradiation

Gestational related changes in the deep venous system of the lower limb on light reflection rheography in pregnancy and the puerperium

Energy Technology Data Exchange (ETDEWEB)

Calderwood, C.J. [St John' s Hospital, Livingston and Simpson Centre for Reproductive Health, Royal Infirmary of Edinburgh, Edinburgh (United Kingdom)], E-mail: catherine.calderwood@luht.scot.nhs.uk; Jamieson, R. [Princess Royal Maternity Unit, Glasgow Royal Infirmary, Glasgow (United Kingdom); Greer, I.A. [Hull York Medical School, University of York, York (United Kingdom)

2007-12-15

Objective: To assess whether light reflection rheography testing is affected by the changes that occur in the deep venous system of the lower limb in pregnancy and the puerperium. Methods: Twenty five women with a singleton pregnancy were recruited to undergo duplex Doppler ultrasound examinations of the common femoral vein to measure the vessel diameter and the blood flow velocity. Light reflection rheography testing was subsequently performed and the rate of venous emptying in the lower limb calculated. Serial measurements using both techniques were made at 15, 28, 36 weeks, and term gestation and at 2 days and 6 weeks postpartum. Results: Duplex Doppler ultrasound confirmed that there is progressive dilatation of the deep venous system in pregnancy, which reaches a maximum at term and reverses after delivery. There is an accompanying reduction in blood flow velocity, which reaches a nadir at term and increases after delivery. The rate of venous emptying as measured by light reflection rheography decreases with increasing gestation, but did not fall to a level consistent with venous occlusion by a deep venous thrombosis. Conclusions: Light reflection rheography has been shown to provide reliable results in pregnancy and the puerperium. Therefore, it is a potential tool for screening for deep venous thrombosis in this population.

Gestational related changes in the deep venous system of the lower limb on light reflection rheography in pregnancy and the puerperium

International Nuclear Information System (INIS)

Calderwood, C.J.; Jamieson, R.; Greer, I.A.

2007-01-01

Objective: To assess whether light reflection rheography testing is affected by the changes that occur in the deep venous system of the lower limb in pregnancy and the puerperium. Methods: Twenty five women with a singleton pregnancy were recruited to undergo duplex Doppler ultrasound examinations of the common femoral vein to measure the vessel diameter and the blood flow velocity. Light reflection rheography testing was subsequently performed and the rate of venous emptying in the lower limb calculated. Serial measurements using both techniques were made at 15, 28, 36 weeks, and term gestation and at 2 days and 6 weeks postpartum. Results: Duplex Doppler ultrasound confirmed that there is progressive dilatation of the deep venous system in pregnancy, which reaches a maximum at term and reverses after delivery. There is an accompanying reduction in blood flow velocity, which reaches a nadir at term and increases after delivery. The rate of venous emptying as measured by light reflection rheography decreases with increasing gestation, but did not fall to a level consistent with venous occlusion by a deep venous thrombosis. Conclusions: Light reflection rheography has been shown to provide reliable results in pregnancy and the puerperium. Therefore, it is a potential tool for screening for deep venous thrombosis in this population

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.

High-throughput isotropic mapping of whole mouse brain using multi-view light-sheet microscopy

Science.gov (United States)

Nie, Jun; Li, Yusha; Zhao, Fang; Ping, Junyu; Liu, Sa; Yu, Tingting; Zhu, Dan; Fei, Peng

2018-02-01

Light-sheet fluorescence microscopy (LSFM) uses an additional laser-sheet to illuminate selective planes of the sample, thereby enabling three-dimensional imaging at high spatial-temporal resolution. These advantages make LSFM a promising tool for high-quality brain visualization. However, even by the use of LSFM, the spatial resolution remains insufficient to resolve the neural structures across a mesoscale whole mouse brain in three dimensions. At the same time, the thick-tissue scattering prevents a clear observation from the deep of brain. Here we use multi-view LSFM strategy to solve this challenge, surpassing the resolution limit of standard light-sheet microscope under a large field-of-view (FOV). As demonstrated by the imaging of optically-cleared mouse brain labelled with thy1-GFP, we achieve a brain-wide, isotropic cellular resolution of 3μm. Besides the resolution enhancement, multi-view braining imaging can also recover complete signals from deep tissue scattering and attenuation. The identification of long distance neural projections across encephalic regions can be identified and annotated as a result.

Application of super-resolution optical microscopy in biology

International Nuclear Information System (INIS)

Mao Xiuhai; Du Jiancong; Huang Qing; Fan Chunhai; Deng Suhui

2013-01-01

Background: A noninvasive, real-time far-field optical microscopy is needed to study the dynamic function inside cells and proteins. However, the resolution limit of traditional optical microscope is about 200 nm due to the diffraction limit of light. So, it's hard to directly observe the subcellular structures. Over the past several years of microscopy development, the diffraction limit of fluorescence microscopy has been overcome and its resolution limit is about tens of nanometers. Methods: To overcome the diffraction limit of light, many super-resolution fluoresce microscopes, including stimulated emission of depletion microscopy (STED), photoactivation localization microscopy (PALM) and stochastic optical reconstruction microscopy (STORM), have been developed. Conclusions: These methods have been applied in cell biology, microbiology and neurobiology, and the technology of super-resolution provides a new insight into the life science. (authors)

Super-resolved linear fluorescence localization microscopy using photostable fluorophores: A virtual microscopy study

Science.gov (United States)

Birk, Udo; Szczurek, Aleksander; Cremer, Christoph

2017-12-01

Current approaches to overcome the conventional limit of the resolution potential of light microscopy (of about 200 nm for visible light), often suffer from non-linear effects, which render the quantification of the image intensities in the reconstructions difficult, and also affect the quantification of the biological structure under investigation. As an attempt to face these difficulties, we discuss a particular method of localization microscopy which is based on photostable fluorescent dyes. The proposed method can potentially be implemented as a fast alternative for quantitative localization microscopy, circumventing the need for the acquisition of thousands of image frames and complex, highly dye-specific imaging buffers. Although the need for calibration remains in order to extract quantitative data (such as the number of emitters), multispectral approaches are largely facilitated due to the much less stringent requirements on imaging buffers. Furthermore, multispectral acquisitions can be readily obtained using commercial instrumentation such as e.g. the conventional confocal laser scanning microscope.

Photoluminescence and reflectivity studies of high energy light ions irradiated polymethyl methacrylate films

Science.gov (United States)

Bharti, Madhu Lata; Singh, Fouran; Ramola, R. C.; Joshi, Veena

2017-11-01

The self-standing films of non-conducting polymethyl methacrylate (PMMA) were irradiated in vacuum using high energy light ions (HELIs) of 50 MeV Lithium (Li+3) and 80 MeV Carbon (C+5) at various ion dose to induce the optical changes in the films. Upon HELI irradiation, films exhibit a significant enhancement in optical reflectivity at the highest dose. Interestingly, the photoluminescence (PL) emission band with green light at (514.5 nm) shows a noticeable increase in the intensity with increasing ion dose for both ions. However, the rate of increase in PL intensity is different for both HELI and can be correlated with the linear energy transfer by these ions in the films. Origin of PL is attributed to the formation of carbon cluster and hydrogenated amorphous carbon in the polymer films. HAC clusters act as PL active centres with optical reflectivity. Most of the harmful radiation like UV are absorbed by the material and is becoming opaque after irradiation and this PL active material are useful in fabrication of optoelectronic devices, UV-filter, back-lit components in liquid crystal display systems, micro-components for integrate optical circuits, diffractive elements, advanced materials and are also applicable to the post irradiation laser treatment by means of ion irradiation.

Separation of ballistic and diffusive fluorescence photons in confocal Light-Sheet Microscopy of Arabidopsis roots

Science.gov (United States)

Meinert, Tobias; Tietz, Olaf; Palme, Klaus J.; Rohrbach, Alexander

2016-01-01

Image quality in light-sheet fluorescence microscopy is strongly affected by the shape of the illuminating laser beam inside embryos, plants or tissue. While the phase of Gaussian or Bessel beams propagating through thousands of cells can be partly controlled holographically, the propagation of fluorescence light to the detector is difficult to control. With each scatter process a fluorescence photon loses information necessary for the image generation. Using Arabidopsis root tips we demonstrate that ballistic and diffusive fluorescence photons can be separated by analyzing the image spectra in each plane without a priori knowledge. We introduce a theoretical model allowing to extract typical scattering parameters of the biological material. This allows to attenuate image contributions from diffusive photons and to amplify the relevant image contributions from ballistic photons through a depth dependent deconvolution. In consequence, image contrast and resolution are significantly increased and scattering artefacts are minimized especially for Bessel beams with confocal line detection. PMID:27553506

Separation of ballistic and diffusive fluorescence photons in confocal Light-Sheet Microscopy of Arabidopsis roots.

Science.gov (United States)

Meinert, Tobias; Tietz, Olaf; Palme, Klaus J; Rohrbach, Alexander

2016-08-24

Image quality in light-sheet fluorescence microscopy is strongly affected by the shape of the illuminating laser beam inside embryos, plants or tissue. While the phase of Gaussian or Bessel beams propagating through thousands of cells can be partly controlled holographically, the propagation of fluorescence light to the detector is difficult to control. With each scatter process a fluorescence photon loses information necessary for the image generation. Using Arabidopsis root tips we demonstrate that ballistic and diffusive fluorescence photons can be separated by analyzing the image spectra in each plane without a priori knowledge. We introduce a theoretical model allowing to extract typical scattering parameters of the biological material. This allows to attenuate image contributions from diffusive photons and to amplify the relevant image contributions from ballistic photons through a depth dependent deconvolution. In consequence, image contrast and resolution are significantly increased and scattering artefacts are minimized especially for Bessel beams with confocal line detection.

Electronic Blending in Virtual Microscopy

Science.gov (United States)

Maybury, Terrence S.; Farah, Camile S.

2010-01-01

Virtual microscopy (VM) is a relatively new technology that transforms the computer into a microscope. In essence, VM allows for the scanning and transfer of glass slides from light microscopy technology to the digital environment of the computer. This transition is also a function of the change from print knowledge to electronic knowledge, or as…

The FTIR study of uranium oxides by the method of light pipe reflection spectroscopy

International Nuclear Information System (INIS)

Bao Zhu Yu; Hansen, W.N.

1988-01-01

Light pipe infrared reflection spectra of UO 2 , UO 3 , U 3 O 8 have been studied by using an FTIR spectrometer. The uranium oxide powders were ground to ensure fine particle size and distributed on the inner surface of a straight glass pipe with gold coating. The infrared beam from the inter-ferometer was focused into one end of the pipe at 45 0 incidence and then the transmitted beam was refocused by a pair of Cassegrainian type mirrors. The resultant spectra show the infrared characteristics of the ...-U-O-U-O-..., uranyl ion UO 2 2+ bond vibration and the active lattice vibrations predicted by group theory calculations. In comparison to the transmission spectra measured by authors or reported in literature, this 45 0 incident light pipe method as well as the previous light pipe method offer advantages of sensitivity, ease of acquisition and interpretation, and require a very small sample. It confirms the power of the light pipe method for studying powders and its special utility for the infrared studies of hazardous materials. (Author)

Quantitative surface topography determination by Nomarski reflection microscopy. 2: Microscope modification, calibration, and planar sample experiments

International Nuclear Information System (INIS)

Hartman, J.S.; Gordon, R.L.; Lessor, D.L.

1980-01-01

The application of reflective Nomarski differential interference contrast microscopy for the determination of quantitative sample topography data is presented. The discussion includes a review of key theoretical results presented previously plus the experimental implementation of the concepts using a commercial Momarski microscope. The experimental work included the modification and characterization of a commercial microscope to allow its use for obtaining quantitative sample topography data. System usage for the measurement of slopes on flat planar samples is also discussed. The discussion has been designed to provide the theoretical basis, a physical insight, and a cookbook procedure for implementation to allow these results to be of value to both those interested in the microscope theory and its practical usage in the metallography laboratory

Combined reflectance confocal microscopy-optical coherence tomography for delineation of basal cell carcinoma margins: an ex vivo study

Science.gov (United States)

Iftimia, Nicusor; Peterson, Gary; Chang, Ernest W.; Maguluri, Gopi; Fox, William; Rajadhyaksha, Milind

2016-01-01

We present a combined reflectance confocal microscopy (RCM) and optical coherence tomography (OCT) approach, integrated within a single optical layout, for diagnosis of basal cell carcinomas (BCCs) and delineation of margins. While RCM imaging detects BCC presence (diagnoses) and its lateral spreading (margins) with measured resolution of ˜1 μm, OCT imaging delineates BCC depth spreading (margins) with resolution of ˜7 μm. When delineating margins in 20 specimens of superficial and nodular BCCs, depth could be reliably determined down to ˜600 μm, and agreement with histology was within about ±50 μm.

The cuticle modulates ultraviolet reflectance of avian eggshells

Directory of Open Access Journals (Sweden)

Daphne C. Fecheyr-Lippens

2015-07-01

Full Text Available Avian eggshells are variedly coloured, yet only two pigments, biliverdin and protoporphyrin IX, are known to contribute to the dramatic diversity of their colours. By contrast, the contributions of structural or other chemical components of the eggshell are poorly understood. For example, unpigmented eggshells, which appear white to the human eye, vary in their ultraviolet (UV reflectance, which may be detectable by birds. We investigated the proximate mechanisms for the variation in UV-reflectance of unpigmented bird eggshells using spectrophotometry, electron microscopy, chemical analyses, and experimental manipulations. We specifically tested how UV-reflectance is affected by the eggshell cuticle, the outermost layer of most avian eggshells. The chemical dissolution of the outer eggshell layers, including the cuticle, increased UV-reflectance for only eggshells that contained a cuticle. Our findings demonstrate that the outer eggshell layers, including the cuticle, absorb UV-light, probably because they contain higher levels of organic components and other chemicals, such as calcium phosphates, compared to the predominantly calcite-based eggshell matrix. These data highlight the need to examine factors other than the known pigments in studies of avian eggshell colour.

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

Phase contrast scanning transmission electron microscopy imaging of light and heavy atoms at the limit of contrast and resolution.

Science.gov (United States)

Yücelen, Emrah; Lazić, Ivan; Bosch, Eric G T

2018-02-08

Using state of the art scanning transmission electron microscopy (STEM) it is nowadays possible to directly image single atomic columns at sub-Å resolution. In standard (high angle) annular dark field STEM ((HA)ADF-STEM), however, light elements are usually invisible when imaged together with heavier elements in one image. Here we demonstrate the capability of the recently introduced Integrated Differential Phase Contrast STEM (iDPC-STEM) technique to image both light and heavy atoms in a thin sample at sub-Å resolution. We use the technique to resolve both the Gallium and Nitrogen dumbbells in a GaN crystal in [[Formula: see text

Scanning Capacitance Microscopy | Materials Science | NREL

Science.gov (United States)

obtained using scanning capacitance microscopy. Top Right: Image of p-type and n-type material, obtained 'fingers' of light-colored n-type material on a yellow and blue background representing p-type material material, obtained using scanning capacitance microscopy, in a sample semiconductor device; the image shows

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.

Masked rhodamine dyes of five principal colors revealed by photolysis of a 2-diazo-1-indanone caging group: synthesis, photophysics, and light microscopy applications.

OpenAIRE

Belov, V.; Mitronova, G.; Bossi, M.; Boyarski, V.; Hebisch, E.; Geisler, C.; Kolmakov, K.; Wurm, C.; Willig, K.; Hell, S.

2014-01-01

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...

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

Despite nanoparticles being used in many different products and applications, the effects and fate in the environment are still not well understood. Uptake of nanoparticles into cells has been shown in vitro and in vivo. However, it is challenging to find suitable methods to identify uptake...... 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...... to the particles through the diet or the water phase in a series of separate experiments. In the dietary exposure experiments Artemia salina were exposed to 1 mg Au/L for 24h before being fed to D. rerio. For exposure through the water phase 1 mg Au/L was added directly to aquaria holding the fish and non...

Reflectance measurements of PTFE, Kapton, and PEEK for xenon scintillation light for the LZ detector.

Science.gov (United States)

Arthurs, M.; Batista, E.; Haefner, J.; Lorenzon, W.; Morton, D.; Neff, A.; Okunawo, M.; Pushkin, K.; Sander, A.; Stephenson, S.; Wang, Y.; LZ Collaboration

2017-01-01

LZ (LUX-Zeplin) is an international collaboration that will look for dark matter candidates, WIMPs (Weakly Interacting Massive Particles), through direct detection by dual-phase time projection chamber (TPC) using liquid xenon. The LZ detector will be located nearly a mile underground at SURF, South Dakota, shielded from cosmic background radiation. Seven tons active mass of liquid xenon will be used for detecting the weak interaction of WIMPs with ordinary matter. Over three years of operation it is expected to reach the ultimate sensitivity of 2x10-48 cm2 for a WIMP mass of 50 GeV. As for many other rare event searches, high light collection efficiency is essential for LZ detector. Moreover, in order to achieve greater active volume for detection as well as reduce potential backgrounds, thinner detector walls without significant loss in reflectance are desired. Reflectance measurements of polytetrafluoroethylene (PTFE), Kapton, and PEEK for xenon scintillation light (178 nm), conducted at the University of Michigan using the Michigan Xenon Detector (MiX) will be presented. The University of Michigan, LZ Collaboration, The US Department of Energy.

Frustrated total internal reflection in organic light-emitting diodes employing sphere cavity embedded in polystyrene

International Nuclear Information System (INIS)

Zhu, Peifen

2016-01-01

The light extraction efficiency of top-emitting organic light-emitting diodes (OLEDs) is numerically investigated employing the finite-difference time-domain method. The periodic nanostructures formed by embedding the sphere arrays in polystyrene (PS) are placed on top of OLED to frustrate the total internal reflection at the interface between OLED and free space. These nanostructures serve as an intermediate medium to extract the light out of OLED devices. Efficiently coupling both evanescent waves and propagation waves into spheres and subsequently extracting these light waves out of the sphere is key to achieving high extraction efficiency. By tuning the thickness of PS layer, both of the in-coupling efficiency and out-coupling efficiency are optimized for achieving high light extraction efficiency. Thicker PS layer results in higher in-coupling efficiency in sphere while the thinner PS layer leads to higher out-coupling efficiency. Thus the maximum light extraction is a trade-off between the in-coupling efficiency and out-coupling efficiency. The study shows that light extraction efficiency of 89% can be achieved by embedding 0.90 μm TiO 2 sphere in 0.30 μm PS layer with optimized in-coupling efficiency, out-coupling efficiency and cavity effect. (paper)

Comprehensive optical and data management infrastructure for high-throughput light-sheet microscopy of whole mouse brains.

Science.gov (United States)

Müllenbroich, M Caroline; Silvestri, Ludovico; Onofri, Leonardo; Costantini, Irene; Hoff, Marcel Van't; Sacconi, Leonardo; Iannello, Giulio; Pavone, Francesco S

2015-10-01

Comprehensive mapping and quantification of neuronal projections in the central nervous system requires high-throughput imaging of large volumes with microscopic resolution. To this end, we have developed a confocal light-sheet microscope that has been optimized for three-dimensional (3-D) imaging of structurally intact clarified whole-mount mouse brains. We describe the optical and electromechanical arrangement of the microscope and give details on the organization of the microscope management software. The software orchestrates all components of the microscope, coordinates critical timing and synchronization, and has been written in a versatile and modular structure using the LabVIEW language. It can easily be adapted and integrated to other microscope systems and has been made freely available to the light-sheet community. The tremendous amount of data routinely generated by light-sheet microscopy further requires novel strategies for data handling and storage. To complete the full imaging pipeline of our high-throughput microscope, we further elaborate on big data management from streaming of raw images up to stitching of 3-D datasets. The mesoscale neuroanatomy imaged at micron-scale resolution in those datasets allows characterization and quantification of neuronal projections in unsectioned mouse brains.

Light reflection from crystal platelets in iridophores determines green or brown skin coloration in Takydromus lizards.

Science.gov (United States)

Kuriyama, Takeo; Esashi, Jyunko; Hasegawa, Masami

2017-04-01

Brown and green are the most commonly imitated colors in prey animals because both colors occur in a range of habitats. Many researchers have evaluated survival with respect to background color matching, but the pigment cell mechanisms underlying such coloration are not known. Dorsal coloration of East Asian Takydromus lizards has shifted from green to brown or from brown to green on multiple occasions during the diversification of the genus, thus giving us an opportunity to examine the cellular mechanisms of background color matching. Brown and green skin were found to differ with respect to the morphological characteristics of iridophores, with different thicknesses of the reflecting platelets and the cytoplasmic spacing between platelets, despite a shared vertical arrangement of pigment cells, i.e., xanthophores in the upper layer, iridophores in the middle layer, and melanophores at the bottom of the dermal layer, among the different Takydromus lizards. Iridophores of brown skin reflected longer wavelengths of light than those of green skin, which may be attributed to the thicker platelets and longer distances between platelets in brown skin. We discuss the potential role of genetic and intracellular mechanisms explaining the thickness and orientation of the light-reflecting platelets of iridophores in Takydromus lizards. Copyright © 2016 Elsevier GmbH. All rights reserved.

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.

Sol-Gel-Hydrothermal Synthesis of the Heterostructured TiO2/N-Bi2WO6 Composite with High-Visible-Light- and Ultraviolet-Light-Induced Photocatalytic Performances

Directory of Open Access Journals (Sweden)

Jiang Zhang

2012-01-01

Full Text Available The heterostructured TiO2/N-Bi2WO6 composites were prepared by a facile sol-gel-hydrothermal method. The phase structures, morphologies, and optical properties of the samples were characterized by using X-ray powder diffraction (XRD, scanning electron microscopy (SEM, high-resolution transmission electron microscopy (HRTEM, energy dispersive spectroscopy (EDS, and UV-vis diffuse reflectance spectroscopy. The photocatalytic activities for rhodamine B of the as-prepared products were measured under visible and ultraviolet light irradiation at room temperature. The TiO2/N-Bi2WO6 composites exhibited much higher photocatalytic performances than TiO2 as well as Bi2WO6. The enhancement in the visible light photocatalytic performance of the TiO2/N-Bi2WO6 composites could be attributed to the effective electron-hole separations at the interfaces of the two semiconductors, which facilitate the transfer of the photoinduced carriers.

Nonspecular reflection of light at an inhomogeneous interface between two media and in a nanostructured layer with a quasi-zero refractive index

International Nuclear Information System (INIS)

Gadomsky, O. N.; Gadomskaya, I. V.

2015-01-01

We have derived formulas for the amplitudes of light reflection and refraction at an inhomogeneous interface between two media and in a nanostructured layer with a quasi-zero refractive index. These formulas are applied to explain the experimental spectra of nonspecular light reflection using a nanostructured (PMMA + Ag) layer with silver nanoparticles on a silicon surface as an example. We show that a surface wave is formed in the nanostructured layer at various angles of light incidence and the layer with a quasi-zero refractive index is an antireflection coating that provides uniform 5% silicon antireflection in the wavelength range from 450 to 1000 nm

Progress in reflectance confocal microscopy for imaging oral tissues in vivo

Science.gov (United States)

Peterson, Gary; Zanoni, Daniella K.; Migliacci, Jocelyn; Cordova, Miguel; Rajadhyaksha, Milind; Patel, Snehal

2016-02-01

We report progress in development and feasibility testing of reflectance confocal microscopy (RCM) for imaging in the oral cavity of humans. We adapted a small rigid relay telescope (120mm long x 14mm diameter) and a small water immersion objective lens (12mm diameter, NA 0.7) to a commercial handheld RCM scanner (Vivascope 3000, Caliber ID, Rochester NY). This scanner is designed for imaging skin but we adapted the front end (the objective lens and the stepper motor that axially translates) for intra-oral use. This adaption required a new approach to address the loss of the automated stepper motor for acquisition of images in depth. A helical spring-like cap (with a coverslip to contact tissue) was designed for approximately 150 um of travel. Additionally other methods for focusing optics were designed and evaluated. The relay telescope optics is being tested in a clinical setting. With the capture of video and "video-mosaicing", extended areas can be imaged. The feasibility of imaging oral tissues was initially investigated in volunteers. RCM imaging in buccal mucosa in vivo shows nuclear and cellular detail in the epithelium and epithelial junction, and connective tissue and blood flow in the underlying lamina propria. Similar detail, including filiform and fungiform papillae, can be seen on the tongue in vivo. Clinical testing during head and neck surgery is now in progress and patients are being imaged for both normal tissue and cancerous margins in lip and tongue mucosa.

An Ingenious Super Light Trapping Surface Templated from Butterfly Wing Scales

Science.gov (United States)

Han, Zhiwu; Li, Bo; Mu, Zhengzhi; Yang, Meng; Niu, Shichao; Zhang, Junqiu; Ren, Luquan

2015-08-01

Based on the super light trapping property of butterfly Trogonoptera brookiana wings, the SiO2 replica of this bionic functional surface was successfully synthesized using a simple and highly effective synthesis method combining a sol-gel process and subsequent selective etching. Firstly, the reflectivity of butterfly wing scales was carefully examined. It was found that the whole reflectance spectroscopy of the butterfly wings showed a lower level (less than 10 %) in the visible spectrum. Thus, it was confirmed that the butterfly wings possessed a super light trapping effect. Afterwards, the morphologies and detailed architectures of the butterfly wing scales were carefully investigated using the ultra-depth three-dimensional (3D) microscope and field emission scanning electronic microscopy (FESEM). It was composed by the parallel ridges and quasi-honeycomb-like structure between them. Based on the biological properties and function above, an exact SiO2 negative replica was fabricated through a synthesis method combining a sol-gel process and subsequent selective etching. At last, the comparative analysis of morphology feature size and the reflectance spectroscopy between the SiO2 negative replica and the flat plate was conducted. It could be concluded that the SiO2 negative replica inherited not only the original super light trapping architectures, but also the super light trapping characteristics of bio-template. This work may open up an avenue for the design and fabrication of super light trapping materials and encourage people to look for more super light trapping architectures in nature.

A Comparative Study of Sample Preparation for Staining and Immunodetection of Plant Cell Walls by Light Microscopy

Science.gov (United States)

Verhertbruggen, Yves; Walker, Jesse L.; Guillon, Fabienne; Scheller, Henrik V.

2017-01-01

Staining and immunodetection by light microscopy are methods widely used to investigate plant cell walls. The two techniques have been crucial to study the cell wall architecture in planta, its deconstruction by chemicals or cell wall-degrading enzymes. They have been instrumental in detecting the presence of cell types, in deciphering plant cell wall evolution and in characterizing plant mutants and transformants. The success of immunolabeling relies on how plant materials are embedded and sectioned. Agarose coating, wax and resin embedding are, respectively, associated with vibratome, microtome and ultramicrotome sectioning. Here, we have systematically carried out a comparative analysis of these three methods of sample preparation when they are applied for cell wall staining and cell wall immunomicroscopy. In order to help the plant community in understanding and selecting adequate methods of embedding and sectioning for cell wall immunodetection, we review in this article the advantages and limitations of these three methods. Moreover, we offer detailed protocols of embedding for studying plant materials through microscopy. PMID:28900439

Nanocrystals of [Cu3(btc)2] (HKUST-1): a combined time-resolved light scattering and scanning electron microscopy study.

Science.gov (United States)

Zacher, Denise; Liu, Jianing; Huber, Klaus; Fischer, Roland A

2009-03-07

The formation of [Cu(3)(btc)(2)] (HKUST-1; btc = 1,3,5-benzenetricarboxylate) nanocrystals from a super-saturated mother solution at room temperature was monitored by time-resolved light scattering (TLS); the system is characterized by a rapid growth up to a size limit of 200 nm within a few minutes, and the size and shape of the crystallites were also determined by scanning electron microscopy (SEM).

Structural and biochemical alterations of human diabetic dermis studied by 3H-lysine incorporation and microscopy

International Nuclear Information System (INIS)

Moczar, M.; Allard, R.; Ouzilou, J.; Robert, L.; Pieraggi, M.-T.; Bouissou, H.; Julian, M.

1976-01-01

The alteration of the structural organization of dermal connective tissue was studied by light and electron microscopy and by biochemical techniques in normal human and in diabetic patients using skin biopsies. Part of the tissue was used for light and electron microscopy, the rest was incubated in the presence of 3 H-lysine for four hours. The 3 H-lysine labelled biopsies were submitted to a sequential extraction procedure in order to obtain representative macromolecular fractions containing the matrix macromolecules. The extracts were analyzed for their chemical composition and radioactivity. Electron microscopy revealed microstructural modifications of the fibroblasts, of the collagen and elastic fibers in the diabetic dermis. The incorporation pattern of 3 H-lysine into the macromolecular fractions was different in the normal and diabetic skin biopsies. The percentage of total radioactivity incorporated increased significantly in the 1M CaCl 2 extractable fraction and in the 6M urea extractable fraction and decreased significantly in the collagenase and elastase extracts in diabetic skin biopsy. These results demonstrate the existence of morphological and biochemical alterations in diabetic connective tissue (dermis) reflecting alterations in the relative rates of synthesis and/or degradation of the intercellular matrix macromolecules as well as of their microarchitectural arrangement

Magnetic force microscopy : Quantitative issues in biomaterials

NARCIS (Netherlands)

Passeri, D.; Dong, C.; Reggente, M.; Angeloni, L.; Barteri, M.; Scaramuzzo, F.A.; De Angelis, F.; Marinelli, F.; Antonelli, F.; Rinaldi, F.; Marianecci, C.; Carafa, M.; Sorbo, A.; Sordi, D.; Arends, I.W.C.E.; Rossi, M.

2014-01-01

Magnetic force microscopy (MFM) is an atomic force microscopy (AFM) based technique in which an AFM tip with a magnetic coating is used to probe local magnetic fields with the typical AFM spatial resolution, thus allowing one to acquire images reflecting the local magnetic properties of the samples

Anti-reflective coating for visible light using a silver nanodisc metasurface with a refractive index of less than 1.0

Science.gov (United States)

Yasuda, Hideki; Matsuno, Ryo; Koito, Naoki; Hosoda, Hidemasa; Tani, Takeharu; Naya, Masayuki

2017-12-01

Suppression of visible-light reflection from material surfaces is an important technology for many applications such as flat-panel displays, camera lenses, and solar panels. In this study, we developed an anti-reflective coating design based on a silver nanodisc metasurface. The effective refractive index of a 10-nm-thick monolayer of silver nanodiscs was less than 1.0, which enabled strong suppression of reflection from the underlying substrate. The nanodisc structure was easy to fabricate using a conventional roll-to-roll wet-coating method. The anti-reflective structure was fabricated over a large area.

Phase resolved and coherence gated en face reflection imaging of multilayered embryonal carcinoma cells

Science.gov (United States)

Yamauchi, Toyohiko; Fukami, Tadashi; Iwai, Hidenao; Yamashita, Yutaka

2012-03-01

Embryonal carcinoma (EC) cells, which are cell lines derived from teratocarcinomas, have characteristics in common with stem cells and differentiate into many kinds of functional cells. Similar to embryonic stem (ES) cells, undifferentiated EC cells form multi-layered spheroids. In order to visualize the three-dimensional structure of multilayered EC cells without labeling, we employed full-field interference microscopy with the aid of a low-coherence quantitative phase microscope, which is a reflection-type interference microscope employing the digital holographic technique with a low-coherent light source. Owing to the low-coherency of the light-source (halogen lamp), only the light reflected from reflective surface at a specific sectioning height generates an interference image on the CCD camera. P19CL6 EC cells, derived from mouse teratocarcinomas, formed spheroids that are about 50 to 200 micrometers in diameter. Since the height of each cell is around 10 micrometers, it is assumed that each spheroid has 5 to 20 cell layers. The P19CL6 spheroids were imaged in an upright configuration and the horizontally sectioned reflection images of the sample were obtained by sequentially and vertically scanning the zero-path-length height. Our results show the threedimensional structure of the spheroids, in which plasma and nuclear membranes were distinguishably imaged. The results imply that our technique is further capable of imaging induced pluripotent stem (iPS) cells for the assessment of cell properties including their pluripotency.

Confocal Raman Microscopy

CERN Document Server

Dieing, Thomas; Toporski, Jan

2011-01-01

Confocal Raman Microscopy is a relatively new technique that allows chemical imaging without specific sample preparation. By integrating a sensitive Raman spectrometer within a state-of-the-art microscope, Raman microscopy with a spatial resolution down to 200nm laterally and 500nm vertically can be achieved using visible light excitation. Recent developments in detector and computer technology as well as optimized instrument design have reduced integration times of Raman spectra by orders of magnitude, so that complete images consisting of tens of thousands of Raman spectra can be acquired in seconds or minutes rather than hours, which used to be standard just one decade ago. The purpose of this book is to provide the reader a comprehensive overview of the rapidly developing field of Confocal Raman Microscopy and its applications.

Searching for reflected light from τ Bootis b with high-resolution ground-based spectroscopy: Approaching the 10-5 contrast barrier

Science.gov (United States)

Hoeijmakers, H. J.; Snellen, I. A. G.; van Terwisga, S. E.

2018-02-01

Context. It is challenging to measure the starlight reflected from exoplanets because of the extreme contrast with their host stars. For hot Jupiters, this contrast is in the range of 10-6 to 10-4, depending on their albedo, radius and orbital distance. Searches for reflected light have been performed since the first hot Jupiters were discovered, but with very limited success because hot Jupiters tend to have low albedo values due to the general absence of reflective cloud decks. Aim. The aim of this study is to search for reflected light from τ Boo b, a hot Jupiter with one of the brightest host stars. Since its discovery in 1997, it has been the subject of several reflected-light searches using high-dispersion spectroscopy. Here we aim to combine these data in to a single meta-analysis. Methods: We analysed more than 2000 archival high-dispersion spectra obtained with the UVES, ESPaDOnS, NARVAL UES and HARPS-N spectrographs during various epochs between 1998 and 2013. Each spectrum was first cleaned of the stellar spectrum and subsequently cross-correlated with a PHOENIX model spectrum. These were then Doppler shifted to the planet rest-frame and co-added in time, weighted according to the expected signal-to-noise of the planet signal. Results: We reach a 3σ upper limit of the planet-to-star contrast of 1.5 × 10-5. Assuming a planet radius of 1.15 RJ, this corresponds to an optical albedo of between 400-700 nm. A low albedo is in line with secondary eclipse and phase curve observations of other hot Jupiters using space-based observatories, as well as theoretical predictions of their reflective properties.

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.

Light mirror reflection combined with heating/cooling curves as a method of studying phase transitions in transparent and opaque petroleum products: Apparatus and theory

International Nuclear Information System (INIS)

Shishkin, Yu.L.

2007-01-01

A portable low weight low cost apparatus 'Phasafot' and method for determining pour and cloud points of petroleum products, as well as precipitation and melting temperatures of paraffins in both transparent (diesel fuels), semi-transparent (lube oils) and opaque (crude oils) samples are described. The method consists in illuminating the surface of a sample with an oblique light beam and registering the intensity of specularly reflected light while heating/cooling the sample in the temperature range of its structural transitions. The mirror reflection of a light beam from an ideally smooth liquid surface falls in intensity when the surface becomes rough (dim) due to crystal formation. Simultaneous recording of the temperature ramp curve and the mirror reflection curve enables the determination of the beginning and end of crystallization of paraffins in both transparent and opaque petroleum products. Besides, their rheological properties can be accurately determined by rocking or tilting the instrument while monitoring the sample movement via its mirror reflection

Extending Whole Slide Imaging: Color Darkfield Internal Reflection Illumination (DIRI for Biological Applications.

Directory of Open Access Journals (Sweden)

Yoshihiro Kawano

Full Text Available Whole slide imaging (WSI is a useful tool for multi-modal imaging, and in our work, we have often combined WSI with darkfield microscopy. However, traditional darkfield microscopy cannot use a single condenser to support high- and low-numerical-aperture objectives, which limits the modality of WSI. To overcome this limitation, we previously developed a darkfield internal reflection illumination (DIRI microscope using white light-emitting diodes (LEDs. Although the developed DIRI is useful for biological applications, substantial problems remain to be resolved. In this study, we propose a novel illumination technique called color DIRI. The use of three-color LEDs dramatically improves the capability of the system, such that color DIRI (1 enables optimization of the illumination color; (2 can be combined with an oil objective lens; (3 can produce fluorescence excitation illumination; (4 can adjust the wavelength of light to avoid cell damage or reactions; and (5 can be used as a photostimulator. These results clearly illustrate that the proposed color DIRI can significantly extend WSI modalities for biological applications.

Super-resolution optical microscopy for studying membrane structure and dynamics.

Science.gov (United States)

Sezgin, Erdinc

2017-07-12

Investigation of cell membrane structure and dynamics requires high spatial and temporal resolution. The spatial resolution of conventional light microscopy is limited due to the diffraction of light. However, recent developments in microscopy enabled us to access the nano-scale regime spatially, thus to elucidate the nanoscopic structures in the cellular membranes. In this review, we will explain the resolution limit, address the working principles of the most commonly used super-resolution microscopy techniques and summarise their recent applications in the biomembrane field.

Virtual Microscopy in Histopathology Training: Changing Student Attitudes in 3 Successive Academic Years.

Science.gov (United States)

Bertram, Christof A; Firsching, Theresa; Klopfleisch, Robert

2018-01-01

Several veterinary faculties have integrated virtual microscopy into their curricula in recent years to improve and refine their teaching techniques. The many advantages of this recent technology are described in the literature, including remote access and an equal and constant slide quality for all students. However, no study has analyzed the change of perception toward virtual microscopy at different time points of students' academic educations. In the present study, veterinary students in 3 academic years were asked for their perspectives and attitudes toward virtual microscopy and conventional light microscopy. Third-, fourth-, and fifth-year veterinary students filled out a questionnaire with 12 questions. The answers revealed that virtual microscopy was overall well accepted by students of all academic years. Most students even suggested that virtual microscopy be implemented more extensively as the modality for final histopathology examinations. Nevertheless, training in the use of light microscopy and associated skills was surprisingly well appreciated. Regardless of their academic year, most students considered these skills important and necessary, and they felt that light microscopy should not be completely replaced. The reasons for this view differed depending on academic year, as the perceived main disadvantage of virtual microscopy varied. Third-year students feared that they would not acquire sufficient light microscopy skills. Fifth-year students considered technical difficulties (i.e., insufficient transmission speed) to be the main disadvantage of this newer teaching modality.

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

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...... 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...

Quantification of protein based on single-molecule counting by total internal reflection fluorescence microscopy with adsorption equilibrium

International Nuclear Information System (INIS)

Wang Lei; Xu Guang; Shi Zhikun; Jiang Wei; Jin Wenrui

2007-01-01

We developed a sensitive single-molecule imaging method for quantification of protein by total internal reflection fluorescence microscopy with adsorption equilibrium. In this method, the adsorption equilibrium of protein was achieved between solution and glass substrate. Then, fluorescence images of protein molecules in a evanescent wave field were taken by a highly sensitive electron multiplying charge coupled device. Finally, the number of fluorescent spots corresponding to the protein molecules in the images was counted. Alexa Fluor 488-labeled goat anti-rat IgG(H + L) was chosen as the model protein. The spot number showed an excellent linear relationship with protein concentration. The concentration linear range was 5.4 x 10 -11 to 8.1 x 10 -10 mol L -1

Ultra-compact silicon nitride grating coupler for microscopy systems

OpenAIRE

Zhu, Yunpeng; Wang, Jie; Xie, Weiqiang; Tian, Bin; Li, Yanlu; Brainis, Edouard; Jiao, Yuqing; Van Thourhout, Dries

2017-01-01

Grating couplers have been widely used for coupling light between photonic chips and optical fibers. For various quantum-optics and bio-optics experiments, on the other hand, there is a need to achieve good light coupling between photonic chips and microscopy systems. Here, we propose an ultra-compact silicon nitride (SiN) grating coupler optimized for coupling light from a waveguide to a microscopy system. The grating coupler is about 4 by 2 mu m(2) in size and a 116 nm 1 dB bandwidth can be...

A simple procedure to analyze positions of interest in infectious cell cultures by correlative light and electron microscopy.

Science.gov (United States)

Madela, Kazimierz; Banhart, Sebastian; Zimmermann, Anja; Piesker, Janett; Bannert, Norbert; Laue, Michael

2014-01-01

Plastic cell culture dishes that contain a thin bottom of highest optical quality including an imprinted finder grid (μ-Dish Grid-500) are optimally suited for routine correlative light and electron microscopy using chemical fixation. Such dishes allow high-resolution fluorescence and bright-field imaging using fixed and living cells and are compatible with standard protocols for scanning and transmission electron microscopy. Ease of use during cell culture and imaging, as well as a tight cover render the dishes particularly suitable for working with infectious organisms up to the highest biosafety level. Detailed protocols are provided and demonstrated by showing two examples: monitoring the production of virus-like particles of the Human Endogenous Retrovirus HERV-K(HML-2) by HeLa cells and investigation of Rab11-positive membrane-compartments of HeLa cells after infection with Chlamydia trachomatis. © 2014 Elsevier Inc. All rights reserved.

Simultaneous differential spinning disk fluorescence optical sectioning microscopy and nanomechanical mapping atomic force microscopy

International Nuclear Information System (INIS)

Miranda, Adelaide; De Beule, Pieter A. A.; Martins, Marco

2015-01-01

Combined microscopy techniques offer the life science research community a powerful tool to investigate complex biological systems and their interactions. Here, we present a new combined microscopy platform based on fluorescence optical sectioning microscopy through aperture correlation microscopy with a Differential Spinning Disk (DSD) and nanomechanical mapping with an Atomic Force Microscope (AFM). The illumination scheme of the DSD microscope unit, contrary to standard single or multi-point confocal microscopes, provides a time-independent illumination of the AFM cantilever. This enables a distortion-free simultaneous operation of fluorescence optical sectioning microscopy and atomic force microscopy with standard probes. In this context, we discuss sample heating due to AFM cantilever illumination with fluorescence excitation light. Integration of a DSD fluorescence optical sectioning unit with an AFM platform requires mitigation of mechanical noise transfer of the spinning disk. We identify and present two solutions to almost annul this noise in the AFM measurement process. The new combined microscopy platform is applied to the characterization of a DOPC/DOPS (4:1) lipid structures labelled with a lipophilic cationic indocarbocyanine dye deposited on a mica substrate

Simultaneous differential spinning disk fluorescence optical sectioning microscopy and nanomechanical mapping atomic force microscopy

Energy Technology Data Exchange (ETDEWEB)

Miranda, Adelaide; De Beule, Pieter A. A., E-mail: pieter.de-beule@inl.int [Applied Nano-Optics Laboratory, International Iberian Nanotechnology Laboratory, Avenida Mestre José Veiga, s/n, 4715-330 Braga (Portugal); Martins, Marco [Nano-ICs Group, International Iberian Nanotechnology Laboratory, Avenida Mestre José Veiga, s/n, 4715-330 Braga (Portugal)

2015-09-15

Combined microscopy techniques offer the life science research community a powerful tool to investigate complex biological systems and their interactions. Here, we present a new combined microscopy platform based on fluorescence optical sectioning microscopy through aperture correlation microscopy with a Differential Spinning Disk (DSD) and nanomechanical mapping with an Atomic Force Microscope (AFM). The illumination scheme of the DSD microscope unit, contrary to standard single or multi-point confocal microscopes, provides a time-independent illumination of the AFM cantilever. This enables a distortion-free simultaneous operation of fluorescence optical sectioning microscopy and atomic force microscopy with standard probes. In this context, we discuss sample heating due to AFM cantilever illumination with fluorescence excitation light. Integration of a DSD fluorescence optical sectioning unit with an AFM platform requires mitigation of mechanical noise transfer of the spinning disk. We identify and present two solutions to almost annul this noise in the AFM measurement process. The new combined microscopy platform is applied to the characterization of a DOPC/DOPS (4:1) lipid structures labelled with a lipophilic cationic indocarbocyanine dye deposited on a mica substrate.

Examining Thermally Sprayed Coats By Fluorescence Microscopy

Science.gov (United States)

Street, Kenneth W., Jr.; Leonhardt, Todd A.

1994-01-01

True flaws distinquished from those induced by preparation of specimens. Fluorescence microscopy reveals debonding, porosity, cracks, and other flaws in specimens of thermally sprayed coating materials. Specimen illuminated, and dye it contains fluoresces, emitting light at different wavelength. Filters emphasize contrast between excitation light and emission light. Specimen viewed directly or photographed on color film.

In situ atomic force microscopy studies of reversible light-induced switching of surface roughness and adhesion in azobenzene-containing PMMA films

International Nuclear Information System (INIS)

Mueller, M.; Gonzalez-Garcia, Y.; Pakula, C.; Zaporojtchenko, V.; Strunskus, T.; Faupel, F.; Herges, R.; Zargarani, D.; Magnussen, O.M.

2011-01-01

Thin films in the range 40-80 nm of a blend of PMMA with an azobenzene derivative have been studied directly during UV and blue light irradiation by atomic force microscopy (AFM), revealing highly reversible changes in the surface roughness and the film adhesion. UV light induces an ∼80% increase in surface roughness, whereas illumination by blue light completely reverses these changes. Based on the observed surface topography and transition kinetics a reversible mass flow mechanisms is suggested, where the polarity changes upon switching trigger a wetting-dewetting transition in a surface segregation layer of the chromophore. Similar AFM measurements of the pull-off force indicate a decrease upon UV and an increase after blue light illumination with a complex kinetic behavior: a rapid initial change, attributed to the change in the cis isomer fraction of the azobenzene derivative, and a more gradual change, indicative of slow structural reorganization.

Spatiotemporal closure of fractional laser-ablated channels imaged by optical coherence tomography and reflectance confocal microscopy

DEFF Research Database (Denmark)

Banzhaf, Christina A.; Wind, Bas S.; Mogensen, Mette

2016-01-01

Background and Objective Optical coherence tomography (OCT) and reflectance confocal microscopy (RCM) offer high-resolution optical imaging of the skin, which may provide benefit in the context of laser-assisted drug delivery. We aimed to characterize postoperative healing of ablative fractional...... laser (AFXL)-induced channels and dynamics in their spatiotemporal closure using in vivo OCT and RCM techniques. Study design/Materials and Methods The inner forearm of healthy subjects (n = 6) was exposed to 10,600 nm fractional CO2 laser using 5 and 25% densities, 120 μm beam diameter, 5, 15, and 25 m......J/microbeam. Treatment sites were scanned with OCT to evaluate closure of AFXL-channels and RCM to evaluate subsequent re-epithelialization. Results OCT and RCM identified laser channels in epidermis and upper dermis as black, ablated tissue defects surrounded by characteristic hyper-and hyporeflective zones. OCT imaged...

Bidirectional reflectance distribution function /BRDF/ measurements of stray light suppression coatings for the Space Telescope /ST/

Science.gov (United States)

Griner, D. B.

1979-01-01

The paper considers the bidirectional reflectance distribution function (BRDF) of black coatings used on stray light suppression systems for the Space Telescope (ST). The ST stray light suppression requirement is to reduce earth, moon, and sun light in the focal plane to a level equivalent to one 23 Mv star per square arcsecond, an attenuation of 14 orders of magnitude. It is impractical to verify the performance of a proposed baffle system design by full scale tests because of the large size of the ST, so that a computer analysis is used to select the design. Accurate computer analysis requires a knowledge of the diffuse scatter at all angles from the surface of the coatings, for all angles of incident light. During the early phases of the ST program a BRDF scanner was built at the Marshall Space Flight Center to study the scatter from black materials; the measurement system is described and the results of measurements on samples proposed for use on the ST are presented.

Identification of light elements in silicon nitride by aberration-corrected scanning transmission electron microscopy

Energy Technology Data Exchange (ETDEWEB)

Idrobo, Juan C., E-mail: idrobojc@ornl.gov [Materials Science and Technology Division, Oak Ridge National Laboratory, Oak Ridge, TN 37831 (United States); Walkosz, Weronika [Materials Science Division, Argonne National Laboratory, Argonne, IL 60439 (United States); Department of Physics, University of Illinois at Chicago, Chicago, IL 60607 (United States); Klie, Robert F.; Oeguet, Serdar [Department of Physics, University of Illinois at Chicago, Chicago, IL 60607 (United States)

2012-12-15

In silicon nitride structural ceramics, the overall mechanical and thermal properties are controlled by the atomic and electronic structures at the interface between the ceramic grains and the amorphous intergranular films (IGFs) formed by various sintering additives. In the last ten years the atomic arrangements of heavy elements (rare-earths) at the Si{sub 3}N{sub 4}/IGF interfaces have been resolved. However, the atomic position of light elements, without which it is not possible to obtain a complete description of the interfaces, has been lacking. This review article details the authors' efforts to identify the atomic arrangement of light elements such as nitrogen and oxygen at the Si{sub 3}N{sub 4}/SiO{sub 2} interface and in bulk Si{sub 3}N{sub 4} using aberration-corrected scanning transmission electron microscopy. -- Highlights: Black-Right-Pointing-Pointer Revealing the atomic structure of the {alpha}-Si{sub 3}N{sub 4}/SiO{sub 2} interface. Black-Right-Pointing-Pointer Identification and lattice location of oxygen impurities in bulk {alpha}-Si{sub 3}N{sub 4}. Black-Right-Pointing-Pointer Short range ordering of nitrogen and oxygen at the {beta}-Si{sub 3}N{sub 4}/SiO{sub 2} interface.

Using the combination refraction-reflection solid to design omni-directional light source used in underwater wireless optical communication

Science.gov (United States)

Rao, Jionghui; Yao, Wenming; Wen, Linqiang

2015-10-01

Underwater wireless optical communication is a communication technology which uses laser as an information carrier and transmits data through water. Underwater wireless optical communication has some good features such as broader bandwidth, high transmission rate, better security, anti—interference performance. Therefore, it is promising to be widely used in the civil and military communication domains. It is also suitable for high-speed, short-range communication between underwater mobile vehicles. This paper presents a design approach of omni-directional light source used in underwater wireless optical communication, using TRACEPRO simulation tool to help design a combination solid composed of the lens, conical reflector and parabolic reflector, and using the modulated DPSS green laser in the transmitter module to output the laser beam in small divergence angles, after expanded by the combination refraction-reflection solid, the angle turns into a space divergence angle of 2π, achieving the omni-directional light source of hemisphere space, and test in the air and underwater, the result shows that the effect is fine. This paper analyzes the experimental test in the air and water, in order to make further improvement of the uniformity of light distribution, we optimize the reflector surface parameters of combination refraction-reflection solid and test in the air and water. The result shows that omni-directional light source used in underwater wireless optical communication optimized could achieve the uniformity of light distribution of underwater space divergence angle of 2π. Omni-directional light source used in underwater wireless optical communication designed in this paper has the characteristics of small size and uniformity of light distribution, it is suitable for application between UUVs, AUVs, Swimmer Delivery Vehicles (SDVs) and other underwater vehicle fleet, it realizes point-to-multipoint communications.

Synthesis of CdSe-TiO_2 Photocatalyst and Their Enhanced Photocatalytic Activities under UV and Visible Light

International Nuclear Information System (INIS)

Lim, Chang Sung; Chen, Ming Liang; Oh, Won Chun

2011-01-01

In this study, CdSe-TiO_2 photocatalyst were synthesized by a facile solvothermal method and characterized by X-ray diffraction (XRD), Scanning electron microscopy (SEM) with energy dispersive X-ray (EDX) analysis and UV-vis diffuse reflectance spectrophotometer. The photocatalytic activity was investigated by degrading methylene blue (MB) in aqueous solution under irradiation of UV light as well as visible light. The absorbance of degraded MB solution was determined by UV-vis spectrophotometer. The results revealed that the CdSe- TiO_2 photocatalyst exhibited much higher photocatalytic activity than TiO_2 both under irradiation of UV light as well as visible light

Advanced spinning disk-TIRF microscopy for faster imaging of the cell interior and the plasma membrane.

Science.gov (United States)

Zobiak, Bernd; Failla, Antonio Virgilio

2018-03-01

Understanding the cellular processes that occur between the cytosol and the plasma membrane is an important task for biological research. Till now, however, it was not possible to combine fast and high-resolution imaging of both the isolated plasma membrane and the surrounding intracellular volume. Here, we demonstrate the combination of fast high-resolution spinning disk (SD) and total internal reflection fluorescence (TIRF) microscopy for specific imaging of the plasma membrane. A customised SD-TIRF microscope was used with specific design of the light paths that allowed, for the first time, live SD-TIRF experiments at high acquisition rates. A series of experiments is shown to demonstrate the feasibility and performance of our setup. © 2017 The Authors. Journal of Microscopy published by JohnWiley & Sons Ltd on behalf of Royal Microscopical Society.

Light-emitting Si films formed by neutral cluster deposition in a thin O2 gas

International Nuclear Information System (INIS)

Honda, Y.; Takei, M.; Ohno, H.; Shida, S.; Goda, K.

2005-01-01

We have fabricated the light-emitting Si-rich and oxygen-rich amorphous SiO 2 (a-SiO 2 ) films using the neutral cluster deposition (NCD) method without and with oxygen gas admitted, respectively, and demonstrate for the first time that these films show a photoluminescent feature. The Si thin films were observed by atomic force microscopy and high-resolution transmission electron microscopy, and analyzed by means of X-ray photoelectron spectroscopy, photoluminescence (PL) and FTIR-attenuated total reflection measurements. All of the PL spectra show mountainous distribution with a peak around 620 nm. It is found that the increase in the oxygen termination in the a-SiO 2 films evidently makes the PL intensity increase. It is demonstrated that NCD technique is one of the hopeful methods to fabricate light-emitting Si thin films

Low-temperature relative reflectivity measurements of reflective and scintillating foils used in rare event searches

Science.gov (United States)

Langenkämper, A.; Ulrich, A.; Defay, X.; Feilitzsch, F. v.; Lanfranchi, J.-C.; Mondragón, E.; Münster, A.; Oppenheimer, C.; Potzel, W.; Roth, S.; Schönert, S.; Steiger, H.; Trinh Thi, H. H.; Wawoczny, S.; Willers, M.; Zöller, A.

2018-03-01

In this work we investigate the reflectivity of highly reflective multilayer polymer foils used in the CRESST experiment. The CRESST experiment searches directly for dark matter via operating scintillating CaWO4 crystals as targets for elastic dark matter-nucleon scattering. In order to suppress background events, the experiment employs the so-called phonon-light technique which is based on the simultaneous measurement of the heat signal in the main CaWO4 target crystal and of the emitted scintillation light with a separate cryogenic light detector. Both detectors are surrounded by a highly reflective and scintillating multilayer polymer foil to increase the light collection efficiency and to veto surface backgrounds. While this study is motivated by the CRESST experiment, the results are also relevant for other rare event searches using scintillating cryogenic bolometers in the field of the search of dark matter and neutrinoless double beta decay (0 νββ). In this work a dedicated experiment has been set up to determine the relative reflectivity at 300 K and 20 K of three multilayer foils ("VM2000", "VM2002", "Vikuiti") produced by the company 3M. The intensity of a light beam reflected off the foil is measured with a CCD camera. The ratio of the intensities at 300 K and 20 K corresponds to the relative reflectivity change. The measurements performed in this work show no variation of the reflectivity with temperature at a level of ∼1%.

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. Published by Oxford University Press on behalf of Entomological Society of America 2016. This work is written by US Government employees and is in the public domain in the US.

Light-reflection random-target method for measurement of the modulation transfer function of a digital video-camera

Czech Academy of Sciences Publication Activity Database

Pospíšil, Jaroslav; Jakubík, P.; Machala, L.

2005-01-01

Roč. 116, - (2005), s. 573-585 ISSN 0030-4026 Institutional research plan: CEZ:AV0Z10100522 Keywords : random-target measuring method * light-reflection white - noise target * digital video camera * modulation transfer function * power spectral density Subject RIV: BH - Optics, Masers, Lasers Impact factor: 0.395, year: 2005

The 2015 super-resolution microscopy roadmap

International Nuclear Information System (INIS)

Hell, Stefan W; Sahl, Steffen J; Bates, Mark; Jakobs, Stefan; Zhuang, Xiaowei; Heintzmann, Rainer; Booth, Martin J; Bewersdorf, Joerg; Shtengel, Gleb; Hess, Harald; Tinnefeld, Philip; Honigmann, Alf; Testa, Ilaria; Cognet, Laurent; Lounis, Brahim; Ewers, Helge; Davis, Simon J; Eggeling, Christian; Klenerman, David; Willig, Katrin I

2015-01-01

Far-field optical microscopy using focused light is an important tool in a number of scientific disciplines including chemical, (bio)physical and biomedical research, particularly with respect to the study of living cells and organisms. Unfortunately, the applicability of the optical microscope is limited, since the diffraction of light imposes limitations on the spatial resolution of the image. Consequently the details of, for example, cellular protein distributions, can be visualized only to a certain extent. Fortunately, recent years have witnessed the development of ‘super-resolution’ far-field optical microscopy (nanoscopy) techniques such as stimulated emission depletion (STED), ground state depletion (GSD), reversible saturated optical (fluorescence) transitions (RESOLFT), photoactivation localization microscopy (PALM), stochastic optical reconstruction microscopy (STORM), structured illumination microscopy (SIM) or saturated structured illumination microscopy (SSIM), all in one way or another addressing the problem of the limited spatial resolution of far-field optical microscopy. While SIM achieves a two-fold improvement in spatial resolution compared to conventional optical microscopy, STED, RESOLFT, PALM/STORM, or SSIM have all gone beyond, pushing the limits of optical image resolution to the nanometer scale. Consequently, all super-resolution techniques open new avenues of biomedical research. Because the field is so young, the potential capabilities of different super-resolution microscopy approaches have yet to be fully explored, and uncertainties remain when considering the best choice of methodology. Thus, even for experts, the road to the future is sometimes shrouded in mist. The super-resolution optical microscopy roadmap of Journal of Physics D: Applied Physics addresses this need for clarity. It provides guidance to the outstanding questions through a collection of short review articles from experts in the field, giving a thorough

Design of a dynamic biofilm imaging cell for white-light interferometric microscopy

Science.gov (United States)

Larimer, Curtis; Brann, Michelle; Suter, Jonathan D.; Addleman, R. Shane

2017-11-01

In microbiology research, there is a strong need for next-generation imaging and sensing instrumentation that will enable minimally invasive and label-free investigation of soft, hydrated structures, such as in bacterial biofilms. White-light interferometry (WLI) can provide high-resolution images of surface topology without the use of fluorescent labels but is not typically used to image biofilms because there is insufficient refractive index contrast to induce reflection from the biofilm's interface. The soft structure and water-like bulk properties of hydrated biofilms make them difficult to characterize in situ, especially in a nondestructive manner. We build on our prior description of static biofilm imaging and describe the design of a dynamic growth flow cell that enables monitoring of the thickness and topology of live biofilms over time using a WLI microscope. The microfluidic system is designed to grow biofilms in dynamic conditions and to create a reflective interface on the surface while minimizing disruption of fragile structures. The imaging cell was also designed to accommodate limitations imposed by the depth of focus of the microscope's objective lens. Example images of live biofilm samples are shown to illustrate the ability of the flow cell and WLI instrument to (1) support bacterial growth and biofilm development, (2) image biofilm structure that reflects growth in flow conditions, and (3) monitor biofilm development over time nondestructively. In future work, the apparatus described here will enable surface metrology measurements (roughness, surface area, etc.) of biofilms and may be used to observe changes in biofilm structure in response to changes in environmental conditions (e.g., flow velocity, availability of nutrients, and presence of biocides). This development will open opportunities for the use of WLI in bioimaging.

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...

How Can Polarization States of Reflected Light from Snow Surfaces Inform Us on Surface Normals and Ultimately Snow Grain Size Measurements?

Science.gov (United States)

Schneider, A. M.; Flanner, M.; Yang, P.; Yi, B.; Huang, X.; Feldman, D.

2016-12-01

The Snow Grain Size and Pollution (SGSP) algorithm is a method applied to Moderate Resolution Imaging Spectroradiometer data to estimate snow grain size from space-borne measurements. Previous studies validate and quantify potential sources of error in this method, but because it assumes flat snow surfaces, however, large scale variations in surface normals can cause biases in its estimates due to its dependence on solar and observation zenith angles. To address these variations, we apply the Monte Carlo method for photon transport using data containing the single scattering properties of different ice crystals to calculate polarization states of reflected monochromatic light at 1500nm from modeled snow surfaces. We evaluate the dependence of these polarization states on solar and observation geometry at 1500nm because multiple scattering is generally a mechanism for depolarization and the ice crystals are relatively absorptive at this wavelength. Using 1500nm thus results in a higher number of reflected photons undergoing fewer scattering events, increasing the likelihood of reflected light having higher degrees of polarization. In evaluating the validity of the model, we find agreement with previous studies pertaining to near-infrared spectral directional hemispherical reflectance (i.e. black-sky albedo) and similarities in measured bidirectional reflectance factors, but few studies exist modeling polarization states of reflected light from snow surfaces. Here, we present novel results pertaining to calculated polarization states and compare dependences on solar and observation geometry for different idealized snow surfaces. If these dependencies are consistent across different ice particle shapes and sizes, then these findings could inform the SGSP algorithm by providing useful relationships between measurable physical quantities and solar and observation geometry to better understand variations in snow surface normals from remote sensing observations.

Selective sensitivity in Kerr microscopy.

Science.gov (United States)

Soldatov, I V; Schäfer, R

2017-07-01

A new technique for contrast separation in wide-field magneto-optical Kerr microscopy is introduced. Utilizing the light from eight light emitting diodes, guided to the microscope by glass fibers and being switched synchronously with the camera exposure, domain images with orthogonal in-plane sensitivity can be displayed simultaneously at real-time, and images with pure in-plane or polar contrast can be obtained. The benefit of this new method of contrast separation is demonstrated for Permalloy films, a NdFeB sinter magnet, and a cobalt crystal. Moreover, the new technique is shown to strongly enhance the sensitivity of Kerr microscopy by eliminating parasitic contrast contributions occurring in conventional setups. A doubling of the in-plane domain contrast and a sensitivity to Kerr rotations as low as 0.6 mdeg is demonstrated.

Selective sensitivity in Kerr microscopy

Science.gov (United States)

Soldatov, I. V.; Schäfer, R.

2017-07-01

A new technique for contrast separation in wide-field magneto-optical Kerr microscopy is introduced. Utilizing the light from eight light emitting diodes, guided to the microscope by glass fibers and being switched synchronously with the camera exposure, domain images with orthogonal in-plane sensitivity can be displayed simultaneously at real-time, and images with pure in-plane or polar contrast can be obtained. The benefit of this new method of contrast separation is demonstrated for Permalloy films, a NdFeB sinter magnet, and a cobalt crystal. Moreover, the new technique is shown to strongly enhance the sensitivity of Kerr microscopy by eliminating parasitic contrast contributions occurring in conventional setups. A doubling of the in-plane domain contrast and a sensitivity to Kerr rotations as low as 0.6 mdeg is demonstrated.

A comparison of atomic force microscopy (AFM) and dynamic light scattering (DLS) methods to characterize nanoparticle size distributions

International Nuclear Information System (INIS)

Hoo, Christopher M.; Starostin, Natasha; West, Paul; Mecartney, Martha L.

2008-01-01

This paper compares the accuracy of conventional dynamic light scattering (DLS) and atomic force microscopy (AFM) for characterizing size distributions of polystyrene nanoparticles in the size range of 20-100 nm. Average DLS values for monosize dispersed particles are slightly higher than the nominal values whereas AFM values were slightly lower than nominal values. Bimodal distributions were easily identified with AFM, but DLS results were skewed toward larger particles. AFM characterization of nanoparticles using automated analysis software provides an accurate and rapid analysis for nanoparticle characterization and has advantages over DLS for non-monodispersed solutions.

Circumventing photodamage in live-cell microscopy

Science.gov (United States)

Magidson, Valentin; Khodjakov, Alexey

2013-01-01

Fluorescence microscopy has become an essential tool in cell biology. This technique allows researchers to visualize the dynamics of tissue, cells, individual organelles and macromolecular assemblies inside the cell. Unfortunately, fluorescence microscopy is not completely ‘non-invasive’ as the high-intensity excitation light required for excitation of fluorophores is inherently toxic for live cells. Physiological changes induced by excessive illumination can lead to artifacts and abnormal responses. In this chapter we review major factors that contribute to phototoxicity and discuss practical solutions for circumventing photodamage. These solutions include the proper choice of image acquisition parameters, optimization of filter sets, hardware synchronization, and the use of intelligent illumination to avoid unnecessary light exposure. PMID:23931522

UmUTracker: A versatile MATLAB program for automated particle tracking of 2D light microscopy or 3D digital holography data

Science.gov (United States)

Zhang, Hanqing; Stangner, Tim; Wiklund, Krister; Rodriguez, Alvaro; Andersson, Magnus

2017-10-01

We present a versatile and fast MATLAB program (UmUTracker) that automatically detects and tracks particles by analyzing video sequences acquired by either light microscopy or digital in-line holographic microscopy. Our program detects the 2D lateral positions of particles with an algorithm based on the isosceles triangle transform, and reconstructs their 3D axial positions by a fast implementation of the Rayleigh-Sommerfeld model using a radial intensity profile. To validate the accuracy and performance of our program, we first track the 2D position of polystyrene particles using bright field and digital holographic microscopy. Second, we determine the 3D particle position by analyzing synthetic and experimentally acquired holograms. Finally, to highlight the full program features, we profile the microfluidic flow in a 100 μm high flow chamber. This result agrees with computational fluid dynamic simulations. On a regular desktop computer UmUTracker can detect, analyze, and track multiple particles at 5 frames per second for a template size of 201 ×201 in a 1024 × 1024 image. To enhance usability and to make it easy to implement new functions we used object-oriented programming. UmUTracker is suitable for studies related to: particle dynamics, cell localization, colloids and microfluidic flow measurement. Program Files doi : http://dx.doi.org/10.17632/fkprs4s6xp.1 Licensing provisions : Creative Commons by 4.0 (CC by 4.0) Programming language : MATLAB Nature of problem: 3D multi-particle tracking is a common technique in physics, chemistry and biology. However, in terms of accuracy, reliable particle tracking is a challenging task since results depend on sample illumination, particle overlap, motion blur and noise from recording sensors. Additionally, the computational performance is also an issue if, for example, a computationally expensive process is executed, such as axial particle position reconstruction from digital holographic microscopy data. Versatile

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

Science.gov (United States)

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

2015-05-01

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

[In vivo reflectance confocal microscopy in dermatology: a proposal concerning French terminology].

Science.gov (United States)

Kanitakis, J; Bahadoran, P; Braun, R; Debarbieux, S; Labeille, B; Perrot, J-L; Vabres, P

2013-11-01

Reflectance confocal microscopy (RCM) is a recently introduced non-invasive imaging technique allowing real-time examination of the skin in vivo. Whereas a substantial literature concerning RCM exists in English, so far there is no official terminology in French, despite the fact that an ever-growing number of French-speaking dermatologists now use this new imaging technique. The aim of the present study is to propose a French terminology for RCM in order to allow French-speaking dermatologists to communicate in a precise and homogeneous language on this topic. A group of French-speaking dermatologists with solid experience of RCM, members of the Non-invasive Cutaneous Imaging group of the French Society of Dermatology, endeavored to suggest terms in French concerning RCM. Each group member dealt with a specific paragraph. The members exchanged comments via email and the terminology was finalized during a meeting of the group members in Paris in June 2012. Descriptive terms referring to the RCM aspects of normal and diseased skin were proposed. Some of these already existed, being used in routine dermatopathology, while other specific terms were created or adapted from the English terminology. This terminology will allow French-speaking dermatologists using RCM to communicate their findings in a homogeneous language. It may be enriched in the future by the introduction of additional terms describing new aspects of both normal and, especially, diseased skin. Copyright © 2013 Elsevier Masson SAS. All rights reserved.

Optical Reflectance Measurements for Commonly Used Reflectors

Science.gov (United States)

Janecek, Martin; Moses, William W.

2008-08-01

When simulating light collection in scintillators, modeling the angular distribution of optical light reflectance from surfaces is very important. Since light reflectance is poorly understood, either purely specular or purely diffuse reflectance is generally assumed. In this paper we measure the optical reflectance distribution for eleven commonly used reflectors. A 440 nm, output power stabilized, un-polarized laser is shone onto a reflector at a fixed angle of incidence. The reflected light's angular distribution is measured by an array of silicon photodiodes. The photodiodes are movable to cover 2pi of solid angle. The light-induced current is, through a multiplexer, read out with a digital multimeter. A LabVIEW program controls the motion of the laser and the photodiode array, the multiplexer, and the data collection. The laser can be positioned at any angle with a position accuracy of 10 arc minutes. Each photodiode subtends 6.3deg, and the photodiode array can be positioned at any angle with up to 10 arc minute angular resolution. The dynamic range for the current measurements is 10 5:1. The measured light reflectance distribution was measured to be specular for several ESR films as well as for aluminum foil, mostly diffuse for polytetrafluoroethylene (PTFE) tape and titanium dioxide paint, and neither specular nor diffuse for Lumirrorreg, Melinexreg and Tyvekreg. Instead, a more complicated light distribution was measured for these three materials.

Visible light induced electron transfer process over nitrogen doped TiO2 nanocrystals prepared by oxidation of titanium nitride

International Nuclear Information System (INIS)

Wu Zhongbiao; Dong Fan; Zhao Weirong; Guo Sen

2008-01-01

Nitrogen doped TiO 2 nanocrystals with anatase and rutile mixed phases were prepared by incomplete oxidation of titanium nitride at different temperatures. The as-prepared samples were characterized by X-ray diffraction (XRD), transmission electron microscopy (TEM), high resolution transmission electron microscopy (HRTEM), core level X-ray photoelectron spectroscopy (CL XPS), valence band X-ray photoelectron spectroscopy (VB XPS), UV-vis diffuse reflectance spectra (UV-vis DRS), and visible light excited photoluminescence (PL). The photocatalytic activity was evaluated for photocatalytic degradation of toluene in gas phase under visible light irradiation. The visible light absorption and photoactivities of these nitrogen doped TiO 2 nanocrystals can be clearly attributed to the change of the additional electronic (N - ) states above the valence band of TiO 2 modified by N dopant as revealed by the VB XPS and visible light induced PL. A band gap structure model was established to explain the electron transfer process over nitrogen doped TiO 2 nanocrystals under visible light irradiation, which was consistent with the previous theoretical and experimental results. This model can also be applied to understand visible light induced photocatalysis over other nonmetal doped TiO 2

Mechanisms of fine extinction band development in vein quartz: new insights from correlative light and electron microscopy

Science.gov (United States)

Derez, Tine; Van Der Donck, Tom; Plümper, Oliver; Muchez, Philippe; Pennock, Gill; Drury, Martyn R.; Sintubin, Manuel

2017-07-01

Fine extinction bands (FEBs) (also known as deformation lamellae) visible with polarized light microscopy in quartz consist of a range of nanostructures, inferring different formation processes. Previous transmission electron microscopy studies have shown that most FEB nanostructures in naturally deformed quartz are elongated subgrains formed by recovery of dislocation slip bands. Here we show that three types of FEB nanostructure occur in naturally deformed vein quartz from the low-grade metamorphic High-Ardenne slate belt (Belgium). Prismatic oriented FEBs are defined by bands of dislocation walls. Dauphiné twin boundaries present along the FEB boundaries probably formed after FEB formation. In an example of two sub-rhombohedral oriented FEBs, developed as two sets in one grain, the finer FEB set consists of elongated subgrains, similar to FEBs described in previous transmission electron microscopy studies. The second wider FEB set consists of bands with different dislocation density and fluid-inclusion content. The wider FEB set is interpreted as bands with different plastic strain associated with the primary growth banding of the vein quartz grain. The nanometre-scale fluid inclusions are interpreted to have formed from structurally bounded hydroxyl groups that moreover facilitated formation of the elongate subgrains. Larger fluid inclusions aligned along FEBs are explained by fluid-inclusion redistribution along dislocation cores. The prismatic FEB nanostructure and the relation between FEBs and growth bands have not been recognized before, although related structures have been reported in experimentally deformed quartz.

Determination of the synthesis site of the infections flacherie virus-RNA by light microscopy-autoradiography

International Nuclear Information System (INIS)

Almeida, I.M.G. de; Silva, D.M.

1981-01-01

The site of the RNA synthesis of the infectious flacherie virus in the midgut epithelial cells of the silkworm, Bombyx mori L., 1758 (Lep., Bombycidae), has been investigated using both autoradiography and light microscopy techniques. The density or ratio between silver grain and the respective cell structure (silver grain/μm 2 ) has been used as criteria to identify the site of the viral RNA synthesis. Actinomycin D selectively blocked about 60% of the cell RNA synthesis without affecting the virus RNA synthesis. The obtained data indicated that the viral RNA synthesis occurs in the nucleus of the midgut epithelial cells of the silkworm larvae. Some evidence about the viral RNA translocation from nucleus to cytoplasm and inhibition of the synthesis of normal RNA by the virus were observed. (Author) [pt

Chromatic confocal microscopy for multi-depth imaging of epithelial tissue

Science.gov (United States)

Olsovsky, Cory; Shelton, Ryan; Carrasco-Zevallos, Oscar; Applegate, Brian E.; Maitland, Kristen C.

2013-01-01

We present a novel chromatic confocal microscope capable of volumetric reflectance imaging of microstructure in non-transparent tissue. Our design takes advantage of the chromatic aberration of aspheric lenses that are otherwise well corrected. Strong chromatic aberration, generated by multiple aspheres, longitudinally disperses supercontinuum light onto the sample. The backscattered light detected with a spectrometer is therefore wavelength encoded and each spectrum corresponds to a line image. This approach obviates the need for traditional axial mechanical scanning techniques that are difficult to implement for endoscopy and susceptible to motion artifact. A wavelength range of 590-775 nm yielded a >150 µm imaging depth with ~3 µm axial resolution. The system was further demonstrated by capturing volumetric images of buccal mucosa. We believe these represent the first microstructural images in non-transparent biological tissue using chromatic confocal microscopy that exhibit long imaging depth while maintaining acceptable resolution for resolving cell morphology. Miniaturization of this optical system could bring enhanced speed and accuracy to endomicroscopic in vivo volumetric imaging of epithelial tissue. PMID:23667789

Reflective and refractive objects for mixed reality.

Science.gov (United States)

Knecht, Martin; Traxler, Christoph; Winklhofer, Christoph; Wimmer, Michael

2013-04-01

In this paper, we present a novel rendering method which integrates reflective or refractive objects into a differential instant radiosity (DIR) framework usable for mixed-reality (MR) applications. This kind of objects are very special from the light interaction point of view, as they reflect and refract incident rays. Therefore they may cause high-frequency lighting effects known as caustics. Using instant-radiosity (IR) methods to approximate these high-frequency lighting effects would require a large amount of virtual point lights (VPLs) and is therefore not desirable due to real-time constraints. Instead, our approach combines differential instant radiosity with three other methods. One method handles more accurate reflections compared to simple cubemaps by using impostors. Another method is able to calculate two refractions in real-time, and the third method uses small quads to create caustic effects. Our proposed method replaces parts in light paths that belong to reflective or refractive objects using these three methods and thus tightly integrates into DIR. In contrast to previous methods which introduce reflective or refractive objects into MR scenarios, our method produces caustics that also emit additional indirect light. The method runs at real-time frame rates, and the results show that reflective and refractive objects with caustics improve the overall impression for MR scenarios.

Mutual transformation of light waves by reflection holograms in photorefractive crystals of the 4-bar 3m symmetry

Energy Technology Data Exchange (ETDEWEB)

Naunyka, V. N.; Shepelevich, V. V., E-mail: vasshep@inbox.ru [Mozyr State Pedagogical University (Belarus)

2011-05-15

The mutual transformation of light waves in the case of their simultaneous diffraction from a bulk reflection phase hologram, which was formed in a cubic photorefractive crystal of the 4-bar 3m symmetry class, has been studied. The indicator surfaces of the polarization-optimized values of the relative intensity of the object wave, which make it possible to determine the amplification of this wave for any crystal cut, are constructed. The linear polarization azimuths at which the energy exchange between the light waves reaches a maximum are found numerically for crystals of different cuts.

Probing neural tissue with airy light-sheet microscopy: investigation of imaging performance at depth within turbid media

Science.gov (United States)

Nylk, Jonathan; McCluskey, Kaley; Aggarwal, Sanya; Tello, Javier A.; Dholakia, Kishan

2017-02-01

Light-sheet microscopy (LSM) has received great interest for fluorescent imaging applications in biomedicine as it facilitates three-dimensional visualisation of large sample volumes with high spatiotemporal resolution whilst minimising irradiation of, and photo-damage to the specimen. Despite these advantages, LSM can only visualize superficial layers of turbid tissues, such as mammalian neural tissue. Propagation-invariant light modes have played a key role in the development of high-resolution LSM techniques as they overcome the natural divergence of a Gaussian beam, enabling uniform and thin light-sheets over large distances. Most notably, Bessel and Airy beam-based light-sheet imaging modalities have been demonstrated. In the single-photon excitation regime and in lightly scattering specimens, Airy-LSM has given competitive performance with advanced Bessel-LSM techniques. Airy and Bessel beams share the property of self-healing, the ability of the beam to regenerate its transverse beam profile after propagation around an obstacle. Bessel-LSM techniques have been shown to increase the penetration-depth of the illumination into turbid specimens but this effect has been understudied in biologically relevant tissues, particularly for Airy beams. It is expected that Airy-LSM will give a similar enhancement over Gaussian-LSM. In this paper, we report on the comparison of Airy-LSM and Gaussian-LSM imaging modalities within cleared and non-cleared mouse brain tissue. In particular, we examine image quality versus tissue depth by quantitative spatial Fourier analysis of neural structures in virally transduced fluorescent tissue sections, showing a three-fold enhancement at 50 μm depth into non-cleared tissue with Airy-LSM. Complimentary analysis is performed by resolution measurements in bead-injected tissue sections.

Spectroscopic refractometer for transparent and absorbing liquids by reflection of white light near the critical angle

International Nuclear Information System (INIS)

Sánchez-Pérez, C.; García-Valenzuela, A.

2012-01-01

We propose and evaluate a spectroscopic refractometer device to measure the refractive index dispersion of transparent and absorbing solutions. The angle-dependent reflectivity of a white beam of light in an internal reflection configuration around the critical angle is spectrally analyzed. The refractive index in a wavelength range from 400 nm to 900 nm is obtained from the angle-reflectivity curve around the critical angle at each wavelength. The device does not use angle scanning mechanisms, decreasing considerably the complexity of the instrument in comparison to previous proposals. As a result, the measurements are obtained relatively fast. Nevertheless, a good experimental resolution in refractive index of about Δn ≈ 10 −4 at all the wavelengths is achieved in the case of transparent solutions. The calibration procedure of the device is discussed in detail. We also present measurements of the refractive index dispersion of rhodamine 6G-methanol solutions, which has a strong absorption band in the visible spectra.

Nanoimprint lithography of light trapping patterns in sol-gel coatings for thin film silicon solar cells

Energy Technology Data Exchange (ETDEWEB)

Heijna, M.; Loffler, J.; Van Aken, B.B.; Soppe, W.J. [ECN Solar Energy, Petten (Netherlands); Borg, H.; Peeters, P. [OM and T, Eindhoven (Netherlands)

2008-04-15

For thin-film silicon solar cells, light trapping schemes are of uppermost importance to harvest all available sunlight. Typically, randomly textured TCO front layers are used to scatter the light diffusively in p-i-n cells on glass. Here, we investigate methods to texture the back contact with both random and periodic textures, for use in n-i-p cells on opaque foil. We applied an electrically insulating SiOx-polymer coating on a stainless steel substrate, and textured this barrier layer by nanoimprint. On this barrier layer the back contact is deposited for further use in the solar cell stack. Replication of masters with various random and periodic patterns was tested, and, using scanning electron microscopy, replicas were found to compare well with the originals. Masters with U-grooves of various sub micrometer widths have been used to investigate the optimal dimensions of regular patterns for light trapping in the silicon layers. Angular reflection distributions were measured to evaluate the light scattering properties of both periodic and random patterns. Diffraction gratings show promising results in scattering the light to specific angles, enhancing the total internal reflection in the solar cell.

Quantitative analysis of phosphoinositide 3-kinase (PI3K) signaling using live-cell total internal reflection fluorescence (TIRF) microscopy.

Science.gov (United States)

Johnson, Heath E; Haugh, Jason M

2013-12-02

This unit focuses on the use of total internal reflection fluorescence (TIRF) microscopy and image analysis methods to study the dynamics of signal transduction mediated by class I phosphoinositide 3-kinases (PI3Ks) in mammalian cells. The first four protocols cover live-cell imaging experiments, image acquisition parameters, and basic image processing and segmentation. These methods are generally applicable to live-cell TIRF experiments. The remaining protocols outline more advanced image analysis methods, which were developed in our laboratory for the purpose of characterizing the spatiotemporal dynamics of PI3K signaling. These methods may be extended to analyze other cellular processes monitored using fluorescent biosensors. Copyright © 2013 John Wiley & Sons, Inc.

Transmission-type angle deviation microscopy

International Nuclear Information System (INIS)

Chiu, M.-H.; Lai, C.-W.; Tan, C.-T.; Lai, C.-F.

2008-01-01

We present a new microscopy technique that we call transmission angle deviation microscopy (TADM). It is based on common-path heterodyne interferometry and geometrical optics. An ultrahigh sensitivity surface plasmon resonance (SPR) angular sensor is used to expand dynamic measurement ranges and to improve the axial resolution in three-dimensional optical microscopy. When transmitted light is incident upon a specimen, the beam converges or diverges because of refractive and/or surface height variations. Advantages include high axial resolution (∼32 nm), nondestructive and noncontact measurement, and larger measurement ranges (± 80 μm) for a numerical aperture of 0.21in a transparent measurement medium. The technique can be used without conductivity and pretreatment

Reflection Matrix Method for Controlling Light After Reflection From a Diffuse Scattering Surface

Science.gov (United States)

2016-12-22

of Philosophy Kenneth W. Burgi, BS, MS Major, USAF 22 December 2016 DISTRIBUTION STATEMENT A APPROVED FOR PUBLIC RELEASE; DISTRIBUTION UNLIMITED. AFIT...refocusing light through thin films of a turbid medium. When coherent light is trans- mitted through a stationary diffuser (i.e. a turbid medium), a fine...resultant light scatter [14, 15, 21, 23]. Transmission matrices were measured with microscopic objectives and thin films of turbid media, resulting in

An introduction to optical super-resolution microscopy for the adventurous biologist

Science.gov (United States)

Vangindertael, J.; Camacho, R.; Sempels, W.; Mizuno, H.; Dedecker, P.; Janssen, K. P. F.

2018-04-01

Ever since the inception of light microscopy, the laws of physics have seemingly thwarted every attempt to visualize the processes of life at its most fundamental, sub-cellular, level. The diffraction limit has restricted our view to length scales well above 250 nm and in doing so, severely compromised our ability to gain true insights into many biological systems. Fortunately, continuous advancements in optics, electronics and mathematics have since provided the means to once again make physics work to our advantage. Even though some of the fundamental concepts enabling super-resolution light microscopy have been known for quite some time, practically feasible implementations have long remained elusive. It should therefore not come as a surprise that the 2014 Nobel Prize in Chemistry was awarded to the scientists who, each in their own way, contributed to transforming super-resolution microscopy from a technological tour de force to a staple of the biologist’s toolkit. By overcoming the diffraction barrier, light microscopy could once again be established as an indispensable tool in an age where the importance of understanding life at the molecular level cannot be overstated. This review strives to provide the aspiring life science researcher with an introduction to optical microscopy, starting from the fundamental concepts governing compound and fluorescent confocal microscopy to the current state-of-the-art of super-resolution microscopy techniques and their applications.

Shedding new light on lipid functions with CARS and SRS microscopy

Science.gov (United States)

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

2014-01-01

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. PMID:24576891

Application of Confocal Laser Scanning Microscopy in Biology and Medicine

OpenAIRE

I. A. Volkov; N. V. Frigo; L. F. Znamenskaya; O. R. Katunina

2014-01-01

Fluorescence confocal laser scanning microscopy and reflectance confocal laser scanning microscopy are up-to-date highend study methods. Confocal microscopy is used in cell biology and medicine. By using confocal microscopy, it is possible to study bioplasts and localization of protein molecules and other compounds relative to cell or tissue structures, and to monitor dynamic cell processes. Confocal microscopes enable layer-by-layer scanning of test items to create demonstrable 3D models. As...

ultraLM and miniLM: Locator tools for smart tracking of fluorescent cells in correlative light and electron microscopy.

Science.gov (United States)

Brama, Elisabeth; Peddie, Christopher J; Wilkes, Gary; Gu, Yan; Collinson, Lucy M; Jones, Martin L

2016-12-13

In-resin fluorescence (IRF) protocols preserve fluorescent proteins in resin-embedded cells and tissues for correlative light and electron microscopy, aiding interpretation of macromolecular function within the complex cellular landscape. Dual-contrast IRF samples can be imaged in separate fluorescence and electron microscopes, or in dual-modality integrated microscopes for high resolution correlation of fluorophore to organelle. IRF samples also offer a unique opportunity to automate correlative imaging workflows. Here we present two new locator tools for finding and following fluorescent cells in IRF blocks, enabling future automation of correlative imaging. The ultraLM is a fluorescence microscope that integrates with an ultramicrotome, which enables 'smart collection' of ultrathin sections containing fluorescent cells or tissues for subsequent transmission electron microscopy or array tomography. The miniLM is a fluorescence microscope that integrates with serial block face scanning electron microscopes, which enables 'smart tracking' of fluorescent structures during automated serial electron image acquisition from large cell and tissue volumes.

A four-phase strategy for the implementation of reflectance confocal microscopy in dermatology.

Science.gov (United States)

Hoogedoorn, L; Gerritsen, M J P; Wolberink, E A W; Peppelman, M; van de Kerkhof, P C M; van Erp, P E J

2016-08-01

Reflectance confocal microscopy (RCM) is gradually implemented in dermatology. Strategies for further implementation and practical 'hands on' guidelines are lacking. The primary outcome was to conduct a general strategy for further implementation of RCM. The secondary outcome was the diagnosis of psoriasis and differentiation of stable from unstable psoriatic plaques by means of the 'hands on' protocol, derived from the strategy. We used a four-phased model; an exploring phase, a systematic literature search, a clinical approach and, finally, an integration phase to develop a clinical guideline for RCM in psoriasis. Receiver operating characteristic curve statistics was applied to define the accuracy for the diagnosis of unstable psoriasis. A general strategy for further implementation of RCM and practical approach was developed to examine psoriasis by RCM and to distinguish stable from unstable psoriasis. Unstable psoriasis was diagnosed by epidermal inflammatory cell counts with a sensitivity and specificity of 91.7% and 98.3%, respectively, and with an accuracy of 0.92 (area under the curve). In addition, a monitoring model was proposed. This is the first study that shows a method for implementation of RCM in dermatology. The strategy and hands on protocol for psoriasis may serve as a model for other dermatological entities and additionally may lead to specialized ready-to-use RCM protocols for clinical dermatological practice. © 2016 European Academy of Dermatology and Venereology.

High-resolution light microscopy using luminescent nanoparticles.

Science.gov (United States)

Ohulchanskyy, Tymish Y; Roy, Indrajit; Yong, Ken-Tye; Pudavar, Haridas E; Prasad, Paras N

2010-01-01

This review presents recent progress in the development of the luminescent nanoparticles for confocal and multiphoton microscopy. Four classes of nanomaterials are discussed: (1) silica-based nanoparticles doped with fluorescent molecules, (2) gold nanoparticles, (3) semiconductor nanocrystals (quantum dots/rods), and (4) nanophosphors. Special considerations are given to recently developed imaging nanoprobes, such as (1) organically modified silica (ORMOSIL) nanoparticles doped with two-photon absorbing fluorophores, which exhibit aggregation-enhanced fluorescence (AEF), and (2) nanophosphors (ceramic nanoparticles containing luminescent lanthanoid ions). Advantages and disadvantages of every class of nanomaterials and their specific applications are briefly discussed.

Chemical and structural composition study through transmission and reflection electron microscopy and X-ray microanalysis of damaged duramater cardiac valves

International Nuclear Information System (INIS)

Verginelli, G.; Didio, L.J.A.; Puig, L.B.; Allen, D.J.; Highinson, G.H.; Zerbini, E.J.

1982-01-01

Ten malfunctioning durameter aortic prosthesis, excised surgically after 44 to 54 months of implantation were studied through transmission and reflection electron microscopy and X-ray microanalysis. Duramater extracted at necropsy but not used for valve prosthesis and duramater aortic prosthesis not implanted in patients were used as controls. It was concluded that homologous duramater valves present subcellular changes following implantation, with degenerating and proliferating areas which could correspond either to normal adaptation or consequence of degeneration - rebuilding which begins with implantation; it is also emphazised the need for ultramicroscopic studies in evaluating biological tissues and establishing its applicability in cardiac surgery. (M.A.C.) [pt

Deleterious phases precipitation on superduplex stainless steel UNS S32750: characterization by light optical and scanning electron microscopy

Directory of Open Access Journals (Sweden)

Juan Manuel Pardal

2010-09-01

Full Text Available Deleterious phases precipitation in superduplex stainless steels is the main concern in fabrication by welding and hot forming of this class of material. Sigma, chi and secondary austenite phases are considered deleterious phases because they produce negative effects on corrosion resistance. Besides, sigma and chi phases also promote strong decrease of toughness. In the present work, the precipitations of sigma, chi and secondary austenite under aging in the 800-950 °C interval were studied in two UNS S32750 steels with different grain sizes. The deleterious phases could be quantified by light optical microscopy, with no distinction between them. Scanning electron microscopy was used to distinguish the individual phases in various aging conditions. The results elucidate the influence of the aging temperature and grain size on the kinetics precipitation and morphology of deleterious phases. The kinetics of deleterious phases is higher in the fine grained material in the initial stage of aging, but the maximum amount of deleterious phases is higher in the coarse grained steel.

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...

Angularly symmetric splitting of a light beam upon reflection and refraction at an air-dielectric plane boundary: comment.

Science.gov (United States)

Andersen, Torben B

2016-05-01

In a recent paper, conditions for achieving equal and opposite angular deflections of a light beam by reflection and refraction at an interface between air and a dielectric were determined [J. Opt. Soc. Am. A32, 2436 (2015)JOAOD60740-323210.1364/JOSAA.32.002436]. The paper gives plots of angles of incidence and refraction as a function of the prism refractive index as well as plots of reflectances and incident linear-polarization azimuth angles as functions of the refractive index. We show here that it is possible to express these quantities as simple algebraic functions of the refractive index.

Accuracy of Mobile Phone and Handheld Light Microscopy for the Diagnosis of Schistosomiasis and Intestinal Protozoa Infections in Côte 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

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

Application of multiphoton microscopy in dermatological studies: A mini-review

Directory of Open Access Journals (Sweden)

Elijah Yew

2014-09-01

Full Text Available This review summarizes the historical and more recent developments of multiphoton microscopy, as applied to dermatology. Multiphoton microscopy offers several advantages over competing microscopy techniques: there is an inherent axial sectioning, penetration depths that compete well with confocal microscopy on account of the use of near-infrared light, and many two-photon contrast mechanisms, such as second-harmonic generation, have no analogue in one-photon microscopy. While the penetration depths of photons into tissue are typically limited on the order of hundreds of microns, this is of less concern in dermatology, as the skin is thin and readily accessible. As a result, multiphoton microscopy in dermatology has generated a great deal of interest, much of which is summarized here. The review covers the interaction of light and tissue, as well as the various considerations that must be made when designing an instrument. The state of multiphoton microscopy in imaging skin cancer and various other diseases is also discussed, along with the investigation of aging and regeneration phenomena, and finally, the use of multiphoton microscopy to analyze the transdermal transport of drugs, cosmetics and other agents is summarized. The review concludes with a look at potential future research directions, especially those that are necessary to push these techniques into widespread clinical acceptance.

Graphene-spindle shaped TiO2 mesocrystal composites: Facile synthesis and enhanced visible light photocatalytic performance

International Nuclear Information System (INIS)

Yang, Xiaofei; Qin, Jieling; Li, Yang; Zhang, Rongxian; Tang, Hua

2013-01-01

Highlights: • Graphene-TiO 2 mesocrystal composites were fabricated via a facile approach. • Graphene sheets were decorated with spindle-like TiO 2 mesocrystals. • Graphene causes enhanced light absorbance and visible light photocatalytic activity. • Oxygen-containing radicals are believed to responsible for its improved activity. -- Abstract: Graphene (GR)-TiO 2 mesocrystal composites were prepared by a facile template-free process based on the combination of sol–gel and solvothermal methods, and were characterized using field emission scanning electron microscopy (FE-SEM), transmission electron microscopy (TEM), high-resolution transmission electron microscopy (HRTEM), X-ray diffraction (XRD), Raman spectroscopy, UV–vis diffuse reflectance spectroscopy (UV–vis DRS), nitrogen absorption and electron spin resonance (ESR). Visible light photocatalytic performance of GR-TiO 2 composites was evaluated for photocatalytic degradation of organic dye Rhodamine B. It was found that the amount of graphene oxide (GO) added obviously affects morphologies of TiO 2 mesocrystals and photocatalytic activities of as-prepared nanocomposites. Composites prepared in the presence of different amounts of GO all exhibit higher photocatalytic activity than pure TiO 2 mesocrystals and P25, the composite obtained by using 20 mg GO presents the most uniform TiO 2 mesocrystals in the composite and shows the highest photocatalytic efficiency. The mechanism for the generation of TiO 2 mesocrystals in the GR-TiO 2 composite is proposed and possible reasons for the enhancement in visible light photocatalytic efficiency are also discussed

Single-wavelength functional photoacoustic microscopy in biological tissue

OpenAIRE

Danielli, Amos; Favazza, Christopher P.; Maslov, Konstantin; Wang, Lihong V.

2011-01-01

Recently, we developed a reflection-mode relaxation photoacoustic microscope, based on saturation intensity, to measure picosecond relaxation times using a nanosecond laser. Here, using the different relaxation times of oxygenated and deoxygenated hemoglobin molecules, both possessing extremely low fluorescence quantum yields, the oxygen saturation was quantified in vivo with single-wavelength photoacoustic microscopy. All previous functional photoacoustic microscopy measurements required ima...

Fluorescence confocal polarizing microscopy

Indian Academy of Sciences (India)

Much of the modern understanding of orientational order in liquid crystals (LCs) is based on polarizing microscopy (PM). A PM image bears only two-dimensional (2D) information, integrating the 3D pattern of optical birefringence over the path of light. Recently, we proposed a technique to image 3D director patterns by ...

Concurrent Reflectance Confocal Microscopy and Laser Doppler Flowmetry to Improve Skin Cancer Imaging: A Monte Carlo Model and Experimental Validation

Directory of Open Access Journals (Sweden)

Alireza Mowla

2016-09-01

Full Text Available Optical interrogation of suspicious skin lesions is standard care in the management of skin cancer worldwide. Morphological and functional markers of malignancy are often combined to improve expert human diagnostic power. We propose the evaluation of the combination of two independent optical biomarkers of skin tumours concurrently. The morphological modality of reflectance confocal microscopy (RCM is combined with the functional modality of laser Doppler flowmetry, which is capable of quantifying tissue perfusion. To realize the idea, we propose laser feedback interferometry as an implementation of RCM, which is able to detect the Doppler signal in addition to the confocal reflectance signal. Based on the proposed technique, we study numerical models of skin tissue incorporating two optical biomarkers of malignancy: (i abnormal red blood cell velocities and concentrations and (ii anomalous optical properties manifested through tissue confocal reflectance, using Monte Carlo simulation. We also conduct a laboratory experiment on a microfluidic channel containing a dynamic turbid medium, to validate the efficacy of the technique. We quantify the performance of the technique by examining a signal to background ratio (SBR in both the numerical and experimental models, and it is shown that both simulated and experimental SBRs improve consistently using this technique. This work indicates the feasibility of an optical instrument, which may have a role in enhanced imaging of skin malignancies.

Angularly symmetric splitting of a light beam upon reflection and refraction at an air-dielectric plane boundary.

Science.gov (United States)

Azzam, R M A

2015-12-01

Conditions for achieving equal and opposite angular deflections of a light beam by reflection and refraction at an air-dielectric boundary are determined. Such angularly symmetric beam splitting (ASBS) is possible only if the angle of incidence is >60° by exactly one third of the angle of refraction. This simple law, plus Snell's law, leads to several analytical results that clarify all aspects of this phenomenon. In particular, it is shown that the intensities of the two symmetrically deflected beams can be equalized by proper choice of the prism refractive index and the azimuth of incident linearly polarized light. ASBS enables a geometrically attractive layout of optical systems that employ multiple prism beam splitters.

A correlative optical microscopy and scanning electron microscopy approach to locating nanoparticles in brain tumors.

Science.gov (United States)

Kempen, Paul J; Kircher, Moritz F; de la Zerda, Adam; Zavaleta, Cristina L; Jokerst, Jesse V; Mellinghoff, Ingo K; Gambhir, Sanjiv S; Sinclair, Robert

2015-01-01

The growing use of nanoparticles in biomedical applications, including cancer diagnosis and treatment, demands the capability to exactly locate them within complex biological systems. In this work a correlative optical and scanning electron microscopy technique was developed to locate and observe multi-modal gold core nanoparticle accumulation in brain tumor models. Entire brain sections from mice containing orthotopic brain tumors injected intravenously with nanoparticles were imaged using both optical microscopy to identify the brain tumor, and scanning electron microscopy to identify the individual nanoparticles. Gold-based nanoparticles were readily identified in the scanning electron microscope using backscattered electron imaging as bright spots against a darker background. This information was then correlated to determine the exact location of the nanoparticles within the brain tissue. The nanoparticles were located only in areas that contained tumor cells, and not in the surrounding healthy brain tissue. This correlative technique provides a powerful method to relate the macro- and micro-scale features visible in light microscopy with the nanoscale features resolvable in scanning electron microscopy. Copyright © 2014 Elsevier Ltd. All rights reserved.

Study on activities of vanadium (IV/V) doped TiO2(R) nanorods induced by UV and visible light

International Nuclear Information System (INIS)

Li, Li; Liu Chunyan; Liu Yun

2009-01-01

Vanadium (IV/V) doped rutile TiO 2 naonorods had been successfully synthesized through a single step hydrothermal method. The photocatalyst was characterized by transmission electron microscopy (TEM), selected area electron diffraction (SAED), high-resolution transmission electron microscopy (HRTEM), X-ray diffraction (XRD), UV-vis diffusive reflectance spectroscopy (DRS) and X-ray photoelectron spectroscopy (XPS). The results showed that the doping of V ions had significant influence on the band gap energy and the surface state of TiO 2 . The photo-activities of the new catalysts were investigated under ultraviolet (UV) and visible light. The UV-photocatalytic activity of the as-prepared catalysts was hardly influenced by doping V ions; while under visible light, the samples with 1 wt% and 0.1 wt% V exhibited enhanced activity to the oxidation of methylene blue (MB) and the reduction of Cr (VI), respectively

The generalized vectorial laws of reflection and refraction

International Nuclear Information System (INIS)

Bhattacharjee, Pramode Ranjan

2005-01-01

This paper discloses two important discoveries. These are: (i) discovery of ambiguity in the well-established laws of reflection and refraction of light which have been in regular use for many years, and (ii) discovery of generalized vectorial laws of reflection and refraction of light. The existing definitions of angle of incidence, angle of reflection and angle of refraction are considered first. Each of these definitions is found to be ambiguous, not in compliance with the fundamental definition of angle in geometry. Two typical questions (one in the case of reflection and the other for refraction) have been addressed, which cannot be dealt with by using the existing laws of reflection and refraction of light. Thus, the existing laws of reflection and refraction of light seem to be ambiguous in respect of generality and their validity in a broad sense is questionable. With a view to removing the ambiguities, proper definitions of the above three angles are given first and then the statement of the generalized vectorial law of reflection (as well as that of refraction) has been offered

High reflectance Cr/C multilayer at 250 eV for soft X-ray polarimetry

Energy Technology Data Exchange (ETDEWEB)

Wen, Mingwu; Jiang, Li; Zhang, Zhong; Huang, Qiushi [MOE Key Laboratory of Advanced Micro-Structured Materials, Institute of Precision Optical Engineering (IPOE), School of Physics Science and Engineering, Tongji University, Shanghai 200092 (China); Wang, Zhanshan, E-mail: wangzs@tongji.edu.cn [MOE Key Laboratory of Advanced Micro-Structured Materials, Institute of Precision Optical Engineering (IPOE), School of Physics Science and Engineering, Tongji University, Shanghai 200092 (China); She, Rui; Feng, Hua [Department of Engineering Physics, Tsinghua University, Beijing (China); Wang, Hongchang [Diamond Light Source, Harwell Science and Innovation Campus, Didcot, Oxfordshire OX11 0DE (United Kingdom)

2015-10-01

X-ray reflection near 45° via multilayer mirrors can be used for astronomical polarization measurements. A Cr/C multilayer mirror (designed for X-ray polarimetry at 250 eV), with a period thickness of 3.86 nm and a bi-layer number of 100, was fabricated using direct current magnetron sputtering. Grazing incidence X-ray reflectometry at 8 keV and transmission electron microscopy were used to investigate the multilayer structure. Different models were introduced to fit the hard X-ray reflectivity curve, which indicates that the layer thickness of two materials slightly drifts from the bottom to the top of the stack. Both the chromium and carbon layers are amorphous with asymmetric interfaces, while the Cr-on-C interface is slightly wider. Based on the good quality of the multilayer structure, a high reflectivity of 21.8% for the s-polarized light was obtained at 250 eV at a grazing incidence angle of 40.7°. The fabricated Cr/C multilayer mirror exhibits high reflectivity and polarization levels in the energy region of 240 eV–260 eV. - Highlights: • We fabricated Cr/C multilayer with 3.8 nm d-spacing. • X-ray reflectometry was used to determine the exact structure of Cr/C multilayer. • A high reflectivity of 21.8% for the s-polarized light was obtained at 250 eV. • Both Cr and C were found to be amorphous with slightly asymmetric interfaces. • A 4-layer model was used to fit and explain the results.

3D imaging of cleared human skin biopsies using light-sheet microscopy: A new way to visualize in-depth skin structure.

Science.gov (United States)

Abadie, S; Jardet, C; Colombelli, J; Chaput, B; David, A; Grolleau, J-L; Bedos, P; Lobjois, V; Descargues, P; Rouquette, J

2018-05-01

Human skin is composed of the superimposition of tissue layers of various thicknesses and components. Histological staining of skin sections is the benchmark approach to analyse the organization and integrity of human skin biopsies; however, this approach does not allow 3D tissue visualization. Alternatively, confocal or two-photon microscopy is an effective approach to perform fluorescent-based 3D imaging. However, owing to light scattering, these methods display limited light penetration in depth. The objectives of this study were therefore to combine optical clearing and light-sheet fluorescence microscopy (LSFM) to perform in-depth optical sectioning of 5 mm-thick human skin biopsies and generate 3D images of entire human skin biopsies. A benzyl alcohol and benzyl benzoate solution was used to successfully optically clear entire formalin fixed human skin biopsies, making them transparent. In-depth optical sectioning was performed with LSFM on the basis of tissue-autofluorescence observations. 3D image analysis of optical sections generated with LSFM was performed by using the Amira ® software. This new approach allowed us to observe in situ the different layers and compartments of human skin, such as the stratum corneum, the dermis and epidermal appendages. With this approach, we easily performed 3D reconstruction to visualise an entire human skin biopsy. Finally, we demonstrated that this method is useful to visualise and quantify histological anomalies, such as epidermal hyperplasia. The combination of optical clearing and LSFM has new applications in dermatology and dermatological research by allowing 3D visualization and analysis of whole human skin biopsies. © 2018 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

Dual light-emitting diode-based multichannel microscopy for whole-slide multiplane, multispectral and phase imaging.

Science.gov (United States)

Liao, Jun; Wang, Zhe; Zhang, Zibang; Bian, Zichao; Guo, Kaikai; Nambiar, Aparna; Jiang, Yutong; Jiang, Shaowei; Zhong, Jingang; Choma, Michael; Zheng, Guoan

2018-02-01

We report the development of a multichannel microscopy for whole-slide multiplane, multispectral and phase imaging. We use trinocular heads to split the beam path into 6 independent channels and employ a camera array for parallel data acquisition, achieving a maximum data throughput of approximately 1 gigapixel per second. To perform single-frame rapid autofocusing, we place 2 near-infrared light-emitting diodes (LEDs) at the back focal plane of the condenser lens to illuminate the sample from 2 different incident angles. A hot mirror is used to direct the near-infrared light to an autofocusing camera. For multiplane whole-slide imaging (WSI), we acquire 6 different focal planes of a thick specimen simultaneously. For multispectral WSI, we relay the 6 independent image planes to the same focal position and simultaneously acquire information at 6 spectral bands. For whole-slide phase imaging, we acquire images at 3 focal positions simultaneously and use the transport-of-intensity equation to recover the phase information. We also provide an open-source design to further increase the number of channels from 6 to 15. The reported platform provides a simple solution for multiplexed fluorescence imaging and multimodal WSI. Acquiring an instant focal stack without z-scanning may also enable fast 3-dimensional dynamic tracking of various biological samples. © 2017 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Tilted Light Sheet Microscopy with 3D Point Spread Functions for Single-Molecule Super-Resolution Imaging in Mammalian Cells.

Science.gov (United States)

Gustavsson, Anna-Karin; Petrov, Petar N; Lee, Maurice Y; Shechtman, Yoav; Moerner, W E

2018-02-01

To obtain a complete picture of subcellular nanostructures, cells must be imaged with high resolution in all three dimensions (3D). Here, we present tilted light sheet microscopy with 3D point spread functions (TILT3D), an imaging platform that combines a novel, tilted light sheet illumination strategy with engineered long axial range point spread functions (PSFs) for low-background, 3D super localization of single molecules as well as 3D super-resolution imaging in thick cells. TILT3D is built upon a standard inverted microscope and has minimal custom parts. The axial positions of the single molecules are encoded in the shape of the PSF rather than in the position or thickness of the light sheet, and the light sheet can therefore be formed using simple optics. The result is flexible and user-friendly 3D super-resolution imaging with tens of nm localization precision throughout thick mammalian cells. We validated TILT3D for 3D super-resolution imaging in mammalian cells by imaging mitochondria and the full nuclear lamina using the double-helix PSF for single-molecule detection and the recently developed Tetrapod PSF for fiducial bead tracking and live axial drift correction. We envision TILT3D to become an important tool not only for 3D super-resolution imaging, but also for live whole-cell single-particle and single-molecule tracking.

Tilted light sheet microscopy with 3D point spread functions for single-molecule super-resolution imaging in mammalian cells

Science.gov (United States)

Gustavsson, Anna-Karin; Petrov, Petar N.; Lee, Maurice Y.; Shechtman, Yoav; Moerner, W. E.

2018-02-01

To obtain a complete picture of subcellular nanostructures, cells must be imaged with high resolution in all three dimensions (3D). Here, we present tilted light sheet microscopy with 3D point spread functions (TILT3D), an imaging platform that combines a novel, tilted light sheet illumination strategy with engineered long axial range point spread functions (PSFs) for low-background, 3D super localization of single molecules as well as 3D super-resolution imaging in thick cells. TILT3D is built upon a standard inverted microscope and has minimal custom parts. The axial positions of the single molecules are encoded in the shape of the PSF rather than in the position or thickness of the light sheet, and the light sheet can therefore be formed using simple optics. The result is flexible and user-friendly 3D super-resolution imaging with tens of nm localization precision throughout thick mammalian cells. We validated TILT3D for 3D superresolution imaging in mammalian cells by imaging mitochondria and the full nuclear lamina using the double-helix PSF for single-molecule detection and the recently developed Tetrapod PSF for fiducial bead tracking and live axial drift correction. We envision TILT3D to become an important tool not only for 3D super-resolution imaging, but also for live whole-cell single-particle and single-molecule tracking.

Doppler optical coherence microscopy and tomography applied to inner ear mechanics

Energy Technology Data Exchange (ETDEWEB)

Page, Scott; Freeman, Dennis M. [Research Laboratory of Electronics, Massachusetts Institute of Technology, Cambridge, Massachusetts (United States); Department of Electrical Engineering and Computer Science, Massachusetts Institute of Technology, Cambridge, Massachusetts (United States); Ghaffari, Roozbeh [Research Laboratory of Electronics, Massachusetts Institute of Technology, Cambridge, Massachusetts (United States)

2015-12-31

While it is clear that cochlear traveling waves underlie the extraordinary sensitivity, frequency selectivity, and dynamic range of mammalian hearing, the underlying micromechanical mechanisms remain unresolved. Recent advances in low coherence measurement techniques show promise over traditional laser Doppler vibrometry and video microscopy, which are limited by low reflectivities of cochlear structures and restricted optical access. Doppler optical coherence tomography (DOCT) and Doppler optical coherence microscopy (DOCM) both utilize a broadband source to limit constructive interference of scattered light to a small axial depth called a coherence gate. The coherence gate can be swept axially to image and measure sub-nanometer motions of cochlear structures throughout the cochlear partition. The coherence gate of DOCT is generally narrower than the confocal gate of the focusing optics, enabling increased axial resolution (typically 15 μm) within optical sections of the cochlear partition. DOCM, frequently implemented in the time domain, centers the coherence gate on the focal plane, achieving enhanced lateral and axial resolution when the confocal gate is narrower than the coherence gate. We compare these two complementary systems and demonstrate their utility in studying cellular and micromechanical mechanisms involved in mammalian hearing.

High-resolution imaging of basal cell carcinoma: a comparison between multiphoton microscopy with fluorescence lifetime imaging and reflectance confocal microscopy.

Science.gov (United States)

Manfredini, Marco; Arginelli, Federica; Dunsby, Christopher; French, Paul; Talbot, Clifford; König, Karsten; Pellacani, Giovanni; Ponti, Giovanni; Seidenari, Stefania

2013-02-01

The aim of this study was to compare morphological aspects of basal cell carcinoma (BCC) as assessed by two different imaging methods: in vivo reflectance confocal microscopy (RCM) and multiphoton tomography with fluorescence lifetime imaging implementation (MPT-FLIM). The study comprised 16 BCCs for which a complete set of RCM and MPT-FLIM images were available. The presence of seven MPT-FLIM descriptors was evaluated. The presence of seven RCM equivalent parameters was scored in accordance to their extension. Chi-squared test with Fisher's exact test and Spearman's rank correlation coefficient were determined between MPT-FLIM scores and adjusted-RCM scores. MPT-FLIM and RCM descriptors of BCC were coupled to match the descriptors that define the same pathological structures. The comparison included: Streaming and Aligned elongated cells, Streaming with multiple directions and Double alignment, Palisading (RCM) and Palisading (MPT-FLIM), Typical tumor islands, and Cell islands surrounded by fibers, Dark silhouettes and Phantom islands, Plump bright cells and Melanophages, Vessels (RCM), and Vessels (MPT-FLIM). The parameters that were significantly correlated were Melanophages/Plump Bright Cells, Aligned elongated cells/Streaming, Double alignment/Streaming with multiple directions, and Palisading (MPT-FLIM)/Palisading (RCM). According to our data, both methods are suitable to image BCC's features. The concordance between MPT-FLIM and RCM is high, with some limitations due to the technical differences between the two devices. The hardest difficulty when comparing the images generated by the two imaging modalities is represented by their different field of view. © 2012 John Wiley & Sons A/S.

Applications of microscopy in Salmonella research.

Science.gov (United States)

Malt, Layla M; Perrett, Charlotte A; Humphrey, Suzanne; Jepson, Mark A

2015-01-01

Salmonella enterica is a Gram-negative enteropathogen that can cause localized infections, typically resulting in gastroenteritis, or systemic infection, e.g., typhoid fever, in humans and many other animals. Understanding the mechanisms by which Salmonella induces disease has been the focus of intensive research. This has revealed that Salmonella invasion requires dynamic cross-talk between the microbe and host cells, in which bacterial adherence rapidly leads to a complex sequence of cellular responses initiated by proteins translocated into the host cell by a type 3 secretion system. Once these Salmonella-induced responses have resulted in bacterial invasion, proteins translocated by a second type 3 secretion system initiate further modulation of cellular activities to enable survival and replication of the invading pathogen. Elucidation of the complex and highly dynamic pathogen-host interactions ultimately requires analysis at the level of single cells and single infection events. To achieve this goal, researchers have applied a diverse range of microscopy techniques to analyze Salmonella infection in models ranging from whole animal to isolated cells and simple eukaryotic organisms. For example, electron microscopy and high-resolution light microscopy techniques such as confocal microscopy can reveal the precise location of Salmonella and its relationship to cellular components. Widefield light microscopy is a simpler approach with which to study the interaction of bacteria with host cells and often has advantages for live cell imaging, enabling detailed analysis of the dynamics of infection and cellular responses. Here we review the use of imaging techniques in Salmonella research and compare the capabilities of different classes of microscope to address specific types of research question. We also provide protocols and notes on some microscopy techniques used routinely in our own research.

Re-scan confocal microscopy (RCM) improves the resolution of confocal microscopy and increases the sensitivity

NARCIS (Netherlands)

de Luca, Giulia; Breedijk, Ronald; Hoebe, Ron; Stallinga, Sjoerd; Manders, Erik

2017-01-01

Re-scan confocal microscopy (RCM) is a new super-resolution technique based on a standard confocal microscope extended with a re-scan unit in the detection path that projects the emitted light onto a sensitive camera. In this paper the fundamental properties of RCM, lateral resolution, axial

Re-scan confocal microscopy (RCM) improves the resolution of confocal microscopy and increases the sensitivity

NARCIS (Netherlands)

De Luca, G.; Breedijk, R.; Hoebe, R.; Stallinga, S.; Manders, E.

Re-scan confocal microscopy (RCM) is a new super-resolution technique based on a standard confocal microscope extended with a re-scan unit in the detection path that projects the emitted light onto a sensitive camera. In this paper the fundamental properties of RCM, lateral resolution, axial

Digital Fourier microscopy for soft matter dynamics

International Nuclear Information System (INIS)

Giavazzi, Fabio; Cerbino, Roberto

2014-01-01

Soft matter is studied with a large portfolio of methods. Light scattering and video microscopy are the most employed at optical wavelengths. Light scattering provides ensemble-averaged information on soft matter in the reciprocal space. The wave-vectors probed correspond to length scales ranging from a few nanometers to fractions of millimetre. Microscopy probes the sample directly in the real space, by offering a unique access to the local properties. However, optical resolution issues limit the access to length scales smaller than approximately 200 nm. We describe recent work that bridges the gap between scattering and microscopy. Several apparently unrelated techniques are found to share a simple basic idea: the correlation properties of the sample can be characterized in the reciprocal space via spatial Fourier analysis of images collected in the real space. We describe the main features of such digital Fourier microscopy (DFM), by providing examples of several possible experimental implementations of it, some of which not yet realized in practice. We also provide an overview of experimental results obtained with DFM for the study of the dynamics of soft materials. Finally, we outline possible future developments of DFM that would ease its adoption as a standard laboratory method. (topical review)

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

Science.gov (United States)

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

2016-08-16

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

Influence of the linear magneto-electric effect on the lateral shift of light reflected from a magneto-electric film

International Nuclear Information System (INIS)

Dadoenkova, Yu S; Petrov, R V; Bichurin, M I; Bentivegna, F F L; Dadoenkova, N N; Lyubchanskii, I L

2016-01-01

We present a theoretical investigation of the lateral shift of an infrared light beam reflected from a magnetic film deposited on a non-magnetic dielectric substrate, taking into account the linear magneto-electric interaction in the magnetic film. We use the stationary phase method to evaluate the lateral shift. It is shown that the magneto-electric coupling leads to a six-fold enhancement of the lateral shift amplitude of a p-(s-) polarized incident beam reflected into a s-(p-) polarized beam. A reversal of the magnetization in the film leads to a nonreciprocal sign change of the lateral shift. (paper)

Electron microscopy of primary cell cultures in solution and correlative optical microscopy using ASEM

International Nuclear Information System (INIS)

Hirano, Kazumi; Kinoshita, Takaaki; Uemura, Takeshi; Motohashi, Hozumi; Watanabe, Yohei; Ebihara, Tatsuhiko; Nishiyama, Hidetoshi; Sato, Mari; Suga, Mitsuo; Maruyama, Yuusuke; Tsuji, Noriko M.; Yamamoto, Masayuki; Nishihara, Shoko; Sato, Chikara

2014-01-01

Correlative light-electron microscopy of cells in a natural environment of aqueous liquid facilitates high-throughput observation of protein complex formation. ASEM allows the inverted SEM to observe the wet sample from below, while an optical microscope observes it from above quasi-simultaneously. The disposable ASEM dish with a silicon nitride (SiN) film window can be coated variously to realize the primary-culture of substrate-sensitive cells in a few milliliters of culture medium in a stable incubator environment. Neuron differentiation, neural networking, proplatelet-formation and phagocytosis were captured by optical or fluorescence microscopy, and imaged at high resolution by gold-labeled immuno-ASEM with/without metal staining. Fas expression on the cell surface was visualized, correlated to the spatial distribution of F-actin. Axonal partitioning was studied using primary-culture neurons, and presynaptic induction by GluRδ2-N-terminus-linked fluorescent magnetic beads was correlated to the presynaptic-marker Bassoon. Further, megakaryocytes secreting proplatelets were captured, and P-selectins with adherence activity were localized to some of the granules present by immuno-ASEM. The phagocytosis of lactic acid bacteria by dendritic cells was also imaged. Based on these studies, ASEM correlative microscopy promises to allow the study of various mesoscopic-scale dynamics in the near future. - Highlights: • In situ correlative light electron microscopy of samples in open solution by ASEM. • Primary cultures for in-solution CLEM by developing SiN-film coating methods • First visualization of fluorescent magnetic beads in aqueous solution by CLEM. • Presynaptic induction of neurons by GluRδ2-N-terminus-coated beads studied by CLEM. • Axonal partitioning, bacterial phagocytosis, platelet formation imaged by CLEM

Electron microscopy of primary cell cultures in solution and correlative optical microscopy using ASEM

Energy Technology Data Exchange (ETDEWEB)

Hirano, Kazumi; Kinoshita, Takaaki [Laboratory of Cell Biology, Department of Bioinformatics, Faculty of Engineering, Soka University, 1-236 Tangi-machi, Hachioji, Tokyo 192-8577 (Japan); Uemura, Takeshi [Department of Molecular Neurobiology and Pharmacology, Graduate School of Medicine, University of Tokyo, 7-3-1 Hongo, Bunkyo-ku, Tokyo 113-0033 (Japan); Department of Molecular and Cellular Physiology, Shinshu University School of Medicine, 3-1-1 Asahi, Matsumoto, Nagano 390-8621 (Japan); Motohashi, Hozumi [Department of Gene Expression Regulation, Institute of Development, Aging and Cancer, Tohoku University, 4-1 Seiryo-cho, Aoba-ku, Sendai 980-8575 (Japan); Watanabe, Yohei; Ebihara, Tatsuhiko [Biomedical Research Institute, National Institute of Industrial Science and Technology (AIST), 1-1-1 Higashi, Tsukuba 305-8566 (Japan); Nishiyama, Hidetoshi [JEOL Ltd., 1-2 Musashino 3-chome, Akishima, Tokyo 196-8558 (Japan); Sato, Mari [Biomedical Research Institute, National Institute of Industrial Science and Technology (AIST), 1-1-1 Higashi, Tsukuba 305-8566 (Japan); Suga, Mitsuo [JEOL Ltd., 1-2 Musashino 3-chome, Akishima, Tokyo 196-8558 (Japan); Maruyama, Yuusuke; Tsuji, Noriko M. [Biomedical Research Institute, National Institute of Industrial Science and Technology (AIST), 1-1-1 Higashi, Tsukuba 305-8566 (Japan); Yamamoto, Masayuki [Department of Medical Biochemistry, Tohoku University Graduate School of Medicine, 2-1 Seiryo-cho, Aoba-ku, Sendai 980-8575 (Japan); Nishihara, Shoko, E-mail: shoko@soka.ac.jp [Laboratory of Cell Biology, Department of Bioinformatics, Faculty of Engineering, Soka University, 1-236 Tangi-machi, Hachioji, Tokyo 192-8577 (Japan); Sato, Chikara, E-mail: ti-sato@aist.go.jp [Biomedical Research Institute, National Institute of Industrial Science and Technology (AIST), 1-1-1 Higashi, Tsukuba 305-8566 (Japan)

2014-08-01

Correlative light-electron microscopy of cells in a natural environment of aqueous liquid facilitates high-throughput observation of protein complex formation. ASEM allows the inverted SEM to observe the wet sample from below, while an optical microscope observes it from above quasi-simultaneously. The disposable ASEM dish with a silicon nitride (SiN) film window can be coated variously to realize the primary-culture of substrate-sensitive cells in a few milliliters of culture medium in a stable incubator environment. Neuron differentiation, neural networking, proplatelet-formation and phagocytosis were captured by optical or fluorescence microscopy, and imaged at high resolution by gold-labeled immuno-ASEM with/without metal staining. Fas expression on the cell surface was visualized, correlated to the spatial distribution of F-actin. Axonal partitioning was studied using primary-culture neurons, and presynaptic induction by GluRδ2-N-terminus-linked fluorescent magnetic beads was correlated to the presynaptic-marker Bassoon. Further, megakaryocytes secreting proplatelets were captured, and P-selectins with adherence activity were localized to some of the granules present by immuno-ASEM. The phagocytosis of lactic acid bacteria by dendritic cells was also imaged. Based on these studies, ASEM correlative microscopy promises to allow the study of various mesoscopic-scale dynamics in the near future. - Highlights: • In situ correlative light electron microscopy of samples in open solution by ASEM. • Primary cultures for in-solution CLEM by developing SiN-film coating methods • First visualization of fluorescent magnetic beads in aqueous solution by CLEM. • Presynaptic induction of neurons by GluRδ2-N-terminus-coated beads studied by CLEM. • Axonal partitioning, bacterial phagocytosis, platelet formation imaged by CLEM.

Implementation of PSF engineering in high-resolution 3D microscopy imaging with a LCoS (reflective) SLM

Science.gov (United States)

King, Sharon V.; Doblas, Ana; Patwary, Nurmohammed; Saavedra, Genaro; Martínez-Corral, Manuel; Preza, Chrysanthe

2014-03-01

Wavefront coding techniques are currently used to engineer unique point spread functions (PSFs) that enhance existing microscope modalities or create new ones. Previous work in this field demonstrated that simulated intensity PSFs encoded with a generalized cubic phase mask (GCPM) are invariant to spherical aberration or misfocus; dependent on parameter selection. Additional work demonstrated that simulated PSFs encoded with a squared cubic phase mask (SQUBIC) produce a depth invariant focal spot for application in confocal scanning microscopy. Implementation of PSF engineering theory with a liquid crystal on silicon (LCoS) spatial light modulator (SLM) enables validation of WFC phase mask designs and parameters by manipulating optical wavefront properties with a programmable diffractive element. To validate and investigate parameters of the GCPM and SQUBIC WFC masks, we implemented PSF engineering in an upright microscope modified with a dual camera port and a LCoS SLM. We present measured WFC PSFs and compare them to simulated PSFs through analysis of their effect on the microscope imaging system properties. Experimentally acquired PSFs show the same intensity distribution as simulation for the GCPM phase mask, the SQUBIC-mask and the well-known and characterized cubic-phase mask (CPM), first applied to high NA microscopy by Arnison et al.10, for extending depth of field. These measurements provide experimental validation of new WFC masks and demonstrate the use of the LCoS SLM as a WFC design tool. Although efficiency improvements are needed, this application of LCoS technology renders the microscope capable of switching among multiple WFC modes.

Out-of-plane reflection and refraction of light by anisotropic optical antenna metasurfaces with phase discontinuities.

Science.gov (United States)

Aieta, Francesco; Genevet, Patrice; Yu, Nanfang; Kats, Mikhail A; Gaburro, Zeno; Capasso, Federico

2012-03-14

Experiments on ultrathin anisotropic arrays of subwavelength optical antennas display out-of-plane refraction. A powerful three-dimensional (3D) extension of the recently demonstrated generalized laws of refraction and reflection shows that the interface imparts a tangential wavevector to the incident light leading to anomalous beams, which in general are noncoplanar with the incident beam. The refracted beam direction can be controlled by varying the angle between the plane of incidence and the antenna array. © 2012 American Chemical Society

Multiphoton Microscopy for Ophthalmic Imaging

Directory of Open Access Journals (Sweden)

Emily A. Gibson

2011-01-01

Full Text Available We review multiphoton microscopy (MPM including two-photon autofluorescence (2PAF, second harmonic generation (SHG, third harmonic generation (THG, fluorescence lifetime (FLIM, and coherent anti-Stokes Raman Scattering (CARS with relevance to clinical applications in ophthalmology. The different imaging modalities are discussed highlighting the particular strength that each has for functional tissue imaging. MPM is compared with current clinical ophthalmological imaging techniques such as reflectance confocal microscopy, optical coherence tomography, and fluorescence imaging. In addition, we discuss the future prospects for MPM in disease detection and clinical monitoring of disease progression, understanding fundamental disease mechanisms, and real-time monitoring of drug delivery.

Signal-enhancement reflective pulse oximeter with Fresnel lens

Science.gov (United States)

Chung, Shuang-Chao; Sun, Ching-Cherng

2016-09-01

In this paper, a new reflective pulse oximeter is proposed and demonstrated with implanting a Fresnel lens, which enhances the reflected signal. An optical simulation model incorporated with human skin characteristics is presented to evaluate the capability of the Fresnel lens. In addition, the distance between the light emitting diode and the photodiode is optimized. Compared with the other reflective oximeters, the reflected signal light detected by the photodiode is enhanced to more than 140%.

Quantitative localization of (/sup 3/H)TCP binding in rat brain by light microscopy autoradiography

Energy Technology Data Exchange (ETDEWEB)

Sircar, R; Zukin, S R

1985-09-30

The anatomical localization of phencyclidine (PCP)/sigma-opiate receptors in rat brain was determined by quantitative light microscopy autoradiography using the new ligand N-(1-(2-thienyl) cyclohexyl(/sup 3/H) piperidine ((/sup 3/H)TCP). TCP is a potent analog of PCP which possesses a higher affinity for PCP/sigma-opiate receptor than does PCP itself. The highest level of (/sup 3/H)TCP binding was detected in the hippocampus. Intermediate levels were found in frontal cortex, striatum, amygdala and cerebellum. Specific (/sup 3/H)TCP binding was undetectable in anterior commissure and corpus callosum. The distribution pattern of (/sup 3/H)TCP binding sites is similar to the pattern obtained with (/sup 3/H)PCP but more sharply defined. On the basis of its greater potency and specificity, (/sup 3/H)TCP may prove superior to (/sup 3/H)PCP as a molecular probe for the study of brain sigma opiate/phencyclidine receptors. 13 refs.; 1 figure; 1 table.

A fiberoptic reflection oximeter.

Science.gov (United States)

Landsman, M L; Knop, N; Kwant, G; Mook, G A; Zijlstra, W G

1978-03-20

A catheter tip oximeter is described consisting of a cardiac catheter containing optical fibers, and incandescent light source, a light detection unit and a processing unit. Half of the optical fibers guide the light to the blood at the tip of the catheter, the other half the backscattered (reflected) light to the detection unit. The detection unit contains a dichroic mirror, transmitting most of the light with lambda less than 800 nm and reflecting most of the light with lambda greater than 900 nm, thus splitting the light into two beams. These pass through interference filters with nominal wavelengths of 640 and 920 nm respectively, and are focused on silicium barrier layer photocells. The photocell signals are amplified and fed into a divider giving the ratio of measuring (R640) and compensating (R920) photocell output. The relationship between log R640/R920 and oxygen saturation is represented by a slightly curved line. The relation may be linearized by subtracting a constant voltage from the divided output before taking the logarithm. The slope of the calibration line is dependent on the total haemoglobin concentration. Nonetheless an average calibration line can be used between 70 and 100% oxygen saturation. For 78 measurements of pig blood samples in this range (haemoglobin concentration between 96 and 161 g.1(-1)), the standard deviation of the difference between the fiberoptic oximeter and a Radiometer OSM1 oxygen saturation meter was 1.9% saturation, for 152 samples over the entire saturation range the standard deviation of the difference was 3.1% saturation. The influence of the flow velocity of blood on the light reflection depends on wavelength as well as on oxygen saturation. Therefore, complete compensation for the flow effect is not possible by simple means.

Single-wavelength functional photoacoustic microscopy in biological tissue.

Science.gov (United States)

Danielli, Amos; Favazza, Christopher P; Maslov, Konstantin; Wang, Lihong V

2011-03-01

Recently, we developed a reflection-mode relaxation photoacoustic microscope, based on saturation intensity, to measure picosecond relaxation times using a nanosecond laser. Here, using the different relaxation times of oxygenated and deoxygenated hemoglobin molecules, both possessing extremely low fluorescence quantum yields, the oxygen saturation was quantified in vivo with single-wavelength photoacoustic microscopy. All previous functional photoacoustic microscopy measurements required imaging with multiple-laser-wavelength measurements to quantify oxygen saturation. Eliminating the need for multiwavelength measurements removes the influence of spectral properties on oxygenation calculations and improves the portability and cost-effectiveness of functional or molecular photoacoustic microscopy.

Deep Learning-Based Banknote Fitness Classification Using the Reflection Images by a Visible-Light One-Dimensional Line Image Sensor

Directory of Open Access Journals (Sweden)

Tuyen Danh Pham

2018-02-01

Full Text Available In automatic paper currency sorting, fitness classification is a technique that assesses the quality of banknotes to determine whether a banknote is suitable for recirculation or should be replaced. Studies on using visible-light reflection images of banknotes for evaluating their usability have been reported. However, most of them were conducted under the assumption that the denomination and input direction of the banknote are predetermined. In other words, a pre-classification of the type of input banknote is required. To address this problem, we proposed a deep learning-based fitness-classification method that recognizes the fitness level of a banknote regardless of the denomination and input direction of the banknote to the system, using the reflection images of banknotes by visible-light one-dimensional line image sensor and a convolutional neural network (CNN. Experimental results on the banknote image databases of the Korean won (KRW and the Indian rupee (INR with three fitness levels, and the Unites States dollar (USD with two fitness levels, showed that our method gives better classification accuracy than other methods.

Deep Learning-Based Banknote Fitness Classification Using the Reflection Images by a Visible-Light One-Dimensional Line Image Sensor.

Science.gov (United States)

Pham, Tuyen Danh; Nguyen, Dat Tien; Kim, Wan; Park, Sung Ho; Park, Kang Ryoung

2018-02-06

In automatic paper currency sorting, fitness classification is a technique that assesses the quality of banknotes to determine whether a banknote is suitable for recirculation or should be replaced. Studies on using visible-light reflection images of banknotes for evaluating their usability have been reported. However, most of them were conducted under the assumption that the denomination and input direction of the banknote are predetermined. In other words, a pre-classification of the type of input banknote is required. To address this problem, we proposed a deep learning-based fitness-classification method that recognizes the fitness level of a banknote regardless of the denomination and input direction of the banknote to the system, using the reflection images of banknotes by visible-light one-dimensional line image sensor and a convolutional neural network (CNN). Experimental results on the banknote image databases of the Korean won (KRW) and the Indian rupee (INR) with three fitness levels, and the Unites States dollar (USD) with two fitness levels, showed that our method gives better classification accuracy than other methods.

X-ray microscopy and spectromicroscopy - tools for environmental studies

International Nuclear Information System (INIS)

Thieme, J.

2002-01-01

Full text: X-ray microscopy achieves a much higher resolution than light microscopy. This is due to the much shorter wavelength of X-rays compared to visible light. The smallest structures that can be seen in an X-ray microscope at present are about 20 nm in size. X-ray microscopy is also capable of imaging specimens directly in aqueous media. By choosing the wavelength of the X-radiation appropriately, it is possible to perform spectromicroscopy studies. Comprising, it is a tool very well suited to study colloidal structures in the environment. X-ray microscopy can be performed with two types of instruments. An X-ray microscope quickly takes high-resolution images of objects to be studied, whereas a scanning X-ray microscope is an analytical instrument for spectromicroscopy. Here, an object can be imaged using energies above and below the absorption edge of an element. Dividing both images gives rise to a map of the distribution of this element in the sample. Using near-edge resonances it is possible to conduct NEXAFS studies. As the X-ray energy is raised to match the absorption edge resonances are found, that reflect the chemical bonding state of the element. Therefore, it is possible with X-ray microscopy to combine high spatial resolution with high spectral resolution for studies of colloidal structures. The Institute of X-ray physics builds up an X-ray microscopy beamline at the electron storage ring BESSY II with both, an X-ray microscope and a scanning X-ray microscope. The status of this beamline will be presented in this talk. Colloidal structures play an important role in the environment. Due to their surface activity they are involved in various processes. Substances can be bound and immobilized or transported, colloids can attach to microorganisms building up microhabitats, and organic substances as humics can flocculate due the interaction with metals. A great variety of colloidal structures from the environment have been studied using X

Method of Determining Reflections of Light

DEFF Research Database (Denmark)

2017-01-01

A method of filtering glints by processing an image of a user's cornea to obtain coordinates of desired glints from a configuration of light sources, comprising processing an image, in a first image space, of a user's cornea to determine coordinates of respective multiple positions of glints...

Nevomelanocytic atypia detection by in vivo reflectance confocal microscopy

Directory of Open Access Journals (Sweden)

Ingrida Vaišnorienė

2014-01-01

Conclusions: Nevus with histopathologically confirmed nevomelanocytic atypia (dysplastic nevus could not be distinguished from nevus without atypia using analyzed in vivo RCM features of melanocytic atypia. More accurate diagnostics by means of in vivo RCM needs further investigation on reflectance of single and nested cutaneous melanocytes in benign and malignant skin lesions.

Imaging bacterial spores by soft-x-ray microscopy

International Nuclear Information System (INIS)

Stead, A.D.; Ford, T.W.; Judge, J.

1997-01-01

Bacterial spores are able to survive dehydration, but neither the physiological nor structural basis of this have been fully elucidated. Furthermore, once hydrated, spores often require activation before they will germinate. Several treatments can be used to activate spores, but in the case of Bacillus subtlis the most effective is heat treatment. The physiological mechanism associated with activation is also not understood, but some workers suggest that the loss of calcium from the spores may be critical. However, just prior to germination, the spores change from being phase bright to phase dark when viewed by light microscopy. Imaging spores by soft x-ray microscopy is possible without fixation. Thus, in contrast to electron microscopy, it is possible to compare the structure of dehydrated and hydrated spores in a manner not possible previously. A further advantage is that it is possible to monitor individual spores by phase contrast light microscopy immediately prior to imaging with soft x-rays; whereas, with both electron microscopy and biochemical studies, it is a population of spores being studied without knowledge of the phase characteristics of individual spores. This study has therefore tried to compare dehydrated and hydrated spores and to determine if there is a mass loss from individual spores as they pass the transition from being phase bright to phase dark

Imaging bacterial spores by soft-x-ray microscopy

Energy Technology Data Exchange (ETDEWEB)

Stead, A.D.; Ford, T.W. [Univ. of London, Surrey (United Kingdom); Judge, J. [Unilever plc, Sharnbrook (United Kingdom)] [and others

1997-04-01

Bacterial spores are able to survive dehydration, but neither the physiological nor structural basis of this have been fully elucidated. Furthermore, once hydrated, spores often require activation before they will germinate. Several treatments can be used to activate spores, but in the case of Bacillus subtlis the most effective is heat treatment. The physiological mechanism associated with activation is also not understood, but some workers suggest that the loss of calcium from the spores may be critical. However, just prior to germination, the spores change from being phase bright to phase dark when viewed by light microscopy. Imaging spores by soft x-ray microscopy is possible without fixation. Thus, in contrast to electron microscopy, it is possible to compare the structure of dehydrated and hydrated spores in a manner not possible previously. A further advantage is that it is possible to monitor individual spores by phase contrast light microscopy immediately prior to imaging with soft x-rays; whereas, with both electron microscopy and biochemical studies, it is a population of spores being studied without knowledge of the phase characteristics of individual spores. This study has therefore tried to compare dehydrated and hydrated spores and to determine if there is a mass loss from individual spores as they pass the transition from being phase bright to phase dark.

Lighting up medicine with lasers

International Nuclear Information System (INIS)

Durrani, M.

2004-01-01

If you have ever shone a torch onto the back of your hand, you will know that your palm glows red. Haemoglobin in the blood absorbs almost all visible radiation at wavelengths below about 600nm, letting only red light pass through. Disappointingly, however, bones and other anatomical structures are impossible to see. Light is scattered so strongly by soft tissue that even a parallel beam becomes diffuse after travelling just a millimetre into the skin. Despite this superficial drawback, light is widely used in modern medicine. It can be reflected, refracted, absorbed, emitted and amplified. Light can be used to detect tumours, to take pictures of your stomach, or - in laser form - to remove cancerous tissue. Light in the 700-1300 nm range, known as the sup t herapeutic window sup , yields molecular information about fat, water and lipids, as well as the oxygenation of haemoglobin. For some, the holy grail in biomedical optics is a portable Star Trek style device that latter-day Dr McCoys can shine over patients to provide a diagnosis and therapy all in one. Optical scientists and engineers have developed a vast range of sophisticated light-based techniques for medical applications. But one of the most common biomedical research tools remains the optical microscope, which is almost the only way of studying biological processes in living tissue at the sub-micron cellular level. Although it might seem rather old-fashioned, microscopy has recently returned to the front line of biological research. In the August issue of Physics World Matin Durrani describes the latest developments in the field of biomedical optics. (U.K.)

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

Specific features of diffuse reflection of human face skin for laser and non-laser sources of visible and near-IR light

International Nuclear Information System (INIS)

Dolotov, L E; Sinichkin, Yu P; Tuchin, Valerii V; Al'tshuler, G B; Yaroslavskii, I V

2011-01-01

The specific features of diffuse reflection from different areas of human face skin for laser and non-laser sources of visible and near-IR light have been investigated to localise the closed-eye (eyelid) region. In the visible spectral range the reflection from the eyelid skin surface can be differentiated by measuring the slope of the spectral dependence of the effective optical density of skin in the wavelength range from 650 to 700nm. In the near-IR spectral range the reflectances of the skin surface at certain wavelengths, normalised to the forehead skin reflectance, can be used as a criterion for differentiating the eyelid skin. In this case, a maximum discrimination is obtained when measuring the skin reflectances at laser wavelengths of 1310 and 1470nm, which correspond to the spectral ranges of maximum and minimum water absorption. (optical technologies in biophysics and medicine)

Fluorescence microscopy for the characterization of structural integrity

Science.gov (United States)

Street, Kenneth W.; Leonhardt, Todd A.

1991-01-01

The absorption characteristics of light and the optical technique of fluorescence microscopy for enhancing metallographic interpretation are presented. Characterization of thermally sprayed coatings by optical microscopy suffers because of the tendency for misidentification of the microstructure produced by metallographic preparation. Gray scale, in bright field microscopy, is frequently the only means of differentiating the actual structural details of porosity, cracking, and debonding of coatings. Fluorescence microscopy is a technique that helps to distinguish the artifacts of metallographic preparation (pullout, cracking, debonding) from the microstructure of the specimen by color contrasting structural differences. Alternative instrumentation and the use of other dye systems are also discussed. The combination of epoxy vacuum infiltration with fluorescence microscopy to verify microstructural defects is an effective means to characterize advanced materials and to assess structural integrity.

Reflection technique for thermal mapping of semiconductors

Science.gov (United States)

Walter, Martin J.

1989-06-20

Semiconductors may be optically tested for their temperatures by illuminating them with tunable monochromatic electromagnetic radiation and observing the light reflected off of them. A transition point will occur when the wavelength of the light corresponds with the actual band gap energy of the semiconductor. At the transition point, the image of the semiconductor will appreciably darken as the light is transmitted through it, rather than being reflected off of it. The wavelength of the light at the transition point corresponds to the actual band gap energy and the actual temperature of the semiconductor.

Reflection measurements for luminescent powders

Science.gov (United States)

Kroon, R. E.

2018-04-01

Luminescent materials are useful in applications varying from lighting and display technologies to document security features and medical research, amongst many others. Measurement of the excitation range is an important consideration, and absorption bands are often determined from a decrease in the measured diffuse reflectance of the material using a ultraviolet-visible (UV-vis) spectrophotometer with an integrating sphere. Such a system may provide questionable results when used to measure the reflectance of a luminescence material, which is demonstrated for a Tb doped silica phosphor, because the system cannot differentiate between the reflected light and luminescence. It is shown that more reliable results are achieved for this phosphor by measuring the reflectance using a synchronous zero-offset scan in a fluorescence spectrometer equipped with an integrating sphere. This method is therefore recommended instead of traditional reflectance measurements using a UV-vis spectrophotometer for luminescent powders.

Recognition of Banknote Fitness Based on a Fuzzy System Using Visible Light Reflection and Near-infrared Light Transmission Images.

Science.gov (United States)

Kwon, Seung Yong; Pham, Tuyen Danh; Park, Kang Ryoung; Jeong, Dae Sik; Yoon, Sungsoo

2016-06-11

Fitness classification is a technique to assess the quality of banknotes in order to determine whether they are usable. Banknote classification techniques are useful in preventing problems that arise from the circulation of substandard banknotes (such as recognition failures, or bill jams in automated teller machines (ATMs) or bank counting machines). By and large, fitness classification continues to be carried out by humans, and this can cause the problem of varying fitness classifications for the same bill by different evaluators, and requires a lot of time. To address these problems, this study proposes a fuzzy system-based method that can reduce the processing time needed for fitness classification, and can determine the fitness of banknotes through an objective, systematic method rather than subjective judgment. Our algorithm was an implementation to actual banknote counting machine. Based on the results of tests on 3856 banknotes in United States currency (USD), 3956 in Korean currency (KRW), and 2300 banknotes in Indian currency (INR) using visible light reflection (VR) and near-infrared light transmission (NIRT) imaging, the proposed method was found to yield higher accuracy than prevalent banknote fitness classification methods. Moreover, it was confirmed that the proposed algorithm can operate in real time, not only in a normal PC environment, but also in an embedded system environment of a banknote counting machine.

Recognition of Banknote Fitness Based on a Fuzzy System Using Visible Light Reflection and Near-infrared Light Transmission Images

Directory of Open Access Journals (Sweden)

Seung Yong Kwon

2016-06-01

Full Text Available Fitness classification is a technique to assess the quality of banknotes in order to determine whether they are usable. Banknote classification techniques are useful in preventing problems that arise from the circulation of substandard banknotes (such as recognition failures, or bill jams in automated teller machines (ATMs or bank counting machines. By and large, fitness classification continues to be carried out by humans, and this can cause the problem of varying fitness classifications for the same bill by different evaluators, and requires a lot of time. To address these problems, this study proposes a fuzzy system-based method that can reduce the processing time needed for fitness classification, and can determine the fitness of banknotes through an objective, systematic method rather than subjective judgment. Our algorithm was an implementation to actual banknote counting machine. Based on the results of tests on 3856 banknotes in United States currency (USD, 3956 in Korean currency (KRW, and 2300 banknotes in Indian currency (INR using visible light reflection (VR and near-infrared light transmission (NIRT imaging, the proposed method was found to yield higher accuracy than prevalent banknote fitness classification methods. Moreover, it was confirmed that the proposed algorithm can operate in real time, not only in a normal PC environment, but also in an embedded system environment of a banknote counting machine.

Comparison of two methods of preparation of tissue to study the internal anatomy of the delphacid Togosodes orizicolus with microscopy of electronic light

International Nuclear Information System (INIS)

Macaya-Lizano, A.V.; Pereira, R.; Espinoza, A.M.

1997-01-01

Two methods of embedding, sectioning and staining were developed to study the internal anatomy of delphacid plant hopper Tagosodes orizicolus, one of the most important plagues of rice in Latin America and the only vector of the white leaf tenuivirus (RHBV), using both light and electron microscopy. The paraffines-hematoxyline-eosin Y method allows color identification of tissues, for example purple for fat tissue, pink for muscles, yellow-brown for exocutile, while the resin-toluidine-blue method preserves better the ultrastructure but do not permit color identification. The information obtained by these procedures is complementary and the material can also be used for in situ studies by immuno microscopy, to assess the changes in cell ultrastructure and the localization and replication of the RHBV during its infection cycle in the insect vector. (author) [es

Reflected Light from Giant Planets in Habitable Zones: Tapping into the Power of the Cross-Correlation Function.

Science.gov (United States)

Martins, J H C; Santos, N C; Figueira, P; Melo, C

2016-11-01

The direct detection of reflected light from exoplanets is an excellent probe for the characterization of their atmospheres. The greatest challenge for this task is the low planet-to-star flux ratio, which even in the most favourable case is of the order of 10 -4 in the optical. This ratio decreases even more for planets in their host's habitable zone, typically lower than 10 -7 . To reach the signal-to-noise level required for such detections, we propose to unleash the power of the Cross Correlation Function in combination with the collecting power of next generation observing facilities. The technique we propose has already yielded positive results by detecting the reflected spectral signature of 51 Pegasi b (see Martins et al. 2015). In this work, we attempted to infer the number of hours required for the detection of several planets in their host's habitable zone using the aforementioned technique from theoretical EELT observations. Our results show that for 5 of the selected planets it should be possible to directly recover their reflected spectral signature.

Virtual Hematoxylin and Eosin Transillumination Microscopy Using Epi-Fluorescence Imaging.

Science.gov (United States)

Giacomelli, Michael G; Husvogt, Lennart; Vardeh, Hilde; Faulkner-Jones, Beverly E; Hornegger, Joachim; Connolly, James L; Fujimoto, James G

2016-01-01

We derive a physically realistic model for the generation of virtual transillumination, white light microscopy images using epi-fluorescence measurements from thick, unsectioned tissue. We demonstrate this technique by generating virtual transillumination H&E images of unsectioned human breast tissue from epi-fluorescence multiphoton microscopy data. The virtual transillumination algorithm is shown to enable improved contrast and color accuracy compared with previous color mapping methods. Finally, we present an open source implementation of the algorithm in OpenGL, enabling real-time GPU-based generation of virtual transillumination microscopy images using conventional fluorescence microscopy systems.

Wide angle light collection with ultralow reflection and super scattering by silicon micro-nanostructures for thin crystalline silicon solar cell applications

International Nuclear Information System (INIS)

Das, Sonali; Kundu, Avra; Saha, Hiranmay; Datta, Swapan K

2016-01-01

Conventional c-Si solar cells employ micron-sized pyramids for achieving reduced reflection (∼10%) and enhanced light trapping by multiple bounces (maximum 3) of the incident light. Alternatively, bio-mimetic, moth-eye sub-wavelength nanostructures offer broadband antireflection properties (∼3%) suitable for solar cell applications in the optical regime. However, such structures do not provide any advantage in the charge carrier extraction process as radial junctions cannot be formed in such sub-wavelength dimensions and they have high surface area causing increased charged carrier recombination. The choice of the geometry for achieving optimum photon–electron harvesting for solar applications is therefore very critical. Cross-fertilization of the conventional solar cell light-trapping techniques and the sub-wavelength nanostructures results in unique micro-nanostructures (structures having sub-wavelength dimensions as well as dimensions of the order of few microns) which provide advanced light management capabilities along with the ability of realizing radial junctions. It is seen that an ultralow reflection along with wide angle light collection is obtained which enables such structures to overcome the morning, evening and winter light losses in solar cells. Further, super-scattering in the structures offer enhanced light trapping not only in the structure itself but also in the substrate housing the structure. Ray and wave optics have been used to understand the optical benefits of the structures. It is seen that the aspect ratio of the structures plays the most significant role for achieving such light management capabilities, and efficiencies as high as 12% can be attained. Experiments have been carried out to fabricate a unique micro-nanomaze-like structure instead of a periodic array of micro-nanostructures with the help of nanosphere lithography and the MacEtch technique. It is seen that randomized micro-nanomaze geometry offers very good

Waves and grains reflections on light and learning

CERN Document Server

Silverman, Mark P

1998-01-01

Mark Silverman has seen light perform many wonders. From the marvel of seeing inside cloudy liquids as a result of his own cutting-edge research to reproducing and examining an unusual diffraction pattern first witnessed by Isaac Newton 300 years ago, he has studied aspects of light that have inspired and puzzled humans for hundreds of years. In this book, he draws on his many experiences as an optical and atomic physicist--and on his consummate skills as a teacher and writer about the mysteries of physics--to present a remarkable tour of the world of light. He explores theoretical, experimental, and historical themes, showing a keen eye for curious and neglected corners of the study of light and a fascination with the human side of scientific discovery. In the course of the book, he covers such questions as how it is possible to achieve magnifications of a millionfold without a single lens or mirror. He asks what all living things have in common that might one day allow the development of a "life-form scann...

Air-void embedded GaN-based light-emitting diodes grown on laser drilling patterned sapphire substrates

Energy Technology Data Exchange (ETDEWEB)

Liu, Hao; Li, Yufeng; Wang, Shuai; Feng, Lungang; Xiong, Han; Yun, Feng, E-mail: fyun2010@mail.xjtu.edu.cn [Key Laboratory of Physical Electronics and Devices of Ministry of Education and Shaanxi Provincial Key Laboratory of Photonics and Information Technology, Xi’an Jiaotong University, Xi’an, Shaanxi 710049 (China); Solid-State Lighting Engineering Research Center, Xi’an Jiaotong University, Xi’an, Shaanxi 710049 (China); Su, Xilin [Shaanxi Supernova Lighting Technology Co., Ltd., Xi’an, Shaanxi 710075 (China)

2016-07-15

Air-void structure was introduced in GaN-based blue light-emitting diodes (LED) with one-step growth on periodic laser drilling patterned sapphire substrate, which free of any photolithography or wet/dry etching process. The influence of filling factors (FF) of air-void on crystal quality and optical performance were investigate. Transmission electron microscopy images and micro-Raman spectroscopy indicated that the dislocation was bended and the partially compressed strain was released. When FF was 55.43%, compared with the LED structure grown on flat sapphire substrate, the incorporation of air-void was observed to reduce the compressed stress of ∼20% and the luminance intensity has improved by 128%. Together with the simulated reflection intensity enhancement by finite difference time-domain (FDTD) method, we attribute the enhanced optical performance to the combined contribution of strong back-side light reflection of air-void and better GaN epitaxial quality. This approach provides a simple replacement to the conventional air-void embedded LED process.

Integration of a high-NA light microscope in a scanning electron microscope.

Science.gov (United States)

Zonnevylle, A C; Van Tol, R F C; Liv, N; Narvaez, A C; Effting, A P J; Kruit, P; Hoogenboom, J P

2013-10-01

We present an integrated light-electron microscope in which an inverted high-NA objective lens is positioned inside a scanning electron microscope (SEM). The SEM objective lens and the light objective lens have a common axis and focal plane, allowing high-resolution optical microscopy and scanning electron microscopy on the same area of a sample simultaneously. Components for light illumination and detection can be mounted outside the vacuum, enabling flexibility in the construction of the light microscope. The light objective lens can be positioned underneath the SEM objective lens during operation for sub-10 μm alignment of the fields of view of the light and electron microscopes. We demonstrate in situ epifluorescence microscopy in the SEM with a numerical aperture of 1.4 using vacuum-compatible immersion oil. For a 40-nm-diameter fluorescent polymer nanoparticle, an intensity profile with a FWHM of 380 nm is measured whereas the SEM performance is uncompromised. The integrated instrument may offer new possibilities for correlative light and electron microscopy in the life sciences as well as in physics and chemistry. © 2013 The Authors Journal of Microscopy © 2013 Royal Microscopical Society.

Contributed Review: Review of integrated correlative light and electron microscopy

NARCIS (Netherlands)

Timmermans, Frank Jan; Otto, Cornelis

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

Fluorescence confocal microscopy for pathologists.

Science.gov (United States)

Ragazzi, Moira; Piana, Simonetta; Longo, Caterina; Castagnetti, Fabio; Foroni, Monica; Ferrari, Guglielmo; Gardini, Giorgio; Pellacani, Giovanni

2014-03-01

Confocal microscopy is a non-invasive method of optical imaging that may provide microscopic images of untreated tissue that correspond almost perfectly to hematoxylin- and eosin-stained slides. Nowadays, following two confocal imaging systems are available: (1) reflectance confocal microscopy, based on the natural differences in refractive indices of subcellular structures within the tissues; (2) fluorescence confocal microscopy, based on the use of fluorochromes, such as acridine orange, to increase the contrast epithelium-stroma. In clinical practice to date, confocal microscopy has been used with the goal of obviating the need for excision biopsies, thereby reducing the need for pathological examination. The aim of our study was to test fluorescence confocal microscopy on different types of surgical specimens, specifically breast, lymph node, thyroid, and colon. The confocal images were correlated to the corresponding histological sections in order to provide a morphologic parallel and to highlight current limitations and possible applications of this technology for surgical pathology practice. As a result, neoplastic tissues were easily distinguishable from normal structures and reactive processes such as fibrosis; the use of fluorescence enhanced contrast and image quality in confocal microscopy without compromising final histologic evaluation. Finally, the fluorescence confocal microscopy images of the adipose tissue were as accurate as those of conventional histology and were devoid of the frozen-section-related artefacts that can compromise intraoperative evaluation. Despite some limitations mainly related to black/white images, which require training in imaging interpretation, this study confirms that fluorescence confocal microscopy may represent an alternative to frozen sections in the assessment of margin status in selected settings or when the conservation of the specimen is crucial. This is the first study to employ fluorescent confocal microscopy on

Use of astronomy filters in fluorescence microscopy.

Science.gov (United States)

Piper, Jörg

2012-02-01

Monochrome astronomy filters are well suited for use as excitation or suppression filters in fluorescence microscopy. Because of their particular optical design, such filters can be combined with standard halogen light sources for excitation in many fluorescent probes. In this "low energy excitation," photobleaching (fading) or other irritations of native specimens are avoided. Photomicrographs can be taken from living motile fluorescent specimens also with a flash so that fluorescence images can be created free from indistinctness caused by movement. Special filter cubes or dichroic mirrors are not needed for our method. By use of suitable astronomy filters, fluorescence microscopy can be carried out with standard laboratory microscopes equipped with condensers for bright-field (BF) and dark-field (DF) illumination in transmitted light. In BF excitation, the background brightness can be modulated in tiny steps up to dark or black. Moreover, standard industry microscopes fitted with a vertical illuminator for examinations of opaque probes in DF or BF illumination based on incident light (wafer inspections, for instance) can also be used for excitation in epi-illumination when adequate astronomy filters are inserted as excitatory and suppression filters in the illuminating and imaging light path. In all variants, transmission bands can be modulated by transmission shift.

Preparation And Characterization of Cu-Fe/ TiO2 Photocatalyst for Visible Light Deep Desulfurization

International Nuclear Information System (INIS)

Hayyiratul Fatimah Mohd Zaid; Kait, C.F.; Mohamed Ibrahim Abdul Mutalib

2016-01-01

A photooxidative system for deep desulfurization of model diesel fuel was explored. Nanoparticles of anatase titania (TiO 2 ) were synthesized via sol-gel hydrothermal method. The TiO 2 was further modified with bimetallic Cu-Fe using wet-impregnation method followed by calcination process in order to extend the activity region of the photocatalyst to visible-light. A series of bimetallic 2.2 wt % Cu-Fe/ TiO 2 photocatalysts with different Cu:Fe mass compositions were characterized for their physical, chemical and optical properties using X-ray diffraction (XRD), field emission scanning electron microscopy (FESEM), high resolution transmission electron microscopy (HRTEM), diffuse reflectance UV-visible spectroscopy (DR-UV-Vis), Fourier Transform Infrared Spectroscopy (FTIR) and Brunauer-Emmet-Teller (BET) surface area analysis. The performance of the photocatalysts was evaluated for photooxidation of dibenzothiophene (DBT) as the sulfur species from model oil in the presence of hydrogen peroxide, H 2 O 2 under 500 W visible light illumination. The highest sulfur conversion of 82.36 % was observed for photocatalyst with 10:1Cu:Fe mass composition. (author)

Nonimaging light concentrator with uniform irradiance

Science.gov (United States)

Winston, Roland; Gee, Randy C.

2003-04-01

A nonimaging light concentrator system including a primary collector of light, an optical mixer disposed near the focal zone for collecting light from the primary collector, the optical mixer having a transparent entrance aperture, an internally reflective housing for substantially total internal reflection of light, a transparent exit aperture and an array of photovoltaic cells disposed near the transparent exit aperture.

Hewlett-Packard's Approaches to Full Color Reflective Displays

Science.gov (United States)

Gibson, Gary

2012-02-01

Reflective displays are desirable in applications requiring low power or daylight readability. However, commercial reflective displays are currently either monochrome or capable of only dim color gamuts. Low cost, high-quality color technology would be rapidly adopted in existing reflective display markets and would enable new solutions in areas such as retail pricing and outdoor digital signage. Technical breakthroughs are required to enable bright color gamuts at reasonable cost. Pixel architectures that rely on pure reflection from a single layer of side-by-side primary-color sub-pixels use only a fraction of the display area to reflect incident light of a given color and are, therefore, unacceptably dark. Reflective devices employing stacked color primaries offer the possibility of a somewhat brighter color gamut but can be more complex to manufacture. In this talk, we describe HP's successes in addressing these fundamental challenges and creating both high performance stacked-primary reflective color displays as well as inexpensive single layer prototypes that provide good color. Our stacked displays utilize a combination of careful light management techniques, proprietary high-contrast electro-optic shutters, and highly transparent active-matrix TFT arrays based on transparent metal oxides. They also offer the possibility of relatively low cost manufacturing through roll-to-roll processing on plastic webs. To create even lower cost color displays with acceptable brightness, we have developed means for utilizing photoluminescence to make more efficient use of ambient light in a single layer device. Existing reflective displays create a desired color by reflecting a portion of the incident spectrum while absorbing undesired wavelengths. We have developed methods for converting the otherwise-wasted absorbed light to desired wavelengths via tailored photoluminescent composites. Here we describe a single active layer prototype display that utilizes these materials

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.

High spatial resolution soft-x-ray microscopy

Energy Technology Data Exchange (ETDEWEB)

Meyer-Ilse, W.; Medecki, H.; Brown, J.T. [Ernest Orlando Lawrence Berkeley National Lab., CA (United States)] [and others

1997-04-01

A new soft x-ray microscope (XM-1) with high spatial resolution has been constructed by the Center for X-ray Optics. It uses bending magnet radiation from beamline 6.1 at the Advanced Light Source, and is used in a variety of projects and applications in the life and physical sciences. Most of these projects are ongoing. The instrument uses zone plate lenses and achieves a resolution of 43 nm, measured over 10% to 90% intensity with a knife edge test sample. X-ray microscopy permits the imaging of relatively thick samples, up to 10 {mu}m thick, in water. XM-1 has an easy to use interface, that utilizes visible light microscopy to precisely position and focus the specimen. The authors describe applications of this device in the biological sciences, as well as in studying industrial applications including structured polymer samples.

From Intensity Profile to Surface Normal: Photometric Stereo for Unknown Light Sources and Isotropic Reflectances.

Science.gov (United States)

Lu, Feng; Matsushita, Yasuyuki; Sato, Imari; Okabe, Takahiro; Sato, Yoichi

2015-10-01

We propose an uncalibrated photometric stereo method that works with general and unknown isotropic reflectances. Our method uses a pixel intensity profile, which is a sequence of radiance intensities recorded at a pixel under unknown varying directional illumination. We show that for general isotropic materials and uniformly distributed light directions, the geodesic distance between intensity profiles is linearly related to the angular difference of their corresponding surface normals, and that the intensity distribution of the intensity profile reveals reflectance properties. Based on these observations, we develop two methods for surface normal estimation; one for a general setting that uses only the recorded intensity profiles, the other for the case where a BRDF database is available while the exact BRDF of the target scene is still unknown. Quantitative and qualitative evaluations are conducted using both synthetic and real-world scenes, which show the state-of-the-art accuracy of smaller than 10 degree without using reference data and 5 degree with reference data for all 100 materials in MERL database.

A review of cellphone microscopy for disease detection.

Science.gov (United States)

Dendere, R; Myburg, N; Douglas, T S

2015-12-01

The expansion in global cellphone network coverage coupled with advances in cellphone imaging capabilities present an opportunity for the advancement of cellphone microscopy as a low-cost alternative to conventional microscopy for disease detection in resource-limited regions. The development of cellphone microscopy has also benefitted from the availability of low-cost miniature microscope components such as low-power light-emitting diodes and ball lenses. As a result, researchers are developing hardware and software techniques that would enable such microscopes to produce high-resolution, diagnostic-quality images. This approach may lead to more widespread delivery of diagnostic services in resource-limited areas where there is a shortage of the skilled labour required for conventional microscopy and where prevalence of infectious and other diseases is still high. In this paper, we review current techniques, clinical applications and challenges faced in the field of cellphone microscopy. © 2015 The Authors Journal of Microscopy © 2015 Royal Microscopical Society.

Effects of leaf age within growth stages of pepper and sorghum plants on leaf thickness, water, chlorophyll, and light reflectance. [in spectral vegetation discrimination

Science.gov (United States)

Gausman, H. W.; Cardenas, R.; Berumen, A.

1974-01-01

Pepper and sorghum plants (characterized by porous and compact leaf mesophylls, respectively) were used to study the influence of leaf age on light reflectance. Measurements were limited to the upper five nodal positions within each growth stage, since upper leaves make up most of the reflectance surfaces remotely sensed. The increase in leaf thickness and water content with increasing leaf age was taken into consideration, since each of these factors affects the reflectance as well as the selection of spectral wavelength intervals for optimum discrimination of vegetation.

Confocal Cornea Microscopy Detects Involvement of Corneal Nerve Fibers in a Patient with Light-Chain Amyloid Neuropathy Caused by Multiple Myeloma: A Case Report

Directory of Open Access Journals (Sweden)

Dietrich Sturm

2016-06-01

Full Text Available Changes in the subbasal corneal plexus detected by confocal cornea microscopy (CCM have been described for various types of neuropathy. An involvement of these nerves within light-chain (AL amyloid neuropathy (a rare cause of polyneuropathy has never been shown. Here, we report on a case of a patient suffering from neuropathy caused by AL amyloidosis and underlying multiple myeloma. Small-fiber damage was detected by CCM.

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

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

2015-12-17

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

Visualizing Ebolavirus Particles Using Single-Particle Interferometric Reflectance Imaging Sensor (SP-IRIS).

Science.gov (United States)

Carter, Erik P; Seymour, Elif Ç; Scherr, Steven M; Daaboul, George G; Freedman, David S; Selim Ünlü, M; Connor, John H

2017-01-01

This chapter describes an approach for the label-free imaging and quantification of intact Ebola virus (EBOV) and EBOV viruslike particles (VLPs) using a light microscopy technique. In this technique, individual virus particles are captured onto a silicon chip that has been printed with spots of virus-specific capture antibodies. These captured virions are then detected using an optical approach called interference reflectance imaging. This approach allows for the detection of each virus particle that is captured on an antibody spot and can resolve the filamentous structure of EBOV VLPs without the need for electron microscopy. Capture of VLPs and virions can be done from a variety of sample types ranging from tissue culture medium to blood. The technique also allows automated quantitative analysis of the number of virions captured. This can be used to identify the virus concentration in an unknown sample. In addition, this technique offers the opportunity to easily image virions captured from native solutions without the need for additional labeling approaches while offering a means of assessing the range of particle sizes and morphologies in a quantitative manner.

Bi-directional reflectance distribution function of a tungsten block for ITER divertor

International Nuclear Information System (INIS)

Iwamae, Atsushi; Ogawa, Hiroaki; Sugie, Tatsuo; Kusama, Yoshinori

2012-02-01

In order to investigate reflection properties on plasma-facing material in ITER, the bi-directional reflectance distribution function (BRDF) of a tungsten block sample has been measured. On the machining surface of the block, one-directional machining lines are engraved. Two laser diodes λ652 nm and λ473 nm were used to simulate H α and H β emissions, respectively. The reflected light is affected by the machining surface. The reflected light traces an arc when the incident light is injected in the parallel direction to the engraved line. On the other hand the reflected light traces a line shape when the incident light is injected in the perpendicular direction to the engraved lines. Ray tracing simulation qualitatively explains the experimental results. (author)

Virtual Hematoxylin and Eosin Transillumination Microscopy Using Epi-Fluorescence Imaging.

Directory of Open Access Journals (Sweden)

Michael G Giacomelli

Full Text Available We derive a physically realistic model for the generation of virtual transillumination, white light microscopy images using epi-fluorescence measurements from thick, unsectioned tissue. We demonstrate this technique by generating virtual transillumination H&E images of unsectioned human breast tissue from epi-fluorescence multiphoton microscopy data. The virtual transillumination algorithm is shown to enable improved contrast and color accuracy compared with previous color mapping methods. Finally, we present an open source implementation of the algorithm in OpenGL, enabling real-time GPU-based generation of virtual transillumination microscopy images using conventional fluorescence microscopy systems.

Polarized light microscopy for 3-dimensional mapping of collagen fiber architecture in ocular tissues.

Science.gov (United States)

Yang, Bin; Jan, Ning-Jiun; Brazile, Bryn; Voorhees, Andrew; Lathrop, Kira L; Sigal, Ian A

2018-04-06

Collagen fibers play a central role in normal eye mechanics and pathology. In ocular tissues, collagen fibers exhibit a complex 3-dimensional (3D) fiber orientation, with both in-plane (IP) and out-of-plane (OP) orientations. Imaging techniques traditionally applied to the study of ocular tissues only quantify IP fiber orientation, providing little information on OP fiber orientation. Accurate description of the complex 3D fiber microstructures of the eye requires quantifying full 3D fiber orientation. Herein, we present 3dPLM, a technique based on polarized light microscopy developed to quantify both IP and OP collagen fiber orientations of ocular tissues. The performance of 3dPLM was examined by simulation and experimental verification and validation. The experiments demonstrated an excellent agreement between extracted and true 3D fiber orientation. Both IP and OP fiber orientations can be extracted from the sclera and the cornea, providing previously unavailable quantitative 3D measures and insight into the tissue microarchitecture. Together, the results demonstrate that 3dPLM is a powerful imaging technique for the analysis of ocular tissues. © 2018 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

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.

Acute radiation nephritis. Light and electron microscopic observations

International Nuclear Information System (INIS)

Kapur, S.; Chandra, R.; Antonovych, T.

1977-01-01

Light and electron microscopy were used to observe acute radiation nephritis. By light microscopy the changes were of fibrinoid necrosis of the arteries and arterioles with segmental necrosis of the glomerular tufts. By electron microscopy the endocapillary cells reacted by hypertrophy and hyperplasia with increase in cytoplasmic organelles. In addition, disruption of endothelial and epithelial cells from the basement membranes were seen. It is concluded that the electron microscopic changes were unique and may be helpful in differentiating the necrotizing glomerulitis seen in other conditions, especially malignant hypertension

Subdiffraction Multicolor Imaging of the Nuclear Periphery with 3D Structured Illumination Microscopy

Science.gov (United States)

Schermelleh, Lothar; Carlton, Peter M.; Haase, Sebastian; Shao, Lin; Winoto, Lukman; Kner, Peter; Burke, Brian; Cardoso, M. Cristina; Agard, David A.; Gustafsson, Mats G. L.; Leonhardt, Heinrich; Sedat, John W.

2010-01-01

Fluorescence light microscopy allows multicolor visualization of cellular components with high specificity, but its utility has until recently been constrained by the intrinsic limit of spatial resolution. We applied three-dimensional structured illumination microscopy (3D-SIM) to circumvent this limit and to study the mammalian nucleus. By simultaneously imaging chromatin, nuclear lamina, and the nuclear pore complex (NPC), we observed several features that escape detection by conventional microscopy. We could resolve single NPCs that colocalized with channels in the lamin network and peripheral heterochromatin. We could differentially localize distinct NPC components and detect double-layered invaginations of the nuclear envelope in prophase as previously seen only by electron microscopy. Multicolor 3D-SIM opens new and facile possibilities to analyze subcellular structures beyond the diffraction limit of the emitted light. PMID:18535242

The differential equation of an arbitrary reflecting surface

Science.gov (United States)

Melka, Richard F.; Berrettini, Vincent D.; Yousif, Hashim A.

2018-05-01

A differential equation describing the reflection of a light ray incident upon an arbitrary reflecting surface is obtained using the law of reflection. The derived equation is written in terms of a parameter and the value of this parameter determines the nature of the reflecting surface. Under various parametric constraints, the solution of the differential equation leads to the various conic surfaces but is not generally solvable. In addition, the dynamics of the light reflections from the conic surfaces are executed in the Mathematica software. Our derivation is the converse of the traditional approach and our analysis assumes a relation between the object distance and the image distance. This leads to the differential equation of the reflecting surface.

The ring structure and organization of light harvesting 2 complexes in a reconstituted lipid bilayer, resolved by atomic force microscopy.

Science.gov (United States)

Stamouli, Amalia; Kafi, Sidig; Klein, Dionne C G; Oosterkamp, Tjerk H; Frenken, Joost W M; Cogdell, Richard J; Aartsma, Thijs J

2003-04-01

The main function of the transmembrane light-harvesting complexes in photosynthetic organisms is the absorption of a light quantum and its subsequent rapid transfer to a reaction center where a charge separation occurs. A combination of freeze-thaw and dialysis methods were used to reconstitute the detergent-solubilized Light Harvesting 2 complex (LH2) of the purple bacterium Rhodopseudomonas acidophila strain 10050 into preformed egg phosphatidylcholine liposomes, without the need for extra chemical agents. The LH2-containing liposomes opened up to a flat bilayer, which were imaged with tapping and contact mode atomic force microscopy under ambient and physiological conditions, respectively. The LH2 complexes were packed in quasicrystalline domains. The endoplasmic and periplasmic sides of the LH2 complexes could be distinguished by the difference in height of the protrusions from the lipid bilayer. The results indicate that the complexes entered in intact liposomes. In addition, it was observed that the most hydrophilic side, the periplasmic, enters first in the membrane. In contact mode the molecular structure of the periplasmic side of the transmembrane pigment-protein complex was observed. Using Föster's theory for describing the distance dependent energy transfer, we estimate the dipole strength for energy transfer between two neighboring LH2s, based on the architecture of the imaged unit cell.