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Sample records for microscopy avaliacao das

  1. Fluorescence Microscopy

    PubMed Central

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

    2016-01-01

    Fluorescence microscopy is a major tool with which to monitor cell physiology. Although the concepts of fluorescence and its optical separation using filters remain similar, microscope design varies with the aim of increasing image contrast and spatial resolution. The basics of wide-field microscopy are outlined to emphasize the selection, advantages, and correct use of laser scanning confocal microscopy, two-photon microscopy, scanning disk confocal microscopy, total internal reflection, and super-resolution microscopy. In addition, the principles of how these microscopes form images are reviewed to appreciate their capabilities, limitations, and constraints for operation. PMID:25275114

  2. DAS performance analysis

    SciTech Connect

    Bates, G.; Bodine, S.; Carroll, T.; Keller, M.

    1984-02-01

    This report begins with an overview of the Data Acquisition System (DAS), which supports several of PPPL's experimental devices. Performance measurements which were taken on DAS and the tools used to make them are then described.

  3. Analytical Microscopy

    SciTech Connect

    Not Available

    2006-06-01

    In the Analytical Microscopy group, within the National Center for Photovoltaic's Measurements and Characterization Division, we combine two complementary areas of analytical microscopy--electron microscopy and proximal-probe techniques--and use a variety of state-of-the-art imaging and analytical tools. We also design and build custom instrumentation and develop novel techniques that provide unique capabilities for studying materials and devices. In our work, we collaborate with you to solve materials- and device-related R&D problems. This sheet summarizes the uses and features of four major tools: transmission electron microscopy, scanning electron microscopy, the dual-beam focused-ion-beam workstation, and scanning probe microscopy.

  4. Correlative Microscopy

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Microscopy and Imaging offers many opportunities to collaborate and cooperate with scientists in many different fields nationally and internationally. Images have proven to be very important components in basic research, product development and understanding structure/function relationships in addit...

  5. Correlative microscopy.

    PubMed

    Loussert Fonta, Céline; Humbel, Bruno M

    2015-09-01

    In recent years correlative microscopy, combining the power and advantages of different imaging system, e.g., light, electrons, X-ray, NMR, etc., has become an important tool for biomedical research. Among all the possible combinations of techniques, light and electron microscopy, have made an especially big step forward and are being implemented in more and more research labs. Electron microscopy profits from the high spatial resolution, the direct recognition of the cellular ultrastructure and identification of the organelles. It, however, has two severe limitations: the restricted field of view and the fact that no live imaging can be done. On the other hand light microscopy has the advantage of live imaging, following a fluorescently tagged molecule in real time and at lower magnifications the large field of view facilitates the identification and location of sparse individual cells in a large context, e.g., tissue. The combination of these two imaging techniques appears to be a valuable approach to dissect biological events at a submicrometer level. Light microscopy can be used to follow a labelled protein of interest, or a visible organelle such as mitochondria, in time, then the sample is fixed and the exactly same region is investigated by electron microscopy. The time resolution is dependent on the speed of penetration and fixation when chemical fixatives are used and on the reaction time of the operator for cryo-fixation. Light microscopy can also be used to identify cells of interest, e.g., a special cell type in tissue or cells that have been modified by either transfections or RNAi, in a large population of non-modified cells. A further application is to find fluorescence labels in cells on a large section to reduce searching time in the electron microscope. Multiple fluorescence labelling of a series of sections can be correlated with the ultrastructure of the individual sections to get 3D information of the distribution of the marked proteins: array

  6. Expansion Microscopy

    PubMed Central

    Chen, Fei; Tillberg, Paul W.; Boyden, Edward S.

    2014-01-01

    In optical microscopy, fine structural details are resolved by using refraction to magnify images of a specimen. Here we report the discovery that, by synthesizing a swellable polymer network within a specimen, it can be physically expanded, resulting in physical magnification. By covalently anchoring specific labels located within the specimen directly to the polymer network, labels spaced closer than the optical diffraction limit can be isotropically separated and optically resolved, a process we call expansion microscopy (ExM). Thus, this process can be used to perform scalable super-resolution microscopy with diffraction-limited microscopes. We demonstrate ExM with effective ~70 nm lateral resolution in both cultured cells and brain tissue, performing three-color super-resolution imaging of ~107 μm3 of the mouse hippocampus with a conventional confocal microscope. PMID:25592419

  7. Positron microscopy

    SciTech Connect

    Hulett, L.D. Jr.; Xu, J.

    1995-02-01

    The negative work function property that some materials have for positrons make possible the development of positron reemission microscopy (PRM). Because of the low energies with which the positrons are emitted, some unique applications, such as the imaging of defects, can be made. The history of the concept of PRM, and its present state of development will be reviewed. The potential of positron microprobe techniques will be discussed also.

  8. DAS User Manual

    NASA Astrophysics Data System (ADS)

    Østensen, Roy

    2008-10-01

    The HELAS Database for AsteroSeismology (DAS) is one of the deliverables of the Work Package NA5: Asteroseismology of the European Coordination Action in Helio- and Asteroseismology (HELAS). The DAS aims to provide easy access to publicly available asteroseismological timeseries data, both photometric and spectroscopic. In particular, the DAS and the HELAS software package FAMIAS are ideally suited to train Master and PhD students in asteroseismic data analysis and to build longterm datasets. The number of stars in the system is still limited and reflects the willingness of data owners to provide their data after publication. Work continues to populate the database with contributions from the community, and at present the number of stars in the database is 82

  9. Das DNA-Puzzle

    NASA Astrophysics Data System (ADS)

    Kirchner, Stefan

    Im Jahre 1953 wurde von James Watson und Francis Crick erstmalig der strukturelle Aufbau der sogenannten DNA (Desoxyribonukleinsäure) beschrieben, welche das Erbgut jedes Lebewesens enthält. Der wesentliche Teil des Erbguts wird dabei durch eine sehr lange Folge der vier Basen Adenin (A), Cytosin (C), Guanin (G) und Thymin (T) codiert. Seit einigen Jahren ist es möglich, die Folge der vier Basen zu einer gegebenen DNA zu bestimmen. Biologen bezeichnen diesen Vorgang als Sequenzierung.

  10. Das Allgemeine Statistische Archiv

    NASA Astrophysics Data System (ADS)

    Rinne, Horst

    Das Allgemeine Statistische Archiv1, nachfolgend kurz Archiv genannt, ist die älteste, ausschließlich der Statistik gewidmete deutschsprachige wissenschaftliche Fachzeitschrift. Sie ist gut zwanzig Jahre älter als die Deutsche Statistische Gesellschaft, zu deren Publikationsorgan sie mit der Gründung der Gesellschaft im Jahre 1911 wurde. Zeitschrift und Gesellschaft blicken auf eine sehr wechselvolle, aber erfolgreiche gemeinsame Geschichte zurück. Die wissenschaftliche Arbeit der Gesellschaft ist im Archiv und seinen Sonderheften für nachfolgende Generationen dokumentiert.

  11. Connecting the Dots in DAS

    ERIC Educational Resources Information Center

    Ford, Tracy

    2012-01-01

    Many institutions implement a distributed antenna system (DAS) as part of a holistic approach to providing better wireless coverage and capacity on campus. A DAS provides wireless service within a particular area or structure via a network of separate antenna nodes that are connected to a common source through fiber or coaxial cable. Because DAS…

  12. Dictionary of Microscopy

    NASA Astrophysics Data System (ADS)

    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.

  13. MyDas, an extensible Java DAS server.

    PubMed

    Salazar, Gustavo A; García, Leyla J; Jones, Philip; Jimenez, Rafael C; Quinn, Antony F; Jenkinson, Andrew M; Mulder, Nicola; Martin, Maria; Hunter, Sarah; Hermjakob, Henning

    2012-01-01

    A large number of diverse, complex, and distributed data resources are currently available in the Bioinformatics domain. The pace of discovery and the diversity of information means that centralised reference databases like UniProt and Ensembl cannot integrate all potentially relevant information sources. From a user perspective however, centralised access to all relevant information concerning a specific query is essential. The Distributed Annotation System (DAS) defines a communication protocol to exchange annotations on genomic and protein sequences; this standardisation enables clients to retrieve data from a myriad of sources, thus offering centralised access to end-users.We introduce MyDas, a web server that facilitates the publishing of biological annotations according to the DAS specification. It deals with the common functionality requirements of making data available, while also providing an extension mechanism in order to implement the specifics of data store interaction. MyDas allows the user to define where the required information is located along with its structure, and is then responsible for the communication protocol details.

  14. Axial Plane Optical Microscopy

    PubMed Central

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

    2014-01-01

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

  15. Light sheet microscopy.

    PubMed

    Weber, Michael; Mickoleit, Michaela; Huisken, Jan

    2014-01-01

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

  16. Lasers for nonlinear microscopy.

    PubMed

    Wise, Frank

    2013-03-01

    Various versions of nonlinear microscopy are revolutionizing the life sciences, almost all of which are made possible because of the development of ultrafast lasers. In this article, the main properties and technical features of short-pulse lasers used in nonlinear microscopy are summarized. Recent research results on fiber lasers that will impact future instruments are also discussed.

  17. Clinical specular microscopy

    SciTech Connect

    Hirst, L.W.; Laing, R.A.

    1987-01-01

    This book provides the general ophthalmologist with a guide to the clinical applications of specular microscopy. Important material is included on laser injury, cataract surgery, corneal transplants, glaucoma, uveitis, and trauma.

  18. Photothermal imaging scanning microscopy

    DOEpatents

    Chinn, Diane; Stolz, Christopher J.; Wu, Zhouling; Huber, Robert; Weinzapfel, Carolyn

    2006-07-11

    Photothermal Imaging Scanning Microscopy produces a rapid, thermal-based, non-destructive characterization apparatus. Also, a photothermal characterization method of surface and subsurface features includes micron and nanoscale spatial resolution of meter-sized optical materials.

  19. Nonlinear vibrational microscopy

    DOEpatents

    Holtom, Gary R.; Xie, Xiaoliang Sunney; Zumbusch, Andreas

    2000-01-01

    The present invention is a method and apparatus for microscopic vibrational imaging using coherent Anti-Stokes Raman Scattering or Sum Frequency Generation. Microscopic imaging with a vibrational spectroscopic contrast is achieved by generating signals in a nonlinear optical process and spatially resolved detection of the signals. The spatial resolution is attained by minimizing the spot size of the optical interrogation beams on the sample. Minimizing the spot size relies upon a. directing at least two substantially co-axial laser beams (interrogation beams) through a microscope objective providing a focal spot on the sample; b. collecting a signal beam together with a residual beam from the at least two co-axial laser beams after passing through the sample; c. removing the residual beam; and d. detecting the signal beam thereby creating said pixel. The method has significantly higher spatial resolution then IR microscopy and higher sensitivity than spontaneous Raman microscopy with much lower average excitation powers. CARS and SFG microscopy does not rely on the presence of fluorophores, but retains the resolution and three-dimensional sectioning capability of confocal and two-photon fluorescence microscopy. Complementary to these techniques, CARS and SFG microscopy provides a contrast mechanism based on vibrational spectroscopy. This vibrational contrast mechanism, combined with an unprecedented high sensitivity at a tolerable laser power level, provides a new approach for microscopic investigations of chemical and biological samples.

  20. Optical imaging. Expansion microscopy.

    PubMed

    Chen, Fei; Tillberg, Paul W; Boyden, Edward S

    2015-01-30

    In optical microscopy, fine structural details are resolved by using refraction to magnify images of a specimen. We discovered that by synthesizing a swellable polymer network within a specimen, it can be physically expanded, resulting in physical magnification. By covalently anchoring specific labels located within the specimen directly to the polymer network, labels spaced closer than the optical diffraction limit can be isotropically separated and optically resolved, a process we call expansion microscopy (ExM). Thus, this process can be used to perform scalable superresolution microscopy with diffraction-limited microscopes. We demonstrate ExM with apparent ~70-nanometer lateral resolution in both cultured cells and brain tissue, performing three-color superresolution imaging of ~10(7) cubic micrometers of the mouse hippocampus with a conventional confocal microscope.

  1. Light microscopy digital imaging.

    PubMed

    Joubert, James; Sharma, Deepak

    2011-10-01

    This unit presents an overview of digital imaging hardware used in light microscopy. CMOS, CCD, and EMCCDs are the primary sensors used. The strengths and weaknesses of each define the primary applications for these sensors. Sensor architecture and formats are also reviewed. Color camera design strategies and sensor window cleaning are also described in the unit.

  2. Video Telescope Operating Microscopy.

    PubMed

    Divers, Stephen J

    2015-09-01

    Exotic pet veterinarians frequently have to operate on small animals, and magnification is commonly used. Existing endoscopy equipment can be used with a mechanical arm and telescope to enable video telescope operating microscopy. The additional equipment items and their specifics are described, and several case examples are provided.

  3. Scanning ultrafast electron microscopy.

    PubMed

    Yang, Ding-Shyue; Mohammed, Omar F; Zewail, Ahmed H

    2010-08-24

    Progress has been made in the development of four-dimensional ultrafast electron microscopy, which enables space-time imaging of structural dynamics in the condensed phase. In ultrafast electron microscopy, the electrons are accelerated, typically to 200 keV, and the microscope operates in the transmission mode. Here, we report the development of scanning ultrafast electron microscopy using a field-emission-source configuration. Scanning of pulses is made in the single-electron mode, for which the pulse contains at most one or a few electrons, thus achieving imaging without the space-charge effect between electrons, and still in ten(s) of seconds. For imaging, the secondary electrons from surface structures are detected, as demonstrated here for material surfaces and biological specimens. By recording backscattered electrons, diffraction patterns from single crystals were also obtained. Scanning pulsed-electron microscopy with the acquired spatiotemporal resolutions, and its efficient heat-dissipation feature, is now poised to provide in situ 4D imaging and with environmental capability.

  4. Photoacoustic computed microscopy

    PubMed Central

    Yao, Lei; Xi, Lei; Jiang, Huabei

    2014-01-01

    Photoacoustic microscopy (PAM) is emerging as a powerful technique for imaging microvasculature at depths beyond the ~1 mm depth limit associated with confocal microscopy, two-photon microscopy and optical coherence tomography. PAM, however, is currently qualitative in nature and cannot quantitatively measure important functional parameters including oxyhemoglobin (HbO2), deoxyhemoglobin (HbR), oxygen saturation (sO2), blood flow (BF) and rate of oxygen metabolism (MRO2). Here we describe a new photoacoustic microscopic method, termed photoacoustic computed microscopy (PACM) that combines current PAM technique with a model-based inverse reconstruction algorithm. We evaluate the PACM approach using tissue-mimicking phantoms and demonstrate its in vivo imaging ability of quantifying HbO2, HbR, sO2, cerebral BF and cerebral MRO2 at the small vessel level in a rodent model. This new technique provides a unique tool for neuroscience research and for visualizing microvasculature dynamics involved in tumor angiogenesis and in inflammatory joint diseases. PMID:24828539

  5. Polarized Light Microscopy

    NASA Technical Reports Server (NTRS)

    Frandsen, Athela F.

    2016-01-01

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

  6. Quantitative deconvolution microscopy.

    PubMed

    Goodwin, Paul C

    2014-01-01

    The light microscope is an essential tool for the study of cells, organelles, biomolecules, and subcellular dynamics. A paradox exists in microscopy whereby the higher the needed lateral resolution, the more the image is degraded by out-of-focus information. This creates a significant need to generate axial contrast whenever high lateral resolution is required. One strategy for generating contrast is to measure or model the optical properties of the microscope and to use that model to algorithmically reverse some of the consequences of high-resolution imaging. Deconvolution microscopy implements model-based methods to enable the full diffraction-limited resolution of the microscope to be exploited even in complex and living specimens.

  7. Ion photon emission microscopy

    NASA Astrophysics Data System (ADS)

    Rossi, P.; Doyle, B. L.; Banks, J. C.; Battistella, A.; Gennaro, G.; McDaniel, F. D.; Mellon, M.; Vittone, E.; Vizkelethy, G.; Wing, N. D.

    2003-09-01

    A new ion-induced emission microscopy has been invented and demonstrated, which is called ion photon emission microscopy (IPEM). It employs a low current, broad ion beam impinging on a sample, previously coated or simply covered with a few microns of a fast, highly efficient phosphor layer. The light produced at the single ion impact point is collected with an optical microscope and projected at high magnification onto a single photon position sensitive detector (PSD). This allows maps of the ion strike effects to be produced, effectively removing the need for a microbeam. Irradiation in air and even the use of alpha particle sources with no accelerator are possible. Potential applications include ion beam induced charge collection studies of semiconducting and insulating materials, single event upset studies on microchips and even biological cells in radiobiological effectiveness experiments. We describe the IPEM setup, including a 60× OM-40 microscope with a 1.5 mm hole for the beam transmission and a Quantar PSD with 60 μm pixel. Bicron plastic scintillator blades of 10 μm were chosen as a phosphor for their nanosecond time resolution, homogeneity, utility and commercial availability. The results given in this paper are for a prototype IPEM system. They indicate a resolution of ˜12 μm, the presence of a spatial halo and a He-ion efficiency of ˜20%. This marks the first time that nuclear microscopy has been performed with a radioactive source.

  8. Multimodal multiphoton microscopy

    NASA Astrophysics Data System (ADS)

    Légaré, François; Pfeffer, Christian P.; Ganikhanov, Feruz

    2009-02-01

    Multiphoton microscopy is a powerful technique for high spatial resolution thick tissue imaging. In its simple version, it uses a high repetition rate femtosecond oscillator laser source focussed and scanned across biological sample that contains fluorophores. However, not every biological structure is inherently fluorescent or can be stained without causing biochemical changes. To circumvent these limitations, other non-invasive nonlinear optical imaging approaches are currently being developed and investigated with regard to different applications. These techniques are: (1) second harmonic generation (SHG), (2) third harmonic generation (THG), and (3) coherent anti-Stokes Raman scattering (CARS) microscopy. The main advantage of the above mentioned techniques is that they derive their imaging contrast from optical nonlinearities that do not involve fluorescence process. As a particular application example we investigated collagen arrays. We show that combining SHG-THG-CARS onto a single imaging platform provides complementary information about the sub-micron architecture of the tissue. SHG microscopy reveals the fibrillar architecture of collagen arrays and confirm a rather high degree of heterogeneity of χ(2) within the focal volume, THG highlights the boundaries between the collagen sheets, and CH2 spectroscopic contrast with CARS.

  9. Data Acquisition System(DAS) Sustaining Engineering

    NASA Technical Reports Server (NTRS)

    1998-01-01

    This paper presents general information describing the Data Acquisition System contract, a summary of objectives, tasks performed and completed. The hardware deliverables which are comprised of: 1) Two ground DAS units; 2) Two flight DAS units; 3) Logistic spares; and 4) Shipping containers are described. Also included are the data requirements and scope of the contract.

  10. Electrochemical force microscopy

    DOEpatents

    Kalinin, Sergei V.; Jesse, Stephen; Collins, Liam F.; Rodriguez, Brian J.

    2017-01-10

    A system and method for electrochemical force microscopy are provided. The system and method are based on a multidimensional detection scheme that is sensitive to forces experienced by a biased electrode in a solution. The multidimensional approach allows separation of fast processes, such as double layer charging, and charge relaxation, and slow processes, such as diffusion and faradaic reactions, as well as capturing the bias dependence of the response. The time-resolved and bias measurements can also allow probing both linear (small bias range) and non-linear (large bias range) electrochemical regimes and potentially the de-convolution of charge dynamics and diffusion processes from steric effects and electrochemical reactivity.

  11. Fourier plane imaging microscopy

    SciTech Connect

    Dominguez, Daniel Peralta, Luis Grave de; Alharbi, Nouf; Alhusain, Mdhaoui; Bernussi, Ayrton A.

    2014-09-14

    We show how the image of an unresolved photonic crystal can be reconstructed using a single Fourier plane (FP) image obtained with a second camera that was added to a traditional compound microscope. We discuss how Fourier plane imaging microscopy is an application of a remarkable property of the obtained FP images: they contain more information about the photonic crystals than the images recorded by the camera commonly placed at the real plane of the microscope. We argue that the experimental results support the hypothesis that surface waves, contributing to enhanced resolution abilities, were optically excited in the studied photonic crystals.

  12. Multi-pass microscopy

    NASA Astrophysics Data System (ADS)

    Juffmann, Thomas; Klopfer, Brannon B.; Frankort, Timmo L. I.; Haslinger, Philipp; Kasevich, Mark A.

    2016-09-01

    Microscopy of biological specimens often requires low light levels to avoid damage. This yields images impaired by shot noise. An improved measurement accuracy at the Heisenberg limit can be achieved exploiting quantum correlations. If sample damage is the limiting resource, an equivalent limit can be reached by passing photons through a specimen multiple times sequentially. Here we use self-imaging cavities and employ a temporal post-selection scheme to present full-field multi-pass polarization and transmission micrographs with variance reductions of 4.4+/-0.8 dB (11.6+/-0.8 dB in a lossless setup) and 4.8+/-0.8 dB, respectively, compared with the single-pass shot-noise limit. If the accuracy is limited by the number of detected probe particles, our measurements show a variance reduction of 25.9+/-0.9 dB. The contrast enhancement capabilities in imaging and in diffraction studies are demonstrated with nanostructured samples and with embryonic kidney 293T cells. This approach to Heisenberg-limited microscopy does not rely on quantum state engineering.

  13. Multi-pass microscopy

    PubMed Central

    Juffmann, Thomas; Klopfer, Brannon B.; Frankort, Timmo L.I.; Haslinger, Philipp; Kasevich, Mark A.

    2016-01-01

    Microscopy of biological specimens often requires low light levels to avoid damage. This yields images impaired by shot noise. An improved measurement accuracy at the Heisenberg limit can be achieved exploiting quantum correlations. If sample damage is the limiting resource, an equivalent limit can be reached by passing photons through a specimen multiple times sequentially. Here we use self-imaging cavities and employ a temporal post-selection scheme to present full-field multi-pass polarization and transmission micrographs with variance reductions of 4.4±0.8 dB (11.6±0.8 dB in a lossless setup) and 4.8±0.8 dB, respectively, compared with the single-pass shot-noise limit. If the accuracy is limited by the number of detected probe particles, our measurements show a variance reduction of 25.9±0.9 dB. The contrast enhancement capabilities in imaging and in diffraction studies are demonstrated with nanostructured samples and with embryonic kidney 293T cells. This approach to Heisenberg-limited microscopy does not rely on quantum state engineering. PMID:27670525

  14. Magnetic force microscopy

    PubMed Central

    Passeri, Daniele; Dong, Chunhua; Reggente, Melania; Angeloni, Livia; Barteri, Mario; Scaramuzzo, Francesca A; De Angelis, Francesca; Marinelli, Fiorenzo; Antonelli, Flavia; Rinaldi, Federica; Marianecci, Carlotta; Carafa, Maria; Sorbo, Angela; Sordi, Daniela; Arends, Isabel WCE; Rossi, Marco

    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 at the nanoscale. Being a well established tool for the characterization of magnetic recording media, superconductors and magnetic nanomaterials, MFM is finding constantly increasing application in the study of magnetic properties of materials and systems of biological and biomedical interest. After reviewing these latter applications, three case studies are presented in which MFM is used to characterize: (i) magnetoferritin synthesized using apoferritin as molecular reactor; (ii) magnetic nanoparticles loaded niosomes to be used as nanocarriers for drug delivery; (iii) leukemic cells labeled using folic acid-coated core-shell superparamagnetic nanoparticles in order to exploit the presence of folate receptors on the cell membrane surface. In these examples, MFM data are quantitatively analyzed evidencing the limits of the simple analytical models currently used. Provided that suitable models are used to simulate the MFM response, MFM can be used to evaluate the magnetic momentum of the core of magnetoferritin, the iron entrapment efficiency in single vesicles, or the uptake of magnetic nanoparticles into cells. PMID:25050758

  15. Silver stain for electron microscopy

    NASA Technical Reports Server (NTRS)

    Corbett, R. L.

    1972-01-01

    Ammoniacal silver stain used for light microscopy was adapted advantageously for use with very thin biological sections required for electron microscopy. Silver stain can be performed in short time, has more contrast, and is especially useful for low power electron microscopy.

  16. Image Force Microscopy

    NASA Astrophysics Data System (ADS)

    Rajapaksa, Indrajith

    In this thesis we describe an enhancement to the Atomic force microscope (AFM) to simultaneously gather topographic features and spectroscopic information .Compared to the current state of the art of near-field excitation and far-field detection AFM imaging techniques our system uses a radical new approach near-field excitation and near-field detection. By placing the detector in the near-field we achieve high signal to noise and single molecular resolution. The origin of our near-field detector signal is the image force gradient due to the interaction of the stimulated molecular dipole with its image on the metal probe. We designed and built an optical and electronic system to capture this signal and simultaneously image nano-scale surface topography and optical image force gradient. By varying the wavelength of the excitation beam we measure the induced optical image force gradient spectra of molecules on surface. These spectra show good agreement with the absorption spectra of the bulk molecules measured by conventional absorption spectroscopy. We show that image force gradient is directly proportional to the optical absorption dipole strength. Using Finite Element 3D electromagnetic simulations and using Lorentz model for the excited molecular dipole we showed that the image force gradient has a decay length of 1nm, making the theoretical resolution of this microscopy technique approximately 1 nm. This rapid decay was measured experimentally .This resolution was seen by the high contrasting spectroscopic images of molecules on the surface. In follow on experiments this technique was extended to provide surface Raman spectroscopy and microscopy at molecular resolution. We create an image force gradient interaction through optical parametric down conversion between stimulated Raman excited molecules on a surface and a cantilevered nanometer scale probe brought very close to it. Spectroscopy and microscopy on clusters of molecules have been performed. Single

  17. Applications of subsurface microscopy.

    PubMed

    Tetard, Laurene; Passian, Ali; Farahi, Rubye H; Voy, Brynn H; Thundat, Thomas

    2012-01-01

    Exploring the interior of a cell is of tremendous importance in order to assess the effects of nanomaterials on biological systems. Outside of a controlled laboratory environment, nanomaterials will most likely not be conveniently labeled or tagged so that their translocation within a biological system cannot be easily identified and quantified. Ideally, the characterization of nanomaterials within a cell requires a nondestructive, label-free, and subsurface approach. Subsurface nanoscale imaging represents a real challenge for instrumentation. Indeed the tools available for high resolution characterization, including optical, electron or scanning probe microscopies, mainly provide topography images or require taggants that fluoresce. Although the intercellular environment holds a great deal of information, subsurface visualization remains a poorly explored area. Recently, it was discovered that by mechanically perturbing a sample, it was possible to observe its response in time with nanoscale resolution by probing the surface with a micro-resonator such as a microcantilever probe. Microcantilevers are used as the force-sensing probes in atomic force microscopy (AFM), where the nanometer-scale probe tip on the microcantilever interacts with the sample in a highly controlled manner to produce high-resolution raster-scanned information of the sample surface. Taking advantage of the existing capabilities of AFM, we present a novel technique, mode synthesizing atomic force microscopy (MSAFM), which has the ability to probe subsurface structures such as non-labeled nanoparticles embedded in a cell. In MSAFM mechanical actuators (PZTs) excite the probe and the sample at different frequencies as depicted in the first figure of this chapter. The nonlinear nature of the tip-sample interaction, at the point of contact of the probe and the surface of the sample, in the contact mode AFM configuration permits the mixing of the elastic waves. The new dynamic system comprises new

  18. Hyperspectral light sheet microscopy

    NASA Astrophysics Data System (ADS)

    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.

  19. Sensitivity of photoacoustic microscopy

    PubMed Central

    Yao, Junjie; Wang, Lihong V.

    2014-01-01

    Building on its high spatial resolution, deep penetration depth and excellent image contrast, 3D photoacoustic microscopy (PAM) has grown tremendously since its first publication in 2005. Integrating optical excitation and acoustic detection, PAM has broken through both the optical diffusion and optical diffraction limits. PAM has 100% relative sensitivity to optical absorption (i.e., a given percentage change in the optical absorption coefficient yields the same percentage change in the photoacoustic amplitude), and its ultimate detection sensitivity is limited only by thermal noise. Focusing on the engineering aspects of PAM, this Review discusses the detection sensitivity of PAM, compares the detection efficiency of different PAM designs, and summarizes the imaging performance of various endogenous and exogenous contrast agents. It then describes representative PAM applications with high detection sensitivity, and outlines paths to further improvement. PMID:25302158

  20. Snapshot Hyperspectral Volumetric Microscopy

    NASA Astrophysics Data System (ADS)

    Wu, Jiamin; Xiong, Bo; Lin, Xing; He, Jijun; Suo, Jinli; Dai, Qionghai

    2016-04-01

    The comprehensive analysis of biological specimens brings about the demand for capturing the spatial, temporal and spectral dimensions of visual information together. However, such high-dimensional video acquisition faces major challenges in developing large data throughput and effective multiplexing techniques. Here, we report the snapshot hyperspectral volumetric microscopy that computationally reconstructs hyperspectral profiles for high-resolution volumes of ~1000 μm × 1000 μm × 500 μm at video rate by a novel four-dimensional (4D) deconvolution algorithm. We validated the proposed approach with both numerical simulations for quantitative evaluation and various real experimental results on the prototype system. Different applications such as biological component analysis in bright field and spectral unmixing of multiple fluorescence are demonstrated. The experiments on moving fluorescent beads and GFP labelled drosophila larvae indicate the great potential of our method for observing multiple fluorescent markers in dynamic specimens.

  1. Snapshot Hyperspectral Volumetric Microscopy

    PubMed Central

    Wu, Jiamin; Xiong, Bo; Lin, Xing; He, Jijun; Suo, Jinli; Dai, Qionghai

    2016-01-01

    The comprehensive analysis of biological specimens brings about the demand for capturing the spatial, temporal and spectral dimensions of visual information together. However, such high-dimensional video acquisition faces major challenges in developing large data throughput and effective multiplexing techniques. Here, we report the snapshot hyperspectral volumetric microscopy that computationally reconstructs hyperspectral profiles for high-resolution volumes of ~1000 μm × 1000 μm × 500 μm at video rate by a novel four-dimensional (4D) deconvolution algorithm. We validated the proposed approach with both numerical simulations for quantitative evaluation and various real experimental results on the prototype system. Different applications such as biological component analysis in bright field and spectral unmixing of multiple fluorescence are demonstrated. The experiments on moving fluorescent beads and GFP labelled drosophila larvae indicate the great potential of our method for observing multiple fluorescent markers in dynamic specimens. PMID:27103155

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

    SciTech Connect

    Ramanathan, Nathan Muruganathan; Darling, Seth B.

    2015-01-01

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

  3. Extraterrestrial optical microscopy.

    PubMed

    Soffen, G A

    1969-07-01

    An examination of the literature concerned with the use of microscopy for planetary investigation reveals a serious deficiency of current efforts. Many scientists have recommended the use of a microscope for planetary investigation [Biology and the Exploration of Mars, C. S. Pittendrigh, W. Vishniac, and J. P. T. Pearman, Eds. (National Academy of Science-National Research Council, Washington, D. C., 1966), (a) D. Mazia, p. 31; (b) J. Lederberg, p. 137; (c) S. Fox, pp. 219, 226; (d) D. Glaser, p. 326; (e) D. Glaser, J. McCarthy, and M. Minsky, pp. 333, 341; (f) D. G. Rea, pp. 347-426; (g) P. G. Conger, pp. 409-414; (h) M. H. Fernandez, pp. 414-425; (i) D. Schwartz, pp.425-426 . H. P. Klein, Some Biological Problems in the Search for Extraterrestrial Life (American Astronautical Society, Washington, D. C., 1968).] but few are involved in developing the experiment. Since this is a particularly timely period for the preparation of planetary lander experiments, the reasons for this lack of effort would appear to be limited resources or an unclear course of action, rather than lack of interest. Microscopy used for planetary investigation is chiefly the interest of the biologist and the mineralogist. In both cases the desire to use magnifying optics in order to observe objects of submillimeter size is based upon the rich body of knowledge we have acquired from observing the terrestrial microcosm. In addition to purely imaging, certain special optical techniques, e.g., polarimetry, colorimetry, phase contrast, etc., can be used to enhance the interpretation of microscopic imaging data. This interaction of the optical with the chemical or structural aspects of nature can be used to great advantage in the exploration of extraterrestrial biology and mineralogy.

  4. Ultrasonic Force Microscopies

    NASA Astrophysics Data System (ADS)

    Kolosov, Oleg; Briggs, Andrew

    Ultrasonic Force Microscopy, or UFM, allows combination of two apparently mutually exclusive requirements for the nanomechanical probe—high stiffness for the efficient indentation and high mechanical compliance that brings force sensitivity. Somewhat inventively, UFM allows to combine these two virtues in the same cantilever by using indention of the sample at high frequency, when cantilever is very rigid, but detecting the result of this indention at much lower frequency. That is made possible due to the extreme nonlinearity of the nanoscale tip-surface junction force-distance dependence, that acts as "mechanical diode" detecting ultrasound in AFM. After introducing UFM principles, we discuss features of experimental UFM implementation, and the theory of contrast in this mode, progressing to quantitative measurements of contact stiffness. A variety of UFM applications ranging from semiconductor quantum nanostructures, graphene, very large scale integrated circuits, and reinforced ceramics to polymer composites and biological materials is presented via comprehensive imaging gallery accompanied by the guidance for the optimal UFM measurements of these materials. We also address effects of adhesion and topography on the elasticity imaging and the approaches for reducing artifacts connected with these effects. This is complemented by another extremely useful feature of UFM—ultrasound induced superlubricity that allows damage free imaging of materials ranging from stiff solid state devices and graphene to biological materials. Finally, we proceed to the exploration of time-resolved nanoscale phenomena using nonlinear mixing of multiple vibration frequencies in ultrasonic AFM—Heterodyne Force Microscopy, or HFM, that also include mixing of ultrasonic vibration with other periodic physical excitations, eg. electrical, photothermal, etc. Significant section of the chapter analyzes the ability of UFM and HFM to detect subsurface mechanical inhomogeneities, as well as

  5. Virtual microscopy in pathology education.

    PubMed

    Dee, Fred R

    2009-08-01

    Technology for acquisition of virtual slides was developed in 1985; however, it was not until the late 1990s that desktop computers had enough processing speed to commercialize virtual microscopy and apply the technology to education. By 2000, the progressive decrease in use of traditional microscopy in medical student education had set the stage for the entry of virtual microscopy into medical schools. Since that time, it has been successfully implemented into many pathology courses in the United States and around the world, with surveys indicating that about 50% of pathology courses already have or expect to implement virtual microscopy. Over the last decade, in addition to an increasing ability to emulate traditional microscopy, virtual microscopy has allowed educators to take advantage of the accessibility, efficiency, and pedagogic versatility of the computer and the Internet. The cost of virtual microscopy in education is now quite reasonable after taking into account replacement cost for microscopes, maintenance of glass slides, and the fact that 1-dimensional microscope space can be converted to multiuse computer laboratories or research. Although the current technology for implementation of virtual microscopy in histopathology education is very good, it could be further improved upon by better low-power screen resolution and depth of field. Nevertheless, virtual microscopy is beginning to play an increasing role in continuing education, house staff education, and evaluation of competency in histopathology. As Z-axis viewing (focusing) becomes more efficient, virtual microscopy will also become integrated into education in cytology, hematology, microbiology, and urinalysis.

  6. Intrinsic Friction Microscopy

    NASA Astrophysics Data System (ADS)

    Knorr, Daniel; Overney, Rene

    2008-03-01

    A novel scanning probe methodology based on lateral force microscopy is presented wherein kinetic friction measurements, obtained as a function of velocity for various temperatures, are used to deduce apparent Arrhenius-type activation energies for surface and subsurface molecular mobilities. Depending on the coupling strength (cooperativity) between molecular mobilities involved the dissipation energy can carry a significant entropic energy contribution, accounting for the majority of the apparent Arrhenius activation energy. The intrinsic friction methodology also provides a means of directly separating enthalpic energy contributions from entropic ones by employing absolute rate theory. As such, the degree of cooperativity in the system is readily apparent. This methodology is illustrated with nanoscale tribological experiments on two systems, (1) monodisperse, atactic polystyrene and (2) self assembling molecular glassy chromophores. In polystyrene, dissipation was found to be a discrete function of loading, where the γ-relaxation (phenyl group rotation) was recovered for ultra low loads and the β-relaxation (local backbone translation) for higher loads in the same temperature range, indicating sensitivity to surface and subsurface mobilities. For self assembling glassy chromophores, the degree of intermolecular cooperativity was deduced using the methodology, resulting in an increased understanding of the interactions between self assembling molecules.

  7. Ultrafast scanning probe microscopy

    DOEpatents

    Weiss, Shimon; Chemla, Daniel S.; Ogletree, D. Frank; Botkin, David

    1995-01-01

    An ultrafast scanning probe microscopy method for achieving subpicosecond-temporal resolution and submicron-spatial resolution of an observation sample. In one embodiment of the present claimed invention, a single short optical pulse is generated and is split into first and second pulses. One of the pulses is delayed using variable time delay means. The first pulse is then directed at an observation sample located proximate to the probe of a scanning probe microscope. The scanning probe microscope produces probe-sample signals indicative of the response of the probe to characteristics of the sample. The second pulse is used to modulate the probe of the scanning probe microscope. The time delay between the first and second pulses is then varied. The probe-sample response signal is recorded at each of the various time delays created between the first and second pulses. The probe-sample response signal is then plotted as a function of time delay to produce a cross-correlation of the probe sample response. In so doing, the present invention provides simultaneous subpicosecond-temporal resolution and submicron-spatial resolution of the sample.

  8. Ultrafast scanning tunneling microscopy

    SciTech Connect

    Botkin, D.A. |

    1995-09-01

    I have developed an ultrafast scanning tunneling microscope (USTM) based on uniting stroboscopic methods of ultrafast optics and scanned probe microscopy to obtain nanometer spatial resolution and sub-picosecond temporal resolution. USTM increases the achievable time resolution of a STM by more than 6 orders of magnitude; this should enable exploration of mesoscopic and nanometer size systems on time scales corresponding to the period or decay of fundamental excitations. USTM consists of a photoconductive switch with subpicosecond response time in series with the tip of a STM. An optical pulse from a modelocked laser activates the switch to create a gate for the tunneling current, while a second laser pulse on the sample initiates a dynamic process which affects the tunneling current. By sending a large sequence of identical pulse pairs and measuring the average tunnel current as a function of the relative time delay between the pulses in each pair, one can map the time evolution of the surface process. USTM was used to measure the broadband response of the STM`s atomic size tunnel barrier in frequencies from tens to hundreds of GHz. The USTM signal amplitude decays linearly with the tunnel junction conductance, so the spatial resolution of the time-resolved signal is comparable to that of a conventional STM. Geometrical capacitance of the junction does not appear to play an important role in the measurement, but a capacitive effect intimately related to tunneling contributes to the measured signals and may limit the ultimate resolution of the USTM.

  9. NMR imaging microscopy

    SciTech Connect

    Not Available

    1986-10-01

    In the past several years, proton nuclear magnetic resonance (NMR) imaging has become an established technique in diagnostic medicine and biomedical research. Although much of the work in this field has been directed toward development of whole-body imagers, James Aguayo, Stephen Blackband, and Joseph Schoeninger of the Johns Hopkins University School of Medicine working with Markus Hintermann and Mark Mattingly of Bruker Medical Instruments, recently developed a small-bore NMR microscope with sufficient resolution to image a single African clawed toad cell (Nature 1986, 322, 190-91). This improved resolution should lead to increased use of NMR imaging for chemical, as well as biological or physiological, applications. The future of NMR microscopy, like that of many other newly emerging techniques, is ripe with possibilities. Because of its high cost, however, it is likely to remain primarily a research tool for some time. ''It's like having a camera,'' says Smith. ''You've got a way to look at things at very fine levels, and people are going to find lots of uses for it. But it is a very expensive technique - it costs $100,000 to add imaging capability once you have a high-resolution NMR, which itself is at least a $300,000 instrument. If it can answer even a few questions that can't be answered any other way, though, it may be well worth the cost.''

  10. Mueller polarimetric microscopy

    NASA Astrophysics Data System (ADS)

    Laude-Boulesteix, Blandine; De Martino, Antonello; Le Naour, Gilles; Genestie, Catherine; Schwartz, Laurent; Garcia-Caurel, Enric; Drevillon, Bernard

    2004-07-01

    We present a multispectral polarimetric imaging system well suited for complete Mueller matrix microscopy. The source is a spectrally filtered halogen light bulb, and the image is formed on a fast CCD camera The light polarization is modulated before the sample and analyzed after the sample by using nematic liquid crystal modulators.. The whole Mueller matrix image of the sample is typically measured over 5 seconds for a good signal-to-noise ratio. The instrument design, together with an original and easy-to-operate calibration procedure provides a high polarimetric accuracy over wide ranges of wavelengths and magnifications. Mueller polarimetry provides separate images of scalar and vector retardation and dichroism of the sample, together with its depolarizing power, while all these effects do contribute simultaneously to the contrasts observed in standard polarized microsopy. Polarimetric images of several samples, namely an unstained rabbit cornea, a picrosirius red stained hepatic biopsy, and a rat artery specifically stained for collagen III are shown and discussed

  11. In vivo microscopy.

    PubMed

    Peti-Peterdi, János

    2016-04-01

    This article summarizes the past, present, and future promise of multiphoton excitation fluorescence microscopy for intravital kidney imaging. During the past 15years, several high-power visual research approaches have been developed using multiphoton imaging to study the normal functions of the healthy, intact, living kidney, and the various molecular and cellular mechanisms of the development of kidney diseases. In this review, the main focus will be on intravital multiphoton imaging of the glomerulus, the structure and function of the glomerular filtration barrier, especially the podocyte. Examples will be given for the combination of two powerful research tools, in vivo multiphoton imaging and mouse genetics using commercially available whole animal models for the detailed characterization of glomerular cell types, their function and fate, and for the better understanding of the molecular mechanisms of glomerular pathologies. One of the new modalities of multiphoton imaging, serial imaging of the same glomerulus in the same animal over several days will be emphasized for its potential for further advancing the field of nephrology research.

  12. Microscopy of semiconducting materials

    NASA Astrophysics Data System (ADS)

    Pennycook, S. J.

    1991-04-01

    The purpose of the trip was to present an invited talk at the 7th Oxford Conference on Microscopy of Semiconducting Materials entitled, High-Resolution Z-Contrast Imaging of Heterostructures and Superlattices, (Oxford, United Kingdom) and to visit VG Microscopes, East Grinstead, for discussions on the progress of the Oak Ridge National Laboratory (ORNL) 300-kV high-resolution scanning transmission electron microscope (STEM), which is currently on order. The traveler also visited three other institutions with 100-kV STEMs that either have or intend to purchase the necessary modifications to provide Z-contrast capability similar to that of the existing ORNL machine. Specifically, Max-Planck Institut fuer Metallforschung (Stuttgart, Germany); Cambridge University, Department of Materials Science and Metallurgy (Cambridge, United Kingdom); and Cavendish Laboratory, Cambridge University (Cambridge, United Kingdom) were visited. In addition, discussions were held with C. Humphreys on the possibility of obtaining joint funding for collaborative research involving electron beam writing and Z-contrast imaging in the Cambridge and Oak Ridge STEMs, respectively.

  13. Ultrafast scanning probe microscopy

    DOEpatents

    Weiss, S.; Chemla, D.S.; Ogletree, D.F.; Botkin, D.

    1995-05-16

    An ultrafast scanning probe microscopy method is described for achieving subpicosecond-temporal resolution and submicron-spatial resolution of an observation sample. In one embodiment of the present claimed invention, a single short optical pulse is generated and is split into first and second pulses. One of the pulses is delayed using variable time delay means. The first pulse is then directed at an observation sample located proximate to the probe of a scanning probe microscope. The scanning probe microscope produces probe-sample signals indicative of the response of the probe to characteristics of the sample. The second pulse is used to modulate the probe of the scanning probe microscope. The time delay between the first and second pulses is then varied. The probe-sample response signal is recorded at each of the various time delays created between the first and second pulses. The probe-sample response signal is then plotted as a function of time delay to produce a cross-correlation of the probe sample response. In so doing, the present invention provides simultaneous subpicosecond-temporal resolution and submicron-spatial resolution of the sample. 6 Figs.

  14. DAS28-CRP and DAS28-ESR cut-offs for high disease activity in rheumatoid arthritis are not interchangeable

    PubMed Central

    Fleischmann, Roy M; van der Heijde, Désirée; Gardiner, Philip V; Szumski, Annette; Marshall, Lisa; Bananis, Eustratios

    2017-01-01

    Background In most patients with rheumatoid arthritis (RA), Disease Activity Score 28-joint count C reactive protein (DAS28-CRP) is lower than DAS28 erythrocyte sedimentation rate (DAS28-ESR), suggesting that use of the DAS28-ESR cut-off to assess high disease activity (HDA) with DAS28-CRP may underestimate the number of patients with HDA. We determined the DAS28-CRP value corresponding to the validated DAS28-ESR cut-off for HDA. Methods Baseline data were pooled from 2 clinical studies evaluating etanercept (ETN) plus methotrexate (MTX) or MTX in early RA; DAS28-CRP and DAS28-ESR were obtained, allowing the determination of the DAS28-CRP HDA value best corresponding to the DAS28-ESR cut-off of >5.1. Results At baseline, as expected, fewer patients had HDA by DAS28-CRP than DAS28-ESR; DAS28-CRP>5.1 and DAS28-ESR>5.1 had only modest agreement (κ coefficients 0.45–0.54). Mean DAS28-CRP and DAS28-ESR were 5.7 and 6.2, respectively, in the ETN+MTX group (n=571), and 6.0 and 6.5 in the MTX group (n=262). A DAS28-CRP cut-off of 4.6 corresponded to a DAS28-ESR cut-off of 5.1. Conclusions We have shown that a DAS28-CRP of 4.6 corresponds to 5.1 for DAS28-ESR. Since this is substantially lower than the DAS28-ESR cut-off of 5.1, using 5.1 as the cut-off for DAS28-CRP underestimates disease activity in RA. Trial registration number NCT00195494; NCT00913458. PMID:28255449

  15. Sample preparation for STED microscopy.

    PubMed

    Wurm, Christian A; Neumann, Daniel; Schmidt, Roman; Egner, Alexander; Jakobs, Stefan

    2010-01-01

    Since the discovery of the diffraction barrier in the late nineteenth century, it has been commonly accepted that with far-field optical microscopy it is not possible to resolve structural details considerably finer than half the wavelength of light. The emergence of STED microscopy showed that, at least for fluorescence imaging, these limits can be overcome. Since STED microscopy is a far-field technique, in principle, the same sample preparation as for conventional confocal microscopy may be utilized. The increased resolution, however, requires additional precautions to ensure the structural preservation of the specimen. We present robust protocols to generate test samples for STED microscopy. These protocols for bead samples and immunolabeled mammalian cells may be used as starting points to adapt existing labeling strategies for the requirements of sub-diffraction resolution microscopy.

  16. Dasty3, a WEB framework for DAS

    PubMed Central

    Villaveces, Jose M.; Jimenez, Rafael C.; Garcia, Leyla J.; Salazar, Gustavo A.; Gel, Bernat; Mulder, Nicola; Martin, Maria; Garcia, Alexander; Hermjakob, Henning

    2011-01-01

    Motivation: Dasty3 is a highly interactive and extensible Web-based framework. It provides a rich Application Programming Interface upon which it is possible to develop specialized clients capable of retrieving information from DAS sources as well as from data providers not using the DAS protocol. Dasty3 provides significant improvements on previous Web-based frameworks and is implemented using the 1.6 DAS specification. Availability: Dasty3 is an open-source tool freely available at http://www.ebi.ac.uk/dasty/ under the terms of the GNU General public license. Source and documentation can be found at http://code.google.com/p/dasty/. Contact: hhe@ebi.ac.uk PMID:21798964

  17. Electronic Blending in Virtual Microscopy

    ERIC Educational Resources Information Center

    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…

  18. Multi-contrast Photoacoustic Microscopy

    NASA Astrophysics Data System (ADS)

    Yao, Junjie

    Photoacoustic microscopy is a hybrid imaging modality with high spatial resolution, moderate imaging depth, excellent imaging contrast and functional imaging capability. Taking full advantage of this powerful weapon, we have investigated different anatomical, functional, flow dynamic and metabolic parameter measurements using photoacoustic microscopy. Specifically, Evans-blue dye was used to enhance photoacoustic microscopy of capillaries; label-free transverse and axial blood flow was measured based on bandwidth broadening and time shift of the photoacoustic signals; metabolic rate of oxygen was quantified in vivo from all the five parameters measured by photoacoustic microcopy; whole cross-sectional imaging of small intestine was achieved on a double-illumination photoacoustic microscopy with extended depth of focus and imaging depth; hemodynamic imaging was performed on a MEMS-mirror enhanced photoacoustic microscopy with a cross-sectional imaging rate of 400 Hz. As a maturing imaging technique, PAM is expected to find new applications in both fundamental life science and clinical practice.

  19. Studies in scanning probe microscopy

    NASA Astrophysics Data System (ADS)

    Sarid, Dror

    1995-06-01

    The following is a final report on our work in the field of Scanning Probe Microscopy (SPM), which has been funded by the AFOSR under Contract #F49620-92-J-0164. The AFOSR funding was instrumental in the establishment of a multi-lab facility at the Optical Sciences Center, which performs research in SPM using two ultrahigh vacuum (UHV) STM facilities, and several Atomic Force Microscopy (AFM) facilities. The fabrication and characterization work performed in the SPM Laboratory is supplemented by infrared (IR) spectroscopy, high resolution transmission electron microscopy (HRTEM), and scanning electron microscopy (SEM), available in other departments on campus. The report covers the following areas: (1) GaAs and CdSe Structures, (2) Optical Interactions on a nm and nsec Scales, (3) Fullerenes on Gold, (4) Fullerenes on MoS2, (5) Fullerenes on Si, (6) SiC, (7) Nanotubes, (8) Scanning Force Microscopy, and (9) Biology.

  20. Chip-scale microscopy imaging.

    PubMed

    Zheng, Guoan

    2012-08-01

    Chip-scale microscopy imaging platforms are pivotal for improving the efficiency of modern biomedical and bioscience experiments. Their integration with other lab-on-a-chip techniques would allow rapid, reliable and high-throughput sample analysis for applications in diverse disciplines. In typical chip-scale microscopy imaging platforms, the light path can be generalized to the following steps: photons leave the light source, interact with the sample and finally are detected by the sensor. Based on the light path of these platforms, the current review aims to provide some insights on design strategies for chip-scale microscopy. Specifically, we analyze current chip-scale microscopy approaches from three aspects: illumination design, sample manipulation and substrate/imager modification. We also discuss some opportunities for future developments of chip-scale microscopy, such as time multiplexed structured illumination and hydrodynamic focusing for high throughput sample manipulation.

  1. Atomic force microscopy combined with optical microscopy for cells investigation.

    PubMed

    Cascione, Mariafrancesca; de Matteis, Valeria; Rinaldi, Rosaria; Leporatti, Stefano

    2017-01-01

    This review reports on the combined use of the atomic force microscopy (AFM) and several type of optical/fluorescence/laser scanning microscopy for investigating cells. It is shown that the hybrid systems of AFM with optical-derived microscopies enable to study in detail cell surface properties (such as topography), their mechanical properties (e.g., Young's modulus) mechanotransduction phenomena and allow to gain insight into biological-related pathways and mechanisms in the complex nanoworld of cells. Microsc. Res. Tech. 80:109-123, 2017. © 2016 Wiley Periodicals, Inc.

  2. Microscopy techniques in flavivirus research.

    PubMed

    Chong, Mun Keat; Chua, Anthony Jin Shun; Tan, Terence Tze Tong; Tan, Suat Hoon; Ng, Mah Lee

    2014-04-01

    The Flavivirus genus is composed of many medically important viruses that cause high morbidity and mortality, which include Dengue and West Nile viruses. Various molecular and biochemical techniques have been developed in the endeavour to study flaviviruses. However, microscopy techniques still have irreplaceable roles in the identification of novel virus pathogens and characterization of morphological changes in virus-infected cells. Fluorescence microscopy contributes greatly in understanding the fundamental viral protein localizations and virus-host protein interactions during infection. Electron microscopy remains the gold standard for visualizing ultra-structural features of virus particles and infected cells. New imaging techniques and combinatory applications are continuously being developed to push the limit of resolution and extract more quantitative data. Currently, correlative live cell imaging and high resolution three-dimensional imaging have already been achieved through the tandem use of optical and electron microscopy in analyzing biological specimens. Microscopy techniques are also used to measure protein binding affinities and determine the mobility pattern of proteins in cells. This chapter will consolidate on the applications of various well-established microscopy techniques in flavivirus research, and discuss how recently developed microscopy techniques can potentially help advance our understanding in these membrane viruses.

  3. Physik gestern und heute Das Eiskalorimeter

    NASA Astrophysics Data System (ADS)

    Heering, P.

    2003-07-01

    Kalorimetrische Messungen gehören heute zum experimentellen Standardrepertoire im Bereich der Thermodynamik und der physikalischen Chemie. Das erste Gerät für derartige Messungen entwickelten Ende des 18. Jahrhunderts die französischen Wissenschaftler Antoine Laurent Lavoisier und Pierre Simon de Laplace.

  4. GHRSST-14 DAS-TAG Report

    NASA Technical Reports Server (NTRS)

    Armstrong, Edward; Piolle, Jean Francois

    2013-01-01

    The DAS-TAG provides the informatics and data management expertise in emerging information technologies for the GHRSST community. It provides expertise in data and metadata formats and standards, fosters improvements for GHRSST data curation, experiments with new data processing paradigms, and evaluates services and tools for data usage. It provides a forum for producer and distributor data management issues and coordination.

  5. Horizontal microscopy in square capillaries

    NASA Astrophysics Data System (ADS)

    Moroz, Pavel E.

    1992-07-01

    Intracellular protoplasmic movements may, due to gravity, have a vertical component greater or different from the horizontal one. This makes horizontal microscopy indispensable in the search for the cellular sensor of gravity. The possibility of the latter being a cell organelle assigns special significance to high-resolution microscopy. A horizontal suction device for picking up a cell and its high-resolution horizontal microscopy in a rectangular capillary may be helpful for detection of gravity-related shifts of cellular organelles in vivo.

  6. Research With Scanning Tip Microscopy

    DTIC Science & Technology

    1991-12-31

    08ro P noiwe bae?041Le Research With Scanning Tip Microscopy AFOSR-89-0498 V AUTHOS)i Professor Dror Sarid 7. PFOUImNG 00ANIZATION NAMEIS) AND...forces and (b) surfaces. UNCLASS UNCLASS UNCLASS UL FINAL REPORT TO THE AFOSR ൱-, to J4ti. r Aat io Research in Scanning Tip Microscopy Dror Sarid Dtst...microscopy have been used to investigate (a) forces and (b) surfaces. a. Forces 1. Dror Sarid , Douglas lams, Volker Weissenberger, and L. Stephen Bell

  7. Soil microstructure and electron microscopy

    NASA Technical Reports Server (NTRS)

    Smart, P.; Fryer, J. R.

    1988-01-01

    As part of the process of comparing Martian soils with terrestial soils, high resolution electron microscopy and associated techniques should be used to examine the finer soil particles, and various techniques of electron and optical microscopy should be used to examine the undisturbed structure of Martian soils. To examine the structure of fine grained portions of the soil, transmission electron microscopy may be required. A striking feature of many Martian soils is their red color. Although the present-day Martian climate appears to be cold, this color is reminiscent of terrestial tropical red clays. Their chemical contents are broadly similar.

  8. Studies in Scanning Probe Microscopy.

    DTIC Science & Technology

    2007-11-02

    refereed journals, as well as two books titled Scanning Force Microscopy, With Applications to Electric, Magnetic, and Atomic Forces published by Oxford University Press in 1991 and a revised edition in 1994.

  9. Fluorescence Microscopy of Single Molecules

    ERIC Educational Resources Information Center

    Zimmermann, Jan; van Dorp, Arthur; Renn, Alois

    2004-01-01

    The investigation of photochemistry and photophysics of individual quantum systems is described with the help of a wide-field fluorescence microscopy approach. The fluorescence single molecules are observed in real time.

  10. En face coherence microscopy [Invited

    PubMed Central

    Thouvenin, Olivier; Grieve, Kate; Xiao, Peng; Apelian, Clement; Boccara, A. Claude

    2017-01-01

    En face coherence microscopy or flying spot or full field optical coherence tomography or microscopy (FF-OCT/FF-OCM) belongs to the OCT family because the sectioning ability is mostly linked to the source coherence length. In this article we will focus our attention on the advantages and the drawbacks of the following approaches: en face versus B scan tomography in terms of resolution, coherent versus incoherent illumination and influence of aberrations, and scanning versus full field imaging. We then show some examples to illustrate the diverse applications of en face coherent microscopy and show that endogenous or exogenous contrasts can add valuable information to the standard morphological image. To conclude we discuss a few domains that appear promising for future development of en face coherence microscopy. PMID:28270972

  11. Computer microscopy in lymphoma diagnostics

    NASA Astrophysics Data System (ADS)

    Mozhenkova, A. V.; Tupitsin, N. N.; Frenkel, M. A.; Falaleeva, N. A.; Nikitaev, V. G.; Polyakov, E. V.

    2017-01-01

    The article describes the application of computer microscopy with multi-spectral camera for the comparative characteristics of normal lymphocytes and lymphoid cells in follicular lymphoma. Wavelet functions are used to quantify parameters of the cells nuclei images.

  12. Vertically scanned laser sheet microscopy.

    PubMed

    Dong, Di; Arranz, Alicia; Zhu, Shouping; Yang, Yujie; Shi, Liangliang; Wang, Jun; Shen, Chen; Tian, Jie; Ripoll, Jorge

    2014-01-01

    Laser sheet microscopy is a widely used imaging technique for imaging the three-dimensional distribution of a fluorescence signal in fixed tissue or small organisms. In laser sheet microscopy, the stripe artifacts caused by high absorption or high scattering structures are very common, greatly affecting image quality. To solve this problem, we report here a two-step procedure which consists of continuously acquiring laser sheet images while vertically displacing the sample, and then using the variational stationary noise remover (VSNR) method to further reduce the remaining stripes. Images from a cleared murine colon acquired with a vertical scan are compared with common stitching procedures demonstrating that vertically scanned light sheet microscopy greatly improves the performance of current light sheet microscopy approaches without the need for complex changes to the imaging setup and allows imaging of elongated samples, extending the field of view in the vertical direction.

  13. Magnetic Force Microscopy in Liquids.

    PubMed

    Ares, Pablo; Jaafar, Miriam; Gil, Adriana; Gómez-Herrero, Julio; Asenjo, Agustina

    2015-09-01

    In this work, the use of magnetic force microscopy (MFM) to acquire images of magnetic nanostructures in liquid environments is presented. Optimization of the MFM signal acquisition in liquid media is performed and it is applied to characterize the magnetic signal of magnetite nanoparticles. The ability for detecting magnetic nanostructures along with the well-known capabilities of atomic force microscopy in liquids suggests potential applications in fields such as nanomedicine, nanobiotechnology, or nanocatalysis.

  14. Confocal microscopy in transmitted light

    NASA Astrophysics Data System (ADS)

    Dodt, Hans-Ulrich; Becker, Klaus

    2003-10-01

    We developed a confocal microscope for transmitted light to visualize fine details in phase objects like unstained biological specimens. The main difficulty of confocal microscopy in transmission is the alignment of illumination and detector pinholes. This alignment was achieved by using "electronic pinholes" on the detector side. As a first step, we were able to image cells in onion skin at greater depths and with higher resolution than by using conventional microscopy.

  15. Low Voltage Scanning Electron Microscopy

    DTIC Science & Technology

    1988-10-01

    Microscopy List of Keywords ,Scanning electron microscopy SEM X -ray .Micoranalysis EDX/EDS -%Low voltage , High resolution -Ceramic surfaces Supported...energy component normal to the surface). (a) Applications to x -ray microanalysis The essential problem leading to the specification of a LVSEM is...illustrated (Fig.l), for a conventional microprobe operated with 20nA probe current, by the contrast of the alumunium (K) x -ray signal as the probe is scanned

  16. Structured line illumination Raman microscopy

    PubMed Central

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

    2015-01-01

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

  17. Computational 'microscopy' of cellular membranes.

    PubMed

    Ingólfsson, Helgi I; Arnarez, Clément; Periole, Xavier; Marrink, Siewert J

    2016-01-15

    Computational 'microscopy' refers to the use of computational resources to simulate the dynamics of a molecular system. Tuned to cell membranes, this computational 'microscopy' technique is able to capture the interplay between lipids and proteins at a spatio-temporal resolution that is unmatched by other methods. Recent advances allow us to zoom out from individual atoms and molecules to supramolecular complexes and subcellular compartments that contain tens of millions of particles, and to capture the complexity of the crowded environment of real cell membranes. This Commentary gives an overview of the main concepts of computational 'microscopy' and describes the state-of-the-art methods used to model cell membrane processes. We illustrate the power of computational modelling approaches by providing a few in-depth examples of large-scale simulations that move up from molecular descriptions into the subcellular arena. We end with an outlook towards modelling a complete cell in silico.

  18. Correlative microscopy of detergent granules.

    PubMed

    van Dalen, G; Nootenboom, P; Heussen, P C M

    2011-03-01

    The microstructure of detergent products for textile cleaning determines to a large extent the physical properties of these products. Correlative microscopy was used to reveal the microstructure by reconciling images obtained by scanning electron microscopy with energy dispersive X-ray analysis, X-ray microtomography and Fourier transform infrared microscopy. These techniques were applied on the same location of a subsample of a spray-dried detergent base powder embedded in polyacrylate. In this way, the three-dimensional internal and external structure of detergent granules could be investigated from milli to nano scale with detailed spatial information about the components present. This will generate knowledge how to design optimal microstructures for laundry products to obtain product properties demanded by the market. This method is also very useful for other powder systems used in a large variety of industries (e.g. for pharmaceutical, food, ceramic and metal industries).

  19. Confocal microscopy and exfoliative cytology

    PubMed Central

    Reddy, Shyam Prasad; Ramani, Pratibha; Nainani, Purshotam

    2013-01-01

    Context: Early detection of potentially malignant lesions and invasive squamous-cell carcinoma in the oral cavity could be greatly improved through techniques that permit visualization of subtle cellular changes indicative of the neoplastic transformation process. One such technique is confocal microscopy. Combining rapidity with reliability, an innovative idea has been put forward using confocal microscope in exfoliative cytology. Aims: The main objective of this study was to assess confocal microscopy for cytological diagnosis and the results were compared with that of the standard PAP stain. Settings and Design: Confocal microscope, acridine orange (AO) stain, PAP (Papanicolaou) stain. The study was designed to assess confocal microscopy for cytological diagnosis. In the process, smears of patients with (clinically diagnosed and/or suspected) oral squamous cell carcinoma as well as those of controls (normal people) were stained with acridine orange and observed under confocal microscope. The results were compared with those of the standard PAP method. Materials and Methods: Samples of buccal mucosa smears from normal patients and squamous cell carcinoma patients were made, fixed in 100% alcohol, followed by AO staining. The corresponding set of smears was stained with PAP stain using rapid PAP stain kit. The results obtained were compared with those obtained with AO confocal microscopy. Results: The study had shown nuclear changes (malignant cells) in the smears of squamous cell carcinoma patients as increased intensity of fluorescence of the nucleus, when observed under confocal microscope. Acridine orange confocal microscopy showed good amount of sensitivity and specificity (93%) in identifying malignant cells in exfoliative cytological smears. Conclusion: Confocal microscopy was found to have good sensitivity in the identification of cancer (malignant) cells in exfoliative cytology, at par with the PAP method. The rapidity of processing and screening a

  20. DHMI: dynamic holographic microscopy interface

    NASA Astrophysics Data System (ADS)

    He, Xuefei; Zheng, Yujie; Lee, Woei Ming

    2016-12-01

    Digital holographic microscopy (DHM) is a powerful in-vitro biological imaging tool. In this paper, we report a fully automated off-axis digital holographic microscopy system completed with a graphical user interface in the Matlab environment. The interface primarily includes Fourier domain processing, phase reconstruction, aberration compensation and autofocusing. A variety of imaging operations such as region of interest selection, de-noising mode (filtering and averaging), low frame rate imaging for immediate reconstruction and high frame rate imaging routine ( 27 fps) are implemented to facilitate ease of use.

  1. The future of electron microscopy

    SciTech Connect

    Zhu, Yimei; Durr, Hermann

    2015-04-01

    Seeing is believing. So goes the old adage and seen evidence is undoubtedly satisfying because it can be interpreted easily, though not always correctly. For centuries, humans have developed such instruments as telescopes that observe the heavens and microscopes that reveal bacteria and viruses. The 2014 Nobel Prize in Chemistry was awarded to Eric Betzig, Stefan Hell, and William Moerner for their foundational work on superresolution fluorescence microscopy in which they overcame the Abbe diffraction limit for the resolving power of conventional light microscopes. (See Physics Today, December 2014, page 18.) That breakthrough enabled discoveries in biological research and testifies to the importance of modern microscopy.

  2. The future of electron microscopy

    DOE PAGES

    Zhu, Yimei; Durr, Hermann

    2015-04-01

    Seeing is believing. So goes the old adage and seen evidence is undoubtedly satisfying because it can be interpreted easily, though not always correctly. For centuries, humans have developed such instruments as telescopes that observe the heavens and microscopes that reveal bacteria and viruses. The 2014 Nobel Prize in Chemistry was awarded to Eric Betzig, Stefan Hell, and William Moerner for their foundational work on superresolution fluorescence microscopy in which they overcame the Abbe diffraction limit for the resolving power of conventional light microscopes. (See Physics Today, December 2014, page 18.) That breakthrough enabled discoveries in biological research and testifiesmore » to the importance of modern microscopy.« less

  3. Optical microscopy versus scanning electron microscopy in urolithiasis.

    PubMed

    Marickar, Y M Fazil; Lekshmi, P R; Varma, Luxmi; Koshy, Peter

    2009-10-01

    Stone analysis is incompletely done in many clinical centers. Identification of the stone component is essential for deciding future prophylaxis. X-ray diffraction, Fourier transform infrared spectroscopy, and scanning electron microscopy (SEM) still remains a distant dream for routine hospital work. It is in this context that optical microscopy is suggested as an alternate procedure. The objective of this article was to assess the utility of an optical microscope which gives magnification of up to 40x and gives clear picture of the surface of the stones. In order to authenticate the morphological analysis of urinary stones, SEM and elemental distribution analysis were performed. A total of 250 urinary stones of different compositions were collected from stone clinic, photographed, observed under an optical microscope, and optical photographs were taken at different angles. Twenty-five representative samples among these were gold sputtered to make them conductive and were fed into the SEM machine. Photographs of the samples were taken at different angles at magnifications up to 4,000. Elemental distribution analysis (EDAX) was done to confirm the composition. The observations of the two studies were compared. The different appearances of the stones under optical illuminated microscopy were mostly standardized appearances, namely bosselations of pure whewellite, spiculations of weddellite, bright yellow colored appearance of uric acid, and dirty white amorphous appearance of phosphates. SEM and EDAX gave clearer pictures and gave added confirmation of the stone composition. From the references thus obtained, it was possible to confirm the composition by studying the optical microscopic pictures. Higher magnification capacity of the SEM and the EDAX patterns are useful to give reference support for performing optical microscopy work. After standardization, routine analysis can be performed with optical microscopy. The advantage of the optical microscope is that, it

  4. Color-televised medical microscopy

    NASA Technical Reports Server (NTRS)

    Heath, M. A.; Peck, J. C.

    1968-01-01

    Color television microscopy used at laboratory range magnifications, reproduces a slide image with sufficient fidelity for medical laboratory and instructional use. The system is used for instant pathological reporting between operating room and remotely located pathologist viewing a biopsy through this medium.

  5. Interferometric Synthetic Aperture Microscopy: Computed Imaging for Scanned Coherent Microscopy.

    PubMed

    Davis, Brynmor J; Marks, Daniel L; Ralston, Tyler S; Carney, P Scott; Boppart, Stephen A

    2008-06-01

    Three-dimensional image formation in microscopy is greatly enhanced by the use of computed imaging techniques. In particular, Interferometric Synthetic Aperture Microscopy (ISAM) allows the removal of out-of-focus blur in broadband, coherent microscopy. Earlier methods, such as optical coherence tomography (OCT), utilize interferometric ranging, but do not apply computed imaging methods and therefore must scan the focal depth to acquire extended volumetric images. ISAM removes the need to scan the focus by allowing volumetric image reconstruction from data collected at a single focal depth. ISAM signal processing techniques are similar to the Fourier migration methods of seismology and the Fourier reconstruction methods of Synthetic Aperture Radar (SAR). In this article ISAM is described and the close ties between ISAM and SAR are explored. ISAM and a simple strip-map SAR system are placed in a common mathematical framework and compared to OCT and radar respectively. This article is intended to serve as a review of ISAM, and will be especially useful to readers with a background in SAR.

  6. PoroTomo Project - Subatask 6.2: Deploy and Operate DAS and DTS arrays - DAS Earthquake Data

    SciTech Connect

    Kurt Feigl

    2016-03-21

    The submitted data correspond to the vibration caused by a 3.4 M earthquake and captured by the DAS horizontal and vertical arrays during the PoroTomo Experiment. Earthquake information : M 4.3 - 23km ESE of Hawthorne, Nevada Time: 2016-03-21 07:37:10 (UTC) Location: 38.479°N 118.366°W Depth: 9.9 km Files for horizontal DAS array (each file is 30 s long and contain 8700 channels): PoroTomo_iDAS16043_160321073721.sgy PoroTomo_iDAS16043_160321073751.sgy Files for vertical DAS Array (each file is 30 s long and contain 380 channels): PoroTomo_iDAS025_160321073717.sgy PoroTomo_iDAS025_160321073747.sgy

  7. Dynamic imaging with electron microscopy

    ScienceCinema

    Campbell, Geoffrey; McKeown, Joe; Santala, Melissa

    2016-07-12

    Livermore researchers have perfected an electron microscope to study fast-evolving material processes and chemical reactions. By applying engineering, microscopy, and laser expertise to the decades-old technology of electron microscopy, the dynamic transmission electron microscope (DTEM) team has developed a technique that can capture images of phenomena that are both very small and very fast. DTEM uses a precisely timed laser pulse to achieve a short but intense electron beam for imaging. When synchronized with a dynamic event in the microscope's field of view, DTEM allows scientists to record and measure material changes in action. A new movie-mode capability, which earned a 2013 R&D 100 Award from R&D Magazine, uses up to nine laser pulses to sequentially capture fast, irreversible, even one-of-a-kind material changes at the nanometer scale. DTEM projects are advancing basic and applied materials research, including such areas as nanostructure growth, phase transformations, and chemical reactions.

  8. Materials science through electron microscopy

    NASA Astrophysics Data System (ADS)

    Fujita, Hiroshi

    1992-03-01

    Electron microscopy has greatly contributed as a powerful tool in both the characterization and identification of materials in the atomic scale. In these contributions, the most important advantage is it's ability for dynamic study of phenomena, i.e., in situ experiments. This research has been carried out using high voltage electron microscopes, but some results have been obtained with high resolution electron microscopes under critical conditions. Electron microscopy has been improved further to become an indispensable ?Micro-Laboratory? in which formation of various advance materials can also be carried out precisely in the atomic scale. Electron beam science and engineering is a typical example in this research field, and detailed processes of crystalline-amorphous transition and electron irradiation induced foreign atom implantation have been clarified by this method. Recently, new applications to the research fields of non-linear material behavior, such as the behavior of atom clusters and the role of electric dipoles on diffusion, have been carried out.

  9. Electron microscopy of electromagnetic waveforms

    NASA Astrophysics Data System (ADS)

    Ryabov, A.; Baum, P.

    2016-07-01

    Rapidly changing electromagnetic fields are the basis of almost any photonic or electronic device operation. We report how electron microscopy can measure collective carrier motion and fields with subcycle and subwavelength resolution. A collimated beam of femtosecond electron pulses passes through a metamaterial resonator that is previously excited with a single-cycle electromagnetic pulse. If the probing electrons are shorter in duration than half a field cycle, then time-frozen Lorentz forces distort the images quasi-classically and with subcycle time resolution. A pump-probe sequence reveals in a movie the sample’s oscillating electromagnetic field vectors with time, phase, amplitude, and polarization information. This waveform electron microscopy can be used to visualize electrodynamic phenomena in devices as small and fast as available.

  10. Contact microscopy with synchrotron radiation

    SciTech Connect

    Panessa-Warren, B.J.

    1985-10-01

    Soft x-ray contact microscopy with synchrotron radiation offers the biologist and especially the microscopist, a way to morphologically study specimens that could not be imaged by conventional TEM, STEM or SEM methods (i.e. hydrated samples, samples easily damaged by an electron beam, electron dense samples, thick specimens, unstained low contrast specimens) at spatial resolutions approaching those of the TEM, with the additional possibility to obtain compositional (elemental) information about the sample as well. Although flash x-ray sources offer faster exposure times, synchrotron radiation provides a highly collimated, intense radiation that can be tuned to select specific discrete ranges of x-ray wavelengths or specific individual wavelengths which optimize imaging or microanalysis of a specific sample. This paper presents an overview of the applications of x-ray contact microscopy to biological research and some current research results using monochromatic synchrotron radiation to image biological samples. 24 refs., 10 figs.

  11. Dynamic imaging with electron microscopy

    SciTech Connect

    Campbell, Geoffrey; McKeown, Joe; Santala, Melissa

    2014-02-20

    Livermore researchers have perfected an electron microscope to study fast-evolving material processes and chemical reactions. By applying engineering, microscopy, and laser expertise to the decades-old technology of electron microscopy, the dynamic transmission electron microscope (DTEM) team has developed a technique that can capture images of phenomena that are both very small and very fast. DTEM uses a precisely timed laser pulse to achieve a short but intense electron beam for imaging. When synchronized with a dynamic event in the microscope's field of view, DTEM allows scientists to record and measure material changes in action. A new movie-mode capability, which earned a 2013 R&D 100 Award from R&D Magazine, uses up to nine laser pulses to sequentially capture fast, irreversible, even one-of-a-kind material changes at the nanometer scale. DTEM projects are advancing basic and applied materials research, including such areas as nanostructure growth, phase transformations, and chemical reactions.

  12. Rotational scanning atomic force microscopy.

    PubMed

    Ulčinas, A; Vaitekonis, Š

    2017-03-10

    A non-raster scanning technique for atomic force microscopy (AFM) imaging which combines rotational and translational motion is presented. The use of rotational motion for the fast scan axis allows us to significantly increase the scanning speed while imaging a large area (diameter > 30 μm). An image reconstruction algorithm and the factors influencing the resolution of the technique are discussed. The experimental results show the potential of the rotational scanning technique for high-throughput large area AFM investigation.

  13. Hyperspectral holographic Fourier-microscopy

    SciTech Connect

    Kalenkov, G S; Kalenkov, S G; Shtan'ko, A E

    2015-04-30

    A detailed theory of the method of holographic recording of hyperspectral wave fields is developed. New experimentally obtained hyperspectral holographic images of microscopic objects are presented. The possibilities of the method are demonstrated experimentally using the examples of urgent microscopy problems: speckle noise suppression, obtaining hyperspectral image of a microscopic object, as well as synthesis of a colour image and obtaining an optical profile of a phase object. (holography)

  14. Rotational scanning atomic force microscopy

    NASA Astrophysics Data System (ADS)

    Ulčinas, A.; Vaitekonis, Š.

    2017-03-01

    A non-raster scanning technique for atomic force microscopy (AFM) imaging which combines rotational and translational motion is presented. The use of rotational motion for the fast scan axis allows us to significantly increase the scanning speed while imaging a large area (diameter > 30 μm). An image reconstruction algorithm and the factors influencing the resolution of the technique are discussed. The experimental results show the potential of the rotational scanning technique for high-throughput large area AFM investigation.

  15. A history of urine microscopy.

    PubMed

    Cameron, J Stewart

    2015-11-01

    The naked-eye appearance of the urine must have been studied by shamans and healers since the Stone Age, and an elaborate interpretation of so-called Uroscopy began around 600 AD as a form of divination. A 1000 years later, the first primitive monocular and compound microscopes appeared in the Netherlands, and along with many other objects and liquids, urine was studied from around 1680 onwards as the enlightenment evolved. However, the crude early instruments did not permit fine study because of chromatic and linear/spherical blurring. Only after complex multi-glass lenses which avoided these problems had been made and used in the 1820s in London by Lister, and in Paris by Chevalier and Amici, could urinary microscopy become a practical, clinically useful tool in the 1830s. Clinical urinary microscopy was pioneered by Rayer and his pupils in Paris (especially Vigla), in the late 1830s, and spread to UK and Germany in the 1840s, with detailed descriptions and interpretations of cells and formed elements of the urinary sediment by Nasse, Henle, Robinson and Golding Bird. Classes were held, most notably by Donné in Paris. After another 50 years, optical microscopy had reached its apogee, with magnifications of over 1000 times obtainable free of aberration, using immersion techniques. Atlases of the urinary sediment were published in all major European countries and in the US. Polarised light and phase contrast was used also after 1900 to study urine, and by the early 20th century, photomicroscopy (pioneered by Donné and Daguerre 50 years previously, but then ignored) became usual for teaching and recording. In the 1940s electron microscopy began, followed by detection of specific proteins and cells using immunofluorescent antibodies. All this had been using handheld methodology. Around 1980, machine-assisted observations began, and have dominated progress since.

  16. Paleomagnetic Analysis Using SQUID Microscopy

    NASA Technical Reports Server (NTRS)

    Weiss, Benjamin P.; Lima, Eduardo A.; Fong, Luis E.; Baudenbacher, Franz J.

    2007-01-01

    Superconducting quantum interference device (SQUID) microscopes are a new generation of instruments that map magnetic fields with unprecedented spatial resolution and moment sensitivity. Unlike standard rock magnetometers, SQUID microscopes map magnetic fields rather than measuring magnetic moments such that the sample magnetization pattern must be retrieved from source model fits to the measured field data. In this paper, we presented the first direct comparison between paleomagnetic analyses on natural samples using joint measurements from SQUID microscopy and moment magnetometry. We demonstrated that in combination with apriori geologic and petrographic data, SQUID microscopy can accurately characterize the magnetization of lunar glass spherules and Hawaiian basalt. The bulk moment magnitude and direction of these samples inferred from inversions of SQUID microscopy data match direct measurements on the same samples using moment magnetometry. In addition, these inversions provide unique constraints on the magnetization distribution within the sample. These measurements are among the most sensitive and highest resolution quantitative paleomagnetic studies of natural remanent magnetization to date. We expect that this technique will be able to extend many other standard paleomagnetic techniques to previously inaccessible microscale samples.

  17. Multi-photon excitation microscopy

    PubMed Central

    Diaspro, Alberto; Bianchini, Paolo; Vicidomini, Giuseppe; Faretta, Mario; Ramoino, Paola; Usai, Cesare

    2006-01-01

    Multi-photon excitation (MPE) microscopy plays a growing role among microscopical techniques utilized for studying biological matter. In conjunction with confocal microscopy it can be considered the imaging workhorse of life science laboratories. Its roots can be found in a fundamental work written by Maria Goeppert Mayer more than 70 years ago. Nowadays, 2PE and MPE microscopes are expected to increase their impact in areas such biotechnology, neurobiology, embryology, tissue engineering, materials science where imaging can be coupled to the possibility of using the microscopes in an active way, too. As well, 2PE implementations in noninvasive optical bioscopy or laser-based treatments point out to the relevance in clinical applications. Here we report about some basic aspects related to the phenomenon, implications in three-dimensional imaging microscopy, practical aspects related to design and realization of MPE microscopes, and we only give a list of potential applications and variations on the theme in order to offer a starting point for advancing new applications and developments. PMID:16756664

  18. Multi-photon excitation microscopy.

    PubMed

    Diaspro, Alberto; Bianchini, Paolo; Vicidomini, Giuseppe; Faretta, Mario; Ramoino, Paola; Usai, Cesare

    2006-06-06

    Multi-photon excitation (MPE) microscopy plays a growing role among microscopical techniques utilized for studying biological matter. In conjunction with confocal microscopy it can be considered the imaging workhorse of life science laboratories. Its roots can be found in a fundamental work written by Maria Goeppert Mayer more than 70 years ago. Nowadays, 2PE and MPE microscopes are expected to increase their impact in areas such biotechnology, neurobiology, embryology, tissue engineering, materials science where imaging can be coupled to the possibility of using the microscopes in an active way, too. As well, 2PE implementations in noninvasive optical bioscopy or laser-based treatments point out to the relevance in clinical applications. Here we report about some basic aspects related to the phenomenon, implications in three-dimensional imaging microscopy, practical aspects related to design and realization of MPE microscopes, and we only give a list of potential applications and variations on the theme in order to offer a starting point for advancing new applications and developments.

  19. Holographic opto-fluidic microscopy.

    PubMed

    Bishara, Waheb; Zhu, Hongying; Ozcan, Aydogan

    2010-12-20

    Over the last decade microfluidics has created a versatile platform that has significantly advanced the ways in which micro-scale organisms and objects are controlled, processed and investigated, by improving the cost, compactness and throughput aspects of analysis. Microfluidics has also expanded into optics to create reconfigurable and flexible optical devices such as reconfigurable lenses, lasers, waveguides, switches, and on-chip microscopes. Here we present a new opto-fluidic microscopy modality, i.e., Holographic Opto-fluidic Microscopy (HOM), based on lensless holographic imaging. This imaging modality complements the miniaturization provided by microfluidics and would allow the integration of microscopy into existing on-chip microfluidic devices with various functionalities. Our imaging modality utilizes partially coherent in-line holography and pixel super-resolution to create high-resolution amplitude and phase images of the objects flowing within micro-fluidic channels, which we demonstrate by imaging C. elegans, Giardia lamblia, and Mulberry pollen. HOM does not involve complicated fabrication processes or precise alignment, nor does it require a highly uniform flow of objects within microfluidic channels.

  20. Multiphoton microscopy of atheroslcerotic plaques

    NASA Astrophysics Data System (ADS)

    Lilledahl, Magnus B.; de Lange Davies, Catharina; Haugen, Olav A.; Svaasand, Lars O.

    2007-02-01

    Multiphoton microscopy is a techniques that fascilitates three dimensional imaging of intact, unstained tissue. Especially connective tissue has a relatively strong nonlinear optical response and can easily be imaged. Atherosclerosis is a disease where lipids accumulate in the vessel wall and there is a thickening of the intima by growth of a cap of connective tissue. The mechanical strength of this fibrous cap is of clinically importance. If the cap ruptures a thrombosis forms which can block a coronary vessel and therby causing myocardial infarction. Multiphoton microscopy can be used to image the fibrous cap and thereby determine the thickness of the cap and the structure of the connective fibres. This could possibly be developed into a diagnostic and clincal tool to monitor the vulnerability of a plaque and also to better understand the development of a plaque and effects of treatment. We have collected multiphoton microscopy images from atherosclerotic plaque in human aorta, both two photon excited fluorescens and second harmonic generated signal. The feasability of using this technique to determine the state of the plaque is explored.

  1. 3D structured illumination microscopy

    NASA Astrophysics Data System (ADS)

    Dougherty, William M.; Goodwin, Paul C.

    2011-03-01

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

  2. Electron Microscopy of Living Cells During in Situ Fluorescence Microscopy

    PubMed Central

    Liv, Nalan; van Oosten Slingeland, Daan S. B.; Baudoin, Jean-Pierre; Kruit, Pieter; Piston, David W.; Hoogenboom, Jacob P.

    2016-01-01

    We present an approach toward dynamic nanoimaging: live fluorescence of cells encapsulated in a bionanoreactor is complemented with in situ scanning electron microscopy (SEM) on an integrated microscope. This allows us to take SEM snapshots on-demand, that is, at a specific location in time, at a desired region of interest, guided by the dynamic fluorescence imaging. We show that this approach enables direct visualization, with EM resolution, of the distribution of bioconjugated quantum dots on cellular extensions during uptake and internalization. PMID:26580231

  3. Fluorescence-integrated transmission electron microscopy images: integrating fluorescence microscopy with transmission electron microscopy.

    PubMed

    Sims, Paul A; Hardin, Jeff D

    2007-01-01

    This chapter describes high-pressure freezing (HPF) techniques for correlative light and electron microscopy on the same sample. Laser scanning confocal microscopy (LSCM) is exploited for its ability to collect fluorescent, as well as transmitted and back scattered light (BSL) images at the same time. Fluorescent information from a whole mount (preembedding) or from thin sections (post-embedding) can be displayed as a color overlay on transmission electron microscopy (TEM) images. Fluorescence-integrated TEM (F-TEM) images provide a fluorescent perspective to TEM images. The pre-embedding method uses a thin two-part agarose pad to immobilize live Caenorhabditis elegans embryos for LSCM, HPF, and TEM. Pre-embedding F-TEM images display fluorescent information collected from a whole mount of live embryos onto all thin sections collected from that sample. In contrast, the postembedding method uses HPF and freeze substitution with 1% paraformaldehyde in 95% ethanol followed by low-temperature embedding in methacrylate resin. This procedure preserves the structure and function of green fluorescent protein (GFP) as determined by immunogold labeling of GFP, when compared with GFP expression, both demonstrated in the same thin section.

  4. Combining fluorescence and bioluminescence microscopy.

    PubMed

    Goda, Kazuhito; Hatta-Ohashi, Yoko; Akiyoshi, Ryutaro; Sugiyama, Takashi; Sakai, Ikuko; Takahashi, Takeo; Suzuki, Hirobumi

    2015-08-01

    Bioluminescence microscopy has revealed that gene expression in individual cells can respond differently to the same stimulus. To understand this phenomenon, it is important to sequentially observe the series of events from cellular signal transduction to gene expression regulated by specific transcription factors derived from signaling cascades in individual cells. However, these processes have been separately analyzed with fluorescence and bioluminescence microscopy. Furthermore, in culture medium, the background fluorescence of luciferin-a substrate of luciferase in promoter assays of gene expression in cultured cells-confounds the simultaneous observation of fluorescence and bioluminescence. Therefore, we optimized conditions for optical filter sets based on spectral properties and the luciferin concentration based on cell permeability for fluorescence observation combined with bioluminescence microscopy. An excitation and emission filter set (492-506 nm and 524-578 nm) was suitable for green fluorescent protein and yellow fluorescent protein imaging of cells, and >100 μM luciferin was acceptable in culture medium based on kinetic constants and the estimated intracellular concentration. Using these parameters, we present an example of sequential fluorescence and bioluminescence microscopic observation of signal transduction (translocation of protein kinase C alpha from the cytoplasm to the plasma membrane) coupled with activation of gene expression by nuclear factor of kappa light polypeptide B in individual cells and show that the gene expression response is not completely concordant with upstream signaling following stimulation with phorbol-12-myristate-13-acetate. Our technique is a powerful imaging tool for analysis of heterogeneous gene expression together with upstream signaling in live single cells.

  5. Twin-photon confocal microscopy.

    PubMed

    Simon, D S; Sergienko, A V

    2010-10-11

    A recently introduced two-channel confocal microscope with correlated detection promises up to 50% improvement in transverse spatial resolution [Simon, Sergienko, Optics Express 18, 9765 (2010)] via the use of photon correlations. Here we achieve similar results in a different manner, introducing a triple-confocal correlated microscope which exploits the correlations present in optical parametric amplifiers. It is based on tight focusing of pump radiation onto a thin sample positioned in front of a nonlinear crystal, followed by coincidence detection of signal and idler photons, each focused onto a pinhole. This approach offers further resolution enhancement in confocal microscopy.

  6. Superresolved spatially multiplexed interferometric microscopy.

    PubMed

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

    2017-03-01

    Superresolution capability by angular and time multiplexing is implemented onto a regular microscope. The technique, named superresolved spatially multiplexed interferometric microscopy (S2MIM), follows our previously reported SMIM technique [Opt. Express22, 14929 (2014)OPEXFF1094-408710.1364/OE.22.014929, J. Biomed. Opt.21, 106007 (2016)JBOPFO1083-366810.1117/1.JBO.21.10.106007] improved with superresolved imaging. All together, S2MIM updates a commercially available non-holographic microscope into a superresolved holographic one. Validation is presented for an Olympus BX-60 upright microscope with resolution test targets.

  7. Confocal microscopy in microgravity research.

    PubMed

    Goede, A P; Brakenhoff, G J; Woldringh, C L; Aalders, J W; Imhof, J P; van Kralingen, P; Mels, W A; Schreinemakers, P; Zegers, A

    1992-01-01

    We have studied the application and the feasibility of confocal scanning laser microscopy (CSLM) in microgravity research. Its superior spatial resolution and 3D imaging capabilities and its use of light as a probe, render this instrument ideally suited for the study of living biological material on a (sub-)cellular level. In this paper a number of pertinent biological microgravity experiments is listed, concentrating on the direct observation of developing cells and cellular structures under microgravity condition. A conceptual instrument design is also presented, aimed at sounding rocket application followed by Biorack/Biolab application at a later stage.

  8. Scanning probe microscopy in catalysis.

    PubMed

    Yeung, King Lun; Yao, Nan

    2004-09-01

    This review discusses the recent progress in the application of scanning probe microscopy (SPM) in catalysis. SPM proves to be an invaluable technique for imaging catalytic surfaces and interfaces. Most SPM research is related to the structural and morphological transformation associated with catalyst preparation and use. Real-time SPM observation of surface dynamics including adsorption, diffusion and reaction, provides invaluable insights to the mechanism of catalysis. SPM is also used to shape and manipulate surfaces and surface processes. Fabrication of nanostructured catalysts, direct manipulation of adsorbed atoms and molecules and tip-mediated reactions are some examples of new SPM approach in catalyst research.

  9. Visual-servoing optical microscopy

    DOEpatents

    Callahan, Daniel E.; Parvin, Bahram

    2009-06-09

    The present invention provides methods and devices for the knowledge-based discovery and optimization of differences between cell types. In particular, the present invention provides visual servoing optical microscopy, as well as analysis methods. The present invention provides means for the close monitoring of hundreds of individual, living cells over time: quantification of dynamic physiological responses in multiple channels; real-time digital image segmentation and analysis; intelligent, repetitive computer-applied cell stress and cell stimulation; and the ability to return to the same field of cells for long-term studies and observation. The present invention further provides means to optimize culture conditions for specific subpopulations of cells.

  10. Visual-servoing optical microscopy

    DOEpatents

    Callahan, Daniel E; Parvin, Bahram

    2013-10-01

    The present invention provides methods and devices for the knowledge-based discovery and optimization of differences between cell types. In particular, the present invention provides visual servoing optical microscopy, as well as analysis methods. The present invention provides means for the close monitoring of hundreds of individual, living cells over time; quantification of dynamic physiological responses in multiple channels; real-time digital image segmentation and analysis; intelligent, repetitive computer-applied cell stress and cell stimulation; and the ability to return to the same field of cells for long-term studies and observation. The present invention further provides means to optimize culture conditions for specific subpopulations of cells.

  11. Visual-servoing optical microscopy

    DOEpatents

    Callahan, Daniel E.; Parvin, Bahram

    2011-05-24

    The present invention provides methods and devices for the knowledge-based discovery and optimization of differences between cell types. In particular, the present invention provides visual servoing optical microscopy, as well as analysis methods. The present invention provides means for the close monitoring of hundreds of individual, living cells over time; quantification of dynamic physiological responses in multiple channels; real-time digital image segmentation and analysis; intelligent, repetitive computer-applied cell stress and cell stimulation; and the ability to return to the same field of cells for long-term studies and observation. The present invention further provides means to optimize culture conditions for specific subpopulations of cells.

  12. Aperture scanning Fourier ptychographic microscopy

    PubMed Central

    Ou, Xiaoze; Chung, Jaebum; Horstmeyer, Roarke; Yang, Changhuei

    2016-01-01

    Fourier ptychographic microscopy (FPM) is implemented through aperture scanning by an LCOS spatial light modulator at the back focal plane of the objective lens. This FPM configuration enables the capturing of the complex scattered field for a 3D sample both in the transmissive mode and the reflective mode. We further show that by combining with the compressive sensing theory, the reconstructed 2D complex scattered field can be used to recover the 3D sample scattering density. This implementation expands the scope of application for FPM and can be beneficial for areas such as tissue imaging and wafer inspection. PMID:27570705

  13. Image scanning microscopy with radially polarized light

    NASA Astrophysics Data System (ADS)

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

    2017-03-01

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

  14. Virtual intraoperative surgical photoacoustic microscopy

    NASA Astrophysics Data System (ADS)

    Lee, Changho; Lee, Donghyun; Zhou, Qifa; Kim, Jeehyun; Kim, Chulhong

    2015-07-01

    A virtual intraoperative surgical photoacoustic microscopy at 1064 nm wavelength (VISPAM) system was designed and fabricated by integrating a commercial type surgical microscope and laser scanning photoacoustic microscopy (PAM) with a 1064 nm pulsed laser. Based on simple augmented reality device, VISPAM could simultaneously provide 2D depth-resolved photoacoustic and magnified microscope images of surgery regions on the same vision of surgeon via an eyepiece of the microscope. The invisible 1064 nm laser removed the interruption of surgical sight due to visible laser scanning of previous report, and decreased the danger of tissue damage caused by over irradiated laser. In addition, to approach the real practical surgery application, a needle-type transducer was utilized without a water bath for PA signal coupling. In order to verify our system's performance, we conducted needle guiding as ex vivo phantom study and needle guiding and injection of carbon particles mixtures into a melanoma tumor region as in vivo study. We expect that VISPAM can be essential tool of brain and ophthalmic microsurgery.

  15. Developing Photo Activated Localization Microscopy

    NASA Astrophysics Data System (ADS)

    Hess, Harald

    2015-03-01

    Photo Activated Localization Microscopy, PALM, acquires super-resolution images by activating a subset of activatable fluorescent labels and estimating the center of the each molecular label to sub-diffractive accuracy. When this process is repeated thousands of times for different subsets of molecules, then an image can be rendered from all the center coordinates of the molecules. I will describe the circuitous story of its development that began with another super-resolution technique, NSOM, developed by my colleague Eric Betzig, who imaged single molecules at room temperature, and later we spectrally resolved individual luminescent centers of quantum wells. These two observations inspired a generalized path to localization microscopy, but that path was abandoned because no really useful fluorescent labels were available. After a decade of nonacademic industrial pursuits and the subsequent freedom of unemployment, we came across a class of genetically expressible fluorescent proteins that were switchable or convertible that enabled the concept to be implemented and be biologically promising. The past ten years have been very active with many groups exploring applications and enhancements of this concept. Demonstrating significant biological relevance will be the metric if its success.

  16. RGB digital lensless holographic microscopy

    NASA Astrophysics Data System (ADS)

    Garcia-Sucerquia, Jorge

    2013-11-01

    The recent introduction of color digital lensless holographic microscopy (CDLHM) has shown the possibility of imaging microscopic specimens at full color without the need of lenses. Owing to the simplicity, robustness, and compactness of the digital lensless holographic microscopes (DLHM), they have been presented as the ideal candidates to being developed into portable holographic microscopes. However, in the case of CDLHM the utilization of three independent lasers hinders the portability option for this microscope. In this contribution an alternative to reduce the complexity of CDLHM aimed to recover the portability of this microscopy technology is presented. A super-bright white-light light-emitting diode (LED) is spectrally and spatially filtered to produce the needed illumination by CDLHM to work. CDLHM with LED illumination is used to image at full color a section of the head of a drosophila melanogaster fly (fruit fly). The LED-CDLHM method shows the capability of imaging objects of 2μm size in comparison with the micrometer resolution reported for LASER-CDLHM.

  17. Light Sheet Fluorescence Microscopy (LSFM)

    PubMed Central

    Adams, Michael W.; Loftus, Andrew F.; Dunn, Sarah E.; Joens, Matthew S.; Fitzpatrick, James A.J.

    2015-01-01

    The development of confocal microscopy techniques introduced the ability to optically section fluorescent samples in the axial dimension, perpendicular to the image plane. These approaches, via the placement of a pinhole in the conjugate image plane, provided superior resolution in the axial (z) dimension resulting in nearly isotropic optical sections. However, increased axial resolution, via pinhole optics, comes at the cost of both speed and excitation efficiency. Light Sheet Fluorescent Microscopy (LSFM), a century old idea (Siedentopf and Zsigmondy, 1902) made possible with modern developments in both excitation and detection optics, provides sub-cellular resolution and optical sectioning capabilities without compromising speed or excitation efficiency. Over the past decade, several variations of LSFM have been implemented each with its own benefits and deficiencies. Here we discuss LSFM fundamentals and outline the basic principles of several major light sheet based imaging modalities (SPIM, inverted SPIM, multi-view SPIM, Bessel beam SPIM, and stimulated emission depletion SPIM while considering their biological relevance in terms of intrusiveness, temporal resolution, and sample requirements. PMID:25559221

  18. Atomic force microscopy as nanorobot.

    PubMed

    Xi, Ning; Fung, Carmen Kar Man; Yang, Ruiguo; Lai, King Wai Chiu; Wang, Donna H; Seiffert-Sinha, Kristina; Sinha, Animesh A; Li, Guangyong; Liu, Lianqing

    2011-01-01

    Atomic force microscopy (AFM) is a powerful and widely used imaging technique that can visualize single molecules under physiological condition at the nanometer scale. In this chapter, an AFM-based nanorobot for biological studies is introduced. Using the AFM tip as an end effector, the AFM can be modified into a nanorobot that can manipulate biological objects at the single-molecule level. By functionalizing the AFM tip with specific antibodies, the nanorobot is able to identify specific types of receptors on the cell membrane. It is similar to the fluorescent optical microscopy but with higher resolution. By locally updating the AFM image based on interaction force information and objects' model during nanomanipulation, real-time visual feedback is obtained through the augmented reality interface. The development of the AFM-based nanorobotic system enables us to conduct in situ imaging, sensing, and manipulation simultaneously at the nanometer scale (e.g., protein and DNA levels). The AFM-based nanorobotic system offers several advantages and capabilities for studying structure-function relationships of biological specimens. As a result, many biomedical applications can be achieved by the AFM-based nanorobotic system.

  19. Cluster computing for digital microscopy.

    PubMed

    Carrington, Walter A; Lisin, Dimitri

    2004-06-01

    Microscopy is becoming increasingly digital and dependent on computation. Some of the computational tasks in microscopy are computationally intense, such as image restoration (deconvolution), some optical calculations, image segmentation, and image analysis. Several modern microscope technologies enable the acquisition of very large data sets. 3D imaging of live cells over time, multispectral imaging, very large tiled 3D images of thick samples, or images from high throughput biology all can produce extremely large images. These large data sets place a very large burden on laboratory computer resources. This combination of computationally intensive tasks and larger data sizes can easily exceed the capability of single personal computers. The large multiprocessor computers that are the traditional technology for larger tasks are too expensive for most laboratories. An alternative approach is to use a number of inexpensive personal computers as a cluster; that is, use multiple networked computers programmed to run the problem in parallel on all the computers in the cluster. By the use of relatively inexpensive over-the-counter hardware and open source software, this approach can be much more cost effective for many tasks. We discuss the different computer architectures available, and their advantages and disadvantages.

  20. 3D multiplexed immunoplasmonics microscopy

    NASA Astrophysics Data System (ADS)

    Bergeron, Éric; Patskovsky, Sergiy; Rioux, David; Meunier, Michel

    2016-07-01

    Selective labelling, identification and spatial distribution of cell surface biomarkers can provide important clinical information, such as distinction between healthy and diseased cells, evolution of a disease and selection of the optimal patient-specific treatment. Immunofluorescence is the gold standard for efficient detection of biomarkers expressed by cells. However, antibodies (Abs) conjugated to fluorescent dyes remain limited by their photobleaching, high sensitivity to the environment, low light intensity, and wide absorption and emission spectra. Immunoplasmonics is a novel microscopy method based on the visualization of Abs-functionalized plasmonic nanoparticles (fNPs) targeting cell surface biomarkers. Tunable fNPs should provide higher multiplexing capacity than immunofluorescence since NPs are photostable over time, strongly scatter light at their plasmon peak wavelengths and can be easily functionalized. In this article, we experimentally demonstrate accurate multiplexed detection based on the immunoplasmonics approach. First, we achieve the selective labelling of three targeted cell surface biomarkers (cluster of differentiation 44 (CD44), epidermal growth factor receptor (EGFR) and voltage-gated K+ channel subunit KV1.1) on human cancer CD44+ EGFR+ KV1.1+ MDA-MB-231 cells and reference CD44- EGFR- KV1.1+ 661W cells. The labelling efficiency with three stable specific immunoplasmonics labels (functionalized silver nanospheres (CD44-AgNSs), gold (Au) NSs (EGFR-AuNSs) and Au nanorods (KV1.1-AuNRs)) detected by reflected light microscopy (RLM) is similar to the one with immunofluorescence. Second, we introduce an improved method for 3D localization and spectral identification of fNPs based on fast z-scanning by RLM with three spectral filters corresponding to the plasmon peak wavelengths of the immunoplasmonics labels in the cellular environment (500 nm for 80 nm AgNSs, 580 nm for 100 nm AuNSs and 700 nm for 40 nm × 92 nm AuNRs). Third, the developed

  1. Multiphoton fluorescence microscopy in biology

    NASA Astrophysics Data System (ADS)

    Heikal, Ahmed A.; Webb, Watt W.

    2002-11-01

    The inherent advantages of nonlinear excitation make multiphoton fluorescence microscopy (MPFM) awell-suited imaging technique for extracting valuable information from turbid and thick biological samples. These advantages include high three-dimensional spatial resolution, large penetration depth, minimum out-of-focus cellular photodamage, and high signal-to-noise contrast. We have investigated the nonlinear spectroscopy of biologically important molecules such as NADH, flavins, and intrinsically fluorescent proteins. Fundamental understanding of the molecular spectroscopy and dynamics of these biomolecules is essential for advancing their applications in biological research. MPFM has been utilized for monitoring a large spectrum of biological processes including metabolic activity and exocytosis. We will discuss two-photon (2P) redox fluorescence microscopy of NADH, which gives a quantitative measure of the respiratory chain activity, thus allowing functional imaging of energy metabolism in neurons and native brain tissue. Finally, a rational design strategy, based on donor-acceptor-donor configuration, will be elucidated for fluorescent probes with large 2P-excitation cross-section. These dyes are water-soluble, yet possess a high affinity to organic phases with site-specific labeling and Ca+2 sensitivity (Kd ~ 350 nM). A brief account on the biological application of nanocrystals and second harmonic imaging will be reviewed.

  2. Nanocharacterization: Atomic Scale Visualization with Microscopy

    NASA Astrophysics Data System (ADS)

    Broadbridge, Christine

    2007-10-01

    This workshop will include an overview presentation of nanotechnology and nanocharacterization tools (electron microscopy and atomic force microscopy) as well as examples of curricular components for middle and high school teachers. Tours/demonstrations of microscopy facilities in the IMS facility at UConn will be provided.

  3. Kelvin Probe Force Microscopy in liquid using Electrochemical Force Microscopy

    DOE PAGES

    Collins, Liam; Jesse, Stephen; Kilpatrick, J.; ...

    2015-01-01

    Conventional closed loop-Kelvin probe force microscopy (KPFM) has emerged as a powerful technique for probing electric and transport phenomena at the solid-gas interface. The extension of KPFM capabilities to probe electrostatic and electrochemical phenomena at the solid–liquid interface is of interest for a broad range of applications from energy storage to biological systems. However, the operation of KPFM implicitly relies on the presence of a linear lossless dielectric in the probe-sample gap, a condition which is violated for ionically-active liquids (e.g., when diffuse charge dynamics are present). Here, electrostatic and electrochemical measurements are demonstrated in ionically-active (polar isopropanol, milli-Q watermore » and aqueous NaCl) and ionically-inactive (non-polar decane) liquids by electrochemical force microscopy (EcFM), a multidimensional (i.e., bias- and time-resolved) spectroscopy method. In the absence of mobile charges (ambient and non-polar liquids), KPFM and EcFM are both feasible, yielding comparable contact potential difference (CPD) values. In ionically-active liquids, KPFM is not possible and EcFM can be used to measure the dynamic CPD and a rich spectrum of information pertaining to charge screening, ion diffusion, and electrochemical processes (e.g., Faradaic reactions). EcFM measurements conducted in isopropanol and milli-Q water over Au and highly ordered pyrolytic graphite electrodes demonstrate both sample- and solvent-dependent features. Finally, the feasibility of using EcFM as a local force-based mapping technique of material-dependent electrostatic and electrochemical response is investigated. The resultant high dimensional dataset is visualized using a purely statistical approach that does not require a priori physical models, allowing for qualitative mapping of electrostatic and electrochemical material properties at the solid–liquid interface.« less

  4. Kelvin Probe Force Microscopy in liquid using Electrochemical Force Microscopy

    SciTech Connect

    Collins, Liam; Jesse, Stephen; Kilpatrick, J.; Tselev, Alexander; Okatan, Mahmut Baris; Kalinin, Sergei V.; Rodriguez, Brian

    2015-01-01

    Conventional closed loop-Kelvin probe force microscopy (KPFM) has emerged as a powerful technique for probing electric and transport phenomena at the solid-gas interface. The extension of KPFM capabilities to probe electrostatic and electrochemical phenomena at the solid–liquid interface is of interest for a broad range of applications from energy storage to biological systems. However, the operation of KPFM implicitly relies on the presence of a linear lossless dielectric in the probe-sample gap, a condition which is violated for ionically-active liquids (e.g., when diffuse charge dynamics are present). Here, electrostatic and electrochemical measurements are demonstrated in ionically-active (polar isopropanol, milli-Q water and aqueous NaCl) and ionically-inactive (non-polar decane) liquids by electrochemical force microscopy (EcFM), a multidimensional (i.e., bias- and time-resolved) spectroscopy method. In the absence of mobile charges (ambient and non-polar liquids), KPFM and EcFM are both feasible, yielding comparable contact potential difference (CPD) values. In ionically-active liquids, KPFM is not possible and EcFM can be used to measure the dynamic CPD and a rich spectrum of information pertaining to charge screening, ion diffusion, and electrochemical processes (e.g., Faradaic reactions). EcFM measurements conducted in isopropanol and milli-Q water over Au and highly ordered pyrolytic graphite electrodes demonstrate both sample- and solvent-dependent features. Finally, the feasibility of using EcFM as a local force-based mapping technique of material-dependent electrostatic and electrochemical response is investigated. The resultant high dimensional dataset is visualized using a purely statistical approach that does not require a priori physical models, allowing for qualitative mapping of electrostatic and electrochemical material properties at the solid–liquid interface.

  5. "Das Konkrete ist das Abstrakte, an das man sich schließlich gewöhnt hat." (Laurent Schwartz) Über den Ablauf des mathematischen Verstehens

    NASA Astrophysics Data System (ADS)

    Lowsky, Martin

    Die im Titel genannte Aussage findet sich in den Lebenserinnerungen von Laurent Schwartz (1915-2002), einem der fruchtbarsten Mathematiker, Mitglied der Gruppe Bourbaki. Im Original lautet die Aussage: "un objet concret est un objet abstrait auquel on a fini par s'habituer." Schwartz erläutert sie am Beispiel des Integrals über {e^{-1/2{x^2}}} , das den Wert Wurzel aus 2π hat und in dem sich also die Zahlen e und π verknüpfen. Was Schwartz aber vor allem ausdrücken will, ist dies: Das mathematische Verständnisd geht langsam vor sich und es bedarf der Anstrengung. "Es ist eine Frage der Zeit und der Energie", sagt Schwartz, und gerade dies mache es so schwer, die höhere Mathematik unter das Volk zu bringen. Das Lernen und Lehren von Mathematik laufe eben mühevoll und langsam ab.

  6. Pedagogical Basis of DAS Formalism in Engineering Education

    ERIC Educational Resources Information Center

    Hiltunen, J.; Heikkinen, E.-P.; Jaako, J.; Ahola, J.

    2011-01-01

    The paper presents a new approach for a bachelor-level curriculum structure in engineering. The approach is called DAS formalism according to its three phases: description, analysis and synthesis. Although developed specifically for process and environmental engineering, DAS formalism has a generic nature and it could also be used in other…

  7. Evidence of Motor Programming Deficits in Children Diagnosed with DAS.

    ERIC Educational Resources Information Center

    Nijland, Lian; Maassen, Ben; van der Meulen, Sjoeke

    2003-01-01

    Five children with developmental apraxia of speech (DAS), 5 controls (ages 5-6), and 6 adults produced utterances in a normal condition and in a bite-block condition in which the mandible was in a fixed position. In children with DAS, the bite-block had large effects on coarticulatory patterns and vowel quality. (Contains references.) (Author/CR)

  8. Algorithmic methods in diffraction microscopy

    NASA Astrophysics Data System (ADS)

    Thibault, Pierre

    Recent diffraction imaging techniques use properties of coherent sources (most notably x-rays and electrons) to transfer a portion of the imaging task to computer algorithms. "Diffraction microscopy" is a method which consists in reconstructing the image of a specimen from its diffraction pattern. Because only the amplitude of a wavefield incident on a detector is measured, reconstruction of the image entails to recovering the lost phases. This extension of the 'phase problem" commonly met in crystallography is solved only if additional information is available. The main topic of this thesis is the development of algorithmic techniques in diffraction microscopy. In addition to introducing new methods, it is meant to be a review of the algorithmic aspects of the field of diffractive imaging. An overview of the scattering approximations used in the interpretation of diffraction datasets is first given, as well as a numerical propagation tool useful in conditions where known approximations fail. Concepts central to diffraction microscopy---such as oversampling---are then introduced and other similar imaging techniques described. A complete description of iterative reconstruction algorithms follows, with a special emphasis on the difference map, the algorithm used in this thesis. The formalism, based on constraint sets and projection onto these sets, is then defined and explained. Simple projections commonly used in diffraction imaging are then described. The various ways experimental realities can affect reconstruction methods will then be enumerated. Among the diverse sources of algorithmic difficulties, one finds that noise, missing data and partial coherence are typically the most important. Other related difficulties discussed are the detrimental effects of crystalline domains in a specimen, and the convergence problems occurring when the support of a complex-valued specimen is not well known. The last part of this thesis presents reconstruction results; an

  9. Multiphoton microscopy in life sciences.

    PubMed

    König, K

    2000-11-01

    Near infrared (NIR) multiphoton microscopy is becoming a novel optical tool of choice for fluorescence imaging with high spatial and temporal resolution, diagnostics, photochemistry and nanoprocessing within living cells and tissues. Three-dimensional fluorescence imaging based on non-resonant two-photon or three-photon fluorophor excitation requires light intensities in the range of MW cm(-2) to GW cm(-2), which can be derived by diffraction limited focusing of continuous wave and pulsed NIR laser radiation. NIR lasers can be employed as the excitation source for multifluorophor multiphoton excitation and hence multicolour imaging. In combination with fluorescence in situ hybridization (FISH), this novel approach can be used for multi-gene detection (multiphoton multicolour FISH). Owing to the high NIR penetration depth, non-invasive optical biopsies can be obtained from patients and ex vivo tissue by morphological and functional fluorescence imaging of endogenous fluorophores such as NAD(P)H, flavin, lipofuscin, porphyrins, collagen and elastin. Recent botanical applications of multiphoton microscopy include depth-resolved imaging of pigments (chlorophyll) and green fluorescent proteins as well as non-invasive fluorophore loading into single living plant cells. Non-destructive fluorescence imaging with multiphoton microscopes is limited to an optical window. Above certain intensities, multiphoton laser microscopy leads to impaired cellular reproduction, formation of giant cells, oxidative stress and apoptosis-like cell death. Major intracellular targets of photodamage in animal cells are mitochondria as well as the Golgi apparatus. The damage is most likely based on a two-photon excitation process rather than a one-photon or three-photon event. Picosecond and femtosecond laser microscopes therefore provide approximately the same safe relative optical window for two-photon vital cell studies. In labelled cells, additional phototoxic effects may occur via

  10. Differential Multiphoton Laser Scanning Microscopy

    SciTech Connect

    Field, Jeffrey J.; Sheetz, Kraig E.; Chandler, Eric V.; Hoover, Erich E.; Young, Michael D.; Ding, Shi-you; Sylvester, Anne W.; Kleinfeld, David; Squier, Jeff A.

    2012-01-01

    Multifocal multiphoton laser scanning microscopy (mfMPLSM) in the biological and medical sciences has the potential to become a ubiquitous tool for obtaining high-resolution images at video rates. While current implementations of mfMPLSM achieve very high frame rates, they are limited in their applicability to essentially those biological samples that exhibit little or no scattering. In this paper, we report on a method for mfMPLSM in which whole-field detection with a single detector, rather than detection with a matrix of detectors, such as a charge-coupled device (CCD) camera, is implemented. This advance makes mfMPLSM fully compatible for use in imaging through scattering media. Further, we demonstrate that this method makes it possible to simultaneously obtain multiple images and view differences in excitation parameters in a single scan of the specimen.

  11. Superresolution microscopy with transient binding.

    PubMed

    Molle, Julia; Raab, Mario; Holzmeister, Susanne; Schmitt-Monreal, Daniel; Grohmann, Dina; He, Zhike; Tinnefeld, Philip

    2016-06-01

    For single-molecule localization based superresolution, the concentration of fluorescent labels has to be thinned out. This is commonly achieved by photophysically or photochemically deactivating subsets of molecules. Alternatively, apparent switching of molecules can be achieved by transient binding of fluorescent labels. Here, a diffusing dye yields bright fluorescent spots when binding to the structure of interest. As the binding interaction is weak, the labeling is reversible and the dye ligand construct diffuses back into solution. This approach of achieving superresolution by transient binding (STB) is reviewed in this manuscript. Different realizations of STB are discussed and compared to other localization-based superresolution modalities. We propose the development of labeling strategies that will make STB a highly versatile tool for superresolution microscopy at highest resolution.

  12. Scanning Electrochemical Microscopy in Neuroscience

    NASA Astrophysics Data System (ADS)

    Schulte, Albert; Nebel, Michaela; Schuhmann, Wolfgang

    2010-07-01

    This article reviews recent work involving the application of scanning electrochemical microscopy (SECM) to the study of individual cultured living cells, with an emphasis on topographical and functional imaging of neuronal and secretory cells of the nervous and endocrine system. The basic principles of biological SECM and associated negative amperometric-feedback and generator/collector-mode SECM imaging are discussed, and successful use of the methodology for screening soft and fragile membranous objects is outlined. The drawbacks of the constant-height mode of probe movement and the benefits of the constant-distance mode of SECM operation are described. Finally, representative examples of constant-height and constant-distance mode SECM on a variety of live cells are highlighted to demonstrate the current status of single-cell SECM in general and of SECM in neuroscience in particular.

  13. Phase Aberrations in Diffraction Microscopy

    SciTech Connect

    Marchesini, S; Chapman, H N; Barty, A; Howells, M R; Spence, J H; Cui, C; Weierstall, U; Minor, A M

    2005-09-29

    In coherent X-ray diffraction microscopy the diffraction pattern generated by a sample illuminated with coherent x-rays is recorded, and a computer algorithm recovers the unmeasured phases to synthesize an image. By avoiding the use of a lens the resolution is limited, in principle, only by the largest scattering angles recorded. However, the imaging task is shifted from the experiment to the computer, and the algorithm's ability to recover meaningful images in the presence of noise and limited prior knowledge may produce aberrations in the reconstructed image. We analyze the low order aberrations produced by our phase retrieval algorithms. We present two methods to improve the accuracy and stability of reconstructions.

  14. Differential Multiphoton Laser Scanning Microscopy

    PubMed Central

    Field, Jeffrey J.; Sheetz, Kraig E.; Chandler, Eric V.; Hoover, Erich E.; Young, Michael D.; Ding, Shi-you; Sylvester, Anne W.; Kleinfeld, David; Squier, Jeff A.

    2016-01-01

    Multifocal multiphoton microscopy (MMM) in the biological and medical sciences has become an important tool for obtaining high resolution images at video rates. While current implementations of MMM achieve very high frame rates, they are limited in their applicability to essentially those biological samples that exhibit little or no scattering. In this paper, we report on a method for MMM in which imaging detection is not necessary (single element point detection is implemented), and is therefore fully compatible for use in imaging through scattering media. Further, we demonstrate that this method leads to a new type of MMM wherein it is possible to simultaneously obtain multiple images and view differences in excitation parameters in a single shot. PMID:27390511

  15. Multifocal interferometric synthetic aperture microscopy

    PubMed Central

    Xu, Yang; Chng, Xiong Kai Benjamin; Adie, Steven G.; Boppart, Stephen A.; Scott Carney, P.

    2014-01-01

    There is an inherent trade-off between transverse resolution and depth of field (DOF) in optical coherence tomography (OCT) which becomes a limiting factor for certain applications. Multifocal OCT and interferometric synthetic aperture microscopy (ISAM) each provide a distinct solution to the trade-off through modification to the experiment or via post-processing, respectively. In this paper, we have solved the inverse problem of multifocal OCT and present a general algorithm for combining multiple ISAM datasets. Multifocal ISAM (MISAM) uses a regularized combination of the resampled datasets to bring advantages of both multifocal OCT and ISAM to achieve optimal transverse resolution, extended effective DOF and improved signal-to-noise ratio. We present theory, simulation and experimental results. PMID:24977909

  16. Interference techniques in fluorescence microscopy

    NASA Astrophysics Data System (ADS)

    Dogan, Mehmet

    We developed a set of interference-based optical microscopy techniques to study biological structures through nanometer-scale axial localization of fluorescent biomarkers. Spectral self-interference fluorescence microscopy (SSFM) utilizes interference of direct and reflected waves emitted from fluorescent molecules in the vicinity of planar reflectors to reveal the axial position of the molecules. A comprehensive calculation algorithm based on Green's function formalism is presented to verify the validity of approximations used in a far-field approach that describes the emission of fluorescent markers near interfaces. Using the validated model, theoretical limits of axial localization were determined with emphasis given to numerical aperture (NA) dependence of localization uncertainty. SSFM was experimentally demonstrated in conformational analysis of nucleoproteins. In particular, interaction between surface-tethered 75-mer double strand DNA and integration host factor (IHF) protein was probed on Si-SiO2 substrates by determining the axial position of fluorescent labels attached to the free ends of DNA molecules. Despite its sub-nanometer precision axial localization capability, SSFM lacks high lateral resolution due to the low-NA requirement for planar reflectors. We developed a second technique, 4Pi-SSFM, which improves the lateral resolution of a conventional SSFM system by an order of magnitude while achieving nanometer-scale axial localization precision. Using two opposing high-NA objectives, fluorescence signal is interferometrically collected and spectral interference pattern is recorded. Axial position of emitters is found from analysis of the spectra. The 4Pi-SSFM technique was experimentally demonstrated by determining the surface profiles of fabricated glass surfaces and outer membranes of Shigella, a type of Gram-negative bacteria. A further discussion is presented to localize surface O antigen, which is an important oligosaccharide structure in the

  17. Resolution enhancement techniques in microscopy

    NASA Astrophysics Data System (ADS)

    Cremer, Christoph; Masters, Barry R.

    2013-05-01

    We survey the history of resolution enhancement techniques in microscopy and their impact on current research in biomedicine. Often these techniques are labeled superresolution, or enhanced resolution microscopy, or light-optical nanoscopy. First, we introduce the development of diffraction theory in its relation to enhanced resolution; then we explore the foundations of resolution as expounded by the astronomers and the physicists and describe the conditions for which they apply. Then we elucidate Ernst Abbe's theory of optical formation in the microscope, and its experimental verification and dissemination to the world wide microscope communities. Second, we describe and compare the early techniques that can enhance the resolution of the microscope. Third, we present the historical development of various techniques that substantially enhance the optical resolution of the light microscope. These enhanced resolution techniques in their modern form constitute an active area of research with seminal applications in biology and medicine. Our historical survey of the field of resolution enhancement uncovers many examples of reinvention, rediscovery, and independent invention and development of similar proposals, concepts, techniques, and instruments. Attribution of credit is therefore confounded by the fact that for understandable reasons authors stress the achievements from their own research groups and sometimes obfuscate their contributions and the prior art of others. In some cases, attribution of credit is also made more complex by the fact that long term developments are difficult to allocate to a specific individual because of the many mutual connections often existing between sometimes fiercely competing, sometimes strongly collaborating groups. Since applications in biology and medicine have been a major driving force in the development of resolution enhancing approaches, we focus on the contribution of enhanced resolution to these fields.

  18. Scanning Tunneling Optical Resonance Microscopy

    NASA Technical Reports Server (NTRS)

    Bailey, Sheila; Wilt, Dave; Raffaelle, Ryne; Gennett, Tom; Tin, Padetha; Lau, Janice; Castro, Stephanie; Jenkins, Philip; Scheiman, Dave

    2003-01-01

    Scanning tunneling optical resonance microscopy (STORM) is a method, now undergoing development, for measuring optoelectronic properties of materials and devices on the nanoscale by means of a combination of (1) traditional scanning tunneling microscopy (STM) with (2) tunable laser spectroscopy. In STORM, an STM tip probing a semiconductor is illuminated with modulated light at a wavelength in the visible-to-near-infrared range and the resulting photoenhancement of the tunneling current is measured as a function of the illuminating wavelength. The photoenhancement of tunneling current occurs when the laser photon energy is sufficient to excite charge carriers into the conduction band of the semiconductor. Figure 1 schematically depicts a proposed STORM apparatus. The light for illuminating the semiconductor specimen at the STM would be generated by a ring laser that would be tunable across the wavelength range of interest. The laser beam would be chopped by an achromatic liquid-crystal modulator. A polarization-maintaining optical fiber would couple the light to the tip/sample junction of a commercial STM. An STM can be operated in one of two modes: constant height or constant current. A STORM apparatus would be operated in the constant-current mode, in which the height of the tip relative to the specimen would be varied in order to keep the tunneling current constant. In this mode, a feedback control circuit adjusts the voltage applied to a piezoelectric actuator in the STM that adjusts the height of the STM tip to keep the tunneling current constant. The exponential relationship between the tunneling current and tip-to-sample distance makes it relatively easy to implement this mode of operation. The choice of method by which the photoenhanced portion of the tunneling current would be measured depends on choice of the frequency at which the input illumination would be modulated (chopped). If the frequency of modulation were low enough (typically < 10 Hz) that the

  19. Developments in optical coherence microscopy

    NASA Astrophysics Data System (ADS)

    Rolland, J. P.; Meemon, P.; Thompson, K. P.; Murali, S.; Lee, K. S.

    2010-11-01

    Optical Coherence Microscopy (OCM) utilizes a high NA microscope objective in the sample arm to achieve an axially and laterally high resolution OCT image. An increase in NA, however, leads to a dramatically decreased depth of focus (DOF), and hence shortens the imaging depth range so that high lateral resolution is maintained only within a small depth region around the focal plane. One solution to increase the depth of imaging while keeping a high lateral resolution is dynamic-focusing. Utilizing the voltage controlled refocus capability of a liquid lens, we have recently presented a solution for invariant high resolution imaging using the liquid lens embedded within a fixed optics hand-held custom microscope designed specifically for optical imaging systems using a broadband light source centered at 800 nm with a 120 nm bandwidth. Subsequently, we have developed a Gabor-Domain Optical Coherence Microscopy (GD-OCM) that utilizes the high speed imaging of spectral domain OCT, the high lateral resolution of OCM, and the ability of real time refocusing of our custom design variable focus objective. Finally, key developments in Phase-Resolved Doppler OCT (PR-DOCT) are key enablers to combine high-resolution structural imaging with functional imaging. In this paper we review achievements in GD-OCM and detail how portions of in-focus cross-sectional images can be extracted and fused to form an invariant lateral resolution image with multiple cross-sectional images acquired corresponding to a discrete refocusing step along depth enabled by the varifocal device. We demonstrate sub-cellular resolution imaging of an African frog tadpole (Xenopus Laevis) taken from a 500 μm × 500 μm cross-section as well as cellular imaging in in vivo skin. Finally, A novel dual-detection full-range Fourier-domain optical coherence tomography system was developed that provides 7 μm axial resolution (in air) at about 90 kHz axial scan rate for mirror-image phase resolved Doppler imaging

  20. Spectroscopic imaging in electron microscopy

    SciTech Connect

    Pennycook, Stephen J; Colliex, C.

    2012-01-01

    In the scanning transmission electron microscope, multiple signals can be simultaneously collected, including the transmitted and scattered electron signals (bright field and annular dark field or Z-contrast images), along with spectroscopic signals such as inelastically scattered electrons and emitted photons. In the last few years, the successful development of aberration correctors for the electron microscope has transformed the field of electron microscopy, opening up new possibilities for correlating structure to functionality. Aberration correction not only allows for enhanced structural resolution with incident probes into the sub-angstrom range, but can also provide greater probe currents to facilitate mapping of intrinsically weak spectroscopic signals at the nanoscale or even the atomic level. In this issue of MRS Bulletin, we illustrate the power of the new generation of electron microscopes with a combination of imaging and spectroscopy. We show the mapping of elemental distributions at atomic resolution and also the mapping of electronic and optical properties at unprecedented spatial resolution, with applications ranging from graphene to plasmonic nanostructures, and oxide interfaces to biology.

  1. Nanorheology by atomic force microscopy

    SciTech Connect

    Li, Tai-De; Chiu, Hsiang-Chih; Ortiz-Young, Deborah; Riedo, Elisa

    2014-12-15

    We present an Atomic Force Microscopy (AFM) based method to investigate the rheological properties of liquids confined within a nanosize gap formed by an AFM tip apex and a solid substrate. In this method, a conventional AFM cantilever is sheared parallel to a substrate surface by means of a lock-in amplifier while it is approaching and retracting from the substrate in liquid. The normal solvation forces and lateral viscoelastic shear forces experienced by the AFM tip in liquid can be simultaneously measured as a function of the tip-substrate distance with sub-nanometer vertical resolution. A new calibration method is applied to compensate for the linear drift of the piezo transducer and substrate system, leading to a more precise determination of the tip-substrate distance. By monitoring the phase lag between the driving signal and the cantilever response in liquid, the frequency dependent viscoelastic properties of the confined liquid can also be derived. Finally, we discuss the results obtained with this technique from different liquid-solid interfaces. Namely, octamethylcyclotetrasiloxane and water on mica and highly oriented pyrolytic graphite.

  2. Electron microscopy and forensic practice

    NASA Astrophysics Data System (ADS)

    Kotrlý, Marek; Turková, Ivana

    2013-05-01

    Electron microanalysis in forensic practice ranks among basic applications used in investigation of traces (latents, stains, etc.) from crime scenes. Applying electron microscope allows for rapid screening and receiving initial information for a wide range of traces. SEM with EDS/WDS makes it possible to observe topography surface and morphology samples and examination of chemical components. Physical laboratory of the Institute of Criminalistics Prague use SEM especially for examination of inorganic samples, rarely for biology and other material. Recently, possibilities of electron microscopy have been extended considerably using dual systems with focused ion beam. These systems are applied mainly in study of inner micro and nanoparticles , thin layers (intersecting lines in graphical forensic examinations, analysis of layers of functional glass, etc.), study of alloys microdefects, creating 3D particles and aggregates models, etc. Automated mineralogical analyses are a great asset to analysis of mineral phases, particularly soils, similarly it holds for cathode luminescence, predominantly colour one and precise quantitative measurement of their spectral characteristics. Among latest innovations that are becoming to appear also at ordinary laboratories are TOF - SIMS systems and micro Raman spectroscopy with a resolution comparable to EDS/WDS analysis (capable of achieving similar level as through EDS/WDS analysis).

  3. Lensfree microscopy on a cellphone.

    PubMed

    Tseng, Derek; Mudanyali, Onur; Oztoprak, Cetin; Isikman, Serhan O; Sencan, Ikbal; Yaglidere, Oguzhan; Ozcan, Aydogan

    2010-07-21

    We demonstrate lensfree digital microscopy on a cellphone. This compact and light-weight holographic microscope installed on a cellphone does not utilize any lenses, lasers or other bulky optical components and it may offer a cost-effective tool for telemedicine applications to address various global health challenges. Weighing approximately 38 grams (<1.4 ounces), this lensfree imaging platform can be mechanically attached to the camera unit of a cellphone where the samples are loaded from the side, and are vertically illuminated by a simple light-emitting diode (LED). This incoherent LED light is then scattered from each micro-object to coherently interfere with the background light, creating the lensfree hologram of each object on the detector array of the cellphone. These holographic signatures captured by the cellphone permit reconstruction of microscopic images of the objects through rapid digital processing. We report the performance of this lensfree cellphone microscope by imaging various sized micro-particles, as well as red blood cells, white blood cells, platelets and a waterborne parasite (Giardia lamblia).

  4. Disposable optics for microscopy diagnostics.

    PubMed

    Vilmi, Pauliina; Varjo, Sami; Sliz, Rafal; Hannuksela, Jari; Fabritius, Tapio

    2015-11-20

    The point-of-care testing (POCT) is having increasing role on modern health care systems due to a possibility to perform tests for patients conveniently and immediately. POCT includes lot of disposable devices because of the environment they are often used. For a disposable system to be reasonably utilized, it needs to be high in quality but low in price. Optics based POCT systems are interesting approach to be developed, and here we describe a low-cost fabrication process for microlens arrays for microscopy. Lens arrays having average lens diameter of 222 μm with 300 μm lens pitch were fabricated. The lenses were characterized to have standard deviation of 0.06 μm in height and 4.61 μm in diameter. The resolution limit of 3.9μm is demonstrated with real images, and the images were compared with ones made with glass and polycarbonate lens arrays. The image quality is at the same level than with the glass lenses and the manufacturing costs are very low, thus making them suitable for POCT applications.

  5. Liquid Cell Transmission Electron Microscopy.

    PubMed

    Liao, Hong-Gang; Zheng, Haimei

    2016-05-27

    Liquid cell transmission electron microscopy (TEM) has attracted significant interest in recent years. With nanofabricated liquid cells, it has been possible to image through liquids using TEM with subnanometer resolution, and many previously unseen materials dynamics have been revealed. Liquid cell TEM has been applied to many areas of research, ranging from chemistry to physics, materials science, and biology. So far, topics of study include nanoparticle growth and assembly, electrochemical deposition and lithiation for batteries, tracking and manipulation of nanoparticles, catalysis, and imaging of biological materials. In this article, we first review the development of liquid cell TEM and then highlight progress in various areas of research. In the study of nanoparticle growth, the electron beam can serve both as the illumination source for imaging and as the input energy for reactions. However, many other research topics require the control of electron beam effects to minimize electron beam damage. We discuss efforts to understand electron beam-liquid matter interactions. Finally, we provide a perspective on future challenges and opportunities in liquid cell TEM.

  6. Liquid Cell Transmission Electron Microscopy

    NASA Astrophysics Data System (ADS)

    Liao, Hong-Gang; Zheng, Haimei

    2016-05-01

    Liquid cell transmission electron microscopy (TEM) has attracted significant interest in recent years. With nanofabricated liquid cells, it has been possible to image through liquids using TEM with subnanometer resolution, and many previously unseen materials dynamics have been revealed. Liquid cell TEM has been applied to many areas of research, ranging from chemistry to physics, materials science, and biology. So far, topics of study include nanoparticle growth and assembly, electrochemical deposition and lithiation for batteries, tracking and manipulation of nanoparticles, catalysis, and imaging of biological materials. In this article, we first review the development of liquid cell TEM and then highlight progress in various areas of research. In the study of nanoparticle growth, the electron beam can serve both as the illumination source for imaging and as the input energy for reactions. However, many other research topics require the control of electron beam effects to minimize electron beam damage. We discuss efforts to understand electron beam-liquid matter interactions. Finally, we provide a perspective on future challenges and opportunities in liquid cell TEM.

  7. Disposable optics for microscopy diagnostics

    PubMed Central

    Vilmi, Pauliina; Varjo, Sami; Sliz, Rafal; Hannuksela, Jari; Fabritius, Tapio

    2015-01-01

    The point-of-care testing (POCT) is having increasing role on modern health care systems due to a possibility to perform tests for patients conveniently and immediately. POCT includes lot of disposable devices because of the environment they are often used. For a disposable system to be reasonably utilized, it needs to be high in quality but low in price. Optics based POCT systems are interesting approach to be developed, and here we describe a low-cost fabrication process for microlens arrays for microscopy. Lens arrays having average lens diameter of 222 μm with 300 μm lens pitch were fabricated. The lenses were characterized to have standard deviation of 0.06 μm in height and 4.61 μm in diameter. The resolution limit of 3.9μm is demonstrated with real images, and the images were compared with ones made with glass and polycarbonate lens arrays. The image quality is at the same level than with the glass lenses and the manufacturing costs are very low, thus making them suitable for POCT applications. PMID:26586153

  8. Holographic microscopy in low coherence

    NASA Astrophysics Data System (ADS)

    Chmelík, Radim; Petráček, Jiří; Slabá, Michala; Kollárová, Věra; Slabý, Tomáš; Čolláková, Jana; Komrska, Jiří; Dostál, Zbyněk.; Veselý, Pavel

    2016-03-01

    Low coherence of the illumination substantially improves the quality of holographic and quantitative phase imaging (QPI) by elimination of the coherence noise and various artefacts and by improving the lateral resolution compared to the coherent holographic microscopy. Attributes of coherence-controlled holographic microscope (CCHM) designed and built as an off-axis holographic system allowing QPI within the range from complete coherent to incoherent illumination confirmed these expected advantages. Low coherence illumination also furnishes the coherence gating which constraints imaging of some spatial frequencies of an object axially thus forming an optical section in the wide sense. In this way the depth discrimination capability of the microscope is introduced at the price of restricting the axial interval of possible numerical refocusing. We describe theoretically these effects for the whole range of illumination coherence. We also show that the axial refocusing constraints can be overcome using advanced mode of imaging based on mutual lateral shift of reference and object image fields in CCHM. Lowering the spatial coherence of illumination means increasing its numerical aperture. We study how this change of the illumination geometry influences 3D objects QPI and especially the interpretation of live cells QPI in terms of the dry mass density measurement. In this way a strong dependence of the imaging process on the light coherence is demonstrated. The theoretical calculations and numerical simulations are supported by experimental data including a chance of time-lapse watching of live cells even in optically turbid milieu.

  9. Advances in quantitative Kerr microscopy

    NASA Astrophysics Data System (ADS)

    Soldatov, I. V.; Schäfer, R.

    2017-01-01

    An advanced wide-field Kerr microscopy approach to the vector imaging of magnetic domains is demonstrated. Utilizing the light from eight monochrome light emitting diodes, guided to the microscope by glass fibers, and being properly switched in synchronization with the camera exposure, domain images with orthogonal in-plane sensitivity are obtained simultaneously at real time. After calibrating the Kerr contrast under the same orthogonal sensitivity conditions, the magnetization vector field of complete magnetization cycles along the hysteresis loop can be calculated and plotted as a coded color or vector image. In the pulsed mode also parasitic, magnetic field-dependent Faraday rotations in the microscope optics are eliminated, thus increasing the accuracy of the measured magnetization angles to better than 5∘. The method is applied to the investigation of the magnetization process in a patterned Permalloy film element. Furthermore it is shown that the effective magnetic anisotropy axes in a GaMnAs semiconducting film can be quantitatively measured by vectorial analysis of the domain structure.

  10. Nanorheology by atomic force microscopy.

    PubMed

    Li, Tai-De; Chiu, Hsiang-Chih; Ortiz-Young, Deborah; Riedo, Elisa

    2014-12-01

    We present an Atomic Force Microscopy (AFM) based method to investigate the rheological properties of liquids confined within a nanosize gap formed by an AFM tip apex and a solid substrate. In this method, a conventional AFM cantilever is sheared parallel to a substrate surface by means of a lock-in amplifier while it is approaching and retracting from the substrate in liquid. The normal solvation forces and lateral viscoelastic shear forces experienced by the AFM tip in liquid can be simultaneously measured as a function of the tip-substrate distance with sub-nanometer vertical resolution. A new calibration method is applied to compensate for the linear drift of the piezo transducer and substrate system, leading to a more precise determination of the tip-substrate distance. By monitoring the phase lag between the driving signal and the cantilever response in liquid, the frequency dependent viscoelastic properties of the confined liquid can also be derived. Finally, we discuss the results obtained with this technique from different liquid-solid interfaces. Namely, octamethylcyclotetrasiloxane and water on mica and highly oriented pyrolytic graphite.

  11. Photoacoustic microscopy of human teeth

    NASA Astrophysics Data System (ADS)

    Rao, Bin; Cai, Xin; Favazza, Christopher; Yao, Junjie; Li, Li; Duong, Steven; Liaw, Lih-Huei; Holtzman, Jennifer; Wilder-Smith, Petra; Wang, Lihong V.

    2011-03-01

    Photoacoustic microscopy (PAM) utilizes short laser pulses to deposit energy into light absorbers and sensitively detects the ultrasonic waves the absorbers generate in response. PAM directly renders a three-dimensional spatial distribution of sub-surface optical absorbers. Unlike other optical imaging technologies, PAM features label-free optical absorption contrast and excellent imaging depths. Standard dental imaging instruments are limited to X-ray and CCD cameras. Subsurface optical dental imaging is difficult due to the highly-scattering enamel and dentin tissue. Thus, very few imaging methods can detect dental decay or diagnose dental pulp, which is the innermost part of the tooth, containing the nerves, blood vessels, and other cells. Here, we conducted a feasibility study on imaging dental decay and dental pulp with PAM. Our results showed that PAM is sensitive to the color change associated with dental decay. Although the relative PA signal distribution may be affected by surface contours and subsurface reflections from deeper dental tissue, monitoring changes in the PA signals (at the same site) over time is necessary to identify the progress of dental decay. Our results also showed that deep-imaging, near-infrared (NIR) PAM can sensitively image blood in the dental pulp of an in vitro tooth. In conclusion, PAM is a promising tool for imaging both dental decay and dental pulp.

  12. Enhanced effects with scanning force microscopy

    NASA Astrophysics Data System (ADS)

    Howells, S.; Chen, T.; Gallagher, M.; Yi, L.; Sarid, D.

    1991-05-01

    A general theory that describes the operation of scanning force microscopy in the contact force regime is presented. It is shown that force derivatives along the surface of a sample produce images that can be dramatically enhanced relative to those of surface topography. For scanning tunneling microscopy atomic force microscopy configurations, the spring constant of the cantilever and the force derivatives perpendicular to the surface of the sample determine the enhancement, respectively.

  13. Scanning Tip Microscopy With Applications To Biology

    NASA Astrophysics Data System (ADS)

    Sarid, Dror; Thall, Edmond H.; Iams, Douglas A.; Ingle, Jeffery T.; Henson, Tammy D.; Lee, Y. C.; Bell, L. Stephen

    1989-06-01

    Scanning tunneling microscopy and atomic force microscopy, denoted here scanning tip microscopy, are two powerful novel techniques for imaging surfaces with atomic resolution. We describe the underlying principles of these two techniques with special emphasis on an instrument developed in our laboratory that uses a laser diode to detect minute deflections of a tip as it raster scans the surface of a sample. Applications of these techniques to research in biology are assessed and their relative merits discussed.

  14. Wavelength Independent Optical Microscopy and Lithography

    DTIC Science & Technology

    1987-10-31

    Leviatan , Y., J. Appl. Phys. 60, 1577 (1986). 7. Harootunian, A., Near-Field Scanning Optical Microscopy and Raman Microscopy, Cornell University Ph.D...although the approach used may not be valid in the Another potential problem concerns the effect of the near field. More recently, Leviatan 21...Massey, "Microscopy and Pattern Generation With Scanned Evanescent Waves," AppL. Opt. 23, 658 (1984). The authors wish to thank Yehuda Leviatan for 21

  15. Scanning Tunneling Microscopy Studies of Quasicrystals

    NASA Astrophysics Data System (ADS)

    Becker, Russell S.; Kortan, A. Refik

    The following sections are included: * INTRODUCTION * EXPERIMENTAL * X-RAY DIFFRACTION * SCANNING TUNNELING MICROSCOPY * STRUCTURE MODELLING BASED ON STM * COMPARISON WITH MODELS BASED ON BULK STUDIES * CONCLUSION * REFERENCES

  16. Twenty‐eight‐joint counts invalidate the DAS28 remission definition owing to the omission of the lower extremity joints: a comparison with the original DAS remission

    PubMed Central

    Landewé, R; van der Heijde, D; van der Linden, S; Boers, M

    2006-01-01

    Objective To compare 28 joint disease activity score (DAS28) remission with comprehensive joint count DAS remission in rheumatoid arthritis. Methods 620 actually measured paired observations of DAS28 and DAS were analysed in 155 patients. Discordant observations (either DAS or DAS28 below remission cut off level: 1.6 for DAS and 2.6 for DAS28) and concordant observations (both DAS and DAS28 below their remission cut off level) were analysed separately. Results 91 of 620 paired DAS observations (15%) were discordant; 87 (in 53 patients) comprised observations in which the DAS28 remission criterion, but not the DAS remission criterion, was met. The reverse was found in only four observations, which were therefore omitted. With the original DAS as standard, DAS28 sensitivity was 95% and specificity 84%. Probability plots showed a swollen joint count >0 in 75% of discordant pairs v 48% of concordant pairs. The same was found for total joint count (TJC >0 in 90% v 40%; median TJC, 0 v 6) and patient global assessment, but not for ESR. Individual joint analysis showed that 51% of discordant v 18% of concordant observations (p<0.0005) had involvement of lower extremity joints that are not included in the DAS28. Conclusions DAS remission is more conservative than DAS28 remission. Activity (tenderness and swelling) in joints not included in the reduced joint counts (ankles, feet) mainly account for the discrepancy between the two assessments. DAS28 remission at a cut off level of 2.6 has insufficient construct validity and should be used with caution in clinical practice and clinical trials. PMID:16219709

  17. The 2015 super-resolution microscopy roadmap

    NASA Astrophysics Data System (ADS)

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

    2015-11-01

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

  18. Digital microscopy. Bringing new technology into focus.

    PubMed

    2010-06-01

    Digital microscopy enables the scanning of microscope slides so that they can be viewed, analyzed, and archived on a computer. While the technology is not yet widely accepted by pathologists, a switch to digital microscopy systems seems to be inevitable in the near future.

  19. Microscopy & microanalysis 2016 in Columbus, Ohio

    DOE PAGES

    Michael, Joseph R.

    2016-01-08

    The article provides information about an upcoming conference from the program chair. The Microscopy Society of America (MSA), the Microanalysis Society (MAS), and the International Metallographic Society (IMS) invite participation in Microscopy & Microanalysis 2016 in Columbus, Ohio, July 24 through July 28, 2016.

  20. Super-resolution microscopy of mitochondria.

    PubMed

    Jakobs, Stefan; Wurm, Christian A

    2014-06-01

    Mitochondria, the powerhouses of the cell, are essential organelles in eukaryotic cells. With their complex inner architecture featuring a smooth outer and a highly convoluted inner membrane, they are challenging objects for microscopy. The diameter of mitochondria is generally close to the resolution limit of conventional light microscopy, rendering diffraction-unlimited super-resolution light microscopy (nanoscopy) for imaging submitochondrial protein distributions often mandatory. In this review, we discuss what can be expected when imaging mitochondria with conventional diffraction-limited and diffraction-unlimited microscopy. We provide an overview on recent studies using super-resolution microscopy to investigate mitochondria and discuss further developments and challenges in mitochondrial biology that might by addressed with these technologies in the future.

  1. Optical super-resolution microscopy in neurobiology.

    PubMed

    Sigrist, Stephan J; Sabatini, Bernardo L

    2012-02-01

    Understanding the highly plastic nature of neurons requires the dynamic visualization of their molecular and cellular organization in a native context. However, due to the limited resolution of standard light microscopy, many of the structural specializations of neurons cannot be resolved. A recent revolution in light microscopy has given rise to several super-resolution light microscopy methods yielding 2-10-fold higher resolution than conventional microscopy. We here describe the principles behind these techniques as well as their application to the analysis of the molecular architecture of the synapse. Furthermore, we discuss the potential for continued development of super-resolution microscopy as necessary for live imaging of neuronal structure and function in the brain.

  2. DAS: A Data Management System for Instrument Tests and Operations

    NASA Astrophysics Data System (ADS)

    Frailis, M.; Sartor, S.; Zacchei, A.; Lodi, M.; Cirami, R.; Pasian, F.; Trifoglio, M.; Bulgarelli, A.; Gianotti, F.; Franceschi, E.; Nicastro, L.; Conforti, V.; Zoli, A.; Smart, R.; Morbidelli, R.; Dadina, M.

    2014-05-01

    The Data Access System (DAS) is a and data management software system, providing a reusable solution for the storage of data acquired both from telescopes and auxiliary data sources during the instrument development phases and operations. It is part of the Customizable Instrument WorkStation system (CIWS-FW), a framework for the storage, processing and quick-look at the data acquired from scientific instruments. The DAS provides a data access layer mainly targeted to software applications: quick-look displays, pre-processing pipelines and scientific workflows. It is logically organized in three main components: an intuitive and compact Data Definition Language (DAS DDL) in XML format, aimed for user-defined data types; an Application Programming Interface (DAS API), automatically adding classes and methods supporting the DDL data types, and providing an object-oriented query language; a data management component, which maps the metadata of the DDL data types in a relational Data Base Management System (DBMS), and stores the data in a shared (network) file system. With the DAS DDL, developers define the data model for a particular project, specifying for each data type the metadata attributes, the data format and layout (if applicable), and named references to related or aggregated data types. Together with the DDL user-defined data types, the DAS API acts as the only interface to store, query and retrieve the metadata and data in the DAS system, providing both an abstract interface and a data model specific one in C, C++ and Python. The mapping of metadata in the back-end database is automatic and supports several relational DBMSs, including MySQL, Oracle and PostgreSQL.

  3. Gabor domain optical coherence microscopy

    NASA Astrophysics Data System (ADS)

    Murali, Supraja

    Time domain Optical Coherence Tomography (TD-OCT), first reported in 1991, makes use of the low temporal coherence properties of a NIR broadband laser to create depth sectioning of up to 2mm under the surface using optical interferometry and point to point scanning. Prior and ongoing work in OCT in the research community has concentrated on improving axial resolution through the development of broadband sources and speed of image acquisition through new techniques such as Spectral domain OCT (SD-OCT). In SD-OCT, an entire depth scan is acquired at once with a low numerical aperture (NA) objective lens focused at a fixed point within the sample. In this imaging geometry, a longer depth of focus is achieved at the expense of lateral resolution, which is typically limited to 10 to 20 mum. Optical Coherence Microscopy (OCM), introduced in 1994, combined the advantages of high axial resolution obtained in OCT with high lateral resolution obtained by increasing the NA of the microscope placed in the sample arm. However, OCM presented trade-offs caused by the inverse quadratic relationship between the NA and the DOF of the optics used. For applications requiring high lateral resolution, such as cancer diagnostics, several solutions have been proposed including the periodic manual re-focusing of the objective lens in the time domain as well as the spectral domain C-mode configuration in order to overcome the loss in lateral resolution outside the DOF. In this research, we report for the first time, high speed, sub-cellular imaging (lateral resolution of 2 mum) in OCM using a Gabor domain image processing algorithm with a custom designed and fabricated dynamic focus microscope interfaced to a Ti:Sa femtosecond laser centered at 800 nm within an SD-OCM configuration. It is envisioned that this technology will provide a non-invasive replacement for the current practice of multiple biopsies for skin cancer diagnosis. The research reported here presents three important advances

  4. Visualizing quantitative microscopy data: History and challenges.

    PubMed

    Sailem, Heba Z; Cooper, Sam; Bakal, Chris

    2016-01-01

    Data visualization is a fundamental aspect of science. In the context of microscopy-based studies, visualization typically involves presentation of the images themselves. However, data visualization is challenging when microscopy experiments entail imaging of millions of cells, and complex cellular phenotypes are quantified in a high-content manner. Most well-established visualization tools are inappropriate for displaying high-content data, which has driven the development of new visualization methodology. In this review, we discuss how data has been visualized in both classical and high-content microscopy studies; as well as the advantages, and disadvantages, of different visualization methods.

  5. Soft x-ray holographic microscopy

    SciTech Connect

    Stickler, Daniel; Froemter, Robert; Stillrich, Holger; Menk, Christian; Oepen, Hans Peter; Tieg, Carsten; Streit-Nierobisch, Simone; Sprung, Michael; Gutt, Christian; Stadler, Lorenz-M.; Leupold, Olaf; Gruebel, Gerhard

    2010-01-25

    We present a new x-ray microscopy technique based on Fourier transform holography (FTH), where the sample is separate from the optics part of the setup. The sample can be shifted with respect to the holography optics, thus large-scale or randomly distributed objects become accessible. As this extends FTH into a true microscopy technique, we call it x-ray holographic microscopy (XHM). FTH allows nanoscale imaging without the need for nanometer-size beams. Simple Fourier transform yields an unambiguous image reconstruction. We demonstrate XHM by studying the magnetic domain evolution of a Co/Pt multilayer film as function of locally varied iron overlayer thickness.

  6. Latest advances in commercially available STED microscopy

    NASA Astrophysics Data System (ADS)

    Fouquet, Wernher; Giske, Arnold

    2012-02-01

    STimulated Emission Depletion (STED) microscopy enables imaging of biological samples combining significantly improved optical resolution with all benefits of confocal microscopy. Especially, by combining multi-channel image acquisition with high spatial resolution opens up a new understanding of co-localization experiments on nanoscales. Such a microscope provides new insights in various fields of biology, such as cell and membrane biology, neurobiology and physiology. We present new developments and a variety of biological examples for STED microscopy, showing structural details on scales well below 70nm and give an overview of possible field of applications, mainly focused on live cell imaging.

  7. Pedagogical basis of DAS formalism in engineering education

    NASA Astrophysics Data System (ADS)

    Hiltunen, J.; Heikkinen, E.-P.; Jaako, J.; Ahola, J.

    2011-03-01

    The paper presents a new approach for a bachelor-level curriculum structure in engineering. The approach is called DAS formalism according to its three phases: description, analysis and synthesis. Although developed specifically for process and environmental engineering, DAS formalism has a generic nature and it could also be used in other engineering fields. The motivation for this new curriculum structure originates from the urge to solve the problems that engineering education has faced during the past decades, e.g. student recruitment problems and dissatisfactory learning outcomes. The focus of this paper is on the structure of the curriculum but the content is also discussed when it has an effect on the structure and its implementation. The presented structure, i.e. DAS formalism, builds upon the ideas of some classical pedagogical theories, which have regularly been applied at course level but seldom used to solve curriculum-level issues.

  8. Using hydrogels in microscopy: A tutorial.

    PubMed

    Flood, Peter; Page, Henry; Reynaud, Emmanuel G

    2016-05-01

    Sample preparation for microscopy is a crucial step to ensure the best experimental outcome. It often requires the use of specific mounting media that have to be tailored to not just the sample but the chosen microscopy technique. The media must not damage the sample or impair the optical path, and may also have to support the correct physiological function/development of the sample. For decades, researchers have used embedding media such as hydrogels to maintain samples in place. Their ease of use and transparency has promoted them as mainstream mounting media. However, they are not as straightforward to implement as assumed. They can contain contaminants, generate forces on the sample, have complex diffusion and structural properties that are influenced by multiple factors and are generally not designed for microscopy in mind. This short review will discuss the advantages and disadvantages of using hydrogels for microscopy sample preparation and highlight some of the less obvious problems associated with the area.

  9. Rotary-scanning optical resolution photoacoustic microscopy

    NASA Astrophysics Data System (ADS)

    Qi, Weizhi; Xi, Lei

    2016-10-01

    Optical resolution photoacoustic microscopy (ORPAM) is currently one of the fastest evolving photoacoustic imaging modalities. It has a comparable spatial resolution to pure optical microscopic techniques such as epifluorescence microscopy, confocal microscopy, and two-photon microscopy, but also owns a deeper penetration depth. In this paper, we report a rotary-scanning (RS)-ORPAM that utilizes a galvanometer scanner integrated with objective to achieve rotary laser scanning. A 15 MHz cylindrically focused ultrasonic transducer is mounted onto a motorized rotation stage to follow optical scanning traces synchronously. To minimize the loss of signal to noise ratio, the acoustic focus is precisely adjusted to reach confocal with optical focus. Black tapes and carbon fibers are firstly imaged to evaluate the performance of the system, and then in vivo imaging of vasculature networks inside the ears and brains of mice is demonstrated using this system.

  10. Saturated pattern-illuminated Fourier ptychography microscopy

    NASA Astrophysics Data System (ADS)

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

    2017-01-01

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

  11. Lensless Digital Holographic Microscopy for Life Detection

    NASA Astrophysics Data System (ADS)

    Serabyn, E.; Liewer, K.; Wallace, J. K.; Rider, S.; Lindensmith, C.; Nadeau, J.

    2016-10-01

    Microscopy capable of volume imaging can be used to search for microbial life on ocean worlds. Here we discuss our recent digital holographic microscope (DHM) systems, which provide micron-scale resolution in a very compact package.

  12. Atomic resolution 3D electron diffraction microscopy

    SciTech Connect

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

    2002-03-01

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

  13. Multiphoton microscopy: an introduction to gastroenterologists.

    PubMed

    Cho, Hye Jin; Chun, Hoon Jai; Kim, Eun Sun; Cho, Bong Rae

    2011-10-28

    Multiphoton microscopy, relying on the simultaneous absorption of two or more photons by a fluorophore, has come to occupy a prominent place in modern biomedical research with its ability to allow real-time observation of a single cell and molecules in intact tissues. Multiphoton microscopy exhibits nonlinear optical contrast properties, which can make it possible to provide an exceptionally large depth penetration with less phototoxicity. This system becomes more and more an inspiring tool for a non-invasive imaging system to realize "optical biopsy" and to examine the functions of living cells. In this review, we briefly present the physical principles and properties of multiphoton microscopy as well as the current applications in biological fields. In addition, we address what we see as the future potential of multiphoton microscopy for gastroenterologic research.

  14. Theoretical simulation of scanning probe microscopy.

    PubMed

    Tsukada, Masaru

    2011-01-01

    Methods of theoretical simulation of scanning probe microscopy, including scanning tunneling microscopy (STM), atomic force microscopy(AFM) and Kelvin prove force microscopy (KPFM) have been reviewed with recent topics as case studies. For the case of the STM simulation, the importance of the tip electronic states is emphasized and some advanced formalism is presented based on the non-equilibrium Green's function theory beyond Bardeen's perturbation theory. For the simulation of AFM, we show examples of 3D-force map for AFM in water, and theoretical analyses for a nano-mechanical experiment on a protein molecule. An attempt to simulate KPFM images based on the electrostatic multi-pole interaction between a tip and a sample is also introduced.

  15. Multiphoton microscopy in defining liver function

    NASA Astrophysics Data System (ADS)

    Thorling, Camilla A.; Crawford, Darrell; Burczynski, Frank J.; Liu, Xin; Liau, Ian; Roberts, Michael S.

    2014-09-01

    Multiphoton microscopy is the preferred method when in vivo deep-tissue imaging is required. This review presents the application of multiphoton microscopy in defining liver function. In particular, multiphoton microscopy is useful in imaging intracellular events, such as mitochondrial depolarization and cellular metabolism in terms of NAD(P)H changes with fluorescence lifetime imaging microscopy. The morphology of hepatocytes can be visualized without exogenously administered fluorescent dyes by utilizing their autofluorescence and second harmonic generation signal of collagen, which is useful in diagnosing liver disease. More specific imaging, such as studying drug transport in normal and diseased livers are achievable, but require exogenously administered fluorescent dyes. If these techniques can be translated into clinical use to assess liver function, it would greatly improve early diagnosis of organ viability, fibrosis, and cancer.

  16. Polarization-sensitive interferometric synthetic aperture microscopy

    PubMed Central

    South, Fredrick A.; Liu, Yuan-Zhi; Xu, Yang; Shemonski, Nathan D.; Carney, P. Scott; Boppart, Stephen A.

    2015-01-01

    Three-dimensional optical microscopy suffers from the well-known compromise between transverse resolution and depth-of-field. This is true for both structural imaging methods and their functional extensions. Interferometric synthetic aperture microscopy (ISAM) is a solution to the 3D coherent microscopy inverse problem that provides depth-independent transverse resolution. We demonstrate the extension of ISAM to polarization sensitive imaging, termed polarization-sensitive interferometric synthetic aperture microscopy (PS-ISAM). This technique is the first functionalization of the ISAM method and provides improved depth-of-field for polarization-sensitive imaging. The basic assumptions of polarization-sensitive imaging are explored, and refocusing of birefringent structures is experimentally demonstrated. PS-ISAM enables high-resolution volumetric imaging of birefringent materials and tissue. PMID:26648593

  17. Magnetic resonance microscopy in biomedical research.

    PubMed

    Serša, I

    2012-01-01

    Magnetic resonance (MR) microscopy is a special modality of MRI with an emphasis on high spatial resolution. While its main principle is identical to conventional clinical MRI, there are several differences between the two that are mainly associated with a use of stronger magnets and gradients. MR microscopy has numerous interesting applications in material and bio sciences in which high spatial resolution is demanded and long experiment times are allowed.

  18. Coatings and alternatives for SEM microscopy

    SciTech Connect

    Lee, R.H.

    1995-03-01

    Several methods of preparing samples of low electrical conductivity for conventional scanning electron microscopy are reviewed. Two new methods are chromium sputter-coating and low-voltage electron microscopy with a field emission gun. Photomicrographs of different coatings at high magnification show the structure of each coating. Advantages and disadvantages of each material are presented. Results with sputtered coatings are compared to an evaporated carbon coating.

  19. Advanced electron microscopy for advanced materials.

    PubMed

    Van Tendeloo, Gustaaf; Bals, Sara; Van Aert, Sandra; Verbeeck, Jo; Van Dyck, Dirk

    2012-11-08

    The idea of this Review is to introduce newly developed possibilities of advanced electron microscopy to the materials science community. Over the last decade, electron microscopy has evolved into a full analytical tool, able to provide atomic scale information on the position, nature, and even the valency atoms. This information is classically obtained in two dimensions (2D), but can now also be obtained in 3D. We show examples of applications in the field of nanoparticles and interfaces.

  20. Imaging DNA Structure by Atomic Force Microscopy.

    PubMed

    Pyne, Alice L B; Hoogenboom, Bart W

    2016-01-01

    Atomic force microscopy (AFM) is a microscopy technique that uses a sharp probe to trace a sample surface at nanometre resolution. For biological applications, one of its key advantages is its ability to visualize substructure of single molecules and molecular complexes in an aqueous environment. Here, we describe the application of AFM to determine superstructure and secondary structure of surface-bound DNA. The method is also readily applicable to probe DNA-DNA interactions and DNA-protein complexes.

  1. Investigation of wear phenomena by microscopy

    NASA Technical Reports Server (NTRS)

    Buckley, D. H.

    1982-01-01

    The various wear mechanisms involved in the loss of material from metallic and nonmetallic surfaces are discussed. The results presented indicate how various microscopy techniques used in conjunction with other analytical tools can assist in the elucidation of a wear mechanism. Without question, microscopy is the single most important tool for the study of the wear of surfaces, to assess and address inherent mechanisms of the material removal process.

  2. Subwavelength optical microscopy in the far field

    SciTech Connect

    Sun Qingqing; Zubairy, M. Suhail; Al-Amri, M.; Scully, Marlan O.

    2011-06-15

    We present a procedure for subwavelength optical microscopy. The identical atoms are distributed on a plane and shined with a standing wave. We rotate the plane to different angles and record the resonant fluorescence spectra in the far field, from which we can obtain their distance and location information. This procedure also works for atomic separation above one wavelength and therefore provides a seamless microscopy.

  3. The development of the disease activity score (DAS) and the disease activity score using 28 joint counts (DAS28).

    PubMed

    van Riel, P L C M

    2014-01-01

    In rheumatoid arthritis, disease activity cannot be measured using a single variable. The Disease Activity Score (DAS) has been developed as a quantitative index to be able to measure, study and manage disease activity in RA in daily clinical practice, clinical trials, and long term observational studies. The DAS is a continuous measure of RA disease activity that combines information from swollen joints, tender joints, acute phase response and patient self-report of general health. Cut points were developed to classify patients in remission, as well as low, moderate, and severe disease activity in the 1990s. DAS-based EULAR response criteria were primarily developed to be used in clinical trials to classify individual patients as non-, moderate, or good responders, depending on the magnitude of change and absolute level of disease activity at the conclusion of the test.

  4. Bessel light sheet structured illumination microscopy

    NASA Astrophysics Data System (ADS)

    Noshirvani Allahabadi, Golchehr

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

  5. Cryo-scanning electron microscopy and light microscopy for the study of fungi interactions.

    PubMed

    Sempere, F; Santamarina, M P

    2011-03-01

    The application of the cryo-scanning electron microscopy and light microscopy for the study of the interactions at different environmental conditions between Penicillium oxalicum and Fusarium verticillioides is described. A dual microculture was developed for the light microscopy analysis of the interaction. The microscope and macroscopic examinations were compared. Analysis of Petri plates revealed that F. verticillioides was a competitor for space and nutrients while P. oxalicum was a mycoparasite under the microscopic observations.

  6. Exploring Scanning Probe Microscopy with Mathematica

    NASA Astrophysics Data System (ADS)

    Sarid, Dror

    1997-10-01

    This book/software edition provides a complete set of computational models that describe the physical phenomena associated with scanning tunneling microscopy, atomic force microscopy, and related technologies. Its self-contained presentation spares researchers the valuable time spent hunting through the technical literature in search of prior theoretical results required to understand the models presented. Mathematica code for all examples is included both in the book and at the accompanying ftp site, affording the freedom to change, at will, the values and parameters of specific problems or even modify the programs themselves to suit various modeling needs. Exploring Scanning Probe Microscopy with Mathematica is both a solid professional reference and an advanced-level text, beginning with scanning probe microscopy basics and moving on to cutting-edge techniques, experiments, and theory. In the section devoted to atomic force microscopy, Dr. Sarid describes the mechanical properties of cantilevers, atomic force microscope tip-sample interactions, and cantilever vibration characteristics. This is followed by an in-depth treatment of theoretical and practical aspects of tunneling phenomena, including metal-insulator-metal tunneling and Fowler-Nordheim field emission. The final section features chapters covering density of states in arbitrary dimensions, quantum wells and dots, and electrostatics.

  7. X-ray microscopy of human malaria

    SciTech Connect

    Magowan, C.; Brown, J.T.; Mohandas, N.; Meyer-Ilse, W.

    1997-04-01

    Associations between intracellular organisms and host cells are complex and particularly difficult to examine. X-ray microscopy provides transmission images of subcellular structures in intact cells at resolutions superior to available methodologies. The spatial resolution is 50-60nm with a 1 micron depth of focus, superior to anything achievable with light microscopy. Image contrast is generated by differences in photoelectric absorption by the atoms in different areas (i.e. subcellular structures) throughout the full thickness of the sample. Absorption due to carbon dominates among all the elements in the sample at 2.4 nm x-ray wavelength. Thus images show features or structures, in a way not usually seen by other types of microscopy. The authors used soft x-ray microscopy to investigate structural development of Plasmodium falciparum malaria parasites in normal and genetically abnormal erythrocytes, and in infected erythrocytes treated with compounds that have anti-malarial effects. X-ray microscopy showed newly elaborated structures in the cytosol of unstained, intact erythrocytes, redistribution of mass (carbon) in infected erythrocytes, and aberrant parasite morphology. Better understanding of the process of intracellular parasite maturation and the interactions between the parasite and its host erythrocyte can help define new approaches to the control of this deadly disease.

  8. Photoacoustic microscopy: superdepth, superresolution, and superb contrast.

    PubMed

    Yao, Junjie; Song, Liang; Wang, Lihing V

    2015-01-01

    Since its invention in the 17th century, optical microscopy has revolutionized biomedical studies by scrutinizing the biological realm on cellular levels, taking advantage of its excellent light-focusing capability. However, most biological tissues scatter light highly. As light travels in tissue, cumulative scattering events cause the photons to lose their original propagation direction and, thus, their ability to be focused, which has largely limited the penetration depth of optical microscopy. Conventional planar optical microscopy can provide penetration of only ~100 ?m before photons begin to be scattered. The penetration of modern optical microcopy, such as confocal microscopy and multiphoton microscopy, is still limited to approximately the optical diffusion limit (~1 mm in the skin as approximated by one optical transport mean free path), where scattered photons retain a strong memory of the original propagation direction. So far, it still remains a challenge for pure optical methods to achieve high-resolution in vivo imaging beyond the diffusion limit (i.e., superdepth imaging).

  9. Functional photoacoustic microscopy of pH

    NASA Astrophysics Data System (ADS)

    Chatni, M. Rameez; Yao, Junjie; Danielli, Amos; Favazza, Christopher P.; Maslov, Konstantin I.; Wang, Lihong V.

    2012-02-01

    pH is a tightly regulated indicator of metabolic activity. In mammalian systems, imbalance of pH regulation may result from or result in serious illness. Even though the regulation system of pH is very robust, tissue pH can be altered in many diseases such as cancer, osteoporosis and diabetes mellitus. Traditional high-resolution optical imaging techniques, such as confocal microscopy, routinely image pH in cells and tissues using pH sensitive fluorescent dyes, which change their fluorescence properties with the surrounding pH. Since strong optical scattering in biological tissue blurs images at greater depths, high-resolution pH imaging is limited to penetration depths of 1mm. Here, we report photoacoustic microscopy (PAM) of commercially available pH-sensitive fluorescent dye in tissue phantoms. Using both opticalresolution photoacoustic microscopy (OR-PAM), and acoustic resolution photoacoustic microscopy (AR-PAM), we explored the possibility of recovering the pH values in tissue phantoms. In this paper, we demonstrate that PAM was capable of recovering pH values up to a depth of 2 mm, greater than possible with other forms of optical microscopy.

  10. CARS microscopy of Alzheimer's diseased brain tissue

    NASA Astrophysics Data System (ADS)

    Enejder, Annika; Kiskis, Juris; Fink, Helen; Nyberg, Lena; Thyr, Jakob; Li, Jia-Yi

    2014-02-01

    Alzheimer's disease (AD) is a progressive neurodegenerative disorder currently without cure, characterized by the presence of extracellular plaques surrounded by dystrophic neurites. In an effort to understand the underlying mechanisms, biochemical analysis (protein immunoblot) of plaque extracts reveals that they consist of amyloid-beta (Aβ) peptides assembled as oligomers, protofibrils and aggregates. Their spatial distribution has been confirmed by Thioflavin-S or immuno-staining with fluorescence microscopy. However, it is increasingly understood that the protein aggregation is only one of several mechanism that causes neuronal dysfunction and death. This raises the need for a more complete biochemical analysis. In this study, we have complemented 2-photon fluorescence microscopy of Thioflavin-S and Aβ immuno-stained human AD plaques with CARS microscopy. We show that the chemical build-up of AD plaques is more complex and that Aβ staining does not provide the complete picture of the spatial distribution or the molecular composition of AD plaques. CARS images provide important complementary information to that obtained by fluorescence microscopy, motivating a broader introduction of CARS microscopy in the AD research field.

  11. Ultrafast Science Opportunities with Electron Microscopy

    SciTech Connect

    Durr, Hermann

    2016-04-28

    X-rays and electrons are two of the most fundamental probes of matter. When the Linac Coherent Light Source (LCLS), the world’s first x-ray free electron laser, began operation in 2009, it transformed ultrafast science with the ability to generate laser-like x-ray pulses from the manipulation of relativistic electron beams. This document describes a similar future transformation. In Transmission Electron Microscopy, ultrafast relativistic (MeV energy) electron pulses can achieve unsurpassed spatial and temporal resolution. Ultrafast temporal resolution will be the next frontier in electron microscopy and can ideally complement ultrafast x-ray science done with free electron lasers. This document describes the Grand Challenge science opportunities in chemistry, material science, physics and biology that arise from an MeV ultrafast electron diffraction & microscopy facility, especially when coupled with linac-based intense THz and X-ray pump capabilities.

  12. Fluorescence microscopy: A tool to study autophagy

    NASA Astrophysics Data System (ADS)

    Rai, Shashank; Manjithaya, Ravi

    2015-08-01

    Autophagy is a cellular recycling process through which a cell degrades old and damaged cellular components such as organelles and proteins and the degradation products are reused to provide energy and building blocks. Dysfunctional autophagy is reported in several pathological situations. Hence, autophagy plays an important role in both cellular homeostasis and diseased conditions. Autophagy can be studied through various techniques including fluorescence based microscopy. With the advancements of newer technologies in fluorescence microscopy, several novel processes of autophagy have been discovered which makes it an essential tool for autophagy research. Moreover, ability to tag fluorescent proteins with sub cellular targets has enabled us to evaluate autophagy processes in real time under fluorescent microscope. In this article, we demonstrate different aspects of autophagy in two different model organisms i.e. yeast and mammalian cells, with the help of fluorescence microscopy.

  13. Spectroscopy and atomic force microscopy of biomass.

    PubMed

    Tetard, L; Passian, A; Farahi, R H; Kalluri, U C; Davison, B H; Thundat, T

    2010-05-01

    Scanning probe microscopy has emerged as a powerful approach to a broader understanding of the molecular architecture of cell walls, which may shed light on the challenge of efficient cellulosic ethanol production. We have obtained preliminary images of both Populus and switchgrass samples using atomic force microscopy (AFM). The results show distinctive features that are shared by switchgrass and Populus. These features may be attributable to the lignocellulosic cell wall composition, as the collected images exhibit the characteristic macromolecular globule structures attributable to the lignocellulosic systems. Using both AFM and a single case of mode synthesizing atomic force microscopy (MSAFM) to characterize Populus, we obtained images that clearly show the cell wall structure. The results are of importance in providing a better understanding of the characteristic features of both mature cells as well as developing plant cells. In addition, we present spectroscopic investigation of the same samples.

  14. Near-infrared hyperspectral reflective confocal microscopy

    NASA Astrophysics Data System (ADS)

    Huang, Wei; Zhang, Yunhai; Miao, Xin; Xue, Xiaojun; Xiao, Yun

    2016-10-01

    A Near-Infrared HyperSpectral Reflective Confocal Microscopy (NIHS-RCM) is proposed in order to get high resolution images of deep biological tissues such as skin. The microscopy system uses a super-continuum laser for illumination, an acousto-optic tunable filter (AOTF) for rapid selection of near-infrared spectrum, a resonant galvanometer scanner for high speed imaging (15f/s) and near-infrared avalanche diode as detector. Porcine skin and other experiments show that the microscopy system could get deep tissue images (180 μm), and show the different ingredients of tissue with different wavelength of illumination. The system has the ability of selectively imaging of multiple ingredients at deep tissue which can be used in skin diseases diagnosis and other fields.

  15. Cryogenics with cement microscopy redefines cement behavior

    SciTech Connect

    Mehta, S.; Jones, R. ); Caveny, B. )

    1994-10-03

    Cement microscopy (CM), cryogenics, environmental scanning microscopy (ESM), scanning electron microscopy (SEM), and other technologies are leading investigators to change their views on cement gelation, hydration, and retardation. Cement samples frozen in a nitrogen slush and viewed with an SEM present a more accurate picture of the setting process. Observations made through this technique have revolutionized ARCO Exploration and Production Technology's and Halliburton Energy Services' oil field cement procurement and slurry design. Findings from this joint study are expected to lead to: optimized waiting on cement (WOC) times; reduced planning and design time; optimized slurry retarder additions; optimized gel times to fit given situations; especially applicable to squeeze operations; improved cement selection (from vendors) for peak performance; and improved cement manufacture. The paper discusses the measuring methods and the findings on the following: cement voids, cement gelation, and retardation mechanisms. It also briefly discusses the impact these discoveries have on operations.

  16. Coherent Nonlinear Optical Imaging: Beyond Fluorescence Microscopy

    PubMed Central

    Min, Wei; Freudiger, Christian W.; Lu, Sijia; Xie, X. Sunney

    2012-01-01

    The quest for ultrahigh detection sensitivity with spectroscopic contrasts other than fluorescence has led to various novel approaches to optical microscopy of biological systems. Coherent nonlinear optical imaging, especially the recently developed nonlinear dissipation microscopy, including stimulated Raman scattering and two photon absorption, and pump-probe microscopy, including stimulated emission, excited state absorption and ground state depletion, provide distinct and powerful image contrasts for non-fluorescent species. Thanks to high-frequency modulation transfer scheme, they exhibit superb detection sensitivity. By directly interrogating vibrational and/or electronic energy levels of molecules, they offer high molecular specificity. Here we review the underlying principles, excitation and detection schemes, as well as exemplary biomedical applications of this emerging class of molecular imaging techniques. PMID:21453061

  17. Coherent nonlinear optical imaging: beyond fluorescence microscopy.

    PubMed

    Min, Wei; Freudiger, Christian W; Lu, Sijia; Xie, X Sunney

    2011-01-01

    The quest for ultrahigh detection sensitivity with spectroscopic contrasts other than fluorescence has led to various novel approaches to optical microscopy of biological systems. Coherent nonlinear optical imaging, especially the recently developed nonlinear dissipation microscopy (including stimulated Raman scattering and two-photon absorption) and pump-probe microscopy (including excited-state absorption, stimulated emission, and ground-state depletion), provides new image contrasts for nonfluorescent species. Thanks to the high-frequency modulation transfer scheme, these imaging techniques exhibit superb detection sensitivity. By directly interrogating vibrational and/or electronic energy levels of molecules, they offer high molecular specificity. Here we review the underlying principles and excitation and detection schemes, as well as exemplary biomedical applications of this emerging class of molecular imaging techniques.

  18. Ultrahigh resolution photoacoustic microscopy via transient absorption

    PubMed Central

    Shelton, Ryan L.; Applegate, Brian E.

    2010-01-01

    We have developed a novel, hybrid imaging modality, Transient Absorption Ultrasonic Microscopy (TAUM), which takes advantage of the optical nonlinearities afforded by transient absorption to achieve ultrahigh-resolution photoacoustic microscopy. The theoretical point spread function for TAUM is functionally equivalent to confocal and two-photon fluorescence microscopy, potentially enabling cellular/subcellular photoacoustic imaging. A prototype TAUM system was designed, built, and used to image a cross-section through several capillaries in the excised cheek pouch of a Syrian Hamster. The well-resolved capillaries in the TAUM image provided experimental evidence of the spatial resolution. These results suggest that TAUM has excellent potential for producing volumetric images with cellular/subcellular resolution in three dimensions deep inside living tissue. PMID:21258499

  19. Space station microscopy: Beyond the box

    NASA Astrophysics Data System (ADS)

    Hunter, N. R.; Pierson, Duane L.; Mishra, S. K.

    Microscopy aboard Space Station Freedom poses many unique challenges for in-flight investigations. Disciplines such as material processing, plant and animal research, human reseach, enviromental monitoring, health care, and biological processing have diverse microscope requirements. The typical microscope not only does not meet the comprehensive needs of these varied users, but also tends to require excessive crew time. To assess user requirements, a comprehensive survey was conducted among investigators with experiments requiring microscopy. The survey examined requirements such as light sources, objectives, stages, focusing systems, eye pieces, video accessories, etc. The results of this survey and the application of an Intelligent Microscope Imaging System (IMIS) may address these demands for efficient microscopy service in space. The proposed IMIS can accommodate multiple users with varied requirements, operate in several modes, reduce crew time needed for experiments, and take maximum advantage of the restrictive data/ instruction transmission environment on Freedom.

  20. Light microscopy: an ongoing contemporary revolution

    NASA Astrophysics Data System (ADS)

    Weisenburger, Siegfried; Sandoghdar, Vahid

    2015-04-01

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

  1. A near-field optical microscopy nanoarray

    SciTech Connect

    Semin, D.J.; Ambrose, W.P.; Goodwin, P.M.; Kwller, A.; Wendt, J.R.

    1996-12-31

    Multiplexing near-field scanning optical microscopy (NSOM) by the use of a nanoarray with parallel imaging is studied. The fabrication, characterization, and utilization of nanoarrays with {approximately} 100 nm diameter apertures spaced 500 nm center-to- center is presented. Extremely uniform nanoarrays with {approximately} 10{sup 8} apertures were fabricated by electron beam lithography and reactive ion etching. The nanoarrays were characterized by atomic force microscopy (AFM) and scanning electron microscopy (SEM). In this paper we utilize these nanoarrays in a laser-illuminated microscope with parallel detection on a charge- coupled device (CCD). Detection of B-phycoerythrin (B-PE) molecules using near-field illumination is presented. In principle, our system can be used to obtain high lateral resolution NSOM images over a wide-field of view (e.g. 50-100 {mu}m) within seconds.

  2. Environmental scanning electron microscopy in cell biology.

    PubMed

    McGregor, J E; Staniewicz, L T L; Guthrie Neé Kirk, S E; Donald, A M

    2013-01-01

    Environmental scanning electron microscopy (ESEM) (1) is an imaging technique which allows hydrated, insulating samples to be imaged under an electron beam. The resolution afforded by this technique is higher than conventional optical microscopy but lower than conventional scanning electron microscopy (CSEM). The major advantage of the technique is the minimal sample preparation needed, making ESEM quick to use and the images less susceptible to the artifacts that the extensive sample preparation usually required for CSEM may introduce. Careful manipulation of both the humidity in the microscope chamber and the beam energy are nevertheless essential to prevent dehydration and beam damage artifacts. In some circumstances it is possible to image live cells in the ESEM (2).In the following sections we introduce the fundamental principles of ESEM imaging before presenting imaging protocols for plant epidermis, mammalian cells, and bacteria. In the first two cases samples are imaged using the secondary electron (topographic) signal, whereas a transmission technique is employed to image bacteria.

  3. Space station microscopy: Beyond the box

    NASA Technical Reports Server (NTRS)

    Hunter, N. R.; Pierson, Duane L.; Mishra, S. K.

    1993-01-01

    Microscopy aboard Space Station Freedom poses many unique challenges for in-flight investigations. Disciplines such as material processing, plant and animal research, human reseach, enviromental monitoring, health care, and biological processing have diverse microscope requirements. The typical microscope not only does not meet the comprehensive needs of these varied users, but also tends to require excessive crew time. To assess user requirements, a comprehensive survey was conducted among investigators with experiments requiring microscopy. The survey examined requirements such as light sources, objectives, stages, focusing systems, eye pieces, video accessories, etc. The results of this survey and the application of an Intelligent Microscope Imaging System (IMIS) may address these demands for efficient microscopy service in space. The proposed IMIS can accommodate multiple users with varied requirements, operate in several modes, reduce crew time needed for experiments, and take maximum advantage of the restrictive data/ instruction transmission environment on Freedom.

  4. Applications of microscopy in Salmonella research.

    PubMed

    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.

  5. Magnetic exchange force microscopy with atomic resolution.

    PubMed

    Kaiser, Uwe; Schwarz, Alexander; Wiesendanger, Roland

    2007-03-29

    The ordering of neighbouring atomic magnetic moments (spins) leads to important collective phenomena such as ferromagnetism and antiferromagnetism. A full understanding of magnetism on the nanometre scale therefore calls for information on the arrangement of spins in real space and with atomic resolution. Spin-polarized scanning tunnelling microscopy accomplishes this but can probe only conducting materials. Force microscopy can be used on any sample independent of its conductivity. In particular, magnetic force microscopy is well suited to exploring ferromagnetic domain structures. However, atomic resolution cannot be achieved because data acquisition involves the sensing of long-range magnetostatic forces between tip and sample. Magnetic exchange force microscopy has been proposed for overcoming this limitation: by using an atomic force microscope with a magnetic tip, it should be possible to detect the short-range magnetic exchange force between tip and sample spins. Here we show for a prototypical antiferromagnetic insulator, the (001) surface of nickel oxide, that magnetic exchange force microscopy can indeed reveal the arrangement of both surface atoms and their spins simultaneously. In contrast with previous attempts to implement this method, we use an external magnetic field to align the magnetic polarization at the tip apex so as to optimize the interaction between tip and sample spins. This allows us to observe the direct magnetic exchange coupling between the spins of the tip atom and sample atom that are closest to each other, and thereby demonstrate the potential of magnetic exchange force microscopy for investigations of inter-spin interactions at the atomic level.

  6. Advanced Electron Microscopy in Materials Physics

    SciTech Connect

    Zhu, Y.; Jarausch, K.

    2009-06-01

    Aberration correction has opened a new frontier in electron microscopy by overcoming the limitations of conventional round lenses, providing sub-angstrom-sized probes and extending information limits. The imaging and analytical performance of these corrector-equipped microscopes affords an unprecedented opportunity to study structure-property relationships of matter at the atomic scale. This new generation of microscopes is able to retrieve high-quality structural information comparable to neutron and synchrotron x-ray experiments, but with local atomic resolution. These advances in instrumentation are accelerating the research and development of various functional materials ranging from those for energy generation, conversion, transportation and storage to those for catalysis and nano-device applications. The dramatic improvements in electron-beam illumination and detection also present a host of new challenges for the interpretation and optimization of experiments. During 7-9 November 2007, a workshop, entitled 'Aberration Corrected Electron Microscopy in Material Physics', was convened at the Center for Functional Nanomaterials, Brookhaven National Laboratories (BNL) to address these opportunities and challenges. The workshop was co-sponsored by Hitachi High Technologies, a leader in electron microscopy instrumentation, and BNL's Institute of Advanced Electron Microscopy, a leader in materials physics research using electron microscopy. The workshop featured presentations by internationally prominent scientists working at the frontiers of electron microscopy, both on developing instrumentation and applying it in materials physics. The meeting, structured to stimulate scientific exchanges and explore new capabilities, brought together {approx}100 people from over 10 countries. This special issue complies many of the advances in instrument performance and materials physics reported by the invited speakers and attendees at the workshop.

  7. Improved Interference configuration for structured illumination microscopy

    NASA Astrophysics Data System (ADS)

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

    2017-02-01

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

  8. Photobleaching imprinting microscopy: seeing clearer and deeper.

    PubMed

    Gao, Liang; Garcia-Uribe, Alejandro; Liu, Yan; Li, Chiye; Wang, Lihong V

    2014-01-15

    We present a generic sub-diffraction-limited imaging method - photobleaching imprinting microscopy (PIM) - for biological fluorescence imaging. A lateral resolution of 110 nm was measured, more than a twofold improvement over the optical diffraction limit. Unlike other super-resolution imaging techniques, PIM does not require complicated illumination modules or specific fluorescent dyes. PIM is expected to facilitate the conversion of super-resolution imaging into a routine lab tool, making it accessible to a much broader biological research community. Moreover, we show that PIM can increase the image contrast of biological tissue, effectively extending the fundamental depth limit of multi-photon fluorescence microscopy.

  9. Aberration corrected Lorentz scanning transmission electron microscopy.

    PubMed

    McVitie, S; McGrouther, D; McFadzean, S; MacLaren, D A; O'Shea, K J; Benitez, M J

    2015-05-01

    We present results from an aberration corrected scanning transmission electron microscope which has been customised for high resolution quantitative Lorentz microscopy with the sample located in a magnetic field free or low field environment. We discuss the innovations in microscope instrumentation and additional hardware that underpin the imaging improvements in resolution and detection with a focus on developments in differential phase contrast microscopy. Examples from materials possessing nanometre scale variations in magnetisation illustrate the potential for aberration corrected Lorentz imaging as a tool to further our understanding of magnetism on this lengthscale.

  10. Electron Microscopy of Natural and Epitaxial Diamond

    NASA Technical Reports Server (NTRS)

    Posthill, J. B.; George, T.; Malta, D. P.; Humphreys, T. P.; Rudder, R. A.; Hudson, G. C.; Thomas, R. E.; Markunas, R. J.

    1993-01-01

    Semiconducting diamond films have the potential for use as a material in which to build active electronic devices capable of operating at high temperatures or in high radiation environments. Ultimately, it is preferable to use low-defect-density single crystal diamond for device fabrication. We have previously investigated polycrystalline diamond films with transmission electron microscopy (TEM) and scanning electron microscopy (SEM), and homoepitaxial films with SEM-based techniques. This contribution describes some of our most recent observations of the microstructure of natural diamond single crystals and homoepitaxial diamond thin films using TEM.

  11. Phase imaging with intermodulation atomic force microscopy.

    PubMed

    Platz, Daniel; Tholén, Erik A; Hutter, Carsten; von Bieren, Arndt C; Haviland, David B

    2010-05-01

    Intermodulation atomic force microscopy (IMAFM) is a dynamic mode of atomic force microscopy (AFM) with two-tone excitation. The oscillating AFM cantilever in close proximity to a surface experiences the nonlinear tip-sample force which mixes the drive tones and generates new frequency components in the cantilever response known as intermodulation products (IMPs). We present a procedure for extracting the phase at each IMP and demonstrate phase images made by recording this phase while scanning. Amplitude and phase images at intermodulation frequencies exhibit enhanced topographic and material contrast.

  12. Super-resolution optical microscopy: multiple choices.

    PubMed

    Huang, Bo

    2010-02-01

    The recent invention of super-resolution optical microscopy enables the visualization of fine features in biological samples with unprecedented clarity. It creates numerous opportunities in biology because vast amount of previously obscured subcellular processes now can be directly observed. Rapid development in this field in the past two years offers many imaging modalities that address different needs but they also complicates the choice of the 'perfect' method for answering a specific question. Here I will briefly describe the principles of super-resolution optical microscopy techniques and then focus on comparing their characteristics in various aspects of practical applications.

  13. Fidelity imaging for atomic force microscopy

    NASA Astrophysics Data System (ADS)

    Ghosal, Sayan; Salapaka, Murti

    2015-01-01

    Atomic force microscopy is widely employed for imaging material at the nanoscale. However, real-time measures on image reliability are lacking in contemporary atomic force microscopy literature. In this article, we present a real-time technique that provides an image of fidelity for a high bandwidth dynamic mode imaging scheme. The fidelity images define channels that allow the user to have additional authority over the choice of decision threshold that facilitates where the emphasis is desired, on discovering most true features on the sample with the possible detection of high number of false features, or emphasizing minimizing instances of false detections. Simulation and experimental results demonstrate the effectiveness of fidelity imaging.

  14. Two-photon excitation fluorescence microscopy.

    PubMed

    So, P T; Dong, C Y; Masters, B R; Berland, K M

    2000-01-01

    Two-photon fluorescence microscopy is one of the most important recent inventions in biological imaging. This technology enables noninvasive study of biological specimens in three dimensions with submicrometer resolution. Two-photon excitation of fluorophores results from the simultaneous absorption of two photons. This excitation process has a number of unique advantages, such as reduced specimen photodamage and enhanced penetration depth. It also produces higher-contrast images and is a novel method to trigger localized photochemical reactions. Two-photon microscopy continues to find an increasing number of applications in biology and medicine.

  15. Observation of DNA Molecules Using Fluorescence Microscopy and Atomic Force Microscopy

    ERIC Educational Resources Information Center

    Ito, Takashi

    2008-01-01

    This article describes experiments for an undergraduate instrumental analysis laboratory that aim to observe individual double-stranded DNA (dsDNA) molecules using fluorescence microscopy and atomic force microscopy (AFM). dsDNA molecules are observed under several different conditions to discuss their chemical and physical properties. In…

  16. Graphene-enabled electron microscopy and correlated super-resolution microscopy of wet cells.

    PubMed

    Wojcik, Michal; Hauser, Margaret; Li, Wan; Moon, Seonah; Xu, Ke

    2015-06-11

    The application of electron microscopy to hydrated biological samples has been limited by high-vacuum operating conditions. Traditional methods utilize harsh and laborious sample dehydration procedures, often leading to structural artefacts and creating difficulties for correlating results with high-resolution fluorescence microscopy. Here, we utilize graphene, a single-atom-thick carbon meshwork, as the thinnest possible impermeable and conductive membrane to protect animal cells from vacuum, thus enabling high-resolution electron microscopy of wet and untreated whole cells with exceptional ease. Our approach further allows for facile correlative super-resolution and electron microscopy of wet cells directly on the culturing substrate. In particular, individual cytoskeletal actin filaments are resolved in hydrated samples through electron microscopy and well correlated with super-resolution results.

  17. Graphene-enabled electron microscopy and correlated super-resolution microscopy of wet cells

    PubMed Central

    Wojcik, Michal; Hauser, Margaret; Li, Wan; Moon, Seonah; Xu, Ke

    2015-01-01

    The application of electron microscopy to hydrated biological samples has been limited by high-vacuum operating conditions. Traditional methods utilize harsh and laborious sample dehydration procedures, often leading to structural artefacts and creating difficulties for correlating results with high-resolution fluorescence microscopy. Here, we utilize graphene, a single-atom-thick carbon meshwork, as the thinnest possible impermeable and conductive membrane to protect animal cells from vacuum, thus enabling high-resolution electron microscopy of wet and untreated whole cells with exceptional ease. Our approach further allows for facile correlative super-resolution and electron microscopy of wet cells directly on the culturing substrate. In particular, individual cytoskeletal actin filaments are resolved in hydrated samples through electron microscopy and well correlated with super-resolution results. PMID:26066680

  18. (Gene sequencing by scanning molecular exciton microscopy)

    SciTech Connect

    Not Available

    1991-01-01

    This report details progress made in setting up a laboratory for optical microscopy of genes. The apparatus including a fluorescence microscope, a scanning optical microscope, various spectrometers, and supporting computers is described. Results in developing photon and exciton tips, and in preparing samples are presented. (GHH)

  19. Value of Reflected Light Microscopy in Teaching.

    ERIC Educational Resources Information Center

    Pasteris, Jill Dill

    1983-01-01

    Briefly reviews some optical and other physical properties of minerals that can be determined in reflected/incident light. Topics include optical properties of minerals, reflectance, internal reflections, color, bireflectance and reflection pleochroism, anisotropism, zonation, and reflected light microscopy as a teaching tool in undergraduate…

  20. Scanning electron microscopy study of Trichomonas gallinae.

    PubMed

    Tasca, Tiana; De Carli, Geraldo A

    2003-12-01

    A scanning electron microscopy (SEM) study of Trichomonas gallinae (Rivolta, 1878), provided more information about the morphology of this flagellated protozoan. SEM showed the morphological features of the trophozoites; the emergence of the anterior flagella, the structure of the undulating membrane, the position and shape of the pelta, axostyle and posterior flagellum. Of special interest were the pseudocyst forms.

  1. DNA base identification by electron microscopy.

    PubMed

    Bell, David C; Thomas, W Kelley; Murtagh, Katelyn M; Dionne, Cheryl A; Graham, Adam C; Anderson, Jobriah E; Glover, William R

    2012-10-01

    Advances in DNA sequencing, based on fluorescent microscopy, have transformed many areas of biological research. However, only relatively short molecules can be sequenced by these technologies. Dramatic improvements in genomic research will require accurate sequencing of long (>10,000 base-pairs), intact DNA molecules. Our approach directly visualizes the sequence of DNA molecules using electron microscopy. This report represents the first identification of DNA base pairs within intact DNA molecules by electron microscopy. By enzymatically incorporating modified bases, which contain atoms of increased atomic number, direct visualization and identification of individually labeled bases within a synthetic 3,272 base-pair DNA molecule and a 7,249 base-pair viral genome have been accomplished. This proof of principle is made possible by the use of a dUTP nucleotide, substituted with a single mercury atom attached to the nitrogenous base. One of these contrast-enhanced, heavy-atom-labeled bases is paired with each adenosine base in the template molecule and then built into a double-stranded DNA molecule by a template-directed DNA polymerase enzyme. This modification is small enough to allow very long molecules with labels at each A-U position. Image contrast is further enhanced by using annular dark-field scanning transmission electron microscopy (ADF-STEM). Further refinements to identify additional base types and more precisely determine the location of identified bases would allow full sequencing of long, intact DNA molecules, significantly improving the pace of complex genomic discoveries.

  2. Low voltage transmission electron microscopy of graphene.

    PubMed

    Bachmatiuk, Alicja; Zhao, Jiong; Gorantla, Sandeep Madhukar; Martinez, Ignacio Guillermo Gonzalez; Wiedermann, Jerzy; Lee, Changgu; Eckert, Juergen; Rummeli, Mark Hermann

    2015-02-04

    The initial isolation of graphene in 2004 spawned massive interest in this two-dimensional pure sp(2) carbon structure due to its incredible electrical, optical, mechanical, and thermal effects. This in turn led to the rapid development of various characterization tools for graphene. Examples include Raman spectroscopy and scanning tunneling microscopy. However, the one tool with the greatest prowess for characterizing and studying graphene is the transmission electron microscope. State-of-the-art (scanning) transmission electron microscopes enable one to image graphene with atomic resolution, and also to conduct various other characterizations simultaneously. The advent of aberration correctors was timely in that it allowed transmission electron microscopes to operate with reduced acceleration voltages, so that damage to graphene is avoided while still providing atomic resolution. In this comprehensive review, a brief introduction is provided to the technical aspects of transmission electron microscopes relevant to graphene. The reader is then introduced to different specimen preparation techniques for graphene. The different characterization approaches in both transmission electron microscopy and scanning transmission electron microscopy are then discussed, along with the different aspects of electron diffraction and electron energy loss spectroscopy. The use of graphene for other electron microscopy approaches such as in-situ investigations is also presented.

  3. Light Microscopy Module (LMM)-Emulator

    NASA Technical Reports Server (NTRS)

    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.

  4. Atomic force microscopy of biological samples.

    PubMed

    Allison, David P; Mortensen, Ninell P; Sullivan, Claretta J; Doktycz, Mitchel J

    2010-01-01

    The ability to evaluate structural-functional relationships in real time has allowed scanning probe microscopy (SPM) to assume a prominent role in post genomic biological research. In this mini-review, we highlight the development of imaging and ancillary techniques that have allowed SPM to permeate many key areas of contemporary research. We begin by examining the invention of the scanning tunneling microscope (STM) by Binnig and Rohrer in 1982 and discuss how it served to team biologists with physicists to integrate high-resolution microscopy into biological science. We point to the problems of imaging nonconductive biological samples with the STM and relate how this led to the evolution of the atomic force microscope (AFM) developed by Binnig, Quate, and Gerber, in 1986. Commercialization in the late 1980s established SPM as a powerful research tool in the biological research community. Contact mode AFM imaging was soon complemented by the development of non-contact imaging modes. These non-contact modes eventually became the primary focus for further new applications including the development of fast scanning methods. The extreme sensitivity of the AFM cantilever was recognized and has been developed into applications for measuring forces required for indenting biological surfaces and breaking bonds between biomolecules. Further functional augmentation to the cantilever tip allowed development of new and emerging techniques including scanning ion-conductance microscopy (SICM), scanning electrochemical microscope (SECM), Kelvin force microscopy (KFM) and scanning near field ultrasonic holography (SNFUH).

  5. Atomic force microscopy of biological samples

    SciTech Connect

    Doktycz, Mitchel John

    2010-01-01

    The ability to evaluate structural-functional relationships in real time has allowed scanning probe microscopy (SPM) to assume a prominent role in post genomic biological research. In this mini-review, we highlight the development of imaging and ancillary techniques that have allowed SPM to permeate many key areas of contemporary research. We begin by examining the invention of the scanning tunneling microscope (STM) by Binnig and Rohrer in 1982 and discuss how it served to team biologists with physicists to integrate high-resolution microscopy into biological science. We point to the problems of imaging nonconductive biological samples with the STM and relate how this led to the evolution of the atomic force microscope (AFM) developed by Binnig, Quate, and Gerber, in 1986. Commercialization in the late 1980s established SPM as a powerful research tool in the biological research community. Contact mode AFM imaging was soon complemented by the development of non-contact imaging modes. These non-contact modes eventually became the primary focus for further new applications including the development of fast scanning methods. The extreme sensitivity of the AFM cantilever was recognized and has been developed into applications for measuring forces required for indenting biological surfaces and breaking bonds between biomolecules. Further functional augmentation to the cantilever tip allowed development of new and emerging techniques including scanning ion-conductance microscopy (SICM), scanning electrochemical microscope (SECM), Kelvin force microscopy (KFM) and scanning near field ultrasonic holography (SNFUH).

  6. High-aperture cryogenic light microscopy.

    PubMed

    Le Gros, M A; McDermott, G; Uchida, M; Knoechel, C G; Larabell, C A

    2009-07-01

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

  7. Frequency domain photoacoustic and fluorescence microscopy

    PubMed Central

    Langer, Gregor; Buchegger, Bianca; Jacak, Jaroslaw; Klar, Thomas A.; Berer, Thomas

    2016-01-01

    We report on simultaneous frequency domain optical-resolution photoacoustic and fluorescence microscopy with sub-µm lateral resolution. With the help of a blood smear, we show that photoacoustic and fluorescence images provide complementary information. Furthermore, we compare theoretically predicted signal-to-noise ratios of sinusoidal modulation in frequency domain with pulsed excitation in time domain. PMID:27446698

  8. CONFOCAL MICROSCOPY SYSTEM PERFORMANCE: AXIAL RESOLUTION

    EPA Science Inventory

    Abstract

    Confocal Microscopy System Performance: Axial resolution.
    Robert M. Zucker, PhD

    Reproductive Toxicology Division, National Health and Environmental Effects Research Laboratory, Office of Research and Development, U.S. Environmental Protection Agency, Re...

  9. Confocal microscopy imaging of solid tissue

    EPA Science Inventory

    Confocal laser scanning microscopy (CLSM) is a technique that is capable of generating serial sections of whole-mount tissue and then reassembling the computer acquired images as a virtual 3-dimensional structure. In many ways CLSM offers an alternative to traditional sectioning ...

  10. Dark Field Microscopy for Analytical Laboratory Courses

    ERIC Educational Resources Information Center

    Augspurger, Ashley E.; Stender, Anthony S.; Marchuk, Kyle; Greenbowe, Thomas J.; Fang, Ning

    2014-01-01

    An innovative and inexpensive optical microscopy experiment for a quantitative analysis or an instrumental analysis chemistry course is described. The students have hands-on experience with a dark field microscope and investigate the wavelength dependence of localized surface plasmon resonance in gold and silver nanoparticles. Students also…

  11. Scanning Probe Microscopy of Organic Solar Cells

    NASA Astrophysics Data System (ADS)

    Reid, Obadiah G.

    Nanostructured composites of organic semiconductors are a promising class of materials for the manufacture of low-cost solar cells. Understanding how the nanoscale morphology of these materials affects their efficiency as solar energy harvesters is crucial to their eventual potential for large-scale deployment for primary power generation. In this thesis we describe the use of optoelectronic scanning-probe based microscopy methods to study this efficiency-structure relationship with nanoscale resolution. In particular, our objective is to make spatially resolved measurements of each step in the power conversion process from photons to an electric current, including charge generation, transport, and recombination processes, and correlate them with local device structure. We have achieved two aims in this work: first, to develop and apply novel electrically sensitive scanning probe microscopy experiments to study the optoelectronic materials and processes discussed above; and second, to deepen our understanding of the physics underpinning our experimental techniques. In the first case, we have applied conductive-, and photoconductive atomic force (cAFM & pcAFM) microscopy to measure both local photocurrent collection and dark charge transport properties in a variety of model and novel organic solar cell composites, including polymer/fullerene blends, and polymer-nanowire/fullerene blends, finding that local heterogeneity is the rule, and that improvements in the uniformity of specific beneficial nanostructures could lead to large increases in efficiency. We have used scanning Kelvin probe microscopy (SKPM) and time resolved-electrostatic force microscopy (trEFM) to characterize all-polymer blends, quantifying their sensitivity to photochemical degradation and the subsequent formation of local charge traps. We find that while trEFM provides a sensitive measure of local quantum efficiency, SKPM is generally unsuited to measurements of efficiency, less sensitive than tr

  12. Fast Ion Beam Microscopy of Whole Cells

    NASA Astrophysics Data System (ADS)

    Watt, Frank; Chen, Xiao; Chen, Ce-Belle; Udalagama, Chammika Nb; Ren, Minqin; Pastorin, G.; Bettiol, Andrew

    2013-08-01

    The way in which biological cells function is of prime importance, and the determination of such knowledge is highly dependent on probes that can extract information from within the cell. Probing deep inside the cell at high resolutions however is not easy: optical microscopy is limited by fundamental diffraction limits, electron microscopy is not able to maintain spatial resolutions inside a whole cell without slicing the cell into thin sections, and many other new and novel high resolution techniques such as atomic force microscopy (AFM) and near field scanning optical microscopy (NSOM) are essentially surface probes. In this paper we show that microscopy using fast ions has the potential to extract information from inside whole cells in a unique way. This novel fast ion probe utilises the unique characteristic of MeV ion beams, which is the ability to pass through a whole cell while maintaining high spatial resolutions. This paper first addresses the fundamental difference between several types of charged particle probes, more specifically focused beams of electrons and fast ions, as they penetrate organic material. Simulations show that whereas electrons scatter as they penetrate the sample, ions travel in a straight path and therefore maintain spatial resolutions. Also described is a preliminary experiment in which a whole cell is scanned using a low energy (45 keV) helium ion microscope, and the results compared to images obtained using a focused beam of fast (1.2 MeV) helium ions. The results demonstrate the complementarity between imaging using low energy ions, which essentially produce a high resolution image of the cell surface, and high energy ions, which produce an image of the cell interior. The characteristics of the fast ion probe appear to be ideally suited for imaging gold nanoparticles in whole cells. Using scanning transmission ion microscopy (STIM) to image the cell interior, forward scattering transmission ion microscopy (FSTIM) to improve the

  13. Leica solution: CARS microscopy at video rates

    NASA Astrophysics Data System (ADS)

    Lurquin, V.

    2008-02-01

    Confocal and multiphoton microscopy are powerful techniques to study morphology and dynamics in cells and tissue, if fluorescent labeling is possible or autofluorescence is strong. For non-fluorescent molecules, Coherent anti-Stokes Raman scattering (CARS) microscopy provides chemical contrast based on intrinsic and highly specific vibrational properties of molecules eliminating the need for labeling. Just as other multiphoton techniques, CARS microscopy possesses three-dimensional sectioning capabilities. Leica Microsystems has combined the CARS imaging technology with its TCS SP5 confocal microscope to provide several advantages for CARS imaging. For CARS microscopy, two picosecond near-infrared lasers are overlapped spatially and temporally and sent into the scanhead of the confocal system. The software allows programmed, automatic switching between these light sources for multi-modal imaging. Furthermore the Leica TCS SP5 can be equipped with a non-descanned detector which will significantly enhance the signal. The Leica TCS SP5 scanhead combines two technologies in one system: a conventional scanner for maximum resolution and a resonant scanner for high time resolution. The fast scanner allows imaging speeds as high as 25 images/per second at a resolution of 512×512 pixel. This corresponds to true video-rate allowing to follow processes at these time-scales as well as the acquisition of three-dimensional stacks in a few seconds. This time resolution is critical to study live animals or human patients for which heart beat and muscle movements lead to a blurring of the image if the acquisition time is high. Furthermore with the resonant scanhead the sectioning is truly confocal and does not suffer from spatial leakage. In summary, CARS microscopy combined with the tandem scanner makes the Leica TCS SP5 a powerful tool for three-dimensional, label-free imaging of chemical and biological samples in vitro and in vivo.

  14. On the Law of Inertia. Translation of: Ueber das Beharrungsgesetz

    NASA Astrophysics Data System (ADS)

    Lange, Ludwig

    2014-04-01

    This article is a translation of Ludwig Lange: "Ueber das Beharrungsgesetz" in: Berichte ueber Verhandlungen der Koenigl. Saechsischen Gesellschaft der Wissenschaften, math.-physik. Klasse (Leipzig, 1885), SS. 333-351. Translated by Herbert Pfister, Institut für Theoretische Physik, Universität Tübingen, Auf der Morgenstelle 14, 72076 Tübingen, Germany; herbert.pfister@uni-tuebingen.de. Kind assistance by Julian Barbour is acknowledged.

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

    SciTech Connect

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

    2015-09-15

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

  16. Probing cytotoxicity of nanoparticles and organic compounds using scanning proton microscopy, scanning electron microscopy and fluorescence microscopy

    NASA Astrophysics Data System (ADS)

    Tong, Yongpeng; Li, Changming; Liang, Feng; Chen, Jianmin; Zhang, Hong; Liu, Guoqing; Sun, Huibin; Luong, John H. T.

    2008-12-01

    Scanning proton microscopy, scanning electron microscopy (SEM) and fluorescence microscopy have been used to probe the cytotoxicity effect of benzo[a]pyrene (BaP), ethidium bromide (EB) and nanoparticles (ZnO, Al 2O 3 and TiO 2) on a T lymphoblastic leukemia Jurkat cell line. The increased calcium ion (from CaCl 2) in the culture medium stimulated the accumulation of BaP and EB inside the cell, leading to cell death. ZnO, Al 2O 3 and TiO 2 nanoparticles, however, showed a protective effect against these two organic compounds. Such inorganic nanoparticles complexed with BaP or EB which became less toxic to the cell. Fe 2O 3 nanoparticles as an insoluble particle model scavenged by macrophage were investigated in rats. They were scavenged out of the lung tissue about 48 h after infection. This result suggest that some insoluble inorganic nanoparticles of PM (particulate matters) showed protective effects on organic toxins induced acute toxic effects as they can be scavenged by macrophage cells. Whereas, some inorganic ions such as calcium ion in PM may help environmental organic toxins to penetrate cell membrane and induce higher toxic effect.

  17. Advances in Light Microscopy for Neuroscience

    PubMed Central

    Wilt, Brian A.; Burns, Laurie D.; Ho, Eric Tatt Wei; Ghosh, Kunal K.; Mukamel, Eran A.

    2010-01-01

    Since the work of Golgi and Cajal, light microscopy has remained a key tool for neuroscientists to observe cellular properties. Ongoing advances have enabled new experimental capabilities using light to inspect the nervous system across multiple spatial scales, including ultrastructural scales finer than the optical diffraction limit. Other progress permits functional imaging at faster speeds, at greater depths in brain tissue, and over larger tissue volumes than previously possible. Portable, miniaturized fluorescence microscopes now allow brain imaging in freely behaving mice. Complementary progress on animal preparations has enabled imaging in head-restrained behaving animals, as well as time-lapse microscopy studies in the brains of live subjects. Mouse genetic approaches permit mosaic and inducible fluorescence-labeling strategies, whereas intrinsic contrast mechanisms allow in vivo imaging of animals and humans without use of exogenous markers. This review surveys such advances and highlights emerging capabilities of particular interest to neuroscientists. PMID:19555292

  18. Stimulated Raman scattering microscopy for biomedical imaging

    NASA Astrophysics Data System (ADS)

    Min, Wei; Freudiger, Christian W.; Lu, Sijia; He, Chengwei; Kang, Jing X.; Xie, X. Sunney

    2009-02-01

    Label-free chemical contrast is highly desirable in biomedical imaging. Spontaneous Raman microscopy provides specific vibrational signatures of chemical bonds, but is often hindered by low sensitivity. Here we report a 3D multi-photon vibrational imaging technique based on stimulated Raman scattering (SRS). The sensitivity of SRS is significantly greater than that of spontaneous Raman scattering, and is further enhanced by high-frequency (MHz) phase-sensitive detection. SRS microscopy has a major advantage over previous coherent Raman techniques in that it offers background-free and easily interpretable chemical contrast. We show a variety of biomedical applications, such as differentiating distributions of omega-3 fatty acids and saturated lipids in living cells, imaging of brain and skin tissues based on intrinsic lipid contrast.

  19. Advances in light microscopy for neuroscience.

    PubMed

    Wilt, Brian A; Burns, Laurie D; Wei Ho, Eric Tatt; Ghosh, Kunal K; Mukamel, Eran A; Schnitzer, Mark J

    2009-01-01

    Since the work of Golgi and Cajal, light microscopy has remained a key tool for neuroscientists to observe cellular properties. Ongoing advances have enabled new experimental capabilities using light to inspect the nervous system across multiple spatial scales, including ultrastructural scales finer than the optical diffraction limit. Other progress permits functional imaging at faster speeds, at greater depths in brain tissue, and over larger tissue volumes than previously possible. Portable, miniaturized fluorescence microscopes now allow brain imaging in freely behaving mice. Complementary progress on animal preparations has enabled imaging in head-restrained behaving animals, as well as time-lapse microscopy studies in the brains of live subjects. Mouse genetic approaches permit mosaic and inducible fluorescence-labeling strategies, whereas intrinsic contrast mechanisms allow in vivo imaging of animals and humans without use of exogenous markers. This review surveys such advances and highlights emerging capabilities of particular interest to neuroscientists.

  20. Single molecule fluorescence and force microscopy.

    PubMed

    Schütz, G J; Hinterdorfer, P

    2002-12-01

    The investigation of biomolecules has entered a new age since the development of methodologies capable of studies at the level of single molecules. In biology, most molecules show a complex dynamical behavior, with individual motions and transitions between different states occurring highly correlated in space and time within an arrangement of various elements. Recent advances in the development of new microscopy techniques with sensitivity at the single molecule have gained access to essentially new types of information obtainable from imaging biomolecular samples. These methodologies are described here in terms of their applicability to the in vivo detection and visualization of molecular processes on surfaces, membranes, and cells. First examples of single molecule microscopy on cell membranes revealed new basic insight into the lateral organization of the plasma membrane, providing the captivating perspective of an ultra-sensitive methodology as a general tool to study local processes and heterogeneities in living cells.

  1. Scanning electron microscopy of lichen sclerosus*

    PubMed Central

    de Almeida, Hiram Larangeira; Bicca, Eduardo de Barros Coelho; Breunig, Juliano de Avelar; Rocha, Nara Moreira; Silva, Ricardo Marques e

    2013-01-01

    Lichen sclerosus is an acquired inflammatory condition characterized by whitish fibrotic plaques, with a predilection for the genital skin. We performed scanning electron microscopy of the dermis from a lesion of lichen sclerosus. Normal collagen fibers could be easily found in deeper layers of the specimen, as well as the transition to pathologic area, which seems homogenized. With higher magnifications in this transitional area collagen fibers are adherent to each other, and with very high magnifications a pearl chain aspect became evident along the collagen fibers. In the superficial dermis this homogenization is even more evident, collagen fibers are packed together and round structures are also observed. Rupture of collagen fibers and inflammatory cells were not found. These autoimmune changes of the extracellular matrix lead to the aggregation of immune complexes and/or changed matrix proteins along the collagen fibers, the reason why they seem hyalinized when examined by light microscopy. PMID:23739707

  2. Understanding liver immunology using intravital microscopy.

    PubMed

    Marques, Pedro Elias; Oliveira, André Gustavo; Chang, Lynne; Paula-Neto, Heitor Affonso; Menezes, Gustavo Batista

    2015-09-01

    The liver has come a long way since it was considered only a metabolic organ attached to the gastrointestinal tract. The simultaneous ascension of immunology and intravital microscopy evidenced the liver as a central axis in the immune system, controlling immune responses to local and systemic agents as well as disease tolerance. The multiple hepatic cell populations are organized in a vascular environment that promotes intimate cellular interactions, including initiation of innate and adaptive immune responses, rapid leukocyte recruitment, pathogen clearance and production of a variety of immune mediators. In this review, we focus on the advances in liver immunology supported by intravital microscopy in diseases such as isquemia/reperfusion, acute liver injury and infections.

  3. Fast interferometric second harmonic generation microscopy

    PubMed Central

    Bancelin, Stéphane; Couture, Charles-André; Légaré, Katherine; Pinsard, Maxime; Rivard, Maxime; Brown, Cameron; Légaré, François

    2016-01-01

    We report the implementation of fast Interferometric Second Harmonic Generation (I-SHG) microscopy to study the polarity of non-centrosymmetric structures in biological tissues. Using a sample quartz plate, we calibrate the spatially varying phase shift introduced by the laser scanning system. Compensating this phase shift allows us to retrieve the correct phase distribution in periodically poled lithium niobate, used as a model sample. Finally, we used fast interferometric second harmonic generation microscopy to acquire phase images in tendon. Our results show that the method exposed here, using a laser scanning system, allows to recover the polarity of collagen fibrils, similarly to standard I-SHG (using a sample scanning system), but with an imaging time about 40 times shorter. PMID:26977349

  4. Circumventing photodamage in live-cell microscopy

    PubMed Central

    Magidson, Valentin; Khodjakov, Alexey

    2013-01-01

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

  5. Frontiers of in situ electron microscopy

    DOE PAGES

    Zheng, Haimei; Zhu, Yimei; Meng, Shirley Ying

    2015-01-01

    In situ transmission electron microscopy (TEM) has become an increasingly important tool for materials characterization. It provides key information on the structural dynamics of a material during transformations and the correlation between structure and properties of materials. With the recent advances in instrumentation, including aberration corrected optics, sample environment control, the sample stage, and fast and sensitive data acquisition, in situ TEM characterization has become more and more powerful. In this article, a brief review of the current status and future opportunities of in situ TEM is included. It also provides an introduction to the six articles covered by inmore » this issue of MRS Bulletin explore the frontiers of in situ electron microscopy, including liquid and gas environmental TEM, dynamic four-dimensional TEM, nanomechanics, ferroelectric domain switching studied by in situ TEM, and state-of-the-art atomic imaging of light elements (i.e., carbon atoms) and individual defects.« less

  6. Global standardization of scanning probe microscopy

    NASA Astrophysics Data System (ADS)

    Fujita, Daisuke; Itoh, Hiroshi; Ichimura, Shingo; Kurosawa, Tomizo

    2007-02-01

    Recent efforts to achieve global standardization of scanning probe microscopy (SPM) including noncontact atomic force microscopy (NC-AFM), especially through the International Organization for Standardization (ISO) and related research, are surveyed. Since the unification of terminology for SPM is a prerequisite for standardization, it should have the first priority, followed by the unification of data management and treatment, which will enable access to and processing of SPM data collected by different types of instrument. Among the various SPM analytical methods, the dimensional metrology of SPM is regarded to be the first priority for standardization. This requires solving two basic problems: calibrating the x, y, and z coordinate axes with traceability to the SI unit of length, and eliminating the morphological artefacts caused by the shape of the probe tip. Pre-standardization efforts on restoring distorted images and characterizing the tip shape during use are discussed.

  7. Reflectance Confocal Microscopy in Lentigo Maligna.

    PubMed

    Gamo, R; Pampín, A; Floristán, U

    2016-12-01

    Lentigo maligna is the most common type of facial melanoma. Diagnosis is complicated, however, as it shares clinical and dermoscopic characteristics with other cutaneous lesions of the face. Reflectance confocal microscopy is an imaging technique that permits the visualization of characteristic features of lentigo maligna. These include a disrupted honeycomb pattern and pagetoid cells with a tendency to show folliculotropism. These cells typically have a dendritic morphology, although they may also appear as round cells measuring over 20μm with atypical nuclei. Poorly defined dermal papillae and atypical cells may be seen at the dermal-epidermal junction and can form bridges resembling mitochondrial structures. Other characteristic findings include junctional swelling with atypical cells located around the follicles, resembling caput medusae. Reflectance confocal microscopy is a very useful tool for diagnosing lentigo maligna.

  8. Nuclear microscopy of sperm cell elemental structure

    NASA Astrophysics Data System (ADS)

    Bench, Graham S.; Balhorn, Rod; Friz, Alexander M.

    1995-05-01

    Theories suggest there is a link between protamine concentrations in individual sperm and male fertility. Previously, biochemical analyses have used pooled samples containing millions of sperm to determine protamine concentrations. These methods have not been able to determine what percentage of morphologically normal sperm are biochemically defective and potentially infertile. Nuclear microscopy has been utilized to measure elemental profiles at the single sperm level. By measuring the amount of phosphorus and sulfur, the total DNA and protamine content in individual sperm from fertile bull and mouse semen have been determined. These values agree with results obtained from other biochemical analyses. Nuclear microscopy shows promise for measuring elemental profiles in the chromatin of individual sperm. The technique may be able to resolve theories regarding the importance of protamines to male fertility and identify biochemical defects responsible for certain types of male infertility.

  9. Diffractive elements performance in chromatic confocal microscopy

    NASA Astrophysics Data System (ADS)

    Garzón, J.; Duque, D.; Alean, A.; Toledo, M.; Meneses, J.; Gharbi, T.

    2011-01-01

    The Confocal Laser Scanning Microscopy (CLSM) has been widely used in the semiconductor industry and biomedicine because of its depth discrimination capability. Subsequent to this technique has been developed in recent years Chromatic Confocal Microscopy. This method retains the same principle of confocal and offers the added advantage of removing the axial movement of the moving system. This advantage is usually accomplished with an optical element that generates a longitudinal chromatic aberration and a coding system that relates the axial position of each point of the sample with the wavelength that is focused on each. The present paper shows the performance of compact chromatic confocal microscope when some different diffractive elements are used for generation of longitudinal chromatic aberration. Diffractive elements, according to the process and manufacturing parameters, may have different diffraction efficiency and focus a specific wavelength in a specific focal position. The performance assessment is carried out with various light sources which exhibit an incoherent behaviour and a broad spectral width.

  10. Nuclear microscopy of sperm cell elemental structure

    SciTech Connect

    Bench, G.S.; Balhorn, R.; Friz, A.M.; Freeman, S.P.H.T.

    1994-09-28

    Theories suggest there is a link between protamine concentrations in individual sperm and male fertility. Previously, biochemical analyses have used pooled samples containing millions of sperm to determine protamine concentrations. These methods have not been able to determine what percentage of morphologically normal sperm are biochemically defective and potentially infertile. Nuclear microscopy has been utilized to measure elemental profiles at the single sperm level. By measuring the amount of phosphorus and sulfur, the total DNA and protamine content in individual sperm from fertile bull and mouse semen have been determined. These values agree with results obtained from other biochemical analyses. Nuclear microscopy shows promise for measuring elemental profiles in the chromatin of individual sperm. The technique may be able to resolve theories regarding the importance of protamines to male fertility and identify biochemical defects responsible for certain types of male infertility.

  11. Atomically resolved force microscopy at room temperature

    SciTech Connect

    Morita, Seizo

    2014-04-24

    Atomic force microscopy (AFM) can now not only image individual atoms but also construct atom letters using atom manipulation method even at room temperature (RT). Therefore, the AFM is the second generation atomic tool following the scanning tunneling microscopy (STM). However the AFM can image even insulating atoms, and also directly measure/map the atomic force and potential at the atomic scale. Noting these advantages, we have been developing a bottom-up nanostructuring system at RT based on the AFM. It can identify chemical species of individual atoms and then manipulate selected atom species to the predesigned site one-by-one to assemble complex nanostructures consisted of multi atom species at RT. Here we introduce our results toward atom-by-atom assembly of composite nanostructures based on the AFM at RT including the latest result on atom gating of nano-space for atom-by-atom creation of atom clusters at RT for semiconductor surfaces.

  12. Resonance response of scanning force microscopy cantilevers

    SciTech Connect

    Chen, G.Y.; Warmack, R.J.; Thundat, T.; Allison, D.P. ); Huang, A. )

    1994-08-01

    A variational method is used to calculate the deflection and the fundamental and harmonic resonance frequencies of commercial V-shaped and rectangular atomic force microscopy cantilevers. The effective mass of V-shaped cantilevers is roughly half that calculated for the equivalent rectangular cantilevers. Damping by environmental gases, including air, nitrogen, argon, and helium, affects the frequency of maximum response and to a much greater degree the quality factor [ital Q]. Helium has the lowest viscosity, resulting in the highest [ital Q], and thus provides the best sensitivity in noncontact force microscopy. Damping in liquids is dominated by an increase in effective mass of the cantilever due to an added mass of the liquid being dragged with that cantilever.

  13. Interferometric phase microscopy of red blood cells

    NASA Astrophysics Data System (ADS)

    Xue, Liang; Sun, Nan; Tang, Xian; Wang, Yin; Wang, Shouyu

    2013-12-01

    Quantitative phase imaging of cells with high accuracy in a completely noninvasive manner is a challenging task. To provide a proper solution to this important need, interferometric phase microscopy is described which relies on the off-axis interferometry, confocal microscopy and high-speed image capture technology. Phase retrieval from the single interferogram is done by algorithms based on the fast Fourier transform, traditional Hilbert transform and two-step Hilbert transform, respectively. Furthermore, a phase aberrations compensation approach is applied to correct the phase distribution of the red blood cells obtained via the three methods mentioned before without the pre-known knowledge for removing the wave front curvature introduced by the microscope objectives, off-axis imaging, etc., which otherwise hinders the phase reconstruction. The improved results reveal the better inner structures of the red blood cells. The development of quantitative phase imaging technique is shedding light on their future directions and applications for basic and clinical research.

  14. Confocal multiview light-sheet microscopy

    PubMed Central

    Medeiros, Gustavo de; Norlin, Nils; Gunther, Stefan; Albert, Marvin; Panavaite, Laura; Fiuza, Ulla-Maj; Peri, Francesca; Hiiragi, Takashi; Krzic, Uros; Hufnagel, Lars

    2015-01-01

    Selective-plane illumination microscopy has proven to be a powerful imaging technique due to its unsurpassed acquisition speed and gentle optical sectioning. However, even in the case of multiview imaging techniques that illuminate and image the sample from multiple directions, light scattering inside tissues often severely impairs image contrast. Here we combine multiview light-sheet imaging with electronic confocal slit detection implemented on modern camera sensors. In addition to improved imaging quality, the electronic confocal slit detection doubles the acquisition speed in multiview setups with two opposing illumination directions allowing simultaneous dual-sided illumination. Confocal multiview light-sheet microscopy eliminates the need for specimen-specific data fusion algorithms, streamlines image post-processing, easing data handling and storage. PMID:26602977

  15. Speckle-field digital holographic microscopy

    PubMed Central

    Park, YongKeun; Choi, Wonshik; Yaqoob, Zahid; Dasari, Ramachandra; Badizadegan, Kamran; Feld, Michael S.

    2010-01-01

    The use of coherent light in conventional holographic phase microscopy (HPM) poses three major drawbacks: poor spatial resolution, weak depth sectioning, and fixed pattern noise due to unwanted diffraction. Here, we report a technique which can overcome these drawbacks, but maintains the advantage of phase microscopy - high contrast live cell imaging and 3D imaging. A speckle beam of a complex spatial pattern is used for illumination to reduce fixed pattern noise and to improve optical sectioning capability. By recording of the electric field of speckle, we demonstrate high contrast 3D live cell imaging without the need for axial scanning - neither objective lens nor sample stage. This technique has great potential in studying biological samples with improved sensitivity, resolution and optical sectioning capability. PMID:19654630

  16. Three-Dimensional Reflectance Traction Microscopy

    PubMed Central

    Jones, Christopher A. R.; Groves, Nicholas Scott; Sun, Bo

    2016-01-01

    Cells in three-dimensional (3D) environments exhibit very different biochemical and biophysical phenotypes compared to the behavior of cells in two-dimensional (2D) environments. As an important biomechanical measurement, 2D traction force microscopy can not be directly extended into 3D cases. In order to quantitatively characterize the contraction field, we have developed 3D reflectance traction microscopy which combines confocal reflection imaging and partial volume correlation postprocessing. We have measured the deformation field of collagen gel under controlled mechanical stress. We have also characterized the deformation field generated by invasive breast cancer cells of different morphologies in 3D collagen matrix. In contrast to employ dispersed tracing particles or fluorescently-tagged matrix proteins, our methods provide a label-free, computationally effective strategy to study the cell mechanics in native 3D extracellular matrix. PMID:27304456

  17. Reconstruction in interferometric synthetic aperture microscopy: comparison with optical coherence tomography and digital holographic microscopy.

    PubMed

    Sheppard, Colin J R; Kou, Shan Shan; Depeursinge, Christian

    2012-03-01

    It is shown that the spatial frequencies recorded in interferometric synthetic aperture microscopy do not correspond to exact backscattering [as they do in unistatic synthetic aperture radar (SAR)] and that the reconstruction process based on SAR is therefore based on an approximation. The spatial frequency response is developed based on the three-dimensional coherent transfer function approach and compared with that in optical coherence tomography and digital holographic microscopy.

  18. Application perspectives of localization microscopy in virology.

    PubMed

    Cremer, C; Kaufmann, R; Gunkel, M; Polanski, F; Müller, P; Dierkes, R; Degenhard, S; Wege, C; Hausmann, M; Birk, U

    2014-07-01

    Localization microscopy approaches allowing an optical resolution down to the single-molecule level in fluorescence-labeled biostructures have already found a variety of applications in cell biology, as well as in virology. Here, we focus on some perspectives of a special localization microscopy embodiment, spectral precision distance/position determination microscopy (SPDM). SPDM permits the use of conventional fluorophores or fluorescent proteins together with standard sample preparation conditions employing an aqueous buffered milieu and typically monochromatic excitation. This allowed superresolution imaging and studies on the aggregation state of modified tobacco mosaic virus particles on the nanoscale with a single-molecule localization accuracy of better than 8 nm, using standard fluorescent dyes in the visible spectrum. To gain a better understanding of cell entry mechanisms during influenza A virus infection, SPDM was used in conjunction with algorithms for distance and cluster analyses to study changes in the distribution of virus particles themselves or in the distribution of infection-related proteins, the hepatocyte growth factor receptors, in the cell membrane on the single-molecule level. Not requiring TIRF (total internal reflection) illumination, SPDM was also applied to study the molecular arrangement of gp36.5/m164 glycoprotein (essentially associated with murine cytomegalovirus infection) in the endoplasmic reticulum and the nuclear membrane inside cells with single-molecule resolution. On the basis of the experimental evidence so far obtained, we finally discuss additional application perspectives of localization microscopy approaches for the fast detection and identification of viruses by multi-color SPDM and combinatorial oligonucleotide fluorescence in situ hybridization, as well as SPDM techniques for optimization of virus-based nanotools and biodetection devices.

  19. Surface Studies by Scanning Probe Microscopy

    NASA Astrophysics Data System (ADS)

    Kim, Ho-Seob

    The scanning probe microscopy reported here includes scanning tunneling microscopy (STM), scanning tunneling spectroscopy (STS), and atomic force microscopy (AFM). The scanning tunneling microscope is a novel tool which can reveal the atomic structure and electronic properties of surfaces using a probe with a sharp tip. An additional technique, atomic force microscopy has the potential to record geometric structures for both conducting and non -conducting materials. The first AFM designs utilized short range forces between a small stylus and a sample surface to produce high resolution images of defects and structural features of the surface. The current-voltage characteristics were also investigated during dynamic changes of the tunnel current and barrier height with an additional technology, tunneling spectroscopy. An advanced design for an AFM has been developed which utilizes a dielectric tunnel junction to retain the high sensitivity of tunnel current control over force ranges between 10^{-6} and 10 ^{-11}N. This AFM has been successfully applied to physical and biological samples. Scanning probe techniques have been developed and applied to a range of sample types including conductors, semi-conductors and non-conductors. Each technique utilizes the same electronics, computers, and imaging facilities. A fundamental problem of the atomic structure of graphite has existed since the inception of STM images. The experimental and theoretical hypotheses have been considered and a resolution of the problem has been developed as reported in this dissertation. Unprecedented resolving power, greater than 1A, has confirmed our hypothesis and has been correctly correlated with the structure of graphite surface. This dissertation also presents the results from studies of the surface structure of: MoS_2 , Cu, Au, Ag, Si, CdTe, HgTe, Fe_2 O_3, mica, gypsum, purple membranes with protein chains, and an organic photoconducting material, by scanning probe microscopes.

  20. Correlative photoactivated localization and scanning electron microscopy.

    PubMed

    Kopek, Benjamin G; Shtengel, Gleb; Grimm, Jonathan B; Clayton, David A; Hess, Harald F

    2013-01-01

    The ability to localize proteins precisely within subcellular space is crucial to understanding the functioning of biological systems. Recently, we described a protocol that correlates a precise map of fluorescent fusion proteins localized using three-dimensional super-resolution optical microscopy with the fine ultrastructural context of three-dimensional electron micrographs. While it achieved the difficult simultaneous objectives of high photoactivated fluorophore preservation and ultrastructure preservation, it required a super-resolution optical and specialized electron microscope that is not available to many researchers. We present here a faster and more practical protocol with the advantage of a simpler two-dimensional optical (Photoactivated Localization Microscopy (PALM)) and scanning electron microscope (SEM) system that retains the often mutually exclusive attributes of fluorophore preservation and ultrastructure preservation. As before, cryosections were prepared using the Tokuyasu protocol, but the staining protocol was modified to be amenable for use in a standard SEM without the need for focused ion beam ablation. We show the versatility of this technique by labeling different cellular compartments and structures including mitochondrial nucleoids, peroxisomes, and the nuclear lamina. We also demonstrate simultaneous two-color PALM imaging with correlated electron micrographs. Lastly, this technique can be used with small-molecule dyes as demonstrated with actin labeling using phalloidin conjugated to a caged dye. By retaining the dense protein labeling expected for super-resolution microscopy combined with ultrastructural preservation, simplifying the tools required for correlative microscopy, and expanding the number of useful labels we expect this method to be accessible and valuable to a wide variety of researchers.

  1. Photoacoustic microscopy of ceramic turbine blades

    NASA Technical Reports Server (NTRS)

    Khandelwal, P. K.; Kinnick, R. R.; Heitman, P. W.

    1985-01-01

    Scanning photoacoustic microscopy (SPAM) is evaluated as a nondestructive technique for the detection of both surface and subsurface flaws in polycrystalline ceramics, such as those currently under consideration for the high temperature components of small vehicular and industrial gas turbine engines; the fracture strength of these brittle materials is controlled by small, 25-200 micron flaws. Attention is given to the correlation of SPAM-detected flaws with actual, fracture-controlling flaws in ceramic turbine blades.

  2. Orbital angular momentum light in microscopy

    NASA Astrophysics Data System (ADS)

    Ritsch-Marte, Monika

    2017-02-01

    Light with a helical phase has had an impact on optical imaging, pushing the limits of resolution or sensitivity. Here, special emphasis will be given to classical light microscopy of phase samples and to Fourier filtering techniques with a helical phase profile, such as the spiral phase contrast technique in its many variants and areas of application. This article is part of the themed issue 'Optical orbital angular momentum'.

  3. Dark Field Microscopy for Analytical Laboratory Courses

    SciTech Connect

    Augspurger, Ashley E; Stender, Anthony S; Marchuk, Kyle; Greenbowe, Thomas J; Fang, Ning

    2014-06-10

    An innovative and inexpensive optical microscopy experiment for a quantitative analysis or an instrumental analysis chemistry course is described. The students have hands-on experience with a dark field microscope and investigate the wavelength dependence of localized surface plasmon resonance in gold and silver nanoparticles. Students also observe and measure individual crystal growth during a replacement reaction between copper and silver nitrate. The experiment allows for quantitative, qualitative, and image data analyses for undergraduate students.

  4. Reflection Acoustic Microscopy for Micro-NDE.

    DTIC Science & Technology

    1983-02-01

    WORDS (Coni, wu rere side. 14 It noeeeey And Idenify1 by block esife) Nondestructive Evaluation Acoustic Microscopy I Subsurface Imaging Pulsecio Cmrsin... subsurface imaging is presented and it is shown that with such lenses it is possible to obtain good focussing performance over a wide depth range...typically few millimeters at 50 MHz. A major problem in subsurface imaging derives from the large reflection obtained frnm the surface, and the small amount

  5. Biological cryo-electron microscopy in China.

    PubMed

    Wang, Hong-Wei; Lei, Jianlin; Shi, Yigong

    2017-01-01

    Cryo-electron microscopy (cryo-EM) plays an increasingly more important role in structural biology. With the construction of an arm of the Chinese National Protein Science Facility at Tsinghua University, biological cryo-EM has entered a phase of rapid development in China. This article briefly reviews the history of biological cryo-EM in China, describes its current status, comments on its impact on the various biological research fields, and presents future outlook.

  6. Biological cryo‐electron microscopy in China

    PubMed Central

    2016-01-01

    Abstract Cryo‐electron microscopy (cryo‐EM) plays an increasingly more important role in structural biology. With the construction of an arm of the Chinese National Protein Science Facility at Tsinghua University, biological cryo‐EM has entered a phase of rapid development in China. This article briefly reviews the history of biological cryo‐EM in China, describes its current status, comments on its impact on the various biological research fields, and presents future outlook. PMID:27534377

  7. A New Twist on Scanning Thermal Microscopy

    DTIC Science & Technology

    2012-01-25

    Materials and Manufacturing Directorate, Air Force Research Laboratory, WPAFB, Ohio 45433, United States ‡School of Materials Science and Engineering...bimorphs do not suffer from these setbacks. In fact, thermal bimorphs transduced with an atomic force microscopy (AFM) quadrant photodetector have a...AUTHOR(S) 5d. PROJECT NUMBER 5e. TASK NUMBER 5f. WORK UNIT NUMBER 7. PERFORMING ORGANIZATION NAME(S) AND ADDRESS(ES) Air Force Research Laboratory

  8. Quantitative imaging of bilirubin by photoacoustic microscopy

    NASA Astrophysics Data System (ADS)

    Zhou, Yong; Zhang, Chi; Yao, Da-Kang; Wang, Lihong V.

    2013-03-01

    Noninvasive detection of both bilirubin concentration and its distribution is important for disease diagnosis. Here we implemented photoacoustic microscopy (PAM) to detect bilirubin distribution. We first demonstrate that our PAM system can measure the absorption spectra of bilirubin and blood. We also image bilirubin distributions in tissuemimicking samples, both without and with blood mixed. Our results show that PAM has the potential to quantitatively image bilirubin in vivo for clinical applications.

  9. Image Correlation Microscopy for Uniform Illumination

    PubMed Central

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

    2011-01-01

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

  10. Observation of Superlubricity by Scanning Tunneling Microscopy

    NASA Astrophysics Data System (ADS)

    Hirano, Motohisa; Shinjo, Kazumasa; Kaneko, Reizo; Murata, Yoshitada

    1997-02-01

    Experimental evidence of superlubricity, the state of vanishing friction, is obtained by examining systems of sliding atomically clean surfaces by using ultrahigh vacuum scanning tunneling microscopy. The experimental results agree with theoretical predictions: Friction is not observed in the superlubricity regime in measurements capable of resolving a friction force of 3×10-9 N, whereas friction of 8×10-8 N, which is comparable to theoretical values, is observed in the friction regime.

  11. Multidepth imaging by chromatic dispersion confocal microscopy

    NASA Astrophysics Data System (ADS)

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

    2012-03-01

    Confocal microscopy has shown potential as an imaging technique to detect precancer. Imaging cellular features throughout the depth of epithelial tissue may provide useful information for diagnosis. However, the current in vivo axial scanning techniques for confocal microscopy are cumbersome, time-consuming, and restrictive when attempting to reconstruct volumetric images acquired in breathing patients. Chromatic dispersion confocal microscopy (CDCM) exploits severe longitudinal chromatic aberration in the system to axially disperse light from a broadband source and, ultimately, spectrally encode high resolution images along the depth of the object. Hyperchromat lenses are designed to have severe and linear longitudinal chromatic aberration, but have not yet been used in confocal microscopy. We use a hyperchromat lens in a stage scanning confocal microscope to demonstrate the capability to simultaneously capture information at multiple depths without mechanical scanning. A photonic crystal fiber pumped with a 830nm wavelength Ti:Sapphire laser was used as a supercontinuum source, and a spectrometer was used as the detector. The chromatic aberration and magnification in the system give a focal shift of 140μm after the objective lens and an axial resolution of 5.2-7.6μm over the wavelength range from 585nm to 830nm. A 400x400x140μm3 volume of pig cheek epithelium was imaged in a single X-Y scan. Nuclei can be seen at several depths within the epithelium. The capability of this technique to achieve simultaneous high resolution confocal imaging at multiple depths may reduce imaging time and motion artifacts and enable volumetric reconstruction of in vivo confocal images of the epithelium.

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

    PubMed

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

    2015-12-01

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

  13. Efficient illumination for microsecond tracking microscopy.

    PubMed

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

    2014-01-01

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

  14. Invited Review Article: Pump-probe microscopy.

    PubMed

    Fischer, Martin C; Wilson, Jesse W; Robles, Francisco E; Warren, Warren S

    2016-03-01

    Multiphoton microscopy has rapidly gained popularity in biomedical imaging and materials science because of its ability to provide three-dimensional images at high spatial and temporal resolution even in optically scattering environments. Currently the majority of commercial and home-built devices are based on two-photon fluorescence and harmonic generation contrast. These two contrast mechanisms are relatively easy to measure but can access only a limited range of endogenous targets. Recent developments in fast laser pulse generation, pulse shaping, and detection technology have made accessible a wide range of optical contrasts that utilize multiple pulses of different colors. Molecular excitation with multiple pulses offers a large number of adjustable parameters. For example, in two-pulse pump-probe microscopy, one can vary the wavelength of each excitation pulse, the detection wavelength, the timing between the excitation pulses, and the detection gating window after excitation. Such a large parameter space can provide much greater molecular specificity than existing single-color techniques and allow for structural and functional imaging without the need for exogenous dyes and labels, which might interfere with the system under study. In this review, we provide a tutorial overview, covering principles of pump-probe microscopy and experimental setup, challenges associated with signal detection and data processing, and an overview of applications.

  15. Invited Review Article: Pump-probe microscopy

    PubMed Central

    Wilson, Jesse W.; Robles, Francisco E.; Warren, Warren S.

    2016-01-01

    Multiphoton microscopy has rapidly gained popularity in biomedical imaging and materials science because of its ability to provide three-dimensional images at high spatial and temporal resolution even in optically scattering environments. Currently the majority of commercial and home-built devices are based on two-photon fluorescence and harmonic generation contrast. These two contrast mechanisms are relatively easy to measure but can access only a limited range of endogenous targets. Recent developments in fast laser pulse generation, pulse shaping, and detection technology have made accessible a wide range of optical contrasts that utilize multiple pulses of different colors. Molecular excitation with multiple pulses offers a large number of adjustable parameters. For example, in two-pulse pump-probe microscopy, one can vary the wavelength of each excitation pulse, the detection wavelength, the timing between the excitation pulses, and the detection gating window after excitation. Such a large parameter space can provide much greater molecular specificity than existing single-color techniques and allow for structural and functional imaging without the need for exogenous dyes and labels, which might interfere with the system under study. In this review, we provide a tutorial overview, covering principles of pump-probe microscopy and experimental setup, challenges associated with signal detection and data processing, and an overview of applications. PMID:27036751

  16. Non-radiative excitation fluorescence microscopy

    NASA Astrophysics Data System (ADS)

    Riachy, Lina; Vézy, Cyrille; Jaffiol, Rodolphe

    2016-03-01

    Non-radiative Excitation Fluorescence Microscopy (NEFM) constitutes a new way to observe biological samples beyond the diffraction limit. Non-radiative excitation of the samples is achieved by coating the substrate with donor species, such as quantum dots (QDs). Thus the dyes are not excited directly by the laser source, as in common fluorescence microscopy, but through a non-radiative energy transfer. To prevent dewetting of the donor film, we have recently implemented a silanization process to covalently bond the QDs on the substrate. An homogeneous monolayer of QDs was then deposited on only one side of the coverslips. Atomic force microscopy was then used to characterize the QD layer. We highlight the potential of our method through the study of Giant Unilamellar Vesicles (GUVs) labeled with DiD as acceptor, in interaction with surface functionalized with poly-L-lysine. In the presence of GUVs, we observed a quenching of QDs emission, together with an emission of DiD located in the membrane, which clearly indicated that non-radiative energy transfer from QDs to DiD occurs.

  17. Mirror-enhanced super-resolution microscopy.

    PubMed

    Yang, Xusan; Xie, Hao; Alonas, Eric; Liu, Yujia; Chen, Xuanze; Santangelo, Philip J; Ren, Qiushi; Xi, Peng; Jin, Dayong

    Axial excitation confinement beyond the diffraction limit is crucial to the development of next-generation, super-resolution microscopy. STimulated Emission Depletion (STED) nanoscopy offers lateral super-resolution using a donut-beam depletion, but its axial resolution is still over 500 nm. Total internal reflection fluorescence microscopy is widely used for single-molecule localization, but its ability to detect molecules is limited to within the evanescent field of ~ 100 nm from the cell attachment surface. We find here that the axial thickness of the point spread function (PSF) during confocal excitation can be easily improved to 110 nm by replacing the microscopy slide with a mirror. The interference of the local electromagnetic field confined the confocal PSF to a 110-nm spot axially, which enables axial super-resolution with all laser-scanning microscopes. Axial sectioning can be obtained with wavelength modulation or by controlling the spacer between the mirror and the specimen. With no additional complexity, the mirror-assisted excitation confinement enhanced the axial resolution six-fold and the lateral resolution two-fold for STED, which together achieved 19-nm resolution to resolve the inner rim of a nuclear pore complex and to discriminate the contents of 120 nm viral filaments. The ability to increase the lateral resolution and decrease the thickness of an axial section using mirror-enhanced STED without increasing the laser power is of great importance for imaging biological specimens, which cannot tolerate high laser power.

  18. Image Restoration in Cryo-electron Microscopy

    PubMed Central

    Penczek, Pawel A.

    2011-01-01

    Image restoration techniques are used to obtain, given experimental measurements, the best possible approximation of the original object within the limits imposed by instrumental conditions and noise level in the data. In molecular electron microscopy, we are mainly interested in linear methods that preserve the respective relationships between mass densities within the restored map. Here, we describe the methodology of image restoration in structural electron microscopy, and more specifically, we will focus on the problem of the optimum recovery of Fourier amplitudes given electron microscope data collected under various defocus settings. We discuss in detail two classes of commonly used linear methods, the first of which consists of methods based on pseudoinverse restoration, and which is further subdivided into mean-square error, chi-square error, and constrained based restorations, where the methods in the latter two subclasses explicitly incorporates non-white distribution of noise in the data. The second class of methods is based on the Wiener filtration approach. We show that the Wiener filter-based methodology can be used to obtain a solution to the problem of amplitude correction (or “sharpening”) of the electron microscopy map that makes it visually comparable to maps determined by X-ray crystallography, and thus amenable to comparable interpretation. Finally, we present a semi-heuristic Wiener filter-based solution to the problem of image restoration given sets of heterogeneous solutions. We conclude the chapter with a discussion of image restoration protocols implemented in commonly used single particle software packages. PMID:20888957

  19. Mirror-enhanced super-resolution microscopy

    PubMed Central

    Yang, Xusan; Xie, Hao; Alonas, Eric; Liu, Yujia; Chen, Xuanze; Santangelo, Philip J; Ren, Qiushi; Xi, Peng; Jin, Dayong

    2016-01-01

    Axial excitation confinement beyond the diffraction limit is crucial to the development of next-generation, super-resolution microscopy. STimulated Emission Depletion (STED) nanoscopy offers lateral super-resolution using a donut-beam depletion, but its axial resolution is still over 500 nm. Total internal reflection fluorescence microscopy is widely used for single-molecule localization, but its ability to detect molecules is limited to within the evanescent field of ~ 100 nm from the cell attachment surface. We find here that the axial thickness of the point spread function (PSF) during confocal excitation can be easily improved to 110 nm by replacing the microscopy slide with a mirror. The interference of the local electromagnetic field confined the confocal PSF to a 110-nm spot axially, which enables axial super-resolution with all laser-scanning microscopes. Axial sectioning can be obtained with wavelength modulation or by controlling the spacer between the mirror and the specimen. With no additional complexity, the mirror-assisted excitation confinement enhanced the axial resolution six-fold and the lateral resolution two-fold for STED, which together achieved 19-nm resolution to resolve the inner rim of a nuclear pore complex and to discriminate the contents of 120 nm viral filaments. The ability to increase the lateral resolution and decrease the thickness of an axial section using mirror-enhanced STED without increasing the laser power is of great importance for imaging biological specimens, which cannot tolerate high laser power. PMID:27398242

  20. Quantitative Aspects of Single Molecule Microscopy

    PubMed Central

    Ober, Raimund J.; Tahmasbi, Amir; Ram, Sripad; Lin, Zhiping; Ward, E. Sally

    2015-01-01

    Single molecule microscopy is a relatively new optical microscopy technique that allows the detection of individual molecules such as proteins in a cellular context. This technique has generated significant interest among biologists, biophysicists and biochemists, as it holds the promise to provide novel insights into subcellular processes and structures that otherwise cannot be gained through traditional experimental approaches. Single molecule experiments place stringent demands on experimental and algorithmic tools due to the low signal levels and the presence of significant extraneous noise sources. Consequently, this has necessitated the use of advanced statistical signal and image processing techniques for the design and analysis of single molecule experiments. In this tutorial paper, we provide an overview of single molecule microscopy from early works to current applications and challenges. Specific emphasis will be on the quantitative aspects of this imaging modality, in particular single molecule localization and resolvability, which will be discussed from an information theoretic perspective. We review the stochastic framework for image formation, different types of estimation techniques and expressions for the Fisher information matrix. We also discuss several open problems in the field that demand highly non-trivial signal processing algorithms. PMID:26167102

  1. Single cell elemental analysis using nuclear microscopy

    NASA Astrophysics Data System (ADS)

    Ren, M. Q.; Thong, P. S. P.; Kara, U.; Watt, F.

    1999-04-01

    The use of Particle Induced X-ray Emission (PIXE), Rutherford Backscattering Spectrometry (RBS) and Scanning Transmission Ion Microscopy (STIM) to provide quantitative elemental analysis of single cells is an area which has high potential, particularly when the trace elements such as Ca, Fe, Zn and Cu can be monitored. We describe the methodology of sample preparation for two cell types, the procedures of cell imaging using STIM, and the quantitative elemental analysis of single cells using RBS and PIXE. Recent work on single cells at the Nuclear Microscopy Research Centre,National University of Singapore has centred around two research areas: (a) Apoptosis (programmed cell death), which has been recently implicated in a wide range of pathological conditions such as cancer, Parkinson's disease etc, and (b) Malaria (infection of red blood cells by the malaria parasite). Firstly we present results on the elemental analysis of human Chang liver cells (ATTCC CCL 13) where vanadium ions were used to trigger apoptosis, and demonstrate that nuclear microscopy has the capability of monitoring vanadium loading within individual cells. Secondly we present the results of elemental changes taking place in individual mouse red blood cells which have been infected with the malaria parasite and treated with the anti-malaria drug Qinghaosu (QHS).

  2. Intermittent contact hydration scanning probe microscopy.

    PubMed

    Aloisi, G; Bacci, F; Carlà, M; Dolci, D

    2010-07-01

    Hydration scanning probe microscopy is a technique similar to scanning tunneling microscopy, in which the probe current, sustained by the slight surface conduction of a thin hydration layer covering an insulating support surface, is essentially electrochemical in nature instead of electronic tunneling. Such a technique allows the imaging of a great variety of samples, including insulators, provided that they are hydrophilic, as well as the study of molecular samples of biological interest (such as DNA) fixed on a suitable supporting surface. The main problem to obtain stable and reproducible images comes from the very critical determination of the operating conditions under which the probe-hydration layer interaction does not lead to the formation of a relatively large water meniscus. It has been suggested that this issue can be removed by adding a high frequency oscillation to the probe movement, as in tapping atomic force microscopy. Meniscus formation and breakup have been investigated in order to determine the best values for the amplitude and the frequency of the oscillation. Results obtained in this mode are discussed in comparison with the usual continuous contact mode.

  3. Invited Review Article: Pump-probe microscopy

    NASA Astrophysics Data System (ADS)

    Fischer, Martin C.; Wilson, Jesse W.; Robles, Francisco E.; Warren, Warren S.

    2016-03-01

    Multiphoton microscopy has rapidly gained popularity in biomedical imaging and materials science because of its ability to provide three-dimensional images at high spatial and temporal resolution even in optically scattering environments. Currently the majority of commercial and home-built devices are based on two-photon fluorescence and harmonic generation contrast. These two contrast mechanisms are relatively easy to measure but can access only a limited range of endogenous targets. Recent developments in fast laser pulse generation, pulse shaping, and detection technology have made accessible a wide range of optical contrasts that utilize multiple pulses of different colors. Molecular excitation with multiple pulses offers a large number of adjustable parameters. For example, in two-pulse pump-probe microscopy, one can vary the wavelength of each excitation pulse, the detection wavelength, the timing between the excitation pulses, and the detection gating window after excitation. Such a large parameter space can provide much greater molecular specificity than existing single-color techniques and allow for structural and functional imaging without the need for exogenous dyes and labels, which might interfere with the system under study. In this review, we provide a tutorial overview, covering principles of pump-probe microscopy and experimental setup, challenges associated with signal detection and data processing, and an overview of applications.

  4. Optical microscopy beyond the diffraction limit

    PubMed Central

    Smolyaninov, Igor I.

    2008-01-01

    Over the past century the resolution of far-field optical microscopes, which rely on propagating optical modes, was widely believed to be limited because of diffraction to a value on the order of a half-wavelength λ∕2 of the light used. Although immersion microscopes had slightly improved resolution on the order of λ∕2n, the increased resolution was limited by the small range of refractive indices, n, of available transparent materials. We are experiencing quick demolition of the diffraction limit in optical microscopy. Over the past few years numerous nonlinear optical microscopy techniques based on photoswitching and saturation of fluorescence demonstrated far-field resolution of 20 to 30 nm. The latest exciting example of these techniques has been demonstrated by Huang et al. [Science 319, 810–813 (2008)]. Moreover, recent progress in metamaterials indicates that artificial optical media can be created, which do not exhibit the diffraction limit. Resolution of linear “immersion” microscopes based on such metamaterials appears limited only by losses, which can be compensated by gain media. Thus, optical microscopy is quickly moving towards the 10 nm resolution scale, which should bring about numerous revolutionary advances in biomedical imaging. PMID:19404465

  5. Shaping field for deep tissue microscopy

    NASA Astrophysics Data System (ADS)

    Colon, J.; Lim, H.

    2015-05-01

    Information capacity of a lossless image-forming system is a conserved property determined by two imaging parameters - the resolution and the field of view (FOV). Adaptive optics improves the former by manipulating the phase, or wavefront, in the pupil plane. Here we describe a homologous approach, namely adaptive field microscopy, which aims to enhance the FOV by controlling the phase, or defocus, in the focal plane. In deep tissue imaging, the useful FOV can be severely limited if the region of interest is buried in a thick sample and not perpendicular to the optic axis. One must acquire many z-scans and reconstruct by post-processing, which exposes tissue to excessive radiation and is also time consuming. We demonstrate the effective FOV can be substantially enhanced by dynamic control of the image plane. Specifically, the tilt of the image plane is continuously adjusted in situ to match the oblique orientation of the sample plane within tissue. The utility of adaptive field microscopy is tested for imaging tissue with non-planar morphology. Ocular tissue of small animals was imaged by two-photon excited fluorescence. Our results show that adaptive field microscopy can utilize the full FOV. The freedom to adjust the image plane to account for the geometrical variations of sample could be extremely useful for 3D biological imaging. Furthermore, it could facilitate rapid surveillance of cellular features within deep tissue while avoiding photo damages, making it suitable for in vivo imaging.

  6. Electron microscopy of Crotalaria pulmonary hypertension

    PubMed Central

    Kay, J. M.; Smith, Paul; Heath, Donald

    1969-01-01

    The lungs of 11 rats fed on Crotalaria spectabilis seeds for periods ranging from 12 to 61 days were examined by both light and electron microscopy. The findings were compared with those obtained from nine control rats given a normal diet. Eight of the 11 test rats showed morphological evidence of pulmonary arterial hypertension in the form of right ventricular hypertrophy; the exceptions were rats killed after receiving the Crotalaria diet for 12, 22, and 29 days respectively. On light microscopy, all the test rats showed exudative lesions in the lungs consisting of eosinophilic alveolar coagulum, intra-alveolar haemorrhage, interstitial fibrosis, and a proliferation of mast cells. Enlarged and proliferated cells were seen to line the alveolar walls or lie free within the alveolar spaces. Electron microscopy showed these cells to be enlarged granular pneumocytes containing enlarged, electron-dense, lamellar secretory inclusions. Scanty macrophages were also seen in the alveolar spaces, in which excessive numbers of myelin figures and lattices were seen: these structures resembled phospholipid membranes and were probably related to pulmonary surfactant. We think that proliferation of granular pneumocytes is a non-specific reaction of the alveolar walls to injury. The alveolar-capillary wall showed interstitial oedema with the formation of intraluminal endothelial vesicles, probably representing the early ultrastructural phase of pulmonary oedema, and more likely to be an effect of the pulmonary hypertension than its cause. Images PMID:5348317

  7. Laser beam shaping for biomedical microscopy techniques

    NASA Astrophysics Data System (ADS)

    Laskin, Alexander; Kaiser, Peter; Laskin, Vadim; Ostrun, Aleksei

    2016-04-01

    Uniform illumination of a working field is very important in optical systems of confocal microscopy and various implementations of fluorescence microscopy like TIR, SSIM, STORM, PALM to enhance performance of these laser-based research techniques. Widely used TEM00 laser sources are characterized by essentially non-uniform Gaussian intensity profile which leads usually to non-uniform intensity distribution in a microscope working field or in a field of microlenses array of a confocal microscope optical system, this non-uniform illumination results in instability of measuring procedure and reducing precision of quantitative measurements. Therefore transformation of typical Gaussian distribution of a TEM00 laser to flat-top (top hat) profile is an actual technical task, it is solved by applying beam shaping optics. Due to high demands to optical image quality the mentioned techniques have specific requirements to a uniform laser beam: flatness of phase front and extended depth of field, - from this point of view the microscopy techniques are similar to holography and interferometry. There are different refractive and diffractive beam shaping approaches used in laser industrial and scientific applications, but only few of them are capable to fulfil the optimum conditions for beam quality required in discussed microscopy techniques. We suggest applying refractive field mapping beam shapers πShaper, which operational principle presumes almost lossless transformation of Gaussian to flat-top beam with flatness of output wavefront, conserving of beam consistency, providing collimated low divergent output beam, high transmittance, extended depth of field, negligible wave aberration, and achromatic design provides capability to work with several lasers with different wavelengths simultaneously. The main function of a beam shaper is transformation of laser intensity profile, further beam transformation to provide optimum for a particular technique spot size and shape has to

  8. Advances in fiber lasers for nonlinear microscopy

    NASA Astrophysics Data System (ADS)

    Wise, F. W.; Ouzounov, D.; Kieu, K.; Renninger, W.; Chong, A.; Liu, H.

    2008-02-01

    In the past 30 years major advances in medical imaging have been made in areas such as magnetic resonance imaging, computed tomography, and ultrasound. These techniques have become quite effective for structural imaging at the organ or tissue level, but do not address the clear need for imaging technologies that exploit existing knowledge of the genetic and molecular bases of disease. Techniques that can provide similar information on the cellular and molecular scale would be very powerful, and ultimately the extension of such techniques to in vivo measurements will be desired. The availability of these imaging capabilities would allow monitoring of the early stages of disease or therapy, for example. Optical techniques provide excellent imaging capabilities, with sub-micron spatial resolution, and are noninvasive. An overall goal of biomedical imaging is to obtain diagnostic or functional information about biological structures. The difficulty of acquiring high-resolution images of structures deep in tissue presents a major challenge, however, owing to strong scattering of light. As a consequence, optical imaging has been limited to thin (typically ~0.5 mm) samples or superficial tissue. In contrast, techniques such as ultrasound and magnetic resonance provide images of structures centimeters deep in tissue, with ~100-micron resolution. It is desirable to develop techniques that offer the resolution of optics with the depth-penetration of other techniques. Since 1990, a variety of nonlinear microscopies have been demonstrated. These include 2- and 3-photon fluorescence microscopy, and 2nd- and 3rd-harmonic generation microscopies. These typically employ femtosecond-pulse excitation, for maximum peak power (and thus nonlinear excitation) for a given pulse energy. A relative newcomer to the group is CARS microscopy [1], which exploits resonant vibrational excitation of molecules or bonds. The CARS signal contrast arises from intrinsic elements of cells, and thus

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

    NASA Astrophysics Data System (ADS)

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

    2017-03-01

    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 resolution and signal-to-noise ratio, are characterized and compared with properties of standard confocal microscopy. The results show that the lateral resolution of RCM is ~170 nm compared to ~240 nm of confocal microscopy for 488 nm excitation and 1.49 NA. As the theory predicts, this improved lateral resolution is independent of the pinhole diameter. In standard confocal microscopy, the same lateral resolution can only be achieved with an almost closed pinhole and, consequently, with a major loss of signal. We show that the sectioning capabilities of the standard confocal microscope are preserved in RCM and that the axial resolution of RCM is slightly better (~15%) than the standard confocal microscope. Furthermore, the signal-to-noise ratio in RCM is a factor of 2 higher than in standard confocal microscopy, also due to the use of highly sensitive modern cameras. In case the pinhole of a confocal microscope is adjusted in such way that the lateral resolution is comparable to that of RCM, the signal-to-noise ratio in RCM is 4 times higher than standard confocal microscopy. Therefore, RCM offers a good alternative to standard confocal microscopy for higher lateral resolution with the main advantage of strongly improved sensitivity.

  10. Combined frequency modulated atomic force microscopy and scanning tunneling microscopy detection for multi-tip scanning probe microscopy applications

    SciTech Connect

    Morawski, Ireneusz; Spiegelberg, Richard; Korte, Stefan; Voigtländer, Bert

    2015-12-15

    A method which allows scanning tunneling microscopy (STM) tip biasing independent of the sample bias during frequency modulated atomic force microscopy (AFM) operation is presented. The AFM sensor is supplied by an electronic circuit combining both a frequency shift signal and a tunneling current signal by means of an inductive coupling. This solution enables a control of the tip potential independent of the sample potential. Individual tip biasing is specifically important in order to implement multi-tip STM/AFM applications. An extensional quartz sensor (needle sensor) with a conductive tip is applied to record simultaneously topography and conductivity of the sample. The high resonance frequency of the needle sensor (1 MHz) allows scanning of a large area of the surface being investigated in a reasonably short time. A recipe for the amplitude calibration which is based only on the frequency shift signal and does not require the tip being in contact is presented. Additionally, we show spectral measurements of the mechanical vibration noise of the scanning system used in the investigations.

  11. Combined frequency modulated atomic force microscopy and scanning tunneling microscopy detection for multi-tip scanning probe microscopy applications.

    PubMed

    Morawski, Ireneusz; Spiegelberg, Richard; Korte, Stefan; Voigtländer, Bert

    2015-12-01

    A method which allows scanning tunneling microscopy (STM) tip biasing independent of the sample bias during frequency modulated atomic force microscopy (AFM) operation is presented. The AFM sensor is supplied by an electronic circuit combining both a frequency shift signal and a tunneling current signal by means of an inductive coupling. This solution enables a control of the tip potential independent of the sample potential. Individual tip biasing is specifically important in order to implement multi-tip STM/AFM applications. An extensional quartz sensor (needle sensor) with a conductive tip is applied to record simultaneously topography and conductivity of the sample. The high resonance frequency of the needle sensor (1 MHz) allows scanning of a large area of the surface being investigated in a reasonably short time. A recipe for the amplitude calibration which is based only on the frequency shift signal and does not require the tip being in contact is presented. Additionally, we show spectral measurements of the mechanical vibration noise of the scanning system used in the investigations.

  12. Combined frequency modulated atomic force microscopy and scanning tunneling microscopy detection for multi-tip scanning probe microscopy applications

    NASA Astrophysics Data System (ADS)

    Morawski, Ireneusz; Spiegelberg, Richard; Korte, Stefan; Voigtländer, Bert

    2015-12-01

    A method which allows scanning tunneling microscopy (STM) tip biasing independent of the sample bias during frequency modulated atomic force microscopy (AFM) operation is presented. The AFM sensor is supplied by an electronic circuit combining both a frequency shift signal and a tunneling current signal by means of an inductive coupling. This solution enables a control of the tip potential independent of the sample potential. Individual tip biasing is specifically important in order to implement multi-tip STM/AFM applications. An extensional quartz sensor (needle sensor) with a conductive tip is applied to record simultaneously topography and conductivity of the sample. The high resonance frequency of the needle sensor (1 MHz) allows scanning of a large area of the surface being investigated in a reasonably short time. A recipe for the amplitude calibration which is based only on the frequency shift signal and does not require the tip being in contact is presented. Additionally, we show spectral measurements of the mechanical vibration noise of the scanning system used in the investigations.

  13. A Correlative Optical Microscopy and Scanning Electron Microscopy Approach to Locating Nanoparticles in Brain Tumors

    PubMed Central

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

    2014-01-01

    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. PMID:25464144

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

    PubMed

    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.

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

    NASA Astrophysics Data System (ADS)

    Hayashi, Shinichi

    2016-08-01

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

  16. Complete and rapid switch from light microscopy to virtual microscopy for teaching medical histology.

    PubMed

    Krippendorf, Beth B; Lough, John

    2005-07-01

    During the interim between the 2003 and 2004 academic years, the cell and tissue biology and integrated medical neuroscience courses at the Medical College of Wisconsin made a complete and rapid switch from light microscopy- to virtual microscopy-based histology laboratories. This switch was prompted by the difficulties in maintaining and the cost of replacing the college's microscopes and microscope slides, and primarily by the desire to promote and streamline learning for our large classes (n > 200) of first-year medical students. A group of students who used the virtual microscope, another group of students who used the light microscope, and faculty with experience using both tools rated the effectiveness of the virtual microscope for learning and teaching. Also, to determine whether virtual microscopy affected student learning, laboratory examination scores for the 2004 class (n = 209) were compared with those of four previous classes that used light microscopes exclusively (n = 811). The switch from light microscopy to virtual microscopy was very favorably received by both students and faculty. More importantly, data from examination scores and course evaluation surveys indicated that use of the virtual microscope may significantly improve student performance and learning efficiency. Procedures for successfully implementing this change are described.

  17. Scanning Ion Conductance Microscopy of Live Keratinocytes

    NASA Astrophysics Data System (ADS)

    Hegde, V.; Mason, A.; Saliev, T.; Smith, F. J. D.; McLean, W. H. I.; Campbell, P. A.

    2012-07-01

    Scanning ion conductance microscopy (SICM) is perhaps the least well known technique from the scanning probe microscopy (SPM) family of instruments. As with its more familiar counterpart, atomic force microscopy (AFM), the technique provides high-resolution topographic imaging, with the caveat that target structures must be immersed in a conducting solution so that a controllable ion current may be utilised as the basis for feedback. In operation, this non-contact characteristic of SICM makes it ideal for the study of delicate structures, such as live cells. Moreover, the intrinsic architecture of the instrument, incorporating as it does, a scanned micropipette, lends itself to combination approaches with complementary techniques such as patch-clamp electrophysiology: SICM therefore boasts the capability for both structural and functional imaging. For the present observations, an ICnano S system (Ionscope Ltd., Melbourn, UK) operating in 'hopping mode' was used, with the objective of assessing the instrument's utility for imaging live keratinocytes under physiological buffers. In scans employing cultured HaCaT cells (spontaneously immortalised, human keratinocytes), we compared the qualitative differences of live cells imaged with SICM and AFM, and also with their respective counterparts after chemical fixation in 4% paraformaldehyde. Characteristic surface microvilli were particularly prominent in live cell imaging by SICM. Moreover, time lapse SICM imaging on live cells revealed that changes in the pattern of microvilli could be tracked over time. By comparison, AFM imaging on live cells, even at very low contact forces (

  18. Atomic force microscopy study of tooth surfaces.

    PubMed

    Farina, M; Schemmel, A; Weissmüller, G; Cruz, R; Kachar, B; Bisch, P M

    1999-03-01

    Atomic force microscopy (AFM) was used to study tooth surfaces in order to compare the pattern of particle distribution in the outermost layer of the tooth surfaces. Human teeth and teeth from a rodent (Golden hamster), from a fish (piranha), and from a grazing mollusk (chiton) with distinct feeding habits were analyzed in terms of particle arrangement, packing, and size distribution. Scanning electron microscopy and transmission electron microscopy were used for comparison. It was found that AFM gives high-contrast, high-resolution images and is an important tool as a source of complementary and/or new structural information. All teeth were cleaned and some were etched with acidic solutions before analysis. It was observed that human enamel (permanent teeth) presents particles tightly packed in the outer surface, whereas enamel from the hamster (continuously growing teeth) shows particles of less dense packing. The piranha teeth have a thin cuticle covering the long apatite crystals of the underlying enameloid. This cuticle has a rough surface of particles that have a globular appearance after the brief acidic treatment. The similar appearance of the in vivo naturally etched tooth surface suggests that the pattern of globule distribution may be due to the presence of an organic material. Elemental analysis of this cuticle indicated that calcium, phosphorus, and iron are the main components of the structure while electron microdiffraction of pulverized cuticle particles showed a pattern consistent with hydroxyapatite. The chiton mineralized tooth cusp had a smooth surface in an unabraded region and a very rough structure with the magnetite crystals (already known to make part of the structure) protruding from the surface. It was concluded that the structures analyzed are optimized for efficiency in feeding mechanism and life span of the teeth.

  19. Light Microscopy Module Imaging Tested and Demonstrated

    NASA Technical Reports Server (NTRS)

    Gati, Frank

    2004-01-01

    The Fluids Integrated Rack (FIR), a facility-class payload, and the Light Microscopy Module (LMM), a subrack payload, are integrated research facilities that will fly in the U.S. Laboratory module, Destiny, aboard the International Space Station. Both facilities are being engineered, designed, and developed at the NASA Glenn Research Center by Northrop Grumman Information Technology. The FIR is a modular, multiuser scientific research facility that is one of two racks that make up the Fluids and Combustion Facility (the other being the Combustion Integrated Rack). The FIR has a large volume dedicated for experimental hardware; easily reconfigurable diagnostics, power, and data systems that allow for unique experiment configurations; and customizable software. The FIR will also provide imagers, light sources, power management and control, command and data handling for facility and experiment hardware, and data processing and storage. The first payload in the FIR will be the LMM. The LMM integrated with the FIR is a remotely controllable, automated, on-orbit microscope subrack facility, with key diagnostic capabilities for meeting science requirements--including video microscopy to observe microscopic phenonema and dynamic interactions, interferometry to make thin-film measurements with nanometer resolution, laser tweezers to manipulate micrometer-sized particles, confocal microscopy to provide enhanced three-dimensional visualization of structures, and spectrophotometry to measure the photonic properties of materials. Vibration disturbances were identified early in the LMM development phase as a high risk for contaminating the science microgravity environment. An integrated FIR-LMM test was conducted in Glenn's Acoustics Test Laboratory to assess mechanical sources of vibration and their impact to microscopic imaging. The primary purpose of the test was to characterize the LMM response at the sample location, the x-y stage within the microscope, to vibration

  20. Super-resolved spatial light interference microscopy.

    PubMed

    Chu, Kaiqin; Smith, Zachary J; Wachsmann-Hogiu, Sebastian; Lane, Stephen

    2012-03-01

    We report a scheme to achieve resolution beyond the diffraction limit in spatial light interference microscopy (SLIM). By adding a grating to the optical path, the structured illumination technique can be used to improve the resolution by a factor of 2. We show that a direct application of the structured illumination technique, however, has proved to be unsuccessful. Through two crucial modifications, namely, one to the pupil plane of the objective and the other to the demodulation procedure, faithful phase information of the object is recovered and the resolution is improved by a factor of 2.

  1. Cell surface fluctuations studied with defocusing microscopy

    NASA Astrophysics Data System (ADS)

    Agero, U.; Monken, C. H.; Ropert, C.; Gazzinelli, R. T.; Mesquita, O. N.

    2003-05-01

    Phase objects can become visible by slightly defocusing an optical microscope, a technique seldom used as a useful tool. We revisited the theory of defocusing and apply it to our optical microscope with optics corrected at infinity. In our approximation, we obtain that the image contrast is proportional to the two-dimensional (2D) Laplacian of the phase difference introduced by the phase object. If the index of refraction of the phase object is uniform the image obtained from defocusing microscopy is the image of curvature (Laplacian of the local thickness) of the phase object, while standard phase-contrast microscopy gives information about the thickness of the object. We made artificial phase objects and measured image contrasts with defocusing microscopy. Measured contrasts are in excellent agreement with our theoretical model. We use defocusing microscopy to study curvature fluctuations (ruffles) on the surface of macrophages (cell of the innate immune system), and try to correlate mechanical properties of macrophage surface and phagocytosis. We observe large coherent propagating structures: Their shape, speed, density are measured and curvature energy estimated. Inhomogeneities of cytoskeleton refractive index, curvature modulations due to thermal fluctuations and/or periodic changes in cytoskeleton-membrane interactions cause random fluctuations in image contrast. From the temporal and spatial contrast correlation functions, we obtain the decay time and correlation length of such fluctuations that are related to their size and the viscoelastic properties of the cytoskeleton. In order to associate the dynamics of cytoskeleton with the process of phagocytosis, we use an optical tweezers to grab a zymosan particle and put it into contact with the macrophage. We then measure the time for a single phagocytosis event. We add the drug cytochalasin D that depolymerizes the cytoskeleton F-actin network: It inhibits the large propagating coherent fluctuations on the

  2. Nonlinear Optical Microscopy of Single Nanostructures

    NASA Astrophysics Data System (ADS)

    Huang, Libai; Cheng, Ji-Xin

    2013-07-01

    We review recent advances in nonlinear optical (NLO) microscopy studies of single nanostructures. NLO signals are intrinsically sensitive to the electronic, vibrational, and structural properties of such nanostructures. Ultrafast excitation allows for mapping of energy relaxation pathways at the single-particle level. The strong nonlinear response of nanostructures makes them highly attractive for applications as novel NLO imaging agents in biological and biomedical research. NLO modalities based on harmonic generation, multiphoton photoluminescence, four-wave mixing, and pump-probe processes are discussed in detail.

  3. Improvement of image quality in holographic microscopy.

    PubMed

    Budhiraja, C J; Som, S C

    1981-05-15

    A novel technique of noise reduction in holographic microscopy has been experimentally studied. It has been shown that significant improvement in the holomicroscopic images of actual low-contrast continuous tone biological objects can be achieved without trade off in image resolution. The technique makes use of holographically produced multidirectional phase gratings used as diffusers and the continuous addition of subchannel holograms. It has been shown that the self-imaging property of this type of diffuser makes the use of these diffusers ideal for microscopic objects. Experimental results have also been presented to demonstrate real-time image processing capability of this technique.

  4. Scanning electron microscopy study of Tritrichomonas augusta.

    PubMed

    Borges, Fernanda P; Wiltuschnig, Renata C M; Tasca, Tiana; De Carli, Geraldo A

    2004-09-01

    Tritrichomonas augusta is a flagellated protozoan that parasitizes amphibians and reptiles. According to scanning electron microscopy (SEM), the cell shape of T. augusta varies from slender pyriform to ovoidal. Our data show the morphological features of the trophozoites: the emergence of the anterior flagella, the structure of the undulating membrane and the position and shape of the pelta, axostyle and posterior flagellum. In addition, herein we describe spherical forms which are probably pseudocysts. The description of the external structure of T. augusta, as demonstrated by SEM, contributes to the understanding of the biology of this parasite.

  5. System analysis of force feedback microscopy

    SciTech Connect

    Rodrigues, Mario S.; Chevrier, Joël; Comin, Fabio

    2014-02-07

    It was shown recently that the Force Feedback Microscope (FFM) can avoid the jump-to-contact in Atomic force Microscopy even when the cantilevers used are very soft, thus increasing force resolution. In this letter, we explore theoretical aspects of the associated real time control of the tip position. We take into account lever parameters such as the lever characteristics in its environment, spring constant, mass, dissipation coefficient, and the operating conditions such as controller gains and interaction force. We show how the controller parameters are determined so that the FFM functions at its best and estimate the bandwidth of the system under these conditions.

  6. Fibreoptic fluorescent microscopy in studying biological objects

    SciTech Connect

    Morozov, A N; Turchin, Il'ya V; Kamenskii, V A; Fiks, I I; Lazutkin, A A; Bezryadkov, D V; Ivanova, A A; Toptunov, D M; Anokhin, K V

    2010-11-13

    The method of fluorescent microscopy is developed based on employment of a single-mode fibreoptic channel to provide high spatial resolution 3D images of large cleared biological specimens using the 488-nm excitation laser line. The transverse and axial resolution of the setup is 5 and 13 {mu}m, respectively. The transversal sample size under investigation is up to 10 mm. The in-depth scanning range depends on the sample transparency and reaches 4 mm in the experiment. The 3D images of whole mouse organs (heart, lungs, brain) and mouse embryos obtained using autofluorescence or fluorescence of exogenous markers demonstrate a high contrast and cellular-level resolution.

  7. Anal melanosis diagnosed by reflectance confocal microscopy.

    PubMed

    Cinotti, Elisa; Chol, Christelle; Perrot, Jean Luc; Labeille, Bruno; Forest, Fabien; Cambazard, Frédéric

    2014-11-01

    Until now, in vivo reflectance-mode confocal microscopy (IVCM) has been applied only to pigmented lesions of the vulvar and oral mucosa, but not to anal mucosa lesions. We present the first case in which IVCM has been used to diagnose anal melanosis. Clinical and dermoscopic features were of concern while IVCM found the draped pattern already described for genital melanosis. IVCM adds information to the clinical and dermatoscopic examination and allows skin biopsies to be avoided. Further studies are needed to define the IVCM features of anal melanosis and to compare the performance of IVCM with the findings of histological examinations.

  8. Magnetic force microscopy of superparamagnetic nanoparticles.

    PubMed

    Schreiber, Sharon; Savla, Mayur; Pelekhov, Denis V; Iscru, Daniel F; Selcu, Camelia; Hammel, P Chris; Agarwal, Gunjan

    2008-02-01

    The use of magnetic force microscopy (MFM) to detect probe-sample interactions from superparamagnetic nanoparticles in vitro in ambient atmospheric conditions is reported here. By using both magnetic and nonmagnetic probes in dynamic lift-mode imaging and by controlling the direction and magnitude of the external magnetic field applied to the samples, it is possible to detect and identify the presence of superparamagnetic nanoparticles. The experimental results shown here are in agreement with the estimated sensitivity of the MFM technique. The potential and challenges for localizing nanoscale magnetic domains in biological samples is discussed.

  9. Embedment-free section electron microscopy.

    PubMed

    Kondo, Hisatake

    2006-08-01

    Because of potential hindrance of clear viewing in epoxy sections of biological entities having an electron density similar to and lower than that of epoxy resin, the author has stressed that the embedment-free section electron microscopy is necessary for re-examination and/or clarification of biological specimen structures, and that the embedment-free electron microscopy is reliably done by using water-soluble polyethylene glycol (PEG) as a transient embedding media and by critical point-drying of embedment-free sections after de-embedment of PEG by immersion of semithin sections into water. With the embedment-free electron microscopy, the author has presented five major findings: the appearance of microtrabecular lattices with different compactnesses in various cells and in intracellular domains of a given cell, the faithful reproduction of microtrabecula-like strand lattices in vitro with increasing compactnesses from artificial protein solutions at correspondingly increasing concentrations, the appearance of more compact lattices from gelated gelatin than from solated gelatin at a given concentration in vitro, the changeability in compactnesses of the microtrabecular lattices by hyper- or hypotonic shock treatments of cells, and the confined appearance of an intracellular protein in the centripetal demilune of centrifuged ganglion cells which is occupied with the microtrabecular lattices of a substantial compactness. From these findings, several conclusions are drawn: individual strands themselves of the microtrabeculae are meaningless, the appearance of microtrabeculae represents the presence of proteins at a certain concentration, and it is therefore likely that the aqueous cytoplasm is equivalent to the aqueous solution. In addition, it is possible that the appearance of two contiguous lattice domains exhibiting different compactnesses in a given cell may represent the occurrence of a contiguity of sol to gel states of cytoplasmic domains. It is thus

  10. Periodicity in bimodal atomic force microscopy

    SciTech Connect

    Lai, Chia-Yun; Santos, Sergio Chiesa, Matteo; Barcons, Victor

    2015-07-28

    Periodicity is fundamental for quantification and the application of conservation principles of many important systems. Here, we discuss periodicity in the context of bimodal atomic force microscopy (AFM). The relationship between the excited frequencies is shown to affect and control both experimental observables and the main expressions quantified via these observables, i.e., virial and energy transfer expressions, which form the basis of the bimodal AFM theory. The presence of a fundamental frequency further simplifies the theory and leads to close form solutions. Predictions are verified via numerical integration of the equation of motion and experimentally on a mica surface.

  11. Epifluorescence collection in two-photon microscopy

    NASA Astrophysics Data System (ADS)

    Beaurepaire, Emmanuel; Mertz, Jerome

    2002-09-01

    We present a simple model to describe epifluorescence collection in two-photon microscopy when one images in a turbid slab with an objective. Bulk and surface scattering determine the spatial and angular distributions of the outgoing fluorescence photons at the slab surface, and geometrical optics determines how efficiently the photons are collected. The collection optics are parameterized by the objective's numerical aperture and working distance and by an effective collection field of view. We identify the roles of each of these parameters and provide simple rules of thumb for the optimization of the epifluorescence collection efficiency. Analytical results are corroborated by Monte Carlo simulation.

  12. In vivo virtual intraoperative surgical photoacoustic microscopy

    NASA Astrophysics Data System (ADS)

    Han, Seunghoon; Lee, Changho; Kim, Sehui; Jeon, Mansik; Kim, Jeehyun; Kim, Chulhong

    2013-11-01

    We developed a virtual intraoperative surgical photoacoustic microscopy system by combining with a commercial surgical microscope and photoacoustic microscope (PAM). By sharing the common optical path in the microscope and PAM system, we could acquire the PAM and microscope images simultaneously. Moreover, by employing a beam projector to back-project 2D PAM images onto the microscope view plane as augmented reality, the conventional microscopic and 2D cross-sectional PAM images are concurrently mapped on the plane via an ocular lens of the microscope in real-time. Further, we guided needle insertion into phantom ex vivo and mice skins in vivo.

  13. Mudrocks examined by backscattered electron microscopy

    NASA Technical Reports Server (NTRS)

    Pye, K.; Krinsley, D.

    1983-01-01

    A method of studying mudrocks is developed using backscattered electrons (BSE) in scanning electron microscopy. Commercially available detectors are utilized to mix the BSE and secondary electron signals in order to obtain the optimum image for a particular material. Thin sections or polished rock chip surfaces are examined with BSE which provides both the atomic number contrast and topographic contrast. This technique provides very detailed information about the form and composition of individual grains in the mudrock thin sections and can be used in studies of the source, mode of deposition, diagenesis, and tectonic deformational history of mudrocks.

  14. Materials Contrast in Piezoresponse Force Microscopy

    SciTech Connect

    Kalinin, Sergei V; Eliseev, E. A.; Morozovska, A. N.

    2006-01-01

    Piezoresponse force microscopy (PFM) contrast in transversally isotropic material corresponding to the case of c{sup +}-c{sup -} domains in tetragonal ferroelectrics is analyzed using Green's function theory by Felten et al. [J. Appl. Phys. 96, 563 (2004)]. A simplified expression for PFM signal as a linear combination of relevant piezoelectric constant is obtained. This analysis is extended to piezoelectric material of arbitrary symmetry with weak elastic and dielectric anisotropies. These results provide a framework for interpretation of PFM signals for systems with unknown or poorly known local elastic and dielectric properties, including nanocrystalline materials, ferroelectric polymers, and biopolymers.

  15. Optical diffraction microscopy in a teaching laboratory

    NASA Astrophysics Data System (ADS)

    Thibault, Pierre; Rankenburg, Ivan C.

    2007-09-01

    We discuss an optics experiment that reproduces all important aspects of diffraction microscopy or coherent diffractive imaging. This technique is used to reconstruct an object's image from its diffraction pattern. The experimental setup is described in detail and only requires material readily available in a well-equipped optics teaching laboratory. The data analysis procedure is explained, in particular the reconstruction part, for which an iterative phase retrieval algorithm is used. The method is illustrated by showing the complex-valued reconstruction of an insect wing from a diffraction pattern measured with this setup.

  16. Electron Microscopy of Young Candida albicans Chlamydospores

    PubMed Central

    Miller, Sara E.; Spurlock, Ben O.; Michaels, G. E.

    1974-01-01

    One- to three-day-old cultures of Candida albicans bearing chlamydospores were grown and harvested by a special technique, free of agar, and prepared for ultramicrotomy and electron microscopy. These young chlamydospores exhibited a subcellular structure similar to that of the yeast phase, e.g., cytoplasmic membrane, ribosomes, and mitochondria. Other structural characteristics unique to chlamydospores were a very thick, layered cell wall, the outer layer of which was continuous with the outer layer of the suspensor cell wall and was covered by hair-like projections; membrane bound organelles; and large lipoid inclusions. Only young chlamydospores less than 3 to 4 days old exhibited these ultrastructural characteristics. Images PMID:4368664

  17. Tip-modulation scanned gate microscopy.

    PubMed

    Wilson, Neil R; Cobden, David H

    2008-08-01

    We introduce a technique that improves the sensitivity and resolution and eliminates the nonlocal background of scanned gate microscopy (SGM). In conventional SGM, a voltage bias is applied to the atomic force microscope tip and the sample conductance is measured as the tip is scanned. In the new technique, which we call tip-modulation SGM (tmSGM), the biased tip is oscillated and the induced oscillation of the sample conductance is measured. Applied to single-walled carbon nanotube network devices, tmSGM gives sharp, low-noise and background-free images.

  18. [Mobile phone based wireless microscopy imaging technology].

    PubMed

    Yuan, Yucheng; Liu, Jing

    2011-03-01

    This article proposes a new device named "Wireless Cellscope" that combining mobile phone and optical microscope together. The established wireless microscope platform consists of mobile phone, network monitor, miniaturized microscope or high resolution microscope etc. A series of conceptual experiments were performed on microscopic observation of ordinary objects and mice tumor tissue slices. It was demonstrated that, the new method could acquire microscopy images via a wireless way, which is spatially independent. With small size and low cost, the device thus developed has rather wide applicability in non-disturbing investigation of cell/tissue culture and long distance observation of dangerous biological sample etc.

  19. Laser-induced air ionization microscopy

    NASA Astrophysics Data System (ADS)

    Zhao, Y.; Zhang, N.; Yang, J.; Zhu, X.

    2006-06-01

    A nonlinear scanning imaging method is introduced that uses the highly localized air ionization initiated by photoelectrons from the sample surface under irradiation of femtosecond laser pulses as the microprobe. This type of microscopy with realizable subdiffraction spatial resolution has the unique advantages of being highly sensitive to both elemental and topographical properties of the samples of interest. Microscopic images of a femtosecond laser ablated micropattern, the cross section and the side view profile of an optical fiber, and a fresh mulberry leaf are obtained with this imaging technique, which demonstrate this technique's broad applicability in microscopic studies of different materials.

  20. Atomic force microscopy on liquid crystals

    NASA Astrophysics Data System (ADS)

    Bahr, Christian; Schulz, Benjamin

    This chapter provides an introduction to the atomic force microscopy (AFM) on thermotropic liquid crystals. We first give a general introduction to the technique of AFM and then describe the special requirements that have to be met for the imaging of liquid-crystalline surfaces. We also discuss the relation between the quality or reliability of the imaging results and various parameters of the scanning conditions. We briey review the existing work on AFM on liquid crystals and finally describe applications beyond the imaging, such as molecular force spectroscopy or manipulation of surface structures.

  1. In vivo virtual intraoperative surgical photoacoustic microscopy

    SciTech Connect

    Han, Seunghoon Kim, Sehui Kim, Jeehyun E-mail: chulhong@postech.edu; Lee, Changho Jeon, Mansik; Kim, Chulhong E-mail: chulhong@postech.edu

    2013-11-11

    We developed a virtual intraoperative surgical photoacoustic microscopy system by combining with a commercial surgical microscope and photoacoustic microscope (PAM). By sharing the common optical path in the microscope and PAM system, we could acquire the PAM and microscope images simultaneously. Moreover, by employing a beam projector to back-project 2D PAM images onto the microscope view plane as augmented reality, the conventional microscopic and 2D cross-sectional PAM images are concurrently mapped on the plane via an ocular lens of the microscope in real-time. Further, we guided needle insertion into phantom ex vivo and mice skins in vivo.

  2. SESAM: exploring the frontiers of electron microscopy.

    PubMed

    Koch, Christoph T; Sigle, Wilfried; Höschen, Rainer; Rühle, Manfred; Essers, Erik; Benner, Gerd; Matijevic, Marko

    2006-12-01

    We report on the sub-electron-volt-sub-angstrom microscope (SESAM), a high-resolution 200-kV FEG-TEM equipped with a monochromator and an in-column MANDOLINE filter. We report on recent results obtained with this instrument, demonstrating its performance (e.g., 87-meV energy resolution at 10-s exposure time, or a transmissivity of the energy filter of T1 ev = 11,000 nm2). New opportunities to do unique experiments that may advance the frontiers of microscopy in areas such as energy-filtered TEM, spectroscopy, energy-filtered electron diffraction and spectroscopic profiling are also discussed.

  3. Computer vision for microscopy diagnosis of malaria.

    PubMed

    Tek, F Boray; Dempster, Andrew G; Kale, Izzet

    2009-07-13

    This paper reviews computer vision and image analysis studies aiming at automated diagnosis or screening of malaria infection in microscope images of thin blood film smears. Existing works interpret the diagnosis problem differently or propose partial solutions to the problem. A critique of these works is furnished. In addition, a general pattern recognition framework to perform diagnosis, which includes image acquisition, pre-processing, segmentation, and pattern classification components, is described. The open problems are addressed and a perspective of the future work for realization of automated microscopy diagnosis of malaria is provided.

  4. Structured illumination fluorescence Fourier ptychographic microscopy

    NASA Astrophysics Data System (ADS)

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

    2016-12-01

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

  5. Fast electron microscopy via compressive sensing

    SciTech Connect

    Larson, Kurt W; Anderson, Hyrum S; Wheeler, Jason W

    2014-12-09

    Various technologies described herein pertain to compressive sensing electron microscopy. A compressive sensing electron microscope includes a multi-beam generator and a detector. The multi-beam generator emits a sequence of electron patterns over time. Each of the electron patterns can include a plurality of electron beams, where the plurality of electron beams is configured to impart a spatially varying electron density on a sample. Further, the spatially varying electron density varies between each of the electron patterns in the sequence. Moreover, the detector collects signals respectively corresponding to interactions between the sample and each of the electron patterns in the sequence.

  6. Reflectance confocal microscopy features of facial angiofibromas

    PubMed Central

    Millán-Cayetano, José-Francisco; Yélamos, Oriol; Rossi, Anthony M.; Marchetti, Michael A.; Jain, Manu

    2017-01-01

    Facial angiofibromas are benign tumors presenting as firm, dome-shaped, flesh-colored to pink papules, typically on the nose and adjoining central face. Clinically and dermoscopically they can mimic melanocytic nevi or basal cell carcinomas (BCC). Reflectance confocal microscopy (RCM) is a noninvasive imaging tool that is useful in diagnosing melanocytic and non-melanocytic facial lesions. To date no studies have described the RCM features of facial angiofibromas. Herein, we present two cases of facial angiofibromas that were imaged with RCM and revealed tumor island-like structures that mimicked BCC, leading to skin biopsy. PMID:28243496

  7. Statistical Quality Control of Moisture Data in GEOS DAS

    NASA Technical Reports Server (NTRS)

    Dee, D. P.; Rukhovets, L.; Todling, R.

    1999-01-01

    A new statistical quality control algorithm was recently implemented in the Goddard Earth Observing System Data Assimilation System (GEOS DAS). The final step in the algorithm consists of an adaptive buddy check that either accepts or rejects outlier observations based on a local statistical analysis of nearby data. A basic assumption in any such test is that the observed field is spatially coherent, in the sense that nearby data can be expected to confirm each other. However, the buddy check resulted in excessive rejection of moisture data, especially during the Northern Hemisphere summer. The analysis moisture variable in GEOS DAS is water vapor mixing ratio. Observational evidence shows that the distribution of mixing ratio errors is far from normal. Furthermore, spatial correlations among mixing ratio errors are highly anisotropic and difficult to identify. Both factors contribute to the poor performance of the statistical quality control algorithm. To alleviate the problem, we applied the buddy check to relative humidity data instead. This variable explicitly depends on temperature and therefore exhibits a much greater spatial coherence. As a result, reject rates of moisture data are much more reasonable and homogeneous in time and space.

  8. Chemistry Viewed through the Eyes of High-Resolution Microscopy.

    ERIC Educational Resources Information Center

    Beer, Michael; And Others

    1981-01-01

    This special report, prepared by several chemists working in the field of electron microscopy, provides information regarding the most recent developments in transmission and scanning electron microscopy that have chemical significance. (CS)

  9. Is bleach-sedimented smear microscopy an alternative to direct microscopy under programme conditions in India?

    PubMed

    Vishnu, P H; Bhat, P; Bansal, A; Satyanarayana, S; Alavadi, U; Ohri, B S; Shrinivas, M S Rao; Desikan, P; Jaju, J; Rao, V G; Moonan, P K

    2013-03-21

    This cross-sectional multi-centric study compared the yield of and potential benefit for detecting smear-positive pulmonary tuberculosis (PTB) by bleach sedimentation (2% sodium-hypochlorite) versus direct microscopy under programme conditions in India. Among 3168 PTB suspects, 684 (21.6%) were detected by bleach sedimentation vs. 625 (19.7%) by direct microscopy, with a proportional overall agreement of 96% (κ = 0.88). While 594 patients were smear-positive with both methods, 31 patients detected by direct microscopy were missed and an additional 90 patients were detected by bleach sedimentation. Overall, bleach sedimentation increased the yield of smear-positive TB detection; however; it also increased the time to results.

  10. Single-spin stochastic optical reconstruction microscopy

    PubMed Central

    Pfender, Matthias; Aslam, Nabeel; Waldherr, Gerald; Neumann, Philipp; Wrachtrup, Jörg

    2014-01-01

    We experimentally demonstrate precision addressing of single-quantum emitters by combined optical microscopy and spin resonance techniques. To this end, we use nitrogen vacancy (NV) color centers in diamond confined within a few ten nanometers as individually resolvable quantum systems. By developing a stochastic optical reconstruction microscopy (STORM) technique for NV centers, we are able to simultaneously perform sub–diffraction-limit imaging and optically detected spin resonance (ODMR) measurements on NV spins. This allows the assignment of spin resonance spectra to individual NV center locations with nanometer-scale resolution and thus further improves spatial discrimination. For example, we resolved formerly indistinguishable emitters by their spectra. Furthermore, ODMR spectra contain metrology information allowing for sub–diffraction-limit sensing of, for instance, magnetic or electric fields with inherently parallel data acquisition. As an example, we have detected nuclear spins with nanometer-scale precision. Finally, we give prospects of how this technique can evolve into a fully parallel quantum sensor for nanometer resolution imaging of delocalized quantum correlations. PMID:25267655

  11. Deep Imaging: the next frontier in microscopy.

    PubMed

    Roukos, Vassilis; Misteli, Tom

    2014-08-01

    The microscope is the quintessential tool for discovery in cell biology. From its earliest incarnation as a tool to make the unseen visible, microscopes have been at the center of most revolutionizing developments in cell biology, histology and pathology. Major quantum leaps in imaging involved the dramatic improvements in resolution to see increasingly smaller structures, methods to visualize specific molecules inside of cells and tissues, and the ability to peer into living cells to study dynamics of molecules and cellular structures. The latest revolution in microscopy is Deep Imaging-the ability to look at very large numbers of samples by high-throughput microscopy at high spatial and temporal resolution. This approach is rooted in the development of fully automated high-resolution microscopes and the application of advanced computational image analysis and mining methods. Deep Imaging is enabling two novel, powerful approaches in cell biology: the ability to image thousands of samples with high optical precision allows every discernible morphological pattern to be used as a read-out in large-scale imaging-based screens, particularly in conjunction with RNAi-based screening technology; in addition, the capacity to capture large numbers of images, combined with advanced computational image analysis methods, has also opened the door to detect and analyze very rare cellular events. These two applications of Deep Imaging are revolutionizing cell biology.

  12. Edge detection in microscopy images using curvelets

    PubMed Central

    Gebäck, Tobias; Koumoutsakos, Petros

    2009-01-01

    Background Despite significant progress in imaging technologies, the efficient detection of edges and elongated features in images of intracellular and multicellular structures acquired using light or electron microscopy is a challenging and time consuming task in many laboratories. Results We present a novel method, based on the discrete curvelet transform, to extract a directional field from the image that indicates the location and direction of the edges. This directional field is then processed using the non-maximal suppression and thresholding steps of the Canny algorithm to trace along the edges and mark them. Optionally, the edges may then be extended along the directions given by the curvelets to provide a more connected edge map. We compare our scheme to the Canny edge detector and an edge detector based on Gabor filters, and show that our scheme performs better in detecting larger, elongated structures possibly composed of several step or ridge edges. Conclusion The proposed curvelet based edge detection is a novel and competitive approach for imaging problems. We expect that the methodology and the accompanying software will facilitate and improve edge detection in images available using light or electron microscopy. PMID:19257905

  13. Prototype cantilevers for quantitative lateral force microscopy

    SciTech Connect

    Reitsma, Mark G.; Gates, Richard S.; Friedman, Lawrence H.; Cook, Robert F.

    2011-09-15

    Prototype cantilevers are presented that enable quantitative surface force measurements using contact-mode atomic force microscopy (AFM). The ''hammerhead'' cantilevers facilitate precise optical lever system calibrations for cantilever flexure and torsion, enabling quantifiable adhesion measurements and friction measurements by lateral force microscopy (LFM). Critically, a single hammerhead cantilever of known flexural stiffness and probe length dimension can be used to perform both a system calibration as well as surface force measurements in situ, which greatly increases force measurement precision and accuracy. During LFM calibration mode, a hammerhead cantilever allows an optical lever ''torque sensitivity'' to be generated for the quantification of LFM friction forces. Precise calibrations were performed on two different AFM instruments, in which torque sensitivity values were specified with sub-percent relative uncertainty. To examine the potential for accurate lateral force measurements using the prototype cantilevers, finite element analysis predicted measurement errors of a few percent or less, which could be reduced via refinement of calibration methodology or cantilever design. The cantilevers are compatible with commercial AFM instrumentation and can be used for other AFM techniques such as contact imaging and dynamic mode measurements.

  14. Image Quality Ranking Method for Microscopy

    PubMed Central

    Koho, Sami; Fazeli, Elnaz; Eriksson, John E.; Hänninen, Pekka E.

    2016-01-01

    Automated analysis of microscope images is necessitated by the increased need for high-resolution follow up of events in time. Manually finding the right images to be analyzed, or eliminated from data analysis are common day-to-day problems in microscopy research today, and the constantly growing size of image datasets does not help the matter. We propose a simple method and a software tool for sorting images within a dataset, according to their relative quality. We demonstrate the applicability of our method in finding good quality images in a STED microscope sample preparation optimization image dataset. The results are validated by comparisons to subjective opinion scores, as well as five state-of-the-art blind image quality assessment methods. We also show how our method can be applied to eliminate useless out-of-focus images in a High-Content-Screening experiment. We further evaluate the ability of our image quality ranking method to detect out-of-focus images, by extensive simulations, and by comparing its performance against previously published, well-established microscopy autofocus metrics. PMID:27364703

  15. Electrostatic-free piezoresponse force microscopy

    NASA Astrophysics Data System (ADS)

    Kim, Sungho; Seol, Daehee; Lu, Xiaoli; Alexe, Marin; Kim, Yunseok

    2017-01-01

    Contact and non-contact based atomic force microscopy (AFM) approaches have been extensively utilized to explore various nanoscale surface properties. In most AFM-based measurements, a concurrent electrostatic effect between the AFM tip/cantilever and sample surface can occur. This electrostatic effect often hinders accurate measurements. Thus, it is very important to quantify as well as remove the impact of the electrostatic effect on AFM-based measurements. In this study, we examine the impact of the electrostatic effect on the electromechanical (EM) response in piezoresponse force microscopy as a model AFM mode. We quantitatively studied the effects of increasing the external electric field and reducing the spring constant of a cantilever. Further, we explored ways to minimize the electrostatic effect. The results provide broad guidelines for quantitatively analyzing the EM response as well as, eventually, for obtaining the electrostatic-free EM response. The conclusions can be applied to other AFM-based measurements that are subject to a strong electrostatic effect between the AFM tip/cantilever and sample surface, regardless of contact and non-contact modes.

  16. Magnetic force microscopy: quantitative issues in biomaterials.

    PubMed

    Passeri, Daniele; Dong, Chunhua; Reggente, Melania; Angeloni, Livia; Barteri, Mario; Scaramuzzo, Francesca A; De Angelis, Francesca; Marinelli, Fiorenzo; Antonelli, Flavia; Rinaldi, Federica; Marianecci, Carlotta; Carafa, Maria; Sorbo, Angela; Sordi, Daniela; Arends, Isabel Wce; Rossi, Marco

    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 at the nanoscale. Being a well established tool for the characterization of magnetic recording media, superconductors and magnetic nanomaterials, MFM is finding constantly increasing application in the study of magnetic properties of materials and systems of biological and biomedical interest. After reviewing these latter applications, three case studies are presented in which MFM is used to characterize: (i) magnetoferritin synthesized using apoferritin as molecular reactor; (ii) magnetic nanoparticles loaded niosomes to be used as nanocarriers for drug delivery; (iii) leukemic cells labeled using folic acid-coated core-shell superparamagnetic nanoparticles in order to exploit the presence of folate receptors on the cell membrane surface. In these examples, MFM data are quantitatively analyzed evidencing the limits of the simple analytical models currently used. Provided that suitable models are used to simulate the MFM response, MFM can be used to evaluate the magnetic momentum of the core of magnetoferritin, the iron entrapment efficiency in single vesicles, or the uptake of magnetic nanoparticles into cells.

  17. Photorefractive phase-conjugation digital holographic microscopy

    NASA Astrophysics Data System (ADS)

    Chang, Chi-Ching; Chan, Huang-Tian; Shiu, Min-Tzung; Chew, Yang-Kun

    2015-05-01

    In this work, we propose an innovative method for digital holographic microscopy named as photorefractive phaseconjugation digital holographic microscopy (PPCDHM) technique based on the phase conjugation dynamic holographic process in photorefractive BaTiO3 crystal and the retrieval of phase and amplitude of the object wave were performed by a reflection-type digital holographic method. Both amplitude and phase reconstruction benefit from the prior amplification by self-pumped conjugation (SPPC) as they have an increased SNR. The interest of the PPCDHM is great, because its hologram is created by interfered the amplified phase-conjugate wave field generated from a photorefractive phase conjugator (PPC) correcting the phase aberration of the imaging system and the reference wave onto the digital CCD camera. Therefore, a precise three-dimensional description of the object with high SNR can be obtained digitally with only one hologram acquisition. The method requires the acquisition of a single hologram from which the phase distribution can be obtained simultaneously with distribution of intensity at the surface of the object.

  18. Scanning probe microscopy of protein nanowires

    NASA Astrophysics Data System (ADS)

    Walsh, Kathleen Ann

    The bacterium Geobacter sulfurreducens grows electrically-conductive pili, which act as protein nanowires, in order to transfer electrons from the cell to electron acceptors in its environment when direct charge transfer through the cell membrane is not feasible. Understanding the electronic structure of the pili can provide insight into fundamental processes of electron transfer in biological systems. This study investigated the electronic structure of these protein nanowires using the toolbox of scanning probe microscopy, specifically scanning tunneling microscopy and point tunneling spectroscopy. These measurements were performed at 77 K and at room temperature. The measured data are compared to theoretical calculations. Density of states measurements using tunneling spectroscopy show that these pili act as narrow-gap biological semiconductors at 77 K. The onset of nonzero density of states remains within the metabolically-relevant voltage range. At room temperature, spectroscopy of the pili retains a gap-like structure, but this pseudogap is raised to a nonzero density of states at even the smallest applied voltages. These pilus nanowires also exhibit a distinct spatial dependence of the density of states across the breadth of the pili.

  19. Atomic Force Microscopy Based Cell Shape Index

    NASA Astrophysics Data System (ADS)

    Adia-Nimuwa, Usienemfon; Mujdat Tiryaki, Volkan; Hartz, Steven; Xie, Kan; Ayres, Virginia

    2013-03-01

    Stellation is a measure of cell physiology and pathology for several cell groups including neural, liver and pancreatic cells. In the present work, we compare the results of a conventional two-dimensional shape index study of both atomic force microscopy (AFM) and fluorescent microscopy images with the results obtained using a new three-dimensional AFM-based shape index similar to sphericity index. The stellation of astrocytes is investigated on nanofibrillar scaffolds composed of electrospun polyamide nanofibers that has demonstrated promise for central nervous system (CNS) repair. Recent work by our group has given us the ability to clearly segment the cells from nanofibrillar scaffolds in AFM images. The clear-featured AFM images indicated that the astrocyte processes were longer than previously identified at 24h. It was furthermore shown that cell spreading could vary significantly as a function of environmental parameters, and that AFM images could record these variations. The new three-dimensional AFM-based shape index incorporates the new information: longer stellate processes and cell spreading. The support of NSF PHY-095776 is acknowledged.

  20. Deep stroma investigation by confocal microscopy

    NASA Astrophysics Data System (ADS)

    Rossi, Francesca; Tatini, Francesca; Pini, Roberto; Valente, Paola; Ardia, Roberta; Buzzonetti, Luca; Canovetti, Annalisa; Malandrini, Alex; Lenzetti, Ivo; Menabuoni, Luca

    2015-03-01

    Laser assisted keratoplasty is nowadays largely used to perform minimally invasive surgery and partial thickness keratoplasty [1-3]. The use of the femtosecond laser enables to perform a customized surgery, solving the specific problem of the single patient, designing new graft profiles and partial thickness keratoplasty (PTK). The common characteristics of the PTKs and that make them eligible respect to the standard penetrating keratoplasty, are: the preservation of eyeball integrity, a reduced risk of graft rejection, a controlled postoperative astigmatism. On the other hand, the optimal surgical results after these PTKs are related to a correct comprehension of the deep stroma layers morphology, which can help in the identification of the correct cleavage plane during surgeries. In the last years some studies were published, giving new insights about the posterior stroma morphology in adult subjects [4,5]. In this work we present a study performed on two groups of tissues: one group is from 20 adult subjects aged 59 +/- 18 y.o., and the other group is from 15 young subjects, aged 12+/-5 y.o.. The samples were from tissues not suitable for transplant in patients. Confocal microscopy and Environmental Scanning Electron Microscopy (ESEM) were used for the analysis of the deep stroma. The preliminary results of this analysis show the main differences in between young and adult tissues, enabling to improve the knowledge of the morphology and of the biomechanical properties of human cornea, in order to improve the surgical results in partial thickness keratoplasty.

  1. Two-Photon Laser Scanning Microscopy

    NASA Astrophysics Data System (ADS)

    Nimmerjahn, A.; Theer, P.; Helmchen, F.

    Since its inception more than 15 years ago, two-photon laser scanning microscopy (2PLSM) has found widespread use in biological and medical research. Two-photon microscopy is based on simultaneous absorption of two photons by fluorophores and subsequent fluorescence emission, a process which under normal illumination conditions is highly improbable. Theoretically described around 1930 by Maria Göppert-Mayer [1], the first experimental demonstration of two-photon excitation had to await the invention of the laser, which produced sufficiently high light intensities to observe two-photon absorption events [2]. Only after the development of ultrafast lasers providing subpicosecond light pulses with high peak power intensities, however, two-photon-excited fluorescence became practical in a laser-scanning microscope [3]. Since then 2PLSM has developed into the method of choice for high-resolution imaging in living animals (reviewed in [4,5]). One of the main reasons is the low sensitivity of 2PLSM to light scattering, which enables imaging relatively deep inside biological tissue and direct observation of the dynamic behavior of cells in their native environment. In this chapter, we introduce the physical principles governing 2PLSM and briefly describe the key instrument components. We give an overview of fluorescence labeling techniques and how they are combined with 2PLSM for functional imaging and photomanipulation in living tissue. Finally, we discuss limitations and provide some future perspectives.

  2. Intravital Microscopy for THz-Bio Analysis

    NASA Astrophysics Data System (ADS)

    Kim, Pilhan

    Intravital microscopy is a high-resolution imaging technique to observe biological phenomena in living organisms. It often also stated as in vivo microscopy. Literal meaning of in vivo is "within the living" and there is another term, ex vivo of which literal meaning is "out of the living". Both terms are commonly used to describe the status of sample at the moment of biological manipulations or investigations are done. In vivo study is a form of research using whole living organism in experiment to investigate a certain biological phenomenon in its natural environment, whereas ex vivo study uses non-living subjects such as tissues or organs dissected from dead animal. In addition, in vitro of which literal meaning is "within the glass" is another commonly used term. In vitro study is a form of research using small living subject such as cell in a controlled environment such as petri dish or test tube. Cell culture, the process of growing cells in a petri dish, is the most common form of in vitro study. Figure 1 summarizes the status of samples for biological study categorized by in vivo, in vitro and ex vivo.

  3. Use of astronomy filters in fluorescence microscopy.

    PubMed

    Piper, Jörg

    2012-02-01

    Monochrome astronomy filters are well suited for use as excitation or suppression filters in fluorescence microscopy. Because of their particular optical design, such filters can be combined with standard halogen light sources for excitation in many fluorescent probes. In this "low energy excitation," photobleaching (fading) or other irritations of native specimens are avoided. Photomicrographs can be taken from living motile fluorescent specimens also with a flash so that fluorescence images can be created free from indistinctness caused by movement. Special filter cubes or dichroic mirrors are not needed for our method. By use of suitable astronomy filters, fluorescence microscopy can be carried out with standard laboratory microscopes equipped with condensers for bright-field (BF) and dark-field (DF) illumination in transmitted light. In BF excitation, the background brightness can be modulated in tiny steps up to dark or black. Moreover, standard industry microscopes fitted with a vertical illuminator for examinations of opaque probes in DF or BF illumination based on incident light (wafer inspections, for instance) can also be used for excitation in epi-illumination when adequate astronomy filters are inserted as excitatory and suppression filters in the illuminating and imaging light path. In all variants, transmission bands can be modulated by transmission shift.

  4. Nuclear microscopy of rat colon epithelial cells

    NASA Astrophysics Data System (ADS)

    Ren, M.; Rajendran, Reshmi; Ng, Mary; Udalagama, Chammika; Rodrigues, Anna E.; Watt, Frank; Jenner, Andrew Michael

    2011-10-01

    Using Nuclear microscopy, we have investigated iron distributions in the colons of Sprague Dawley rats, in order to elucidate heme uptake. Four groups of five Sprague Dawley rats (mean weight 180 g) were fed different purified diets containing either heme diet (2.5% w/w hemoglobin), high fat diet (HFD) (18% w/w fat, 1% w/w cholesterol), 'western' diet (combination of hemoglobin 2.5% and 18% fat, 1% cholesterol) or control diet (7% w/w fat). After 4 weeks, animals were sacrificed by exsanguination after anaesthesia. Thin sections of frozen colon tissue were taken, freeze dried and scanned using nuclear microscopy utilising the techniques PIXE, RBS and STIM. The new data acquisition system (IonDaq) developed in CIBA was used to obtain high resolution images and line scans were used to map the iron distributions across the colon boundaries. The nuclear microscope results indicate that when HFD is given in addition to heme, the iron content of the epithelial cells that line the colon decreases, and the zinc in the smooth muscle wall increases. This implies that the level of heme and fat in diet has an important role in colon health, possibly by influencing epithelial cells directly or changing luminal composition such as bacterial flora or levels of metabolites and cytotoxins.

  5. Near Field Scanning Optical Microscopy (NSOM)

    PubMed Central

    Betzig, E.; Lewis, A.; Harootunian, A.; Isaacson, M.; Kratschmer, E.

    1986-01-01

    A new method for high-resolution imaging, near-field scanning optical microscopy (NSOM), has been developed. The concepts governing this method are discussed, and the technical challenges encountered in constructing a working NSOM instrument are described. Two distinct methods are presented for the fabrication of well-characterized, highly reproducible, subwavelength apertures. A sample one-dimensional scan is provided and compared to the scanning electron micrograph of a test pattern. From this comparison, a resolution of > 1,500 Å (i.e., ≃λ/3.6) is determined, which represents a significant step towards our eventual goal of 500 Å resolution. Fluorescence has been observed through apertures smaller than 600 Å and signal-to-noise calculations show that fluorescent imaging should be feasible. The application of such imaging is then discussed in reference to specific biological problems. The NSOM method employs nonionizing visible radiation and can be used in air or aqueous environments for nondestructive visualization of functioning biological systems with a resolution comparable to that of scanning electron microscopy. ImagesFIGURE 4FIGURE 7FIGURE 9FIGURE 10 PMID:19431633

  6. Virtual k -Space Modulation Optical Microscopy

    NASA Astrophysics Data System (ADS)

    Kuang, Cuifang; Ma, Ye; Zhou, Renjie; Zheng, Guoan; Fang, Yue; Xu, Yingke; Liu, Xu; So, Peter T. C.

    2016-07-01

    We report a novel superresolution microscopy approach for imaging fluorescence samples. The reported approach, termed virtual k -space modulation optical microscopy (VIKMOM), is able to improve the lateral resolution by a factor of 2, reduce the background level, improve the optical sectioning effect and correct for unknown optical aberrations. In the acquisition process of VIKMOM, we used a scanning confocal microscope setup with a 2D detector array to capture sample information at each scanned x -y position. In the recovery process of VIKMOM, we first modulated the captured data by virtual k -space coding and then employed a ptychography-inspired procedure to recover the sample information and correct for unknown optical aberrations. We demonstrated the performance of the reported approach by imaging fluorescent beads, fixed bovine pulmonary artery endothelial (BPAE) cells, and living human astrocytes (HA). As the VIKMOM approach is fully compatible with conventional confocal microscope setups, it may provide a turn-key solution for imaging biological samples with ˜100 nm lateral resolution, in two or three dimensions, with improved optical sectioning capabilities and aberration correcting.

  7. Photon-induced near field electron microscopy

    NASA Astrophysics Data System (ADS)

    Park, Sang Tae; Zewail, Ahmed H.

    2013-09-01

    Ultrafast electron microscopy in the space and time domains utilizes a pulsed electron probe to directly map structural dynamics of nanomaterials initiated by an optical pump pulse, in imaging, di raction, spectroscopy, and their combinations. It has demonstrated its capability in the studies of phase transitions, mechanical vibrations, and chemical reactions. Moreover, electrons can directly interact with photons via the near eld component of light scattering by nanostructures, and either gain or lose light quanta discretely in energy. By energetically selecting those electrons that exchanged photon energies, we can map this photon-electron interaction, and the technique is termed photon-induced near eld electron microscopy (PINEM). Here, we give an account of the theoretical understanding of PINEM. Experimentally, nanostructures such as a sphere, cylinder, strip, and triangle have been investigated. Theoretically, time-dependent Schrodinger and Dirac equations for an electron under light are directly solved to obtain analytical solutions. The interaction probability is expressed by the mechanical work done by an optical wave on a traveling electron, which can be evaluated analytically by the near eld components of the Rayleigh scattering for small spheres and thin cylinders, and numerically by the discrete dipole approximation for other geometries. Application in visualization of plasmon elds is discussed.

  8. The Bioscience Nuclear Microscopy Program at LLNL

    SciTech Connect

    Bench, G.; Freeman, S.; Roberts, M.; Sideras-Haddad, E.

    1996-12-31

    Since initiation in mid 1994, a bioscience nuclear microscopy program at Livermore has enabled collaboration with bio-scientists on a variety of projects requiring quantitative elemental microanalysis. For microprobe analysis a combination of PIXE and STIM are typically used; respectively generating element distribution maps with micron scale spatial resolution, and projected densities and histological information with sub-micron spatial resolution. Current studies demonstrate the applicability of nuclear microscopy (particularly when combined with other analysis techniques) in environmental tracing, toxicology, carcinogenesis, and structural biology. The program currently uses {approximately}10 percent of the available time on a 10 MV tandem accelerator that is also applied to a variety of Accelerator Mass Spectrometry and other microprobe programs. The completion of a dedicated nuclear microprobe system, using a 5 SDH NEC 1.7 MV tandem accelerator and employing several energy dispersive x-ray detectors to improve x-ray counting rates, promises increased accelerator access, greater sample throughput and continued expansion of the program.

  9. Atomic force microscopy investigation of viruses.

    PubMed

    McPherson, Alexander; Kuznetsov, Yurii G

    2011-01-01

    Atomic force microscopy (AFM) has proven to be a valuable approach to delineate the architectures and detailed structural features of a wide variety of viruses. These have ranged from small plant satellite viruses of only 17 nm to the giant mimivirus of 750 nm diameter, and they have included diverse morphologies such as those represented by HIV, icosahedral particles, vaccinia, and bacteriophages. Because it is a surface technique, it provides images and information that are distinct from those obtained by electron microscopy, and in some cases, at even higher resolution. By enzymatic and chemical dissection of virions, internal structures can be revealed, as well as DNA and RNA. The method is relatively rapid and can be carried out on both fixed and unfixed samples in either air or fluids, including culture media. It is nondestructive and even non-perturbing. It can be applied to individual isolated virus, as well as to infected cells. AFM is still in its early development and holds great promise for further investigation of biological systems at the nanometer scale.

  10. Electrostatic-free piezoresponse force microscopy

    PubMed Central

    Kim, Sungho; Seol, Daehee; Lu, Xiaoli; Alexe, Marin; Kim, Yunseok

    2017-01-01

    Contact and non-contact based atomic force microscopy (AFM) approaches have been extensively utilized to explore various nanoscale surface properties. In most AFM-based measurements, a concurrent electrostatic effect between the AFM tip/cantilever and sample surface can occur. This electrostatic effect often hinders accurate measurements. Thus, it is very important to quantify as well as remove the impact of the electrostatic effect on AFM-based measurements. In this study, we examine the impact of the electrostatic effect on the electromechanical (EM) response in piezoresponse force microscopy as a model AFM mode. We quantitatively studied the effects of increasing the external electric field and reducing the spring constant of a cantilever. Further, we explored ways to minimize the electrostatic effect. The results provide broad guidelines for quantitatively analyzing the EM response as well as, eventually, for obtaining the electrostatic-free EM response. The conclusions can be applied to other AFM-based measurements that are subject to a strong electrostatic effect between the AFM tip/cantilever and sample surface, regardless of contact and non-contact modes. PMID:28139715

  11. Encoded multisite two-photon microscopy

    PubMed Central

    Ducros, Mathieu; Houssen, Yannick Goulam; Bradley, Jonathan; de Sars, Vincent; Charpak, Serge

    2013-01-01

    The advent of scanning two-photon microscopy (2PM) has created a fertile new avenue for noninvasive investigation of brain activity in depth. One principal weakness of this method, however, lies with the limit of scanning speed, which makes optical interrogation of action potential-like activity in a neuronal network problematic. Encoded multisite two-photon microscopy (eMS2PM), a scanless method that allows simultaneous imaging of multiple targets in depth with high temporal resolution, addresses this drawback. eMS2PM uses a liquid crystal spatial light modulator to split a high-power femto-laser beam into multiple subbeams. To distinguish them, a digital micromirror device encodes each subbeam with a specific binary amplitude modulation sequence. Fluorescence signals from all independently targeted sites are then collected simultaneously onto a single photodetector and site-specifically decoded. We demonstrate that eMS2PM can be used to image spike-like voltage transients in cultured cells and fluorescence transients (calcium signals in neurons and red blood cells in capillaries from the cortex) in depth in vivo. These results establish eMS2PM as a unique method for simultaneous acquisition of neuronal network activity. PMID:23798397

  12. Encoded multisite two-photon microscopy.

    PubMed

    Ducros, Mathieu; Goulam Houssen, Yannick; Bradley, Jonathan; de Sars, Vincent; Charpak, Serge

    2013-08-06

    The advent of scanning two-photon microscopy (2PM) has created a fertile new avenue for noninvasive investigation of brain activity in depth. One principal weakness of this method, however, lies with the limit of scanning speed, which makes optical interrogation of action potential-like activity in a neuronal network problematic. Encoded multisite two-photon microscopy (eMS2PM), a scanless method that allows simultaneous imaging of multiple targets in depth with high temporal resolution, addresses this drawback. eMS2PM uses a liquid crystal spatial light modulator to split a high-power femto-laser beam into multiple subbeams. To distinguish them, a digital micromirror device encodes each subbeam with a specific binary amplitude modulation sequence. Fluorescence signals from all independently targeted sites are then collected simultaneously onto a single photodetector and site-specifically decoded. We demonstrate that eMS2PM can be used to image spike-like voltage transients in cultured cells and fluorescence transients (calcium signals in neurons and red blood cells in capillaries from the cortex) in depth in vivo. These results establish eMS2PM as a unique method for simultaneous acquisition of neuronal network activity.

  13. Live cell imaging by multifocal multiphoton microscopy.

    PubMed

    Straub, M; Lodemann, P; Holroyd, P; Jahn, R; Hell, S W

    2000-10-01

    Multifocal multiphoton microscopy (MMM) permits parallel multiphoton excitation by scanning an array of high numerical aperture foci across a plane in the sample. MMM is particularly suitable for live cell investigations since it combines advantages of standard multiphoton microscopy such as optical sectioning and suppression of out-of-focus phototoxicity with high recording speeds. Here we describe several applications of MMM to live cell imaging using the neuroendocrine cell line PC12 and bovine chromaffin cells. Stainings were performed with the acidophilic dye acridine orange and the lipophilic dyes FM1-43 and Fast DiA as well as by transfection of the cells with GFP. In both bovine chromaffin and PC12 cells structural elements of nuclear chromatin and the 3-D distribution of acidic organelles inside the cells were visualized. In PC12 cells differentiated by nerve growth factor examples of neurites were monitored. Stainings of membranes were used to reconstruct the morphology of cells and neurites in three dimensions by volume-rendering and by isosurface plots. 3-D reconstructions were composed from stacks of about 50 images each with a diameter of 30-100 microm that were acquired within a few seconds. We conclude that MMM proves to be a technically simple and very effective method for fast 3-D live cell imaging at high resolution.

  14. Confocal microscopy of Aspergillus fumigatus keratitis

    PubMed Central

    Avunduk, A M; Beuerman, R W; Varnell, E D; Kaufman, H E

    2003-01-01

    Aim: To use a confocal microscope to characterise the treated and untreated courses of fungal keratitis. Methods: In the first experiment, Aspergillus fumigatus stromal keratitis was produced in both eyes of seven New Zealand white rabbits. In the second experiment, keratitis was induced in right eyes of 20 rabbits. Group 1 rabbits were treated with topical fluconazole, group 2 rabbits received oral fluconazole, and group 3 rabbits were used as controls. The rabbits were examined with a slit lamp and confocal microscope 2, 6, 10, 14, and 20 days after inoculation. The corneal cultures were taken on days 2, 14, and 20 and biopsies were taken on days 2 and 22. Results: On days 14 and 22 confocal microscopy was more sensitive than culture technique in both treated and untreated animals, since not all cases of fungal keratitis can be cultured. Conclusion: This study indicates that confocal microscopy is a rapid and sensitive diagnostic tool for both the early diagnosis and non-invasive follow up of fungal keratitis PMID:12642300

  15. Image Quality Ranking Method for Microscopy

    NASA Astrophysics Data System (ADS)

    Koho, Sami; Fazeli, Elnaz; Eriksson, John E.; Hänninen, Pekka E.

    2016-07-01

    Automated analysis of microscope images is necessitated by the increased need for high-resolution follow up of events in time. Manually finding the right images to be analyzed, or eliminated from data analysis are common day-to-day problems in microscopy research today, and the constantly growing size of image datasets does not help the matter. We propose a simple method and a software tool for sorting images within a dataset, according to their relative quality. We demonstrate the applicability of our method in finding good quality images in a STED microscope sample preparation optimization image dataset. The results are validated by comparisons to subjective opinion scores, as well as five state-of-the-art blind image quality assessment methods. We also show how our method can be applied to eliminate useless out-of-focus images in a High-Content-Screening experiment. We further evaluate the ability of our image quality ranking method to detect out-of-focus images, by extensive simulations, and by comparing its performance against previously published, well-established microscopy autofocus metrics.

  16. Sewage coliphages studied by electron microscopy.

    PubMed Central

    Ackermann, H W; Nguyen, T M

    1983-01-01

    Sewage was enriched with 35 Escherichia coli strains, and sediments of enrichment cultures were studied in the electron microscope. They contained up to 10 varieties of morphologically different particles. T-even-type phages predominated in 14 samples. Thirteen phages were enriched, representing the families Myoviridae (seven), Styloviridae (two), Podoviridae (three), and Microviridae (one). Twelve of these corresponded to known enterobacterial phage species, namely, 121, K19, FC3-9, O1, 9266, T2, 16-19, kappa, beta 4, N4, T7, and phi X174. Cubic RNA phages and filamentous phages were not detected. Types 121 and 9266 have previously been observed only in Romania and South Africa. Identification by morphology is usually simple. Our investigative technique is qualitative and will not detect all phages present. Most enrichment strains are polyvalent, and electron microscopy is always required for phage identification. In a general way, electron microscopy seems to be the method of choice for investigation of phage geography and ecology. Images PMID:6847179

  17. Phase-contrast scanning transmission electron microscopy.

    PubMed

    Minoda, Hiroki; Tamai, Takayuki; Iijima, Hirofumi; Hosokawa, Fumio; Kondo, Yukihito

    2015-06-01

    This report introduces the first results obtained using phase-contrast scanning transmission electron microscopy (P-STEM). A carbon-film phase plate (PP) with a small center hole is placed in the condenser aperture plane so that a phase shift is introduced in the incident electron waves except those passing through the center hole. A cosine-type phase-contrast transfer function emerges when the phase-shifted scattered waves interfere with the non-phase-shifted unscattered waves, which passed through the center hole before incidence onto the specimen. The phase contrast resulting in P-STEM is optically identical to that in phase-contrast transmission electron microscopy that is used to provide high contrast for weak phase objects. Therefore, the use of PPs can enhance the phase contrast of the STEM images of specimens in principle. The phase shift resulting from the PP, whose thickness corresponds to a phase shift of π, has been confirmed using interference fringes displayed in the Ronchigram of a silicon single crystal specimen. The interference fringes were found to abruptly shift at the edge of the PP hole by π.

  18. Full information acquisition in piezoresponse force microscopy

    SciTech Connect

    Somnath, Suhas Belianinov, Alexei E-mail: sergei2@ornl.gov Kalinin, Sergei V. E-mail: sergei2@ornl.gov Jesse, Stephen E-mail: sergei2@ornl.gov

    2015-12-28

    The information flow from the tip-surface junction to the detector electronics during the piezoresponse force microscopy (PFM) imaging is explored using the recently developed general mode (G-mode) detection. Information-theory analysis suggests that G-mode PFM in the non-switching regime, close to the first resonance mode, contains a relatively small (100–150) number of components containing significant information. The first two primary components are similar to classical PFM images, suggesting that classical lock-in detection schemes provide high veracity information in this case. At the same time, a number of transient components exhibit contrast associated with surface topography, suggesting pathway to separate the two. The number of significant components increases considerably in the non-linear and switching regimes and approaching cantilever resonances, precluding the use of classical lock-in detection and necessitating the use of band excitation or G-mode detection schemes. The future prospects of full information imaging in scanning probe microscopy are discussed.

  19. Magnetoelectric force microscopy based on magnetic force microscopy with modulated electric field.

    PubMed

    Geng, Yanan; Wu, Weida

    2014-05-01

    We present the realization of a mesoscopic imaging technique, namely, the Magnetoelectric Force Microscopy (MeFM), for visualization of local magnetoelectric effect. The basic principle of MeFM is the lock-in detection of local magnetoelectric response, i.e., the electric field-induced magnetization, using magnetic force microscopy. We demonstrate MeFM capability by visualizing magnetoelectric domains on single crystals of multiferroic hexagonal manganites. Results of several control experiments exclude artifacts or extrinsic origins of the MeFM signal. The parameters are tuned to optimize the signal to noise ratio.

  20. Characterization of Softmagnetic Thin Layers Using Barkhausen Noise Microscopy

    DTIC Science & Technology

    2001-04-01

    Barkhausen Noise Microscopy DISTRIBUTION: Approved for public release, distribution unlimited This paper is part of the following report: TITLE: Applications...Proc. Vol. 674 © 2001 Materials Research Society Characterization of Softmagnetic Thin Layers using Barkhausen Noise Microscopy Jochen Hoffmann...layer system. Barkhausen Noise Microscopy enables the characterization of such thin layers. By cycling the magnetic hysteresis of ferromagnetic material

  1. Accessible Microscopy Workstation for Students and Scientists with Mobility Impairments

    ERIC Educational Resources Information Center

    Duerstock, Bradley S.

    2006-01-01

    An integrated accessible microscopy workstation was designed and developed to allow persons with mobility impairments to control all aspects of light microscopy with minimal human assistance. This system, named AccessScope, is capable of performing brightfield and fluorescence microscopy, image analysis, and tissue morphometry requisite for…

  2. M-DAS: System for multispectral data analysis. [in Saginaw Bay, Michigan

    NASA Technical Reports Server (NTRS)

    Johnson, R. H.

    1975-01-01

    M-DAS is a ground data processing system designed for analysis of multispectral data. M-DAS operates on multispectral data from LANDSAT, S-192, M2S and other sources in CCT form. Interactive training by operator-investigators using a variable cursor on a color display was used to derive optimum processing coefficients and data on cluster separability. An advanced multivariate normal-maximum likelihood processing algorithm was used to produce output in various formats: color-coded film images, geometrically corrected map overlays, moving displays of scene sections, coverage tabulations and categorized CCTs. The analysis procedure for M-DAS involves three phases: (1) screening and training, (2) analysis of training data to compute performance predictions and processing coefficients, and (3) processing of multichannel input data into categorized results. Typical M-DAS applications involve iteration between each of these phases. A series of photographs of the M-DAS display are used to illustrate M-DAS operation.

  3. Nonlinear Polarimetric Microscopy for Biomedical Imaging

    NASA Astrophysics Data System (ADS)

    Samim, Masood

    A framework for the nonlinear optical polarimetry and polarimetric microscopy is developed. Mathematical equations are derived in terms of linear and nonlinear Stokes Mueller formalism, which comprehensively characterize the polarization properties of the incoming and outgoing radiations, and provide structural information about the organization of the investigated materials. The algebraic formalism developed in this thesis simplifies many predictions for a nonlinear polarimetry study and provides an intuitive understanding of various polarization properties for radiations and the intervening medium. For polarimetric microscopy experiments, a custom fast-scanning differential polarization microscope is developed, which is also capable of real-time three-dimensional imaging. The setup is equipped with a pair of high-speed resonant and galvanometric scanning mirrors, and supplemented by advanced adaptive optics and data acquisition modules. The scanning mirrors when combined with the adaptive optics deformable mirror enable fast 3D imaging. Deformable membrane mirrors and genetic algorithm optimization routines are employed to improve the imaging conditions including correcting the optical aberrations, maximizing signal intensities, and minimizing point-spread-functions of the focal volume. A field-programmable-gate array (FPGA) chip is exploited to rapidly acquire and process the multidimensional data. Using the nonlinear optical polarimetry framework and the home-built polarization microscope, a few biologically important tissues are measured and analyzed to gain insight as to their structure and dynamics. The structure and distribution of muscle sarcomere myosins, connective tissue collagen, carbohydrate-rich starch, and fruit fly eye retinal molecules are characterized with revealing polarization studies. In each case, using the theoretical framework, polarization sensitive data are analyzed to decipher the molecular orientations and nonlinear optical

  4. Soft X-Ray Microscopy Radiation Damage On Fixed Cells Investigated With Synchrotron Radiation FTIR Microscopy.

    PubMed

    Gianoncelli, A; Vaccari, L; Kourousias, G; Cassese, D; Bedolla, D E; Kenig, S; Storici, P; Lazzarino, M; Kiskinova, M

    2015-05-14

    Radiation damage of biological samples remains a limiting factor in high resolution X-ray microscopy (XRM). Several studies have attempted to evaluate the extent and the effects of radiation damage, proposing strategies to minimise or prevent it. The present work aims to assess the impact of soft X-rays on formalin fixed cells on a systematic manner. The novelty of this approach resides on investigating the radiation damage not only with XRM, as often reported in relevant literature on the topic, but by coupling it with two additional independent non-destructive microscopy methods: Atomic Force Microscopy (AFM) and FTIR Microscopy (FTIRM). Human Embryonic Kidney 293 cells were exposed to different radiation doses at 1 keV. In order to reveal possible morphological and biochemical changes, the irradiated cells were systematically analysed with AFM and FTIRM before and after. Results reveal that while cell morphology is not substantially affected, cellular biochemical profile changes significantly and progressively when increasing dose, resulting in a severe breakdown of the covalent bonding network. This information impacts most soft XRM studies on fixed cells and adds an in-depth understanding of the radiation damage for developing better prevention strategies.

  5. Improved axial resolution of FINCH fluorescence microscopy when combined with spinning disk confocal microscopy

    PubMed Central

    Siegel, Nisan; Brooker, Gary

    2014-01-01

    FINCH holographic fluorescence microscopy creates super-resolved images with enhanced depth of focus. Addition of a Nipkow disk real-time confocal image scanner is shown to reduce the FINCH depth of focus while improving transverse confocal resolution in a combined method called “CINCH”. PMID:25321701

  6. Photoelectron microscopy and immunofluorescence microscopy of cytoskeletal elements in the same cells.

    PubMed Central

    Nadakavukaren, K K; Chen, L B; Habliston, D L; Griffith, O H

    1983-01-01

    Pt K2 rat kangaroo epithelial cells and Rat-1 fibroblasts were grown on conductive glass discs, fixed, and permeabilized, and the cytoskeletal elements actin, keratin, and vimentin were visualized by indirect immunofluorescence. After the fluorescence microscopy, the cells were postfixed and dehydrated for photoelectron microscopy. The contrast in these photoelectron micrographs is primarily topographical in origin, and the presence of fluorescent dyes at low density does not contribute significantly to the material contrast. By comparison with fluorescence micrographs obtained on the same individual cells, actin-containing stress fibers, keratin filaments, and vimentin filaments were identified in the photoelectron micrographs. The apparent volume occupied by the cytoskeletal network in the cells as judged from the photoelectron micrographs is much less than it appears to be from the fluorescence micrographs because the higher resolution of photoelectron microscopy shows the fibers closer to their true dimensions. Photoelectron microscopy is a surface technique, and the images highlight the exposed cytoskeletal structures and suppress those extending along the substrate below the nuclei. The results reported here show marked improvement in image quality of photoelectron micrographs and that this technique has the potential of contributing to higher resolution studies of cytoskeletal structures. Images PMID:6191327

  7. Application of scanning acoustic microscopy to advanced structural ceramics

    NASA Technical Reports Server (NTRS)

    Vary, Alex; Klima, Stanley J.

    1987-01-01

    A review is presentod of research investigations of several acoustic microscopy techniques for application to structural ceramics for advanced heat engines. Results obtained with scanning acoustic microscopy (SAM), scanning laser acoustic microscopy (SLAM), scanning electron acoustic microscopy (SEAM), and photoacoustic microscopy (PAM) are compared. The techniques were evaluated on research samples of green and sintered monolithic silicon nitrides and silicon carbides in the form of modulus-of-rupture bars containing deliberately introduced flaws. Strengths and limitations of the techniques are described with emphasis on statistics of detectability of flaws that constitute potential fracture origins.

  8. 3D super-resolution imaging by localization microscopy.

    PubMed

    Magenau, Astrid; Gaus, Katharina

    2015-01-01

    Fluorescence microscopy is an important tool in all fields of biology to visualize structures and monitor dynamic processes and distributions. Contrary to conventional microscopy techniques such as confocal microscopy, which are limited by their spatial resolution, super-resolution techniques such as photoactivated localization microscopy (PALM) and stochastic optical reconstruction microscopy (STORM) have made it possible to observe and quantify structure and processes on the single molecule level. Here, we describe a method to image and quantify the molecular distribution of membrane-associated proteins in two and three dimensions with nanometer resolution.

  9. Comparison of multi-mode parallel detection microscopy methods

    NASA Astrophysics Data System (ADS)

    Zhu, Dazhao; Fang, Yue; Chen, Youhua; Hussain, Anwar; Kuang, Cuifang; Ding, Zhihua; Liu, Xu

    2017-03-01

    Four microscopy resolution enhancement methods based on parallel detection were investigated in this study: confocal microscopy with four pinhole sizes, fluorescence emission difference microscopy (FED) based on parallel detection, Airyscan microscopy, and virtual k-vector modulation optical microscopy (Vikmom). These methods use different algorithms to process parallel detection data and achieve resolution improvement. We investigated these methods first by performing simulations and then experimentally. In this report, the basic theories of these methods are briefly introduced. Then, analyses and comparisons of their imaging performances, especially in terms of resolution improvement, imaging speed, and signal-to-noise ratio, are presented. Finally, the results of our comparative study are summarized.

  10. Improved methods for high resolution electron microscopy

    SciTech Connect

    Taylor, J.R.

    1987-04-01

    Existing methods of making support films for high resolution transmission electron microscopy are investigated and novel methods are developed. Existing methods of fabricating fenestrated, metal reinforced specimen supports (microgrids) are evaluated for their potential to reduce beam induced movement of monolamellar crystals of C/sub 44/H/sub 90/ paraffin supported on thin carbon films. Improved methods of producing hydrophobic carbon films by vacuum evaporation, and improved methods of depositing well ordered monolamellar paraffin crystals on carbon films are developed. A novel technique for vacuum evaporation of metals is described which is used to reinforce microgrids. A technique is also developed to bond thin carbon films to microgrids with a polymer bonding agent. Unique biochemical methods are described to accomplish site specific covalent modification of membrane proteins. Protocols are given which covalently convert the carboxy terminus of papain cleaved bacteriorhodopsin to a free thiol. 53 refs., 19 figs., 1 tab.

  11. Nonlinear femtosecond laser induced scanning tunneling microscopy.

    PubMed

    Dey, Shirshendu; Mirell, Daniel; Perez, Alejandro Rodriguez; Lee, Joonhee; Apkarian, V Ara

    2013-04-21

    We demonstrate ultrafast laser driven nonlinear scanning tunneling microscopy (STM), under ambient conditions. The design is an adaptation of the recently introduced cross-polarized double beat method, whereby z-polarized phase modulated fields are tightly focused at a tunneling junction consisting of a sharp tungsten tip and an optically transparent gold film as substrate. We demonstrate the prerequisites for ultrafast time-resolved STM through an operative mechanism of nonlinear laser field-driven tunneling. The spatial resolution of the nonlinear laser driven STM is determined by the local field intensity. Resolution of 0.3 nm-10 nm is demonstrated for the intensity dependent, exponential tunneling range. The demonstration is carried out on a junction consisting of tungsten tip and gold substrate. Nano-structured gold is used for imaging purposes, to highlight junction plasmon controlled tunneling in the conductivity limit.

  12. Thermal expansion recovery microscopy: Practical design considerations

    SciTech Connect

    Mingolo, N. Martínez, O. E.

    2014-01-15

    A detailed study of relevant parameters for the design and operation of a photothermal microscope technique recently introduced is presented. The technique, named thermal expansion recovery microscopy (ThERM) relies in the measurement of the defocusing introduced by a surface that expands and recovers upon the heating from a modulated source. A new two lens design is presented that can be easily adapted to commercial infinite conjugate microscopes and the sensitivity to misalignment is analyzed. The way to determine the beam size by means of a focus scan and the use of that same scan to verify if a thermoreflectance signal is overlapping with the desired ThERM mechanism are discussed. Finally, a method to cancel the thermoreflectance signal by an adequate choice of a nanometric coating is presented.

  13. Local tomographic phase microscopy from differential projections

    NASA Astrophysics Data System (ADS)

    Vishnyakov, G. N.; Levin, G. G.; Minaev, V. L.; Nekrasov, N. A.

    2016-12-01

    It is proposed to use local tomography for optical studies of the internal structure of transparent phase microscopic objects, for example, living cells. From among the many local tomography methods that exist, the algorithms of back projection summation (in which partial derivatives of projections are used as projection data) are chosen. The application of local tomography to living cells is reasonable because, using optical phase microscopy, one can easily obtain projection data in the form of first-order derivatives of projections applying the methods of differential interference contrast and shear interferometry. The mathematical fundamentals of local tomography in differential projections are considered, and a computer simulation of different local tomography methods is performed. A tomographic phase microscope and the results of reconstructing a local tomogram of an erythrocyte from a set of experimental differential projections are described.

  14. Halo-free Phase Contrast Microscopy.

    PubMed

    Nguyen, Tan H; Kandel, Mikhail; Shakir, Haadi M; Best-Popescu, Catherine; Arikkath, Jyothi; Do, Minh N; Popescu, Gabriel

    2017-03-24

    We present a new approach for retrieving halo-free phase contrast microscopy (hfPC) images by upgrading the conventional PC microscope with an external interferometric module, which generates sufficient data for reversing the halo artifact. Acquiring four independent intensity images, our approach first measures haloed phase maps of the sample. We solve for the halo-free sample transmission function by using a physical model of the image formation under partial spatial coherence. Using this halo-free sample transmission, we can numerically generate artifact-free PC images. Furthermore, this transmission can be further used to obtain quantitative information about the sample, e.g., the thickness with known refractive indices, dry mass of live cells during their cycles. We tested our hfPC method on various control samples, e.g., beads, pillars and validated its potential for biological investigation by imaging live HeLa cells, red blood cells, and neurons.

  15. Inherent error in interferometric surface plasmon microscopy

    NASA Astrophysics Data System (ADS)

    Zhang, Bei; Yan, Peng; Gao, Feng; Liu, Yu; Zhang, Qiancheng; Wang, Le

    2016-11-01

    Surface plasmon microscopy (SPRM) usually employs high refractive index prism or high numerical aperture (NA) objective as coupling device to excite surface plasmon. Here we apply high NA oil-immersion objective considering k vector conditions of SPs and localization of SPs which provides better lateral resolution and less cross-talk between adjacent areas. However, performance of an objective based SPRM is often limited by the finite aperture of a physical objective which corresponds to sudden transition and limited bandwidth. Here we give a simplified model of the SPRM and numerically calculate how the sudden transition on the clear aperture edge causes inherent error. Notch filtering algorithm is designed to suppress the noisy ripples. Compared to the pupil function engineering technique, this technique makes both the sacrifice of NA and utilization of spatial light modulator unnecessary and provides a more compact system setup without decreasing the resolution and contrast.

  16. Scanning electron microscopy of tinea nigra.

    PubMed

    Guarenti, Isabelle Maffei; Almeida, Hiram Larangeira de; Leitão, Aline Hatzenberger; Rocha, Nara Moreira; Silva, Ricardo Marques E

    2014-01-01

    Tinea nigra is a rare superficial mycosis caused by Hortaea werneckii. This infection presents as asymptomatic brown to black maculae mostly in palmo-plantar regions. We performed scanning electron microscopy of a superficial shaving of a tinea nigra lesion. The examination of the outer surface of the sample showed the epidermis with corneocytes and hyphae and elimination of fungal filaments. The inner surface of the sample showed important aggregation of hyphae among keratinocytes, which formed small fungal colonies. The ultrastructural findings correlated with those of dermoscopic examination - the small fungal aggregations may be the dark spicules seen on dermoscopy - and also allowed to document the mode of dissemination of tinea nigra, showing how hyphae are eliminated on the surface of the lesion.

  17. Scanning electron microscopy of molluscum contagiosum*

    PubMed Central

    de Almeida Jr, Hiram Larangeira; Abuchaim, Martha Oliveira; Schneider, Maiko Abel; Marques, Leandra; de Castro, Luis Antônio Suíta

    2013-01-01

    Molluscum contagiosum is a disease caused by a poxvirus. It is more prevalent in children up to 5 years of age. There is a second peak of incidence in young adults. In order to examine its ultrastructure, three lesions were curetted without disruption, cut transversely with a scalpel, and routinely processed for scanning electron microscopy (SEM). The oval structure of molluscum contagiosum could be easily identified. In its core, there was a central umbilication and just below this depression, there was a keratinized tunnel. Under higher magnification, a proliferation similar to the epidermis was seen. Moreover, there were areas of cells disposed like a mosaic. Under higher magnification, rounded structures measuring 0.4 micron could be observed at the end of the keratinized tunnel and on the surface of the lesion. PMID:23539009

  18. Noninvasive photoacoustic microscopy of methemoglobin in vivo

    NASA Astrophysics Data System (ADS)

    Tang, Min; Zhou, Yong; Zhang, Ruiying; Wang, Lihong V.

    2015-03-01

    Various causes can lead to methemoglobinemia, and it has the potential to be confused with other diseases. In vivo measurements of methemoglobin have significant applications in the clinics. We quantified the average and the distributed percentage of methemoglobin both in vitro and in vivo using photoacoustic microscopy (PAM). Based on the absorption spectra of methemoglobin, oxyhemoglobin, and deoxyhemoglobin, three wavelengths were chosen to differentiate methemoglobin from the others. We imaged the methemoglobin percentage in microtubes that mimicked blood vessels as a phantom experiment. The methemoglobin concentrations calculated from the photoacoustic signals were in accordance with the preset concentrations. We also demonstrated the ability of PAM to quantitatively image methemoglobin distribution in vivo in a mouse ear.

  19. Atomic Force Microscopy for DNA SNP Identification

    NASA Astrophysics Data System (ADS)

    Valbusa, Ugo; Ierardi, Vincenzo

    The knowledge of the effects of single-nucleotide polymorphisms (SNPs) in the human genome greatly contributes to better comprehension of the relation between genetic factors and diseases. Sequence analysis of genomic DNA in different individuals reveals positions where variations that involve individual base substitutions can occur. Single-nucleotide polymorphisms are highly abundant and can have different consequences at phenotypic level. Several attempts were made to apply atomic force microscopy (AFM) to detect and map SNP sites in DNA strands. The most promising approach is the study of DNA mutations producing heteroduplex DNA strands and identifying the mismatches by means of a protein that labels the mismatches. MutS is a protein that is part of a well-known complex of mismatch repair, which initiates the process of repairing when the MutS binds to the mismatched DNA filament. The position of MutS on the DNA filament can be easily recorded by means of AFM imaging.

  20. Recent applications of superresolution microscopy in neurobiology.

    PubMed

    Willig, Katrin I; Barrantes, Francisco J

    2014-06-01

    Chemical synapses in brain are structural differentiations where excitatory or inhibitory signals are vectorially transmitted between two neurons. Excitatory synapses occur mostly on dendritic spines, submicron sized protrusions of the neuronal dendritic arborizations. Axons establish contacts with these tiny specializations purported to be the smallest functional processing units in the central nervous system. The minute size of synapses and their macromolecular constituents creates an inherent difficulty for imaging but makes them an ideal object for superresolution microscopy. Here we discuss some representative examples of nanoscopy studies, ranging from quantification of receptors and scaffolding proteins in postsynaptic densities and their dynamic behavior, to imaging of synaptic vesicle proteins and dendritic spines in living neurons or even live animals.

  1. Inspection of surface flaws by comparator microscopy.

    PubMed

    Baker, L R

    1988-11-15

    The increasing attention paid in recent years to control of surface quality has exposed the lack of objectivity of existing standards relating to flaws such as digs and scratches. The present requirements of the customer and supplier of optical components for improved standards are discussed, and recent attempts to satisfy these requirements are reviewed. It is concluded that a technique, described as comparator microscopy, in which a width of line or slit is identified which removes from a transmitted or reflected beam the same amount of light as the flaw under examination, has much in its favor in terms of user needs. The method, which is under consideration as the basis of an ISO standard, is described, and results are presented indicating how this concept of line-equivalent width can be used to compare different national standard flaws. Extending this technique to measure flaws and polish on-machine is suggested as a future possibility.

  2. Nuclear microscopy of sperm cell elemental structure

    SciTech Connect

    Bench, G.S.

    1994-12-31

    Theories have suggested that there is a link between protamine concentrations in individual sperm and sperm fertility. At present, biochemical analyses have only been performed on bulk populations and existing methods have not been able to determine what percentage of morphologically normal sperm are biochemically defective and potentially infertile. As part of an investigation into male sperm fertility, nuclear microscopy has been utilized to measure elemental profiles at the single sperm level. By measuring the ratio of Phosphorus to Sulfur the authors have been able to determine the amount of protamine 1 and protamine 2 in individual cells from bulk fertile samples of bull and mouse sperm. Preliminary results show that, for each species, the relative amounts of protamine 1 and protamine 2 in morphologically normal sperm agree well with expected values.

  3. Monitoring photodynamic therapy with photoacoustic microscopy

    NASA Astrophysics Data System (ADS)

    Shao, Peng; Chapman, David W.; Moore, Ronald B.; Zemp, Roger J.

    2015-10-01

    We present our work on examining the feasibility of monitoring photodynamic therapy (PDT)-induced vasculature change with acoustic-resolution photoacoustic microscopy (PAM). Verteporfin, an FDA-approved photosensitizer for clinical PDT, was utilized. With a 60-μm-resolution PAM system, we demonstrated the capability of PAM to monitor PDT-induced vasculature variations in a chick chorioallantoic membrane model with topical application and in a rat ear with intravenous injection of the photosensitizer. We also showed oxygen saturation change in target blood vessels due to PDT. Success of the present approach may potentially lead to the application of PAM imaging in evaluating PDT efficacy, guiding treatment, and predicting responders from nonresponders.

  4. Photoactivated Localization Microscopy (PALM) of Adhesion Complexes

    PubMed Central

    Shroff, Hari; White, Helen; Betzig, Eric

    2017-01-01

    Key to understanding a protein’s biological function is the accurate determination of its spatial distribution inside a cell. Although fluorescent protein markers allow the targeting of specific proteins with molecular precision, much of this information is lost when the resultant fusion proteins are imaged with conventional, diffraction-limited optics. In response, several imaging modalities that are capable of resolution below the diffraction limit (~200 nm) have emerged. Here, both single- and dual-color superresolution imaging of biological structures using photoactivated localization microscopy (PALM) are described. The examples discussed focus on adhesion complexes: dense, protein-filled assemblies that form at the interface between cells and their substrata. A particular emphasis is placed on the instrumentation and photoactivatable fluorescent protein (PA-FP) tags necessary to achieve PALM images at ~20 nm resolution in 5 to 30 min in fixed cells. PMID:23456603

  5. Scanning electron microscopy studies of bacterial cultures

    NASA Astrophysics Data System (ADS)

    Swinger, Tracy; Blust, Brittni; Calabrese, Joseph; Tzolov, Marian

    2012-02-01

    Scanning electron microscopy is a powerful tool to study the morphology of bacteria. We have used conventional scanning electron microscope to follow the modification of the bacterial morphology over the course of the bacterial growth cycle. The bacteria were fixed in vapors of Glutaraldehyde and ruthenium oxide applied in sequence. A gold film of about 5 nm was deposited on top of the samples to avoid charging and to enhance the contrast. We have selected two types of bacteria Alcaligenes faecalis and Kocuria rhizophila. Their development was carefully monitored and samples were taken for imaging in equal time intervals during their cultivation. These studies are supporting our efforts to develop an optical method for identification of the Gram-type of bacterial cultures.

  6. Scanning Tunneling Microscopy Observation of Phonon Condensate.

    PubMed

    Altfeder, Igor; Voevodin, Andrey A; Check, Michael H; Eichfeld, Sarah M; Robinson, Joshua A; Balatsky, Alexander V

    2017-02-22

    Using quantum tunneling of electrons into vibrating surface atoms, phonon oscillations can be observed on the atomic scale. Phonon interference patterns with unusually large signal amplitudes have been revealed by scanning tunneling microscopy in intercalated van der Waals heterostructures. Our results show that the effective radius of these phonon quasi-bound states, the real-space distribution of phonon standing wave amplitudes, the scattering phase shifts, and the nonlinear intermode coupling strongly depend on the presence of defect-induced scattering resonance. The observed coherence of these quasi-bound states most likely arises from phase- and frequency-synchronized dynamics of all phonon modes, and indicates the formation of many-body condensate of optical phonons around resonant defects. We found that increasing the strength of the scattering resonance causes the increase of the condensate droplet radius without affecting the condensate fraction inside it. The condensate can be observed at room temperature.

  7. Three-photon excitation in fluorescence microscopy

    NASA Astrophysics Data System (ADS)

    Hell, Stefan W.; Bahlmann, Karsten; Schrader, Martin; Soini, Aleksi; Malak, Henryk; Gryczynski, Ignacy; Lakowicz, Joseph R.

    1996-01-01

    We show experiments proving the feasibility of scanning fluorescence microscopy by three-photon excitation. Three-photon excitation fluorescence axial images are shown of polystyrene beads stained with the fluorophore 2,5- bis(4-biphenyl)oxazole (BBO). Three-photon excitation is performed at an excitation wavelength of 900 nm and with pulses of 130 fs duration provided by a mode-locked titanium-sapphire laser. Fluorescence is collected between 350 and 450 nm. The fluorescence image signal features a third-order dependence on the excitation power, also providing intrinsic 3-D imaging. The resolution of a three-photon excitation microscope is increased over that of a comparable two-photon excitation microscope.

  8. Scanning Tunneling Microscopy Observation of Phonon Condensate

    PubMed Central

    Altfeder, Igor; Voevodin, Andrey A.; Check, Michael H.; Eichfeld, Sarah M.; Robinson, Joshua A.; Balatsky, Alexander V.

    2017-01-01

    Using quantum tunneling of electrons into vibrating surface atoms, phonon oscillations can be observed on the atomic scale. Phonon interference patterns with unusually large signal amplitudes have been revealed by scanning tunneling microscopy in intercalated van der Waals heterostructures. Our results show that the effective radius of these phonon quasi-bound states, the real-space distribution of phonon standing wave amplitudes, the scattering phase shifts, and the nonlinear intermode coupling strongly depend on the presence of defect-induced scattering resonance. The observed coherence of these quasi-bound states most likely arises from phase- and frequency-synchronized dynamics of all phonon modes, and indicates the formation of many-body condensate of optical phonons around resonant defects. We found that increasing the strength of the scattering resonance causes the increase of the condensate droplet radius without affecting the condensate fraction inside it. The condensate can be observed at room temperature. PMID:28225066

  9. Atomic Force Microscopy of Biological Membranes

    PubMed Central

    Frederix, Patrick L.T.M.; Bosshart, Patrick D.; Engel, Andreas

    2009-01-01

    Abstract Atomic force microscopy (AFM) is an ideal method to study the surface topography of biological membranes. It allows membranes that are adsorbed to flat solid supports to be raster-scanned in physiological solutions with an atomically sharp tip. Therefore, AFM is capable of observing biological molecular machines at work. In addition, the tip can be tethered to the end of a single membrane protein, and forces acting on the tip upon its retraction indicate barriers that occur during the process of protein unfolding. Here we discuss the fundamental limitations of AFM determined by the properties of cantilevers, present aspects of sample preparation, and review results achieved on reconstituted and native biological membranes. PMID:19167286

  10. Improved methods for high resolution electron microscopy

    NASA Astrophysics Data System (ADS)

    Taylor, J. R.

    1987-04-01

    Existing methods of making support films for high resolution transmission electron microscopy are investigated and novel methods are developed. Existing methods of fabricating fenestrated, metal reinforced specimen supports (microgrids) are evaluated for their potential to reduce beam induced movement of monolamellar crystals of C44H90 paraffin supported on thin carbon films. Improved methods of producing hydrophobic carbon films by vacuum evaporation, and improved methods of depositing well ordered monolamellar paraffin crystals on carbon films are developed. A novel technique for vacuum evaporation of metals is described which is used to reinforce microgrids. A technique is also developed to bond thin carbon films to microgrids with a polymer bonding agent. Unique biochemical methods are described to accomplish site specific covalent modification of membrane proteins. Protocols are given which covalently convert the carboxy terminus of papain cleaved bacteriorhodopsin to a free thiol.

  11. 4D electron microscopy: principles and applications.

    PubMed

    Flannigan, David J; Zewail, Ahmed H

    2012-10-16

    The transmission electron microscope (TEM) is a powerful tool enabling the visualization of atoms with length scales smaller than the Bohr radius at a factor of only 20 larger than the relativistic electron wavelength of 2.5 pm at 200 keV. The ability to visualize matter at these scales in a TEM is largely due to the efforts made in correcting for the imperfections in the lens systems which introduce aberrations and ultimately limit the achievable spatial resolution. In addition to the progress made in increasing the spatial resolution, the TEM has become an all-in-one characterization tool. Indeed, most of the properties of a material can be directly mapped in the TEM, including the composition, structure, bonding, morphology, and defects. The scope of applications spans essentially all of the physical sciences and includes biology. Until recently, however, high resolution visualization of structural changes occurring on sub-millisecond time scales was not possible. In order to reach the ultrashort temporal domain within which fundamental atomic motions take place, while simultaneously retaining high spatial resolution, an entirely new approach from that of millisecond-limited TEM cameras had to be conceived. As shown below, the approach is also different from that of nanosecond-limited TEM, whose resolution cannot offer the ultrafast regimes of dynamics. For this reason "ultrafast electron microscopy" is reserved for the field which is concerned with femtosecond to picosecond resolution capability of structural dynamics. In conventional TEMs, electrons are produced by heating a source or by applying a strong extraction field. Both methods result in the stochastic emission of electrons, with no control over temporal spacing or relative arrival time at the specimen. The timing issue can be overcome by exploiting the photoelectric effect and using pulsed lasers to generate precisely timed electron packets of ultrashort duration. The spatial and temporal resolutions

  12. High-frequency multimodal atomic force microscopy

    PubMed Central

    Nievergelt, Adrian P; Adams, Jonathan D; Odermatt, Pascal D

    2014-01-01

    Summary Multifrequency atomic force microscopy imaging has been recently demonstrated as a powerful technique for quickly obtaining information about the mechanical properties of a sample. Combining this development with recent gains in imaging speed through small cantilevers holds the promise of a convenient, high-speed method for obtaining nanoscale topography as well as mechanical properties. Nevertheless, instrument bandwidth limitations on cantilever excitation and readout have restricted the ability of multifrequency techniques to fully benefit from small cantilevers. We present an approach for cantilever excitation and deflection readout with a bandwidth of 20 MHz, enabling multifrequency techniques extended beyond 2 MHz for obtaining materials contrast in liquid and air, as well as soft imaging of delicate biological samples. PMID:25671141

  13. Microlymphatic flow using fast video microscopy

    NASA Astrophysics Data System (ADS)

    Dixon, J. B.; Zawieja, David C.; Gashev, Anatoly A.; Cote, Gerard L.

    2005-03-01

    Despite advances in the measurement of lymphatic flow, little is known about the actual velocities of flow in microlymphatic (~100 um diameter) vessels. In this paper, video microscopy and particle tracking methods were adapted and integrated with an ultra high-speed imaging camera to obtain measurements of high-speed lymph velocities that previous systems were incapable of measuring. In this study, a mesenteric microlymphatic vessel in a loop of the small intestine of a male Sprague-Dawley rat was exteriorized and imaged at a rate of 500 frames per second (fps) for several contraction sequences. Lymph velocity was shown to fluctuate cyclically with the vessel wall contractions and ranged from -1 to 4 mm/sec through a ten second sequence.

  14. Electric fields in Scanning Electron Microscopy simulations

    NASA Astrophysics Data System (ADS)

    Arat, K. T.; Bolten, J.; Klimpel, T.; Unal, N.

    2016-03-01

    The electric field distribution and charging effects in Scanning Electron Microscopy (SEM) were studied by extending a Monte-Carlo based SEM simulator by a fast and accurate multigrid (MG) based 3D electric field solver. The main focus is on enabling short simulation times with maintaining sufficient accuracy, so that SEM simulation can be used in practical applications. The implementation demonstrates a gain in computation speed, when compared to a Gauss-Seidel based reference solver is roughly factor of 40, with negligible differences in the result (~10-6 𝑉). In addition, the simulations were compared with experimental SEM measurements using also complex 3D sample, showing that i) the modelling of e-fields improves the simulation accuracy, and ii) multigrid method provide a significant benefit in terms of simulation time.

  15. Classification of microscopy images of Langerhans islets

    NASA Astrophysics Data System (ADS)

    Å vihlík, Jan; Kybic, Jan; Habart, David; Berková, Zuzana; Girman, Peter; Kříž, Jan; Zacharovová, Klára

    2014-03-01

    Evaluation of images of Langerhans islets is a crucial procedure for planning an islet transplantation, which is a promising diabetes treatment. This paper deals with segmentation of microscopy images of Langerhans islets and evaluation of islet parameters such as area, diameter, or volume (IE). For all the available images, the ground truth and the islet parameters were independently evaluated by four medical experts. We use a pixelwise linear classifier (perceptron algorithm) and SVM (support vector machine) for image segmentation. The volume is estimated based on circle or ellipse fitting to individual islets. The segmentations were compared with the corresponding ground truth. Quantitative islet parameters were also evaluated and compared with parameters given by medical experts. We can conclude that accuracy of the presented fully automatic algorithm is fully comparable with medical experts.

  16. Super-resolution microscopy: a comparative treatment.

    PubMed

    Kasuboski, James M; Sigal, Yury J; Joens, Matthew S; Lillemeier, Bjorn F; Fitzpatrick, James A J

    2012-10-01

    One of the fundamental limitations of optical microscopy is that of diffraction, or in essence, how small a beam of light can be focused by using an optical lens system. This constraint, or barrier if you will, was theoretically described by Ernst Abbe in 1873 and is roughly equal to half the wavelength of light used to probe the system. Many structures, particularly those within cells, are much smaller than this limit and thus are difficult to visualize. Over the last two decades, a new field of super-resolution imaging has been created and been developed into a broad range of techniques that allow routine imaging beyond the far-field diffraction limit of light. In this unit we outline the basic principles of the various super-resolution imaging modalities, paying particular attention to the technical considerations for biological imaging. Furthermore, we discuss their various applications in the imaging of both fixed and live biological samples.

  17. Hexamethyldisilazane for scanning electron microscopy of Gastrotricha.

    PubMed

    Hochberg, R; Litvaitis, M K

    2000-01-01

    We evaluated treatment with hexamethyldisilazane (HMDS) as an alternative to critical-point drying (CPD) for preparing microscopic Gastrotricha for scanning electron microscopy (SEM). We prepared large marine (2 mm) and small freshwater (100 microm) gastrotrichs using HMDS as the primary dehydration solvent and compared the results to earlier investigations using CPD. The results of HMDS dehydration are similar to or better than CPD for resolution of two important taxonomic features: cuticular ornamentation and patterns of ciliation. The body wall of both sculpted (Lepidodermella) and smooth (Dolichodasys) gastrotrichs retained excellent morphology as did the delicate sensory and locomotory cilia. The only unfavorable result of HMDS dehydration was an occasional coagulation of gold residue when the solvent had not fully evaporated before sputter-coating. We consider HMDS an effective alternative for preparing of gastrotrichs for SEM because it saves time and expense compared to CPD.

  18. Contact x-ray microscopy using Asterix

    NASA Astrophysics Data System (ADS)

    Conti, Aldo; Batani, Dimitri; Botto, Cesare; Masini, Alessandra; Bernardinello, A.; Bortolotto, Fulvia; Moret, M.; Poletti, G.; Piccoli, S.; Cotelli, F.; Lora Lamia Donin, C.; Stead, Anthony D.; Marranca, A.; Eidmann, Klaus; Flora, Francesco; Palladino, Libero; Reale, Lucia

    1997-10-01

    The use of a high energy laser source for soft x-ray contact microscopy is discussed. Several different targets were used and their emission spectra compared. The x-ray emission, inside and outside the Water Window, was characterized in detail by means of many diagnostics, including pin hole and streak cameras. Up to 12 samples holders per shot were exposed thanks to the large x-ray flux and the geometry of the interaction chamber. Images of several biological samples were obtained, including Chlamydomonas and Crethidia green algae, fish and boar sperms and Saccharomyces Cerevisiae yeast cells. A 50 nm resolution was reached on the images of boar sperm. Original information concerning the density of inner structures of Crethidia green algae were obtained.

  19. Atomic force microscopy of Precambrian microscopic fossils.

    PubMed

    Kempe, André; Schopf, J William; Altermann, Wladyslaw; Kudryavtsev, Anatoliy B; Heckl, Wolfgang M

    2002-07-09

    Atomic force microscopy (AFM) is a technique used routinely in material science to image substances at a submicron (including nm) scale. We apply this technique to analysis of the fine structure of organic-walled Precambrian fossils, microscopic sphaeromorph acritarchs (cysts of planktonic unicellular protists) permineralized in approximately 650-million-year-old cherts of the Chichkan Formation of southern Kazakhstan. AFM images, backed by laser-Raman spectroscopic analysis of individual specimens, demonstrate that the walls of these petrified fossils are composed of stacked arrays of approximately 200-nm-sized angular platelets of polycyclic aromatic kerogen. Together, AFM and laser-Raman spectroscopy provide means by which to elucidate the submicron-scale structure of individual microscopic fossils, investigate the geochemical maturation of ancient organic matter, and, potentially, distinguish true fossils from pseudofossils and probe the mechanisms of fossil preservation by silica permineralization.

  20. Multiphoton microscopy of cleared mouse organs

    NASA Astrophysics Data System (ADS)

    Parra, Sonia G.; Chia, Thomas H.; Zinter, Joseph P.; Levene, Michael J.

    2010-05-01

    Typical imaging depths with multiphoton microscopy (MPM) are limited to less than 300 μm in many tissues due to light scattering. Optical clearing significantly reduces light scattering by replacing water in the organ tissue with a fluid having a similar index of refraction to that of proteins. We demonstrate MPM of intact, fixed, cleared mouse organs with penetration depths and fields of view in excess of 2 mm. MPM enables the creation of large 3-D data sets with flexibility in pixel format and ready access to intrinsic fluorescence and second-harmonic generation. We present high-resolution images and 3-D image stacks of the brain, small intestine, large intestine, kidney, lung, and testicle with image sizes as large as 4096×4096 pixels.

  1. Automated cellular pathology in noninvasive confocal microscopy

    NASA Astrophysics Data System (ADS)

    Ting, Monica; Krueger, James; Gareau, Daniel

    2014-03-01

    A computer algorithm was developed to automatically identify and count melanocytes and keratinocytes in 3D reflectance confocal microscopy (RCM) images of the skin. Computerized pathology increases our understanding and enables prevention of superficial spreading melanoma (SSM). Machine learning involved looking at the images to measure the size of cells through a 2-D Fourier transform and developing an appropriate mask with the erf() function to model the cells. Implementation involved processing the images to identify cells whose image segments provided the least difference when subtracted from the mask. With further simplification of the algorithm, the program may be directly implemented on the RCM images to indicate the presence of keratinocytes in seconds and to quantify the keratinocytes size in the en face plane as a function of depth. Using this system, the algorithm can identify any irregularities in maturation and differentiation of keratinocytes, thereby signaling the possible presence of cancer.

  2. Pulling angle-dependent force microscopy

    NASA Astrophysics Data System (ADS)

    Grebíková, L.; Gojzewski, H.; Kieviet, B. D.; Klein Gunnewiek, M.; Vancso, G. J.

    2017-03-01

    In this paper, we describe a method allowing one to perform three-dimensional displacement control in force spectroscopy by atomic force microscopy (AFM). Traditionally, AFM force curves are measured in the normal direction of the contacted surface. The method described can be employed to address not only the magnitude of the measured force but also its direction. We demonstrate the technique using a case study of angle-dependent desorption of a single poly(2-hydroxyethyl methacrylate) (PHEMA) chain from a planar silica surface in an aqueous solution. The chains were end-grafted from the AFM tip in high dilution, enabling single macromolecule pull experiments. Our experiments give evidence of angular dependence of the desorption force of single polymer chains and illustrate the added value of introducing force direction control in AFM.

  3. Noninvasive photoacoustic microscopy of methemoglobin in vivo

    PubMed Central

    Tang, Min; Zhou, Yong; Zhang, Ruiying; Wang, Lihong V.

    2015-01-01

    Abstract. Due to the various causes of methemoglobinemia and its potential to be confused with other diseases, in vivo measurements of methemoglobin have significant applications in the clinic. Using photoacoustic microscopy (PAM), we quantified the average and the distributed percentage of methemoglobin both in vitro and in vivo. Based on the absorption spectra of methemoglobin, oxyhemoglobin, and deoxyhemoglobin, three wavelengths were chosen to differentiate methemoglobin from the others. The methemoglobin concentrations calculated from the photoacoustic signals agreed well with the preset concentrations. Then we imaged the methemoglobin percentage in microtubes that mimicked blood vessels. Average percentages calculated for five samples with different methemoglobin concentrations also agreed well with the preset values. Finally, we demonstrated the ability of PAM to detect methemoglobin in vivo in a mouse ear. Our results show that PAM can quantitatively image methemoglobin distribution in vivo. PMID:25760655

  4. Confocal reference free traction force microscopy

    PubMed Central

    Bergert, Martin; Lendenmann, Tobias; Zündel, Manuel; Ehret, Alexander E.; Panozzo, Daniele; Richner, Patrizia; Kim, David K.; Kress, Stephan J. P.; Norris, David J.; Sorkine-Hornung, Olga; Mazza, Edoardo; Poulikakos, Dimos; Ferrari, Aldo

    2016-01-01

    The mechanical wiring between cells and their surroundings is fundamental to the regulation of complex biological processes during tissue development, repair or pathology. Traction force microscopy (TFM) enables determination of the actuating forces. Despite progress, important limitations with intrusion effects in low resolution 2D pillar-based methods or disruptive intermediate steps of cell removal and substrate relaxation in high-resolution continuum TFM methods need to be overcome. Here we introduce a novel method allowing a one-shot (live) acquisition of continuous in- and out-of-plane traction fields with high sensitivity. The method is based on electrohydrodynamic nanodrip-printing of quantum dots into confocal monocrystalline arrays, rendering individually identifiable point light sources on compliant substrates. We demonstrate the undisrupted reference-free acquisition and quantification of high-resolution continuous force fields, and the simultaneous capability of this method to correlatively overlap traction forces with spatial localization of proteins revealed using immunofluorescence methods. PMID:27681958

  5. Atomic force microscopy of Precambrian microscopic fossils

    PubMed Central

    Kempe, André; Schopf, J. William; Altermann, Wladyslaw; Kudryavtsev, Anatoliy B.; Heckl, Wolfgang M.

    2002-01-01

    Atomic force microscopy (AFM) is a technique used routinely in material science to image substances at a submicron (including nm) scale. We apply this technique to analysis of the fine structure of organic-walled Precambrian fossils, microscopic sphaeromorph acritarchs (cysts of planktonic unicellular protists) permineralized in ≈650-million-year-old cherts of the Chichkan Formation of southern Kazakhstan. AFM images, backed by laser-Raman spectroscopic analysis of individual specimens, demonstrate that the walls of these petrified fossils are composed of stacked arrays of ≈200-nm-sized angular platelets of polycyclic aromatic kerogen. Together, AFM and laser-Raman spectroscopy provide means by which to elucidate the submicron-scale structure of individual microscopic fossils, investigate the geochemical maturation of ancient organic matter, and, potentially, distinguish true fossils from pseudofossils and probe the mechanisms of fossil preservation by silica permineralization. PMID:12089337

  6. Optical Third-Harmonic Microscopy of Graphene

    NASA Astrophysics Data System (ADS)

    Dadap, Jerry I.; Hong, Sung-Young; Petrone, Nicholas W.; Yeh, Po-Chun; Hone, James C.; Osgood, Richard M., Jr.

    2013-03-01

    We report strong third-harmonic (TH) generation in monolayer graphene mounted on an amorphous silica substrate using a photon energy that is three-photon resonant with the exciton-shifted van Hove singularity at the M-point of graphene. Our polarization-dependent and azimuthal rotation measurements confirm the expected isotropic symmetry properties for the TH nonlinear optical process in graphene. Since this monolayer graphene TH signal exceeds that of bulk glass by more than two orders of magnitude, the signal contrast permits background-free scanning of graphene and provides structural information that is difficult to obtain via linear optical microscopy. We also discuss the dependence of TH signals on the number of graphene layers and compare the graphene signal strength with that from crystalline Au(111) sample. We acknowledge support from AFOSR MURI Program #FA9550-09-1-0705.

  7. Visualizing individual microtubules by bright field microscopy

    NASA Astrophysics Data System (ADS)

    Gutiérrez-Medina, Braulio; Block, Steven M.

    2010-11-01

    Microtubules are slender (˜25 nm diameter), filamentous polymers involved in cellular structure and organization. Individual microtubules have been visualized via fluorescence imaging of dye-labeled tubulin subunits and by video-enhanced, differential interference-contrast microscopy of unlabeled polymers using sensitive CCD cameras. We demonstrate the imaging of unstained microtubules using a microscope with conventional bright field optics in conjunction with a webcam-type camera and a light-emitting diode illuminator. The light scattered by microtubules is image-processed to remove the background, reduce noise, and enhance contrast. The setup is based on a commercial microscope with a minimal set of inexpensive components, suitable for implementation in a student laboratory. We show how this approach can be used in a demonstration motility assay, tracking the gliding motions of microtubules driven by the motor protein kinesin.

  8. Atomic force microscopy of model lipid membranes.

    PubMed

    Morandat, Sandrine; Azouzi, Slim; Beauvais, Estelle; Mastouri, Amira; El Kirat, Karim

    2013-02-01

    Supported lipid bilayers (SLBs) are biomimetic model systems that are now widely used to address the biophysical and biochemical properties of biological membranes. Two main methods are usually employed to form SLBs: the transfer of two successive monolayers by Langmuir-Blodgett or Langmuir-Schaefer techniques, and the fusion of preformed lipid vesicles. The transfer of lipid films on flat solid substrates offers the possibility to apply a wide range of surface analytical techniques that are very sensitive. Among them, atomic force microscopy (AFM) has opened new opportunities for determining the nanoscale organization of SLBs under physiological conditions. In this review, we first focus on the different protocols generally employed to prepare SLBs. Then, we describe AFM studies on the nanoscale lateral organization and mechanical properties of SLBs. Lastly, we survey recent developments in the AFM monitoring of bilayer alteration, remodeling, or digestion, by incubation with exogenous agents such as drugs, proteins, peptides, and nanoparticles.

  9. Combined Confocal and Magnetic Resonance Microscopy

    SciTech Connect

    Wind, Robert A.; Majors, Paul D.; Minard, Kevin R.; Ackerman, Eric J.; Daly, Don S.; Holtom, Gary R.; Thrall, Brian D.; Weber, Thomas J.

    2002-05-12

    Confocal and magnetic resonance microscopy are both used to study live cells in a minimally invasive way. Both techniques provide complementary information. Therefore, by examining cells simultaneously with both methodologies, more detailed information is obtained than is possible with each of the microscopes individually. In this paper two configurations of a combined confocal and magnetic resonance microscope described. In both cases the sample compartment is part of a temperature regulated perfusion system. The first configuration is capable of studying large single cells or three-dimensional cell agglomerates, whereas with the second configuration monolayers of mammalian cells can be investigated . Combined images are shown of Xenopus laevis frog oocytes, model JB6 tumor spheroids, and a single layer of Chinese hamster ovary cells. Finally, potential applications of the combined microscope are discussed.

  10. Fake turquoises investigated by Raman microscopy.

    PubMed

    Bernardino, Nathália D'Elboux; Izumi, Celly M S; de Faria, Dalva L A

    2016-05-01

    Turquoise is frequently adulterated by unscrupulous dealers and, not rarely, simulants are commercialized as true stones. On the other hand, turquoise is a cryptocrystalline mineral and its use in adornments commonly demands some kind of treatment to facilitate its manipulation, such as impregnation using oil or fats, consolidation with resin and stabilization or reconstitution made with resins. In this work, Raman microscopy was employed in the investigation of turquoise adornments aiming to differentiate processed turquoise from fakes or simulants. Only one out of the five adornment objects analyzed was truly stabilized turquoise (powdered turquoise aggregated with a resin). Another one was constituted of turquoise, calcium carbonate, phthalocyanine blue and resin; the other objects were dyed minerals.

  11. Nanoscale thermometry by scanning thermal microscopy

    NASA Astrophysics Data System (ADS)

    Menges, Fabian; Riel, Heike; Stemmer, Andreas; Gotsmann, Bernd

    2016-07-01

    Measuring temperature is a central challenge in nanoscience and technology. Addressing this challenge, we report the development of a high-vacuum scanning thermal microscope and a method for non-equilibrium scanning probe thermometry. The microscope is built inside an electromagnetically shielded, temperature-stabilized laboratory and features nanoscopic spatial resolution at sub-nanoWatt heat flux sensitivity. The method is a dual signal-sensing technique inferring temperature by probing a total steady-state heat flux simultaneously to a temporally modulated heat flux signal between a self-heated scanning probe sensor and a sample. Contact-related artifacts, which so far limit the reliability of nanoscopic temperature measurements by scanning thermal microscopy, are minimized. We characterize the microscope's performance and demonstrate the benefits of the new thermometry approach by studying hot spots near lithographically defined constrictions in a self-heated metal interconnect.

  12. Photoacoustic microscopy of bilirubin in tissue phantoms

    NASA Astrophysics Data System (ADS)

    Zhou, Yong; Zhang, Chi; Yao, Da-Kang; Wang, Lihong V.

    2012-12-01

    Determining both bilirubin's concentration and its spatial distribution are important in disease diagnosis. Here, for the first time, we applied quantitative multiwavelength photoacoustic microscopy (PAM) to detect bilirubin concentration and distribution simultaneously. By measuring tissue-mimicking phantoms with different bilirubin concentrations, we showed that the root-mean-square error of prediction has reached 0.52 and 0.83 mg/dL for pure bilirubin and for blood-mixed bilirubin detection (with 100% oxygen saturation), respectively. We further demonstrated the capability of the PAM system to image bilirubin distribution both with and without blood. Finally, by underlaying bilirubin phantoms with mouse skins, we showed that bilirubin can be imaged with consistent accuracy down to >400 μm in depth. Our results show that PAM has potential for noninvasive bilirubin monitoring in vivo, as well as for further clinical applications.

  13. Halo-free Phase Contrast Microscopy

    NASA Astrophysics Data System (ADS)

    Nguyen, Tan H.; Kandel, Mikhail; Shakir, Haadi M.; Best-Popescu, Catherine; Arikkath, Jyothi; Do, Minh N.; Popescu, Gabriel

    2017-03-01

    We present a new approach for retrieving halo-free phase contrast microscopy (hfPC) images by upgrading the conventional PC microscope with an external interferometric module, which generates sufficient data for reversing the halo artifact. Acquiring four independent intensity images, our approach first measures haloed phase maps of the sample. We solve for the halo-free sample transmission function by using a physical model of the image formation under partial spatial coherence. Using this halo-free sample transmission, we can numerically generate artifact-free PC images. Furthermore, this transmission can be further used to obtain quantitative information about the sample, e.g., the thickness with known refractive indices, dry mass of live cells during their cycles. We tested our hfPC method on various control samples, e.g., beads, pillars and validated its potential for biological investigation by imaging live HeLa cells, red blood cells, and neurons.

  14. Video-rate tomographic phase microscopy

    NASA Astrophysics Data System (ADS)

    Fang-Yen, Christopher; Choi, Wonshik; Sung, Yongjin; Holbrow, Charles J.; Dasari, Ramachandra R.; Feld, Michael S.

    2011-01-01

    Tomographic phase microscopy measures the 3-D refractive index distribution of cells and tissues by combining the information from a series of angle-dependent interferometric phase images. In the original device, the frame rate was limited to 0.1 frames per second (fps) by the technique used to acquire phase images, preventing measurements of moving or rapidly changing samples. We describe an improved tomographic phase microscope in which phase images are acquired via a spatial fringe pattern demodulation method, enabling a full tomogram acquisition rate of 30 fps. In addition, in this system the refractive index is calculated by a diffraction tomography algorithm that accounts for the effects of diffraction in the 3-D reconstruction. We use the instrument to quantitatively monitor rapid changes in refractive index within defined subregions of cells due to exposure to acetic acid or changes in medium osmolarity.

  15. Electrostatic interaction in atomic force microscopy

    PubMed Central

    Butt, Hans-Jüurgen

    1991-01-01

    In atomic force microscopy, the stylus experiences an electrostatic force when imaging in aqueous medium above a charged surface. This force has been calculated numerically with continuum theory for a silicon nitrite or silicon oxide stylus. For comparison, the Van der Waals force was also calculated. In contrast to the Van der Waals attraction, the electrostatic force is repulsive. At a distance of 0.5 nm the electrostatic force is typically 10-12-10-10 N and thus comparable in strength to the Van der Waals force. The electrostatic force increases with increasing surface charge density and decreases roughly exponentially with distance. It can be reduced by imaging in high salt concentrations. Below surface potentials of ≈50 mV, a simple analytical approximation of the electrostatic force is described. PMID:19431803

  16. Image simulation for biological microscopy: microlith

    PubMed Central

    Mehta, Shalin B.; Oldenbourg, Rudolf

    2014-01-01

    Image simulation remains under-exploited for the most widely used biological phase microscopy methods, because of difficulties in simulating partially coherent illumination. We describe an open-source toolbox, microlith (https://code.google.com/p/microlith), which accurately predicts three-dimensional images of a thin specimen observed with any partially coherent imaging system, as well as images of coherently illuminated and self-luminous incoherent specimens. Its accuracy is demonstrated by comparing simulated and experimental bright-field and dark-field images of well-characterized amplitude and phase targets, respectively. The comparison provides new insights about the sensitivity of the dark-field microscope to mass distributions in isolated or periodic specimens at the length-scale of 10nm. Based on predictions using microlith, we propose a novel approach for detecting nanoscale structural changes in a beating axoneme using a dark-field microscope. PMID:24940543

  17. Photoacoustic microscopy of blood pulse wave

    NASA Astrophysics Data System (ADS)

    Yeh, Chenghung; Hu, Song; Maslov, Konstantin; Wang, Lihong V.

    2012-07-01

    Blood pulse wave velocity (PWV) is an important physiological parameter that characterizes vascular stiffness. In this letter, we present electrocardiogram-synchronized, photoacoustic microscopy for noninvasive quantification of the PWV in the peripheral vessels of living mice. Interestingly, blood pulse wave-induced fluctuations in blood flow speed were clearly observed in arteries and arterioles, but not in veins or venules. Simultaneously recorded electrocardiograms served as references to measure the travel time of the pulse wave between two cross sections of a chosen vessel and vessel segmentation analysis enabled accurate quantification of the travel distance. PWVs were quantified in ten vessel segments from two mice. Statistical analysis shows a linear correlation between the PWV and the vessel diameter which agrees with known physiology.

  18. Halo-free Phase Contrast Microscopy

    PubMed Central

    Nguyen, Tan H.; Kandel, Mikhail; Shakir, Haadi M.; Best-Popescu, Catherine; Arikkath, Jyothi; Do, Minh N.; Popescu, Gabriel

    2017-01-01

    We present a new approach for retrieving halo-free phase contrast microscopy (hfPC) images by upgrading the conventional PC microscope with an external interferometric module, which generates sufficient data for reversing the halo artifact. Acquiring four independent intensity images, our approach first measures haloed phase maps of the sample. We solve for the halo-free sample transmission function by using a physical model of the image formation under partial spatial coherence. Using this halo-free sample transmission, we can numerically generate artifact-free PC images. Furthermore, this transmission can be further used to obtain quantitative information about the sample, e.g., the thickness with known refractive indices, dry mass of live cells during their cycles. We tested our hfPC method on various control samples, e.g., beads, pillars and validated its potential for biological investigation by imaging live HeLa cells, red blood cells, and neurons. PMID:28338086

  19. Lensless fluorescent microscopy on a chip.

    PubMed

    Coskun, Ahmet F; Su, Ting-Wei; Sencan, Ikbal; Ozcan, Aydogan

    2011-08-17

    On-chip lensless imaging in general aims to replace bulky lens-based optical microscopes with simpler and more compact designs, especially for high-throughput screening applications. This emerging technology platform has the potential to eliminate the need for bulky and/or costly optical components through the help of novel theories and digital reconstruction algorithms. Along the same lines, here we demonstrate an on-chip fluorescent microscopy modality that can achieve e.g., <4 μm spatial resolution over an ultra-wide field-of-view (FOV) of >0.6-8 cm(2) without the use of any lenses, mechanical-scanning or thin-film based interference filters. In this technique, fluorescent excitation is achieved through a prism or hemispherical-glass interface illuminated by an incoherent source. After interacting with the entire object volume, this excitation light is rejected by total-internal-reflection (TIR) process that is occurring at the bottom of the sample micro-fluidic chip. The fluorescent emission from the excited objects is then collected by a fiber-optic faceplate or a taper and is delivered to an optoelectronic sensor array such as a charge-coupled-device (CCD). By using a compressive-sampling based decoding algorithm, the acquired lensfree raw fluorescent images of the sample can be rapidly processed to yield e.g., <4 μm resolution over an FOV of >0.6-8 cm(2). Moreover, vertically stacked micro-channels that are separated by e.g., 50-100 μm can also be successfully imaged using the same lensfree on-chip microscopy platform, which further increases the overall throughput of this modality. This compact on-chip fluorescent imaging platform, with a rapid compressive decoder behind it, could be rather valuable for high-throughput cytometry, rare-cell research and microarray-analysis.

  20. Nuclear microscopy of atherosclerotic tissue: A review

    NASA Astrophysics Data System (ADS)

    Watt, Frank; Ren, M. Q.; Xie, J. P.; Tan, B. K. H.; Halliwell, B.

    2001-07-01

    This paper reviews the work carried out in the Research Centre for Nuclear Microscopy, NUS on the role of iron in coronary heart disease, using the technique of nuclear microscopy to determine the levels of iron and other trace elements in the artery wall and lesions. These investigations have indicated that iron may play a significant role in the development of atherosclerosis, probably through the promotion of cytotoxic free radicals leading to the oxidation of low-density lipoprotein (LDL). Using a rabbit model we have observed that early atherosclerotic lesions, induced by feeding the animals on a 1% cholesterol diet, contain increased levels of iron (up to 8 times) compared with the adjacent healthy artery wall. In a follow-up time sequence study, we have shown that iron accumulation occurs at the onset of lesion formation, which takes place around 4-6 weeks after exposure to the 1% cholesterol diet. As the lesions mature, they enlarge to occupy a significant fraction of the artery wall, and at about 16 weeks the lesions begin to show signs of calcification. In an additional experiment, where the cholesterol fed rabbits were kept anaemic through weekly bleeding, the iron content of the artery wall was reduced and the onset of atherogenesis was delayed. In a further investigation, rabbits were fed on a 1% cholesterol diet and after 6 weeks (corresponding to the period of early lesion formation) a test group was subjected to treatment using the iron chelator desferal. Preliminary results indicate that during the treatment with desferal, lesion development was slowed down.

  1. Microscopy image segmentation tool: Robust image data analysis

    SciTech Connect

    Valmianski, Ilya Monton, Carlos; Schuller, Ivan K.

    2014-03-15

    We present a software package called Microscopy Image Segmentation Tool (MIST). MIST is designed for analysis of microscopy images which contain large collections of small regions of interest (ROIs). Originally developed for analysis of porous anodic alumina scanning electron images, MIST capabilities have been expanded to allow use in a large variety of problems including analysis of biological tissue, inorganic and organic film grain structure, as well as nano- and meso-scopic structures. MIST provides a robust segmentation algorithm for the ROIs, includes many useful analysis capabilities, and is highly flexible allowing incorporation of specialized user developed analysis. We describe the unique advantages MIST has over existing analysis software. In addition, we present a number of diverse applications to scanning electron microscopy, atomic force microscopy, magnetic force microscopy, scanning tunneling microscopy, and fluorescent confocal laser scanning microscopy.

  2. Selective plane illumination microscopy techniques in developmental biology

    PubMed Central

    Huisken, Jan; Stainier, Didier Y. R.

    2009-01-01

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

  3. Correlative video-light-electron microscopy: development, impact and perspectives.

    PubMed

    Rizzo, Riccardo; Parashuraman, Seetharaman; Luini, Alberto

    2014-08-01

    Green fluorescent protein (GFP)-based video microscopy can provide profound insight into biological processes by generating information on the 'history,' or dynamics, of the cellular structures involved in such processes in live cells. A crucial limitation of this approach, however, is that many such structures may not be resolved by light microscopy. Like more recent super-resolution techniques, correlative video-light-electron microscopy (CLEM) was developed to overcome this limitation. CLEM integrates GFP-based video microscopy and electron microscopy through a series of ancillary techniques, such as proper fixation, hybrid labeling and retracing, and so provides sufficient resolution as well as, crucially, cellular 'context' to the fluorescent dynamic structures of interest. CLEM 'multiplies' the power of video microscopy and is having an important impact in several areas cell and developmental biology. Here, we discuss potential, limitations and perspectives of correlative approaches aimed at integrating the unique insight generated by video microscopy with information from other forms of imaging.

  4. Microscopy image segmentation tool: robust image data analysis.

    PubMed

    Valmianski, Ilya; Monton, Carlos; Schuller, Ivan K

    2014-03-01

    We present a software package called Microscopy Image Segmentation Tool (MIST). MIST is designed for analysis of microscopy images which contain large collections of small regions of interest (ROIs). Originally developed for analysis of porous anodic alumina scanning electron images, MIST capabilities have been expanded to allow use in a large variety of problems including analysis of biological tissue, inorganic and organic film grain structure, as well as nano- and meso-scopic structures. MIST provides a robust segmentation algorithm for the ROIs, includes many useful analysis capabilities, and is highly flexible allowing incorporation of specialized user developed analysis. We describe the unique advantages MIST has over existing analysis software. In addition, we present a number of diverse applications to scanning electron microscopy, atomic force microscopy, magnetic force microscopy, scanning tunneling microscopy, and fluorescent confocal laser scanning microscopy.

  5. Nano-fEM: protein localization using photo-activated localization microscopy and electron microscopy.

    PubMed

    Watanabe, Shigeki; Richards, Jackson; Hollopeter, Gunther; Hobson, Robert J; Davis, Wayne M; Jorgensen, Erik M

    2012-12-03

    Mapping the distribution of proteins is essential for understanding the function of proteins in a cell. Fluorescence microscopy is extensively used for protein localization, but subcellular context is often absent in fluorescence images. Immuno-electron microscopy, on the other hand, can localize proteins, but the technique is limited by a lack of compatible antibodies, poor preservation of morphology and because most antigens are not exposed to the specimen surface. Correlative approaches can acquire the fluorescence image from a whole cell first, either from immuno-fluorescence or genetically tagged proteins. The sample is then fixed and embedded for electron microscopy, and the images are correlated (1-3). However, the low-resolution fluorescence image and the lack of fiducial markers preclude the precise localization of proteins. Alternatively, fluorescence imaging can be done after preserving the specimen in plastic. In this approach, the block is sectioned, and fluorescence images and electron micrographs of the same section are correlated (4-7). However, the diffraction limit of light in the correlated image obscures the locations of individual molecules, and the fluorescence often extends beyond the boundary of the cell. Nano-resolution fluorescence electron microscopy (nano-fEM) is designed to localize proteins at nano-scale by imaging the same sections using photo-activated localization microscopy (PALM) and electron microscopy. PALM overcomes the diffraction limit by imaging individual fluorescent proteins and subsequently mapping the centroid of each fluorescent spot (8-10). We outline the nano-fEM technique in five steps. First, the sample is fixed and embedded using conditions that preserve the fluorescence of tagged proteins. Second, the resin blocks are sectioned into ultrathin segments (70-80 nm) that are mounted on a cover glass. Third, fluorescence is imaged in these sections using the Zeiss PALM microscope. Fourth, electron dense structures are

  6. High-resolution noncontact atomic force microscopy.

    PubMed

    Pérez, Rubén; García, Ricardo; Schwarz, Udo

    2009-07-01

    Progress in nanoscience and nanotechnology requires tools that enable the imaging and manipulation of matter at the atomic and molecular scale. During the last two decades or so, scanning probe-based techniques have proven to be particularly versatile in this regard. Among the various probe-based approaches, atomic force microscopy (AFM) stands out in many ways, including the total number of citations and the breadth of possible applications, ranging from materials characterization to nanofabrication and biological studies. However, while nanometer scale operation in different environments became routine, atomic resolution imaging remained elusive for a long time. The reason for this initial deficiency was that contact with the sample blunts atomically sharp tips, which are mandatory for successful atomic resolution imaging. This problem was overcome in the mid-1990s with the introduction of noncontact atomic force microscopy (NC-AFM), which represents a version of AFM where the cantilever is oscillated close to the sample surface without actually 'touching' it. This allows the preservation of the atomic sharpness of the tip while interaction-induced changes in the cantilever's resonance frequency are used to quantify the tip-sample distance. Since then, progress has been steady and includes the development of commercial instruments as well as the addition of many new capabilities beyond imaging, such as the identification and manipulation of individual atoms. A series of annual international conferences, starting in Osaka in 1998, have contributed significantly to this outstanding performance. The program of the most recent conference from this series, held in Madrid on 15-19 September 2008, reflects the maturity of this field, with an increasing number of groups developing strong activities that involve novel approaches and applications covering areas well beyond the original vacuum-based imaging. In this special issue of Nanotechnology we present a selection of

  7. Advanced electron microscopy characterization of multimetallic nanoparticles

    NASA Astrophysics Data System (ADS)

    Khanal, Subarna Raj

    Research in noble metal nanoparticles has led to exciting progress in a versatile array of applications. For the purpose of better tailoring of nanoparticles activities and understanding the correlation between their structures and properties, control over the composition, shape, size and architecture of bimetallic and multimetallic nanomaterials plays an important role on revealing their new or enhanced functions for potentials application. Advance electron microscopy techniques were used to provide atomic scale insights into the structure-properties of different materials: PtPd, Au-Au3Cu, Cu-Pt, AgPd/Pt and AuCu/Pt nanoparticles. The objective of this work is to understand the physical and chemical properties of nanomaterials and describe synthesis, characterization, surface properties and growth mechanism of various bimetallic and multimetallic nanoparticles. The findings have provided us with novel and significant insights into the physical and chemical properties of noble metal nanoparticles. Different synthesis routes allowed us to synthesize bimetallic: Pt-Pd, Au-Au3Cu, Cu-Pt and trimetallic: AgPd/Pt, AuCu/Pt, core-shell and alloyed nanoparticles with monodispersed sizes, controlled shapes and tunable surface properties. For example, we have synthesized the polyhedral PtPd core-shell nanoparticles with octahedral, decahedral, and triangular plates. Decahedral PtPd core-shell structures are novel morphologies for this system. For the first time we fabricated that the Au core and Au3Cu alloyed shell nanoparticles passivated with CuS2 surface layers and characterized by Cs-corrected scanning transmission electron microscopy. The analysis of the high-resolution micrographs reveals that these nanoparticles have decahedral structure with shell periodicity, and that each of the particles is composed by Au core and Au3Cu ordered superlattice alloyed shell surrounded by CuS 2 surface layer. Additionally, we have described both experimental and theoretical methods of

  8. The Disease Activity Score (DAS) and the Disease Activity Score using 28 joint counts (DAS28) in the management of rheumatoid arthritis.

    PubMed

    van Riel, Piet L C M; Renskers, Lisanne

    2016-01-01

    In rheumatoid arthritis (RA), disease activity cannot be measured in all individual patients according to a single variable. The Disease Activity Score (DAS) and the DAS28 have been developed to measure disease activity in RA both in daily clinical practice as well as in clinical trials on a group as well as individual level. The DAS/DAS28 is a continuous measure of RA disease activity that combines information from swollen joints, tender joints, acute phase response and general health. The DAS-based EULAR response criteria were primarily developed to be used in clinical trials. The EULAR response criteria classify individual patients as non-, moderate, or good responders, dependent on the magnitude of change and level of disease activity reached. In addition, already in the early nineties, cut points were developed to categorise patients in remission. The DAS28 is incorporated in several electronic patient records and web-based systems for monitoring purposes in daily clinical practice. In addition to this, it is being used in combination with patient-reported outcome measures (PROMs) to facilitate self-monitoring.

  9. Ultrastable Atomic Force Microscopy for Biophysics

    NASA Astrophysics Data System (ADS)

    Churnside, Allison B.

    Atomic force microscopy (AFM) is a multifunctional workhorse of nanoscience and molecular biophysics, but instrumental drift remains a critical issue that limits the precision and duration of experiments. We have significantly reduced the two most important types of drift: in position and in force. The first, position drift, is defined as uncontrolled motion between the tip and the sample, which occurs in all three dimensions. By scattering a laser off the apex of a commercial AFM tip, we locally measured and thereby actively controlled its three-dimensional position above a sample surface to <0.4 A (Deltaf = 0.01--10 Hz) in air at room temperature. With this enhanced stability, we demonstrated atomic-scale (˜1 A) tip-sample stability and registration over tens of minutes with a series of AFM images. The second type of drift is force drift. We found that the primary source of force drift for a popular class of soft cantilevers is their gold coating, even though they are coated on both sides to minimize drift. When the gold coating was removed through a simple chemical etch, this drift in deflection was reduced by more than an order of magnitude over the first 2 hours after wetting the tip. Removing the gold also led to ˜ 10-fold reduction in reflected light, yet short-term (0.1--10 s) force precision improved. With both position and force drift greatly reduced, the utility of the AFM is enhanced. These improvements led to several new AFM abilities, including a five-fold increase in the image signal-to-noise ratio; tip-registered, label-free optical imaging; registered tip return to a particular point on the sample; and dual-detection force spectroscopy, which enables a new extension clamp mode. We have applied these abilities to folding of both membrane and soluble proteins. In principle, the techniques we describe can be fully incorporated into many types of scanning probe microscopy, making this work a general improvement to scanning probe techniques.

  10. Comparison of scanning ion conductance microscopy with atomic force microscopy for cell imaging.

    PubMed

    Rheinlaender, Johannes; Geisse, Nicholas A; Proksch, Roger; Schäffer, Tilman E

    2011-01-18

    We present the first direct comparison of scanning ion conductance microscopy (SICM) with atomic force microscopy (AFM) for cell imaging. By imaging the same fibroblast or myoblast cell with both technologies in series, we highlight their advantages and disadvantages with respect to cell imaging. The finite imaging force applied to the sample in AFM imaging results in a coupling of mechanical sample properties into the measured sample topography. For soft samples such as cells this leads to artifacts in the measured topography and to elastic deformation, which we demonstrate by imaging whole fixed cells and cell extensions at high resolution. SICM imaging, on the other hand, has a noncontact character and can provide the true topography of soft samples at a comparable resolution.

  11. Atomic force microscopy and scanning electron microscopy study of MgO(110) surface faceting

    NASA Astrophysics Data System (ADS)

    Giese, D. R.; Lamelas, F. J.; Owen, H. A.; Plass, R.; Gajdardziska-Josifovska, M.

    2000-06-01

    Phosphoric- and nitric-acid etching of the MgO(110) surface generates vicinal faceting in both the <001> and <110> directions. Vacuum annealing (to 1000°C) does not introduce thermal faceting, and does not alter the chemical-etch morphology. Three types of acid-induced faceting (early-stage pits, later-stage grooves, and inverted trapezoidal pyramids) are seen as a function of etching time. Facet-angle analysis by atomic force microscopy (AFM) and scanning electron microscopy (SEM) shows the etch morphology to be vicinal, with angles in the range of 9° to 23°, not the low-energy {100} planes expected from minimization of surface energy.

  12. A study of hydrogenated carbon fibers by scanning electron microscopy and confocal laser scanning microscopy.

    PubMed

    de la Cal, Antonio Madroñero; Aguado-Serrano, Juan; Rojas-Cervantes, Maria Luisa; Adame, Elena V Rosa; Marron, Belen Sarmiento; Rosende, Africa Castro; Nevshupa, Roman

    2009-06-01

    The hydrogen absorption process is studied in carbonaceous fibers produced from a mixture of methane and hydrogen. The absorption of the hydrogen was examined in two types of fibers, in "as-grown" state and after a process of desorption during an annealing to 1.473 K under vacuum. Later to its production process, the fibers withstand an oxidation in air to 973 K. The fibers were examined by means of scanning electron microscopy (SEM) and confocal microscopy by reflection. Differences in the behavior during the oxidation were observed between the fibers in as-grown state and those subjected to a further annealing. It could be verified that the fibers were really constituted by two different phases. In one of the phases, the storage of the hydrogen absorbed took place, whereas in the other phase there was no alteration. The process of annealing prior to the absorption of the hydrogen has an appreciable effect on the desorption rate of the hydrogen.

  13. FTIR microscopy and confocal Raman microscopy for studying lateral drug diffusion from a semisolid formulation.

    PubMed

    Gotter, B; Faubel, W; Neubert, R H H

    2010-01-01

    Fourier transform infrared (FTIR) microscopy was applied to obtain information on lateral drug diffusion of dithranol in artificial acceptor membranes. Lateral (2D) drug distribution into an artificial membrane was investigated on an area of 300microm x 1000microm with a lateral resolution of 25microm x 25microm by integrating a specific IR band located at 1430cm(-1). The concentration profiles show a heterogeneous distribution of dithranol particles resulting in non-uniform drug diffusion. Use of the FTIR microscope either in the transmission or in the reflection mode was restricted to a thickness of the DDC membrane <15microm. The third dimension (depth profile) was analysed by means of confocal Raman microscopy (CRM). In an artificial membrane, the depth range from a minimum of 1.5microm up to a maximum of 49microm was analysed for dithranol distribution.

  14. High-sensitivity quantitative Kelvin probe microscopy by noncontact ultra-high-vacuum atomic force microscopy

    NASA Astrophysics Data System (ADS)

    Sommerhalter, Ch.; Matthes, Th. W.; Glatzel, Th.; Jäger-Waldau, A.; Lux-Steiner, M. Ch.

    1999-07-01

    We present quantitative measurements of the work function of semiconductor and metal surfaces prepared in ultrahigh vacuum (UHV) using a combination of UHV noncontact atomic force microscopy and Kelvin probe force microscopy. High energetic and lateral resolution is achieved by using the second resonance frequency of the cantilever to measure the electrostatic forces, while the first resonance frequency is used to simultaneously obtain topographic images by the frequency modulation technique. Spatially resolved work-function measurements reveal a reduced work function in the vicinity of steps on highly oriented pyrolytic graphite. On the GaAs(110) surface it could be demonstrated that defect states in the forbidden band gap cause a local pinning of the Fermi level along monolayer steps. On p-WSe2(0001) work-function variations due to the Coulomb potential of single dopant sites were resolved.

  15. GPU-based image registration in aperture correlation microscopy, and reflection mode correlation microscopy

    NASA Astrophysics Data System (ADS)

    Fafchamps, Lionel J.; Neil, Mark A. A.; Juskaitis, Rimas

    2013-02-01

    Aperture Correlation Microscopy (ACM) is a fluorescence microscopy technique capable of depth resolved imaging and enhanced lateral resolution at real-time acquisition rates. It relies on the subtraction of 2 separate images from different cameras which must be registered to the sub-pixel level. In order to achieve real-time registration and subtraction, the graphics processing unit (GPU) is used to apply a transformation from one frame to the other, resulting in a system capable of processing over 200 frames per second on modest hardware (GeForce 330M). Currently, this rate is limited by camera acquision to 16fps. Additionally, a novel reflection mode correlation microscope is introduced which functions on similar principles as the fluorescent system but can be used to examine reflective samples. Images and z-stacks taken with this system are presented here.

  16. Metal particles in a ceramic matrix--scanning electron microscopy and transmission electron microscopy characterization.

    PubMed

    Konopka, K

    2006-09-01

    This paper is concerned with ceramic matrix (Al(2)O(3)) composites with introduced metal particles (Ni, Fe). The composites were obtained via sintering of powders under very high pressure (2.5 GPa). Scanning electron microscopy and transmission electron microscopy were chosen as the tools for the identification and description of the shape, size and distribution of the metal particles. The Al(2)O(3)-Ni composite contained agglomerates of the Ni particles surrounded by ceramic grains and nanometre-size Ni particles located inside the ceramic grains and at the ceramic grain boundaries. In the Al(2)O(3)-Fe composite, the Fe particles were mostly surrounded by ceramic grains. Moreover, holes left by the Fe particles were found. The high pressure used in the fabrication of the composites changed the shape of the metal and ceramic powder grains via plastic deformation.

  17. Investigation of Nematode Diversity using Scanning Electron Microscopy and Fluorescent Microscopy

    NASA Astrophysics Data System (ADS)

    Seacor, Taylor; Howell, Carina

    2013-03-01

    Nematode worms account for the vast majority of the animals in the biosphere. They are colossally important to global public health as parasites, and to agriculture both as pests and as beneficial inhabitants of healthy soil. Amphid neurons are the anterior chemosensory neurons in nematodes, mediating critical behaviors including chemotaxis and mating. We are examining the cellular morphology and external anatomy of amphid neurons, using fluorescence microscopy and scanning electron microscopy, respectively, of a wide range of soil nematodes isolated in the wild. We use both classical systematics (e.g. diagnostic keys) and molecular markers (e.g. ribosomal RNA) to classify these wild isolates. Our ultimate aim is to build a detailed anatomical database in order to dissect genetic pathways of neuronal development and function across phylogeny and ecology. Research supported by NSF grants 092304, 0806660, 1058829 and Lock Haven University FPDC grants

  18. Simultaneous Scanning Ion Conductance Microscopy and Atomic Force Microscopy with Microchanneled Cantilevers

    NASA Astrophysics Data System (ADS)

    Ossola, Dario; Dorwling-Carter, Livie; Dermutz, Harald; Behr, Pascal; Vörös, János; Zambelli, Tomaso

    2015-12-01

    We combined scanning ion conductance microscopy (SICM) and atomic force microscopy (AFM) into a single tool using AFM cantilevers with an embedded microchannel flowing into the nanosized aperture at the apex of the hollow pyramid. An electrode was positioned in the AFM fluidic circuit connected to a second electrode in the bath. We could thus simultaneously measure the ionic current and the cantilever bending (in optical beam deflection mode). First, we quantitatively compared the SICM and AFM contact points on the approach curves. Second, we estimated where the probe in SICM mode touches the sample during scanning on a calibration grid and applied the finding to image a network of neurites on a Petri dish. Finally, we assessed the feasibility of a double controller using both the ionic current and the deflection as input signals of the piezofeedback. The experimental data were rationalized in the framework of finite elements simulations.

  19. Image segmentation for integrated multiphoton microscopy and reflectance confocal microscopy imaging of human skin in vivo

    PubMed Central

    Chen, Guannan; Lui, Harvey

    2015-01-01

    Background Non-invasive cellular imaging of the skin in vivo can be achieved in reflectance confocal microscopy (RCM) and multiphoton microscopy (MPM) modalities to yield complementary images of the skin based on different optical properties. One of the challenges of in vivo microscopy is the delineation (i.e., segmentation) of cellular and subcellular architectural features. Methods In this work we present a method for combining watershed and level-set models for segmentation of multimodality images obtained by an integrated MPM and RCM imaging system from human skin in vivo. Results Firstly, a segmentation model based on watershed is introduced for obtaining the accurate structure of cell borders from the RCM image. Secondly,, a global region based energy level-set model is constructed for extracting the nucleus of each cell from the MPM image. Thirdly, a local region-based Lagrange Continuous level-set approach is used for segmenting cytoplasm from the MPM image. Conclusions Experimental results demonstrated that cell borders from RCM image and boundaries of cytoplasm and nucleus from MPM image can be obtained by our segmentation method with better accuracy and effectiveness. We are planning to use this method to perform quantitative analysis of MPM and RCM images of in vivo human skin to study the variations of cellular parameters such as cell size, nucleus size and other mophormetric features with skin pathologies. PMID:25694949

  20. Confocal microscopy for astrocyte in vivo imaging: Recycle and reuse in microscopy

    PubMed Central

    Pérez-Alvarez, Alberto; Araque, Alfonso; Martín, Eduardo D.

    2013-01-01

    In vivo imaging is one of the ultimate and fundamental approaches for the study of the brain. Two-photon laser scanning microscopy (2PLSM) constitutes the state-of-the-art technique in current neuroscience to address questions regarding brain cell structure, development and function, blood flow regulation and metabolism. This technique evolved from laser scanning confocal microscopy (LSCM), which impacted the field with a major improvement in image resolution of live tissues in the 1980s compared to widefield microscopy. While nowadays some of the unparalleled features of 2PLSM make it the tool of choice for brain studies in vivo, such as the possibility to image deep within a tissue, LSCM can still be useful in this matter. Here we discuss the validity and limitations of LSCM and provide a guide to perform high-resolution in vivo imaging of the brain of live rodents with minimal mechanical disruption employing LSCM. We describe the surgical procedure and experimental setup that allowed us to record intracellular calcium variations in astrocytes evoked by sensory stimulation, and to monitor intact neuronal dendritic spines and astrocytic processes as well as blood vessel dynamics. Therefore, in spite of certain limitations that need to be carefully considered, LSCM constitutes a useful, convenient, and affordable tool for brain studies in vivo. PMID:23658537

  1. Spatial Resolution in Scanning Electron Microscopy and Scanning Transmission Electron Microscopy Without a Specimen Vacuum Chamber.

    PubMed

    Nguyen, Kayla X; Holtz, Megan E; Richmond-Decker, Justin; Muller, David A

    2016-08-01

    A long-standing goal of electron microscopy has been the high-resolution characterization of specimens in their native environment. However, electron optics require high vacuum to maintain an unscattered and focused probe, a challenge for specimens requiring atmospheric or liquid environments. Here, we use an electron-transparent window at the base of a scanning electron microscope's objective lens to separate column vacuum from the specimen, enabling imaging under ambient conditions, without a specimen vacuum chamber. We demonstrate in-air imaging of specimens at nanoscale resolution using backscattered scanning electron microscopy (airSEM) and scanning transmission electron microscopy. We explore resolution and contrast using Monte Carlo simulations and analytical models. We find that nanometer-scale resolution can be obtained at gas path lengths up to 400 μm, although contrast drops with increasing gas path length. As the electron-transparent window scatters considerably more than gas at our operating conditions, we observe that the densities and thicknesses of the electron-transparent window are the dominant limiting factors for image contrast at lower operating voltages. By enabling a variety of detector configurations, the airSEM is applicable to a wide range of environmental experiments including the imaging of hydrated biological specimens and in situ chemical and electrochemical processes.

  2. Detecting Changes Following the Provision of Assistive Devices: Utility of the WHO-DAS II

    ERIC Educational Resources Information Center

    Raggi, Alberto

    2010-01-01

    The World Health Organization Disability Assessment Schedule II (WHO-DAS II) is a non-disease-specific International Classification of Functioning, Disability, and Health-based disability assessment instrument developed to measure activity limitations and restrictions to participation. The aim of this pilot study is to evaluate WHO-DAS II…

  3. Psychometric Properties of the Disability Assessment Schedule (DAS) for Behavior Problems: An Independent Investigation

    ERIC Educational Resources Information Center

    Tsakanikos, Elias; Underwood, Lisa; Sturmey, Peter; Bouras, Nick; McCarthy, Jane

    2011-01-01

    The present study employed the Disability Assessment Schedule (DAS) to assess problem behaviors in a large sample of adults with ID (N = 568) and evaluate the psychometric properties of this instrument. Although the DAS problem behaviors were found to be internally consistent (Cronbach's [alpha] = 0.87), item analysis revealed one weak item…

  4. Functional photoacoustic microscopy of diabetic vasculature.

    PubMed

    Krumholz, Arie; Wang, Lidai; Yao, Junjie; Wang, Lihong V

    2012-06-01

    We used functional photoacoustic microscopy to image diabetes-induced damage to the microvasculature. To produce an animal model for Type 1 diabetes, we used streptozotocin (STZ), which is particularly toxic to the insulin-producing beta cells of the pancreas in mammals. A set number of ND4 Swiss Webster mice received intraperitoneal injections of STZ for five consecutive days at 50 mg/kg. Most mice developed a significant rise in blood glucose level (≈ 400 mg/dL) within three weeks of the first injection. Changes in vasculature and hemodynamics were monitored for six weeks. The mouse ear was imaged with an optical-resolution photoacoustic microscope at a main blood vessel branch from the root of the ear. There are noticeable and measurable changes associated with the disease, including decreased vessel diameter and possible occlusion due to vessel damage and polyurea. We also observed an increase in the blood flow speed in the vein and a decrease in the artery, which could be due to compensation for the dehydration and vessel diameter changes. Functional and metabolic parameters such as hemoglobin oxygen saturation, oxygen extraction fraction, and oxygen consumption rate were also measured, but showed no significant change.

  5. [Membrane protein characterization by photoactivatable localization microscopy].

    PubMed

    Huang, Li; Fang, Weihuan; Yu, Ying; Song, Houhui

    2012-11-01

    The on-site labeling and localization tracking of membrane proteins in pathogenic bacteria are tedious work. In order to develop a novel protein labeling technology at super resolution level (nanometer scale) using the photoactivatable localization microscopy (PALM), the chimeric protein of the outer membrane protein A (OmpA) of Mycobacterium tuberculosis and the photoactivatable mEos2m protein were expressed in the non-pathogenic Mycobacterium smegmatis. The recombinant bacteria were fixed on slide, activated by 405 nm laser and subject to PALM imaging to capture photons released by the fusion protein. Meanwhile, colony and cell morphology were visualized under regular fluorescent stereomicroscope and upright fluorescent microscope to characterize fluorescence conversion and protein localization. The fusion proteins formed a "belt"-like structure on cell membrane of M. smegmatis under PALM, providing direct evidence of on-site imaging of membrane proteins. Expression of fusion protein did not compromise the localization properties of OmpA. Thus, mEos2m could be used as a labeling probe to track localizations of non-oligomer oriented membrane proteins. This indicates non-pathogenic M. smegmatis could be served as a model strain to characterize the function and localization of the proteins derived from pathogenic M. tuberculosis. This is the first report using PALM to characterize localization of membrane proteins.

  6. Multiparallel Three-Dimensional Optical Microscopy

    NASA Technical Reports Server (NTRS)

    Nguyen, Lam K.; Price, Jeffrey H.; Kellner, Albert L.; Bravo-Zanoquera, Miguel

    2010-01-01

    Multiparallel three-dimensional optical microscopy is a method of forming an approximate three-dimensional image of a microscope sample as a collection of images from different depths through the sample. The imaging apparatus includes a single microscope plus an assembly of beam splitters and mirrors that divide the output of the microscope into multiple channels. An imaging array of photodetectors in each channel is located at a different distance along the optical path from the microscope, corresponding to a focal plane at a different depth within the sample. The optical path leading to each photodetector array also includes lenses to compensate for the variation of magnification with distance so that the images ultimately formed on all the photodetector arrays are of the same magnification. The use of optical components common to multiple channels in a simple geometry makes it possible to obtain high light-transmission efficiency with an optically and mechanically simple assembly. In addition, because images can be read out simultaneously from all the photodetector arrays, the apparatus can support three-dimensional imaging at a high scanning rate.

  7. Scanning Tunneling Optical Resonance Microscopy Developed

    NASA Technical Reports Server (NTRS)

    Bailey, Sheila G.; Raffaelle, Ryne P.; Lau, Janis E.; Jenkins, Phillip P.; Castro, Stephanie L.; Tin, Padetha; Wilt, David M.; Pal, Anna Maria; Fahey, Stephen D.

    2004-01-01

    The ability to determine the in situ optoelectronic properties of semiconductor materials has become especially important as the size of device architectures has decreased and the development of complex microsystems has increased. Scanning Tunneling Optical Resonance Microscopy, or STORM, can interrogate the optical bandgap as a function of its position within a semiconductor micro-structure. This technique uses a tunable solidstate titanium-sapphire laser whose output is "chopped" using a spatial light modulator and is coupled by a fiber-optic connector to a scanning tunneling microscope in order to illuminate the tip-sample junction. The photoenhanced portion of the tunneling current is spectroscopically measured using a lock-in technique. The capabilities of this technique were verified using semiconductor microstructure calibration standards that were grown by organometallic vapor-phase epitaxy. Bandgaps characterized by STORM measurements were found to be in good agreement with the bulk values determined by transmission spectroscopy and photoluminescence and with the theoretical values that were based on x-ray diffraction results.

  8. Nanoscale Imaging of RNA with Expansion Microscopy

    PubMed Central

    Chen, Fei; Wassie, Asmamaw T.; Cote, Allison J.; Sinha, Anubhav; Alon, Shahar; Asano, Shoh; Daugharthy, Evan R.; Chang, Jae-Byum; Marblestone, Adam; Church, George M.; Raj, Arjun; Boyden, Edward S.

    2016-01-01

    The ability to image RNA identity and location with nanoscale precision in intact tissues is of great interest for defining cell types and states in normal and pathological biological settings. Here, we present a strategy for expansion microscopy (ExM) of RNA. We developed a small molecule linker that enables RNA to be covalently attached to a swellable polyelectrolyte gel synthesized throughout a biological specimen. Then, post-expansion, fluorescent in situ hybridization (FISH) imaging of RNA can be performed with high yield and specificity, with single molecule precision, in both cultured cells and intact brain tissue. Expansion FISH (ExFISH) de-crowds RNAs and supports amplification of single molecule signals (i.e., via hybridization chain reaction (HCR)) as well as multiplexed RNA FISH readout. ExFISH thus enables super-resolution imaging of RNA structure and location with diffraction-limited microscopes in thick specimens, such as intact brain tissue and other tissues of importance to biology and medicine. PMID:27376770

  9. Laser interference microscopy in erythrocyte study

    NASA Astrophysics Data System (ADS)

    Yusipovich, A. I.; Parshina, E. Yu.; Brysgalova, N. Yu.; Brazhe, A. R.; Brazhe, N. A.; Lomakin, A. G.; Levin, G. G.; Maksimov, G. V.

    2009-05-01

    With the laser interference microscopy (LIM) technique, one can measure phase height of cells—a variable proportional to the cell thickness and the difference in the refractive indices of the cell and the surrounding medium. This makes functional optical cell imaging possible, and estimation of shape, thickness, and area of erythrocytes feasible. In this paper, we studied changes in erythrocyte shape and volume with osmolarity and pH. Obtained from the LIM technique, erythrocyte phase heights and area values, as well as the hematocrit-measured erythrocyte volume, were used to estimate changes in the refractive index with osmolarity and pH. A comparison between the estimated refractive index with the refractive index, calculated in the assumption that it can only depend on the hemoglobin concentration in the cell, indicates that these two estimates are identical in the range of osmolarity (250-1000 mOsm) and pH (4.5-10.0) values. Thus, refractive index changes result exclusively from the changes in hemoglobin concentration with the changes in erythrocyte volume. Under these conditions, it is possible to estimate the amount of hemoglobin in an erythrocyte from its phase height and area, obtained from LIM.

  10. Feature Adaptive Sampling for Scanning Electron Microscopy

    PubMed Central

    Dahmen, Tim; Engstler, Michael; Pauly, Christoph; Trampert, Patrick; de Jonge, Niels; Mücklich, Frank; Slusallek, Philipp

    2016-01-01

    A new method for the image acquisition in scanning electron microscopy (SEM) was introduced. The method used adaptively increased pixel-dwell times to improve the signal-to-noise ratio (SNR) in areas of high detail. In areas of low detail, the electron dose was reduced on a per pixel basis, and a-posteriori image processing techniques were applied to remove the resulting noise. The technique was realized by scanning the sample twice. The first, quick scan used small pixel-dwell times to generate a first, noisy image using a low electron dose. This image was analyzed automatically, and a software algorithm generated a sparse pattern of regions of the image that require additional sampling. A second scan generated a sparse image of only these regions, but using a highly increased electron dose. By applying a selective low-pass filter and combining both datasets, a single image was generated. The resulting image exhibited a factor of ≈3 better SNR than an image acquired with uniform sampling on a Cartesian grid and the same total acquisition time. This result implies that the required electron dose (or acquisition time) for the adaptive scanning method is a factor of ten lower than for uniform scanning. PMID:27150131

  11. Imaging mechanisms of force detected FMR microscopy

    SciTech Connect

    Midzor, M. M.; Wigen, P. E.; Pelekhov, D.; Chen, W.; Hammel, P. C.; Roukes, M. L.

    2000-05-01

    We demonstrate spatial resolution of ferromagnetic resonance in a microscopic sample of YIG using ferromagnetic resonance force microscopy (FMRFM). Measurements were performed on a small single crystal YIG film grown on a GGG substrate, roughly rectangular in shape 20 {mu}mx{approx}150 {mu}m and 3 {mu}m thick. The perpendicular and parallel force geometries of FMRFM, in conjunction with an external bias field both parallel and perpendicular to the film, were used to scan the sample. This enabled the detection of strong signals, even at atmospheric pressure and room temperature. The fundamental and higher-order magnetostatic modes were observed to have 26-29 Gauss separation. The intensity of these modes exhibited spatial variation as the magnetic tip was scanned over the sample, and this behavior is qualitatively explained by DE theory. An improved fabrication method for magnet on cantilever was employed, which yielded a spatial resolution of 15 {mu}m. These results demonstrate the potential of FMRFM for investigating the spatial dependence of ferromagnetic resonance, and for studying the anisotropy fields and exchange coupling effects within multilayer films and small magnetic systems. (c) 2000 American Institute of Physics.

  12. Differential interference contrast microscopy using Savart plates

    NASA Astrophysics Data System (ADS)

    Trịnh, Hưng-Xuân; Lin, Shyh-Tsong; Chen, Liang-Chia; Yeh, Sheng-Lih; Chen, Chin-Sheng

    2017-04-01

    A new differential interference contrast microscopy (DICM), which uses Savart prisms as a shearing plate and a phase-shifting device, is proposed. The system consists of a phase-shifting module (PSM) and a DICM module (DICMM). The PSM has two Savart prisms: the first prism separates the incident beam into two parallel beams, and the second prism recombines these two beams. The optical path difference (OPD) of the two beams, which is represented by a biased OPD, can be adjusted by rotating the angle of the normal surface of the second prism. In the DICMM, the other Savart prism is used to replace the Nomarski prism (NP) in conventional DICM. It combines with an afocal microscopic system (AMS) to produce a Savart-DICM system, which is able to perform a phase-shifting technique by changing the biased OPD to produce a phase shift of π/2 for each step. This paper describes the configuration and measurement theory of the microscope. The experimental results confirm the validity and capability of the proposed microscope.

  13. Stimulated Raman microscopy without ultrafast lasers

    NASA Astrophysics Data System (ADS)

    Meng, Zhaokai; Petrov, Georgi I.; Yakovlev, Vladislav V.

    2013-02-01

    Stimulated Raman scattering (SRS) microscopy is a powerful tool for chemically-sensitive non-invasive optical imaging. However, the short-pulse laser sources, which are currently being employed for this imaging technique, are still expensive and require substantial maintenance to provide temporal and spectral overlap. SRS imaging, which utilizes cw laser sources, has a major advantage over pulsed lasers, as it eliminates the possibility of cell damage due to exposure to high-intensity light radiation, while substantially reducing the cost and complexity of the set-up and keeping a sub-cellular spatial resolution. As a proof-of-principle, we demonstrate microscopic imaging of dimethyl sulfoxide using two independent, commonly used and inexpensive lasers: a diode-pumped, intracavity doubled 532 nm laser and a He-Ne laser operating at 633 nm. In our proof-of-principle experience, dimethyl sulfoxide acts as a contrast agent providing Raman scattering signal. The 532 nm and 633 nm lasers act as excitation and probe sources, respectively [1].

  14. Electron microscopy in the investigation of asthenozoospermia.

    PubMed

    Mobberley, M A

    2010-01-01

    Asthenozoospermia, defined as low sperm motility, is a significant cause of subfertility in men. Its origins are diverse and in some instances cannot be ascertained. However, severely reduced motility can often be associated with abnormalities in the structure of the sperm tails, which can only be detected by transmission electron microscopy (TEM). In this respect, TEM is an important adjunct to the traditional methods of semen analysis. This review examines the development of the current state of knowledge of sperm tail abnormalities. These may be genetic in origin, or they may be acquired as a result of extrinsic factors. At present, consistent molecular markers are not available to characterise many of the genetic defects. However, TEM can distinguish specific defects of genetic origin and the non-specific structural anomalies that are typical of an acquired condition. It can also differentiate sperm structural anomalies from necrospermia, or sperm death, which is another significant cause of asthenozoospermia. In this modern era of assisted reproduction, it is possible in some instances to circumvent the problems of sperm immotility and to achieve fertilisation and pregnancy using intracytoplasmic sperm injection (ICSI). However, because of the possible genetic origin of asthenozoospermia, many scientists working in the field of infertility believe that it is of the utmost importance to investigate the causes of asthenozoospermia. This review considers the continuing relevance of TEM to the evaluation of sperm tail abnormalities in the context of current reproductive techniques.

  15. Imaging Cytoskeleton Components by Electron Microscopy

    PubMed Central

    Svitkina, Tatyana

    2016-01-01

    The cytoskeleton is a complex of detergent-insoluble components of the cytoplasm playing critical roles in cell motility, shape generation, and mechanical properties of a cell. Fibrillar polymers—actin filaments, microtubules, and intermediate filaments—are major constituents of the cytoskeleton, which constantly change their organization during cellular activities. The actin cytoskeleton is especially polymorphic, as actin filaments can form multiple higher order assemblies performing different functions. Structural information about cytoskeleton organization is critical for understanding its functions and mechanisms underlying various forms of cellular activity. Because of the nanometer-scale thickness of cytoskeletal fibers, electron microscopy (EM) is a key tool to determine the structure of the cytoskeleton. This article describes application of rotary shadowing (or metal replica) EM for visualization of the cytoskeleton. The procedure is applicable to thin cultured cells growing on glass coverslips and consists of detergent extraction of cells to expose their cytoskeleton, chemical fixation to provide stability, ethanol dehydration and critical point drying to preserve three-dimensionality, rotary shadowing with platinum to create contrast, and carbon coating to stabilize replicas. This technique provides easily interpretable three-dimensional images, in which individual cytoskeletal fibers are clearly resolved, and individual proteins can be identified by immunogold labeling. More importantly, replica EM is easily compatible with live cell imaging, so that one can correlate the dynamics of a cell or its components, e.g., expressed fluorescent proteins, with high resolution structural organization of the cytoskeleton in the same cell. PMID:26498781

  16. Modulated CMOS camera for fluorescence lifetime microscopy.

    PubMed

    Chen, Hongtao; Holst, Gerhard; Gratton, Enrico

    2015-12-01

    Widefield frequency-domain fluorescence lifetime imaging microscopy (FD-FLIM) is a fast and accurate method to measure the fluorescence lifetime of entire images. However, the complexity and high costs involved in construction of such a system limit the extensive use of this technique. PCO AG recently released the first luminescence lifetime imaging camera based on a high frequency modulated CMOS image sensor, QMFLIM2. Here we tested and provide operational procedures to calibrate the camera and to improve the accuracy using corrections necessary for image analysis. With its flexible input/output options, we are able to use a modulated laser diode or a 20 MHz pulsed white supercontinuum laser as the light source. The output of the camera consists of a stack of modulated images that can be analyzed by the SimFCS software using the phasor approach. The nonuniform system response across the image sensor must be calibrated at the pixel level. This pixel calibration is crucial and needed for every camera settings, e.g. modulation frequency and exposure time. A significant dependency of the modulation signal on the intensity was also observed and hence an additional calibration is needed for each pixel depending on the pixel intensity level. These corrections are important not only for the fundamental frequency, but also for the higher harmonics when using the pulsed supercontinuum laser. With these post data acquisition corrections, the PCO CMOS-FLIM camera can be used for various biomedical applications requiring a large frame and high speed acquisition.

  17. Characterization of nanomaterials with transmission electron microscopy

    NASA Astrophysics Data System (ADS)

    Anjum, D. H.

    2016-08-01

    The field of nanotechnology is about research and development on materials whose at least one dimension is in the range of 1 to 100 nanometers. In recent years, the research activity for developing nano-materials has grown exponentially owing to the fact that they offer better solutions to the challenges faced by various fields such as energy, food, and environment. In this paper, the importance of transmission electron microscopy (TEM) based techniques is demonstrated for investigating the properties of nano-materials. Specifically the nano-materials that are investigated in this report include gold nano-particles (Au-NPs), silver atom-clusters (Ag-ACs), tantalum single-atoms (Ta-SAs), carbon materials functionalized with iron cobalt (Fe-Co) NPs and titania (TiO2) NPs, and platinum loaded Ceria (Pt-CeO2) Nano composite. TEM techniques that are employed to investigate nano-materials include aberration corrected bright-field TEM (BF-TEM), high-angle dark-field scanning TEM (HAADF-STEM), electron energy-loss spectroscopy (EELS), and BF-TEM electron tomography (ET). With the help presented of results in this report, it is proved herein that as many TEM techniques as available in a given instrument are essential for a comprehensive nano-scale analysis of nanomaterials.

  18. Spinning-Spot Shadowless TIRF Microscopy.

    PubMed

    Ellefsen, Kyle L; Dynes, Joseph L; Parker, Ian

    2015-01-01

    Total internal reflection fluorescence (TIRF) microscopy is a powerful tool for visualizing near-membrane cellular structures and processes, including imaging of local Ca2+ transients with single-channel resolution. TIRF is most commonly implemented in epi-fluorescence mode, whereby laser excitation light is introduced at a spot near the periphery of the back focal plane of a high numerical aperture objective lens. However, this approach results in an irregular illumination field, owing to interference fringes and scattering and shadowing by cellular structures. We describe a simple system to circumvent these limitations, utilizing a pair of galvanometer-driven mirrors to rapidly spin the laser spot in a circle at the back focal plane of the objective lens, so that irregularities average out during each camera exposure to produce an effectively uniform field. Computer control of the mirrors enables precise scanning at 200 Hz (5ms camera exposure times) or faster, and the scan radius can be altered on a frame-by-frame basis to achieve near-simultaneous imaging in TIRF, widefield and 'skimming plane' imaging modes. We demonstrate the utility of the system for dynamic recording of local inositol trisphosphate-mediated Ca2+ signals and for imaging the redistribution of STIM and Orai proteins during store-operated Ca2+ entry. We further anticipate that it will be readily applicable for numerous other near-membrane studies, especially those involving fast dynamic processes.

  19. Investigating bioconjugation by atomic force microscopy

    PubMed Central

    2013-01-01

    Nanotechnological applications increasingly exploit the selectivity and processivity of biological molecules. Integration of biomolecules such as proteins or DNA into nano-systems typically requires their conjugation to surfaces, for example of carbon-nanotubes or fluorescent quantum dots. The bioconjugated nanostructures exploit the unique strengths of both their biological and nanoparticle components and are used in diverse, future oriented research areas ranging from nanoelectronics to biosensing and nanomedicine. Atomic force microscopy imaging provides valuable, direct insight for the evaluation of different conjugation approaches at the level of the individual molecules. Recent technical advances have enabled high speed imaging by AFM supporting time resolutions sufficient to follow conformational changes of intricately assembled nanostructures in solution. In addition, integration of AFM with different spectroscopic and imaging approaches provides an enhanced level of information on the investigated sample. Furthermore, the AFM itself can serve as an active tool for the assembly of nanostructures based on bioconjugation. AFM is hence a major workhorse in nanotechnology; it is a powerful tool for the structural investigation of bioconjugation and bioconjugation-induced effects as well as the simultaneous active assembly and analysis of bioconjugation-based nanostructures. PMID:23855448

  20. High-Resolution Traction Force Microscopy

    PubMed Central

    Plotnikov, Sergey V.; Sabass, Benedikt; Schwarz, Ulrich S.; Waterman, Clare M.

    2015-01-01

    Cellular forces generated by the actomyosin cytoskeleton and transmitted to the extracellular matrix (ECM) through discrete, integrin-based protein assemblies, that is, focal adhesions, are critical to developmental morphogenesis and tissue homeostasis, as well as disease progression in cancer. However, quantitative mapping of these forces has been difficult since there has been no experimental technique to visualize nanonewton forces at submicrometer spatial resolution. Here, we provide detailed protocols for measuring cellular forces exerted on two-dimensional elastic substrates with a high-resolution traction force microscopy (TFM) method. We describe fabrication of polyacrylamide substrates labeled with multiple colors of fiducial markers, functionalization of the substrates with ECM proteins, setting up the experiment, and imaging procedures. In addition, we provide the theoretical background of traction reconstruction and experimental considerations important to design a high-resolution TFM experiment. We describe the implementation of a new algorithm for processing of images of fiducial markers that are taken below the surface of the substrate, which significantly improves data quality. We demonstrate the application of the algorithm and explain how to choose a regularization parameter for suppression of the measurement error. A brief discussion of different ways to visualize and analyze the results serves to illustrate possible uses of high-resolution TFM in biomedical research. PMID:24974038

  1. Complete information acquisition in scanning probe microscopy

    DOE PAGES

    Belianinov, Alex; Kalinin, Sergei V.; Jesse, Stephen

    2015-03-13

    In the last three decades, scanning probe microscopy (SPM) has emerged as a primary tool for exploring and controlling the nanoworld. A critical part of the SPM measurements is the information transfer from the tip-surface junction to a macroscopic measurement system. This process reduces the many degrees of freedom of a vibrating cantilever to relatively few parameters recorded as images. Similarly, the details of dynamic cantilever response at sub-microsecond time scales of transients, higher-order eigenmodes and harmonics are averaged out by transitioning to millisecond time scale of pixel acquisition. Hence, the amount of information available to the external observer ismore » severely limited, and its selection is biased by the chosen data processing method. Here, we report a fundamentally new approach for SPM imaging based on information theory-type analysis of the data stream from the detector. This approach allows full exploration of complex tip-surface interactions, spatial mapping of multidimensional variability of material s properties and their mutual interactions, and SPM imaging at the information channel capacity limit.« less

  2. Spectral fusing Gabor domain optical coherence microscopy.

    PubMed

    Meemon, Panomsak; Widjaja, Joewono; Rolland, Jannick P

    2016-02-01

    Gabor domain optical coherence microscopy (GD-OCM) is one of many variations of optical coherence tomography (OCT) techniques that aims for invariant high resolution across a 3D field of view by utilizing the ability to dynamically refocus the imaging optics in the sample arm. GD-OCM acquires multiple cross-sectional images at different focus positions of the objective lens, and then fuses them to obtain an invariant high-resolution 3D image of the sample, which comes with the intrinsic drawback of a longer processing time as compared to conventional Fourier domain OCT. Here, we report on an alternative Gabor fusing algorithm, the spectral-fusion technique, which directly processes each acquired spectrum and combines them prior to the Fourier transformation to obtain a depth profile. The implementation of the spectral-fusion algorithm is presented and its performance is compared to that of the prior GD-OCM spatial-fusion approach. The spectral-fusion approach shows twice the speed of the spatial-fusion approach for a spectrum size of less than 2000 point sampling, which is a commonly used spectrum size in OCT imaging, including GD-OCM.

  3. In vivo confocal microscopy of toxic keratopathy

    PubMed Central

    Chen, Y; Le, Q; Hong, J; Gong, L; Xu, J

    2017-01-01

    Purpose To explore the morphological characteristics of toxic keratopathy (TK), which clinically presented as superficial punctate keratopathy (SPK), with the application of in vivo laser-scanning confocal microscopy (LSCM), and evaluate its potential in the early diagnosis of TK. Patients and methods This was a cross-sectional study involving 16 patients with TK and 16 patients with dry eye (DE), demonstrating SPK under slit-lamp observation, and 10 normal eyes were enrolled in the study. All participants underwent history interviews, fluorescein staining, tear film break-up time (BUT) tests, Schirmer tests, and in vivo LSCM. Results The area grading of corneal fluorescein punctate staining was higher in the TK group than the DE group. Measured by in vivo LSCM, superficial epithelial cell density was lower in the TK group than that of DE group. The sub-basal nerve presented lower density and tortuosity in the TK group than the DE group. Most notably, deposits with a snow-like appearance were observed in the epithelium in 12/16 (75.0%) of the TK cases, but none in the DE group (P<0.05). Conclusion The SPK in TK patients was characterized by more widespread punctate staining, a lower density of superficial epithelial cells and sub-basal nerves, and a typical snow-like pattern of deposits in the epithelium by LSCM. These features might facilitate early diagnosis of TK from other disorders manifested as SPK. PMID:27740620

  4. Intermodulation Atomic Force Microscopy and Spectroscopy

    NASA Astrophysics Data System (ADS)

    Hutter, Carsten; Platz, Daniel; Tholen, Erik; Haviland, David; Hansson, Hans

    2009-03-01

    We present a powerful new method of dynamic AFM, which allows to gain far more information about the tip-surface interaction than standard amplitude or phase imaging, while scanning at comparable speed. Our method, called intermodulation atomic force microscopy (ImAFM), employs the manifestly nonlinear phenomenon of intermodulation to extract information about tip-surface forces. ImAFM uses one eigenmode of a mechanical resonator, the latter driven at two frequencies to produce many spectral peaks near its resonace, where sensitivity is highest [1]. We furthermore present a protocol for decoding the combined information encoded in the spectrum of intermodulation peaks. Our theoretical framework suggests methods to enhance the gained information by using a different parameter regime as compared to Ref. [1]. We also discuss strategies for solving the inverse problem, i.e., for extracting the nonlinear tip-surface interaction from the response, also naming limitations of our theoretical analysis. We will further report on latest progress to experimentally employ our new protocol.[3pt] [1] D. Platz, E. A. Tholen, D. Pesen, and D. B. Haviland, Appl. Phys. Lett. 92, 153106 (2008).

  5. Radio-frequency scanning tunnelling microscopy.

    PubMed

    Kemiktarak, U; Ndukum, T; Schwab, K C; Ekinci, K L

    2007-11-01

    The scanning tunnelling microscope (STM) relies on localized electron tunnelling between a sharp probe tip and a conducting sample to attain atomic-scale spatial resolution. In the 25-year period since its invention, the STM has helped uncover a wealth of phenomena in diverse physical systems--ranging from semiconductors to superconductors to atomic and molecular nanosystems. A severe limitation in scanning tunnelling microscopy is the low temporal resolution, originating from the diminished high-frequency response of the tunnel current readout circuitry. Here we overcome this limitation by measuring the reflection from a resonant inductor-capacitor circuit in which the tunnel junction is embedded, and demonstrate electronic bandwidths as high as 10 MHz. This approximately 100-fold bandwidth improvement on the state of the art translates into fast surface topography as well as delicate measurements in mesoscopic electronics and mechanics. Broadband noise measurements across the tunnel junction using this radio-frequency STM have allowed us to perform thermometry at the nanometre scale. Furthermore, we have detected high-frequency mechanical motion with a sensitivity approaching approximately 15 fm Hz(-1/2). This sensitivity is on par with the highest available from nanoscale optical and electrical displacement detection techniques, and the radio-frequency STM is expected to be capable of quantum-limited position measurements.

  6. High voltage electron microscopy of lunar samples

    NASA Technical Reports Server (NTRS)

    Fernandez-Moran, H.

    1973-01-01

    Lunar pyroxenes from Apollo 11, 12, 14, and 15 were investigated. The iron-rich and magnesium-rich pyroxene specimens were crushed to a grain size of ca. 50 microns and studied by a combination of X-ray and electron diffraction, electron microscopy, 57 Fe Mossbauer spectroscopy and X-ray crystallography techniques. Highly ordered, uniform electron-dense bands, corresponding to exsolution lamellae, with average widths of ca. 230A to 1000A dependent on the source specimen were observed. These were?qr separated by wider, less-dense interband spacings with average widths of ca. 330A to 3100A. In heating experiments, splitting of the dense bands into finer structures, leading finally to obliteration of the exsolution lamellae was recorded. The extensive exsolution is evidence for significantly slower cooling rates, or possibly annealing, at temperatures in the subsolidus range, adding evidence that annealing of rock from the surface of the moon took place at ca. 600 C. Correlation of the band structure with magnetic ordering at low temperatures and iron clustering within the bands was studied.

  7. Scanning electron microscopy of softened enamel.

    PubMed

    Eisenburger, M; Shellis, R P; Addy, M

    2004-01-01

    After exposing enamel specimens to 0.3% citric acid at pH 3.2 for various times, the acid was titrated to pH 7 before rinsing the specimens in water. After freeze-drying the specimens were examined by scanning electron microscopy. This procedure eliminates artefacts due to drying and mineral precipitation. The results showed that the outer region of softened enamel is much more delicate than previously thought, even after short (5- to 20-min) etching times. Mineral was lost from both prism boundaries and the prism bodies, resulting in a surface presenting thin, separate crystal bundles. In further studies, replicas of subsurface pores, created by resin impregnation, showed the softening depth to be several times greater than is suggested by techniques based on removing the softened enamel by physical forces. The results point to a need for improved methods of measuring softening depth. More importantly, it appears that the outer region of the softened layer remaining after an erosive challenge might be too fragile to resist frictional forces in vivo.

  8. Spinning-Spot Shadowless TIRF Microscopy

    PubMed Central

    Ellefsen, Kyle L.; Dynes, Joseph L.; Parker, Ian

    2015-01-01

    Total internal reflection fluorescence (TIRF) microscopy is a powerful tool for visualizing near-membrane cellular structures and processes, including imaging of local Ca2+ transients with single-channel resolution. TIRF is most commonly implemented in epi-fluorescence mode, whereby laser excitation light is introduced at a spot near the periphery of the back focal plane of a high numerical aperture objective lens. However, this approach results in an irregular illumination field, owing to interference fringes and scattering and shadowing by cellular structures. We describe a simple system to circumvent these limitations, utilizing a pair of galvanometer-driven mirrors to rapidly spin the laser spot in a circle at the back focal plane of the objective lens, so that irregularities average out during each camera exposure to produce an effectively uniform field. Computer control of the mirrors enables precise scanning at 200 Hz (5ms camera exposure times) or faster, and the scan radius can be altered on a frame-by-frame basis to achieve near-simultaneous imaging in TIRF, widefield and ‘skimming plane’ imaging modes. We demonstrate the utility of the system for dynamic recording of local inositol trisphosphate-mediated Ca2+ signals and for imaging the redistribution of STIM and Orai proteins during store-operated Ca2+ entry. We further anticipate that it will be readily applicable for numerous other near-membrane studies, especially those involving fast dynamic processes. PMID:26308212

  9. Investigating cell mechanics with atomic force microscopy.

    PubMed

    Haase, Kristina; Pelling, Andrew E

    2015-03-06

    Transmission of mechanical force is crucial for normal cell development and functioning. However, the process of mechanotransduction cannot be studied in isolation from cell mechanics. Thus, in order to understand how cells 'feel', we must first understand how they deform and recover from physical perturbations. Owing to its versatility, atomic force microscopy (AFM) has become a popular tool to study intrinsic cellular mechanical properties. Used to directly manipulate and examine whole and subcellular reactions, AFM allows for top-down and reconstitutive approaches to mechanical characterization. These studies show that the responses of cells and their components are complex, and largely depend on the magnitude and time scale of loading. In this review, we generally describe the mechanotransductive process through discussion of well-known mechanosensors. We then focus on discussion of recent examples where AFM is used to specifically probe the elastic and inelastic responses of single cells undergoing deformation. We present a brief overview of classical and current models often used to characterize observed cellular phenomena in response to force. Both simple mechanistic models and complex nonlinear models have been used to describe the observed cellular behaviours, however a unifying description of cell mechanics has not yet been resolved.

  10. Complete information acquisition in scanning probe microscopy

    SciTech Connect

    Belianinov, Alex; Kalinin, Sergei V.; Jesse, Stephen

    2015-03-13

    In the last three decades, scanning probe microscopy (SPM) has emerged as a primary tool for exploring and controlling the nanoworld. A critical part of the SPM measurements is the information transfer from the tip-surface junction to a macroscopic measurement system. This process reduces the many degrees of freedom of a vibrating cantilever to relatively few parameters recorded as images. Similarly, the details of dynamic cantilever response at sub-microsecond time scales of transients, higher-order eigenmodes and harmonics are averaged out by transitioning to millisecond time scale of pixel acquisition. Hence, the amount of information available to the external observer is severely limited, and its selection is biased by the chosen data processing method. Here, we report a fundamentally new approach for SPM imaging based on information theory-type analysis of the data stream from the detector. This approach allows full exploration of complex tip-surface interactions, spatial mapping of multidimensional variability of material s properties and their mutual interactions, and SPM imaging at the information channel capacity limit.

  11. Nanoscale chemical imaging by photoinduced force microscopy

    PubMed Central

    Nowak, Derek; Morrison, William; Wickramasinghe, H. Kumar; Jahng, Junghoon; Potma, Eric; Wan, Lei; Ruiz, Ricardo; Albrecht, Thomas R.; Schmidt, Kristin; Frommer, Jane; Sanders, Daniel P.; Park, Sung

    2016-01-01

    Correlating spatial chemical information with the morphology of closely packed nanostructures remains a challenge for the scientific community. For example, supramolecular self-assembly, which provides a powerful and low-cost way to create nanoscale patterns and engineered nanostructures, is not easily interrogated in real space via existing nondestructive techniques based on optics or electrons. A novel scanning probe technique called infrared photoinduced force microscopy (IR PiFM) directly measures the photoinduced polarizability of the sample in the near field by detecting the time-integrated force between the tip and the sample. By imaging at multiple IR wavelengths corresponding to absorption peaks of different chemical species, PiFM has demonstrated the ability to spatially map nm-scale patterns of the individual chemical components of two different types of self-assembled block copolymer films. With chemical-specific nanometer-scale imaging, PiFM provides a powerful new analytical method for deepening our understanding of nanomaterials. PMID:27051870

  12. Functional photoacoustic microscopy of diabetic vasculature

    NASA Astrophysics Data System (ADS)

    Krumholz, Arie; Wang, Lidai; Yao, Junjie; Wang, Lihong V.

    2012-06-01

    We used functional photoacoustic microscopy to image diabetes-induced damage to the microvasculature. To produce an animal model for Type 1 diabetes, we used streptozotocin (STZ), which is particularly toxic to the insulin-producing beta cells of the pancreas in mammals. A set number of ND4 Swiss Webster mice received intraperitoneal injections of STZ for five consecutive days at 50 mg/kg. Most mice developed a significant rise in blood glucose level (~400 mg/dL) within three weeks of the first injection. Changes in vasculature and hemodynamics were monitored for six weeks. The mouse ear was imaged with an optical-resolution photoacoustic microscope at a main blood vessel branch from the root of the ear. There are noticeable and measurable changes associated with the disease, including decreased vessel diameter and possible occlusion due to vessel damage and polyurea. We also observed an increase in the blood flow speed in the vein and a decrease in the artery, which could be due to compensation for the dehydration and vessel diameter changes. Functional and metabolic parameters such as hemoglobin oxygen saturation, oxygen extraction fraction, and oxygen consumption rate were also measured, but showed no significant change.

  13. Investigating cell mechanics with atomic force microscopy

    PubMed Central

    Haase, Kristina; Pelling, Andrew E.

    2015-01-01

    Transmission of mechanical force is crucial for normal cell development and functioning. However, the process of mechanotransduction cannot be studied in isolation from cell mechanics. Thus, in order to understand how cells ‘feel’, we must first understand how they deform and recover from physical perturbations. Owing to its versatility, atomic force microscopy (AFM) has become a popular tool to study intrinsic cellular mechanical properties. Used to directly manipulate and examine whole and subcellular reactions, AFM allows for top-down and reconstitutive approaches to mechanical characterization. These studies show that the responses of cells and their components are complex, and largely depend on the magnitude and time scale of loading. In this review, we generally describe the mechanotransductive process through discussion of well-known mechanosensors. We then focus on discussion of recent examples where AFM is used to specifically probe the elastic and inelastic responses of single cells undergoing deformation. We present a brief overview of classical and current models often used to characterize observed cellular phenomena in response to force. Both simple mechanistic models and complex nonlinear models have been used to describe the observed cellular behaviours, however a unifying description of cell mechanics has not yet been resolved. PMID:25589563

  14. Doppler encoded excitation pattern tomographic optical microscopy.

    PubMed

    Feldkhun, Daniel; Wagner, Kelvin H

    2010-12-01

    Most far-field optical imaging systems rely on lenses and spatially resolved detection to probe distinct locations on the object. We describe and demonstrate a high-speed wide-field approach to imaging that instead measures the complex spatial Fourier transform of the object by detecting its spatially integrated response to dynamic acousto-optically synthesized structured illumination. Tomographic filtered backprojection is applied to reconstruct the object in two or three dimensions. This technique decouples depth of field and working distance from resolution, in contrast to conventional imaging, and can be used to image biological and synthetic structures in fluoresced or scattered light employing coherent or broadband illumination. We discuss the electronically programmable transfer function of the optical system and its implications for imaging dynamic processes. We also explore wide-field fluorescence imaging in scattering media by coherence gating. Finally, we present two-dimensional high-resolution tomographic image reconstructions in both scattered and fluoresced light demonstrating a thousandfold improvement in the depth of field compared to conventional lens-based microscopy.

  15. 3D Viscoelastic traction force microscopy.

    PubMed

    Toyjanova, Jennet; Hannen, Erin; Bar-Kochba, Eyal; Darling, Eric M; Henann, David L; Franck, Christian

    2014-10-28

    Native cell-material interactions occur on materials differing in their structural composition, chemistry, and physical compliance. While the last two decades have shown the importance of traction forces during cell-material interactions, they have been almost exclusively presented on purely elastic in vitro materials. Yet, most bodily tissue materials exhibit some level of viscoelasticity, which could play an important role in how cells sense and transduce tractions. To expand the realm of cell traction measurements and to encompass all materials from elastic to viscoelastic, this paper presents a general, and comprehensive approach for quantifying 3D cell tractions in viscoelastic materials. This methodology includes the experimental characterization of the time-dependent material properties for any viscoelastic material with the subsequent mathematical implementation of the determined material model into a 3D traction force microscopy (3D TFM) framework. Utilizing this new 3D viscoelastic TFM (3D VTFM) approach, we quantify the influence of viscosity on the overall material traction calculations and quantify the error associated with omitting time-dependent material effects, as is the case for all other TFM formulations. We anticipate that the 3D VTFM technique will open up new avenues of cell-material investigations on even more physiologically relevant time-dependent materials including collagen and fibrin gels.

  16. Electron microscopy and theoretical modeling of cochleates.

    PubMed

    Nagarsekar, Kalpa; Ashtikar, Mukul; Thamm, Jana; Steiniger, Frank; Schacher, Felix; Fahr, Alfred; May, Sylvio

    2014-11-11

    Cochleates are self-assembled cylindrical condensates that consist of large rolled-up lipid bilayer sheets and represent a novel platform for oral and systemic delivery of therapeutically active medicinal agents. With few preceding investigations, the physical basis of cochleate formation has remained largely unexplored. We address the structure and stability of cochleates in a combined experimental/theoretical approach. Employing different electron microscopy methods, we provide evidence for cochleates consisting of phosphatidylserine and calcium to be hollow tubelike structures with a well-defined constant lamellar repeat distance and statistically varying inner and outer radii. To rationalize the relation between inner and outer radii, we propose a theoretical model. Based on the minimization of a phenomenological free energy expression containing a bending, adhesion, and frustration contribution, we predict the optimal tube dimensions of a cochleate and estimate ratios of material constants for cochleates consisting of phosphatidylserines with varied hydrocarbon chain structures. Knowing and understanding these ratios will ultimately benefit the successful formulation of cochleates for drug delivery applications.

  17. Visualizing the podocyte with multiphoton microscopy

    PubMed Central

    Khoury, Charbel C.; Khayat, Mark F.; Yeo, Tet-Kin; Pyagay, Petr E.; Wang, Amy; Asuncion, Allan M.; Sharma, Kumar; Yu, Weiming; Chen, Sheldon

    2012-01-01

    The podocyte is a highly specialized kidney glomerular epithelial cell that plays an essential role in glomerular filtration and is believed to be the target of numerous glomerular diseases leading to proteinuria. Despite the leaps in our understanding of podocyte biology, new methodologies are needed to facilitate research into the cell. Multiphoton microscopy (MPM) was used to image the nephrin knockout/green fluorescent protein (GFP) knock-in heterozygote (Nphs1tm1Rkl/J) mouse. The nephrin promoter restricts GFP expression to the podocytes that fluoresce green under excitation. From the exterior of an intact kidney, MPM can peer into the renal parenchyma and visualize the podocytes that outline the globular shape of the glomeruli. Details as fine as the podocyte’s secondary processes can be resolved. In contrast, podocytes exhibit no fluorescence in the wildtype mouse and are invisible to MPM. Phenotypically, there are no significant differences between wildtype and Nphs1tm1Rkl/J mice in body weight, urinary albumin excretion, creatinine clearance, or glomerular depth. Interestingly, the glomeruli are closer to the kidney capsule in female mice, making the gender the preferred choice for MPM. For the first time, green fluorescent podocytes in a mouse model free of confounding phenotypes can be visualized unequivocally and in the “positive” by MPM, facilitating intravital studies of the podocyte. PMID:23022193

  18. Catalysis resolved using scanning tunnelling microscopy.

    PubMed

    Bowker, Michael

    2007-10-01

    The technique of scanning tunnelling microscopy has revolutionised our understanding of surface chemistry, due to its ability to image at the atomic and molecular scale, the very realm at which chemistry operates. This critical review focuses on its contribution to the resolution of various problems in heterogeneous catalysis, including surface structure, surface intermediates, active sites and spillover. In the article a number of images of surfaces are shown, many at atomic resolution, and the insights which these give into surface reactivity are invaluable. The article should be of interest to catalytic chemists, surface and materials scientists and those involved with nanotechnology/nanoscience. (129 references.)The graphical abstract shows the reaction between gas phase methanol and oxygen islands on Cu(110), courtesy of Philip Davies of Cardiff University. The added-row island is shown as silver-coloured spheres (copper) and red (oxygen) on the copper surface. Methanol preferentially reacts with the terminal oxygen atoms in the island forming adsorbed methoxy and OH groups. Only the terminal oxygen atoms in the island are active sites for the reaction.

  19. Ultrasensitive Silicon Cantilever for Force Microscopy

    NASA Astrophysics Data System (ADS)

    Stowe, Timothy; Yasumura, Kevin; Thomas, Kenny; Daniel, Rugar

    1996-03-01

    A new type of ultrasensitive high Q scanning force probe has been developed which is capable of measuring forces smaller than 10-16 N in vacuum at a few degK. These single crystal silicon probes are 500 Åto 1700 Åthick and have in-plane tips with a radius of curvature under 90 nm. Because of their low spring constant (10-4 N/m) these cantilevers were placed normal to the sample surface thereby avoiding tip crashes due to snap in and were vibrated in the pendulum mode. Cantilever amplitude was detected using a fiber interferometer. Measurements of the Q vs. distance from sample surfaces in vacuum and nonlinear effects have been obtained. Measurements Q vs. temperature and ways of improving the Q will be discussed. This scanning force probe may be used to extend the spatial resolution and force sensitivity of Magnetic Resonance Force Microscopy. This research has been partially supported by the Fannie-Hertz Foundation.

  20. Multifocal multiphoton microscopy with adaptive optical correction

    NASA Astrophysics Data System (ADS)

    Coelho, Simao; Poland, Simon; Krstajic, Nikola; Li, David; Monypenny, James; Walker, Richard; Tyndall, David; Ng, Tony; Henderson, Robert; Ameer-Beg, Simon

    2013-02-01

    Fluorescence lifetime imaging microscopy (FLIM) is a well established approach for measuring dynamic signalling events inside living cells, including detection of protein-protein interactions. The improvement in optical penetration of infrared light compared with linear excitation due to Rayleigh scattering and low absorption have provided imaging depths of up to 1mm in brain tissue but significant image degradation occurs as samples distort (aberrate) the infrared excitation beam. Multiphoton time-correlated single photon counting (TCSPC) FLIM is a method for obtaining functional, high resolution images of biological structures. In order to achieve good statistical accuracy TCSPC typically requires long acquisition times. We report the development of a multifocal multiphoton microscope (MMM), titled MegaFLI. Beam parallelization performed via a 3D Gerchberg-Saxton (GS) algorithm using a Spatial Light Modulator (SLM), increases TCSPC count rate proportional to the number of beamlets produced. A weighted 3D GS algorithm is employed to improve homogeneity. An added benefit is the implementation of flexible and adaptive optical correction. Adaptive optics performed by means of Zernike polynomials are used to correct for system induced aberrations. Here we present results with significant improvement in throughput obtained using a novel complementary metal-oxide-semiconductor (CMOS) 1024 pixel single-photon avalanche diode (SPAD) array, opening the way to truly high-throughput FLIM.

  1. Reflectance Confocal Microscopy for Inflammatory Skin Diseases.

    PubMed

    Agozzino, M; Gonzalez, S; Ardigò, M

    2016-10-01

    In vivo reflectance confocal microscopy (RCM) is a relatively novel non-invasive tool for microscopic evaluation of the skin used prevalently for diagnosis and management of skin tumour. Its axial resolution, its non-invasive and easy clinical application represents the goals for a large diffusion of this technique. During the last 15 years, RCM has been demonstrated to be able to increase the sensibility and sensitivity of dermoscopy in the diagnosis of skin tumours integrating in real time clinic, dermoscopic and microscopic information useful for the definition of malignancy. Despite to date, no large comparative studies on inflammatory skin diseases has been published in the literature, several papers already showed that RCM has a potential for the evaluation of the descriptive features of the most common inflammatory skin diseases as psoriasis, lupus erythematosus, contact dermatitis and others. The aim of the application of this technique in non-neoplastic skin diseases has been prevalently focused on the possibility of clinical diagnosis confirmation, as well as therapeutic management. Moreover, the use of RCM as driver for an optimised skin biopsy has been also followed in order to reduce the number of unsuccessful histopathological examination. In this review article we describe the confocal features of the major groups of inflammatory skin disorders focusing on psoriasiform dermatitis, interface dermatitis and spongiotic dermatitis.

  2. Assessment of breast pathologies using nonlinear microscopy

    PubMed Central

    Tao, Yuankai K.; Shen, Dejun; Sheikine, Yuri; Ahsen, Osman O.; Wang, Helen H.; Schmolze, Daniel B.; Johnson, Nicole B.; Brooker, Jeffrey S.; Cable, Alex E.; Connolly, James L.; Fujimoto, James G.

    2014-01-01

    Rapid intraoperative assessment of breast excision specimens is clinically important because up to 40% of patients undergoing breast-conserving cancer surgery require reexcision for positive or close margins. We demonstrate nonlinear microscopy (NLM) for the assessment of benign and malignant breast pathologies in fresh surgical specimens. A total of 179 specimens from 50 patients was imaged with NLM using rapid extrinsic nuclear staining with acridine orange and intrinsic second harmonic contrast generation from collagen. Imaging was performed on fresh, intact specimens without the need for fixation, embedding, and sectioning required for conventional histopathology. A visualization method to aid pathological interpretation is presented that maps NLM contrast from two-photon fluorescence and second harmonic signals to features closely resembling histopathology using hematoxylin and eosin staining. Mosaicking is used to overcome trade-offs between resolution and field of view, enabling imaging of subcellular features over square-centimeter specimens. After NLM examination, specimens were processed for standard paraffin-embedded histology using a protocol that coregistered histological sections to NLM images for paired assessment. Blinded NLM reading by three pathologists achieved 95.4% sensitivity and 93.3% specificity, compared with paraffin-embedded histology, for identifying invasive cancer and ductal carcinoma in situ versus benign breast tissue. Interobserver agreement was κ = 0.88 for NLM and κ = 0.89 for histology. These results show that NLM achieves high diagnostic accuracy, can be rapidly performed on unfixed specimens, and is a promising method for intraoperative margin assessment. PMID:25313045

  3. Robust tumor morphometry in multispectral fluorescence microscopy

    NASA Astrophysics Data System (ADS)

    Tabesh, Ali; Vengrenyuk, Yevgen; Teverovskiy, Mikhail; Khan, Faisal M.; Sapir, Marina; Powell, Douglas; Mesa-Tejada, Ricardo; Donovan, Michael J.; Fernandez, Gerardo

    2009-02-01

    Morphological and architectural characteristics of primary tissue compartments, such as epithelial nuclei (EN) and cytoplasm, provide important cues for cancer diagnosis, prognosis, and therapeutic response prediction. We propose two feature sets for the robust quantification of these characteristics in multiplex immunofluorescence (IF) microscopy images of prostate biopsy specimens. To enable feature extraction, EN and cytoplasm regions were first segmented from the IF images. Then, feature sets consisting of the characteristics of the minimum spanning tree (MST) connecting the EN and the fractal dimension (FD) of gland boundaries were obtained from the segmented compartments. We demonstrated the utility of the proposed features in prostate cancer recurrence prediction on a multi-institution cohort of 1027 patients. Univariate analysis revealed that both FD and one of the MST features were highly effective for predicting cancer recurrence (p <= 0.0001). In multivariate analysis, an MST feature was selected for a model incorporating clinical and image features. The model achieved a concordance index (CI) of 0.73 on the validation set, which was significantly higher than the CI of 0.69 for the standard multivariate model based solely on clinical features currently used in clinical practice (p < 0.0001). The contributions of this work are twofold. First, it is the first demonstration of the utility of the proposed features in morphometric analysis of IF images. Second, this is the largest scale study of the efficacy and robustness of the proposed features in prostate cancer prognosis.

  4. Fully hydrated yeast cells imaged with electron microscopy.

    PubMed

    Peckys, Diana B; Mazur, Peter; Gould, Kathleen L; de Jonge, Niels

    2011-05-18

    We demonstrate electron microscopy of fully hydrated eukaryotic cells with nanometer resolution. Living Schizosaccharomyces pombe cells were loaded in a microfluidic chamber and imaged in liquid with scanning transmission electron microscopy (STEM). The native intracellular (ultra)structures of wild-type cells and three different mutants were studied without prior labeling, fixation, or staining. The STEM images revealed various intracellular components that were identified on the basis of their shape, size, location, and mass density. The maximal achieved spatial resolution in this initial study was 32 ± 8 nm, an order of magnitude better than achievable with light microscopy on pristine cells. Light-microscopy images of the same samples were correlated with the corresponding electron-microscopy images. Achieving synergy between the capabilities of light and electron microscopy, we anticipate that liquid STEM will be broadly applied to explore the ultrastructure of live cells.

  5. Review on advances in nanoscale microscopy in cement research.

    PubMed

    Sharif, Ahmed

    2016-01-01

    With the rapid advancement of nanotechnology, manipulation and characterization of materials in nano scale have become an obvious part of construction related technology. This review will focus on some of the nanoscopy techniques that are most frequently used in current research of cement based nanostructured materials. In particular scanning electron microscopy, transmission electron microscopy, atomic force microscopy, scanning tunneling microscopy, tomography, scanning transmission X-ray microscopy and laser scanning confocal microscopy are addressed. A number of case studies related to microscopic characterization of nano materials utilizing the aforementioned techniques from the published literature are discussed. While these approaches are beginning to yield promising insight, continued progress will definitely provide a potential sustainable solution for the design, development and promotion towards nanoscale engineering of cementitious materials.

  6. Super-resolution spectroscopic microscopy via photon localization

    NASA Astrophysics Data System (ADS)

    Dong, Biqin; Almassalha, Luay; Urban, Ben E.; Nguyen, The-Quyen; Khuon, Satya; Chew, Teng-Leong; Backman, Vadim; Sun, Cheng; Zhang, Hao F.

    2016-07-01

    Traditional photon localization microscopy analyses only the spatial distributions of photons emitted by individual molecules to reconstruct super-resolution optical images. Unfortunately, however, the highly valuable spectroscopic information from these photons have been overlooked. Here we report a spectroscopic photon localization microscopy that is capable of capturing the inherent spectroscopic signatures of photons from individual stochastic radiation events. Spectroscopic photon localization microscopy achieved higher spatial resolution than traditional photon localization microscopy through spectral discrimination to identify the photons emitted from individual molecules. As a result, we resolved two fluorescent molecules, which were 15 nm apart, with the corresponding spatial resolution of 10 nm--a four-fold improvement over photon localization microscopy. Using spectroscopic photon localization microscopy, we further demonstrated simultaneous multi-colour super-resolution imaging of microtubules and mitochondria in COS-7 cells and showed that background autofluorescence can be identified through its distinct emission spectra.

  7. Super-resolution spectroscopic microscopy via photon localization.

    PubMed

    Dong, Biqin; Almassalha, Luay; Urban, Ben E; Nguyen, The-Quyen; Khuon, Satya; Chew, Teng-Leong; Backman, Vadim; Sun, Cheng; Zhang, Hao F

    2016-07-25

    Traditional photon localization microscopy analyses only the spatial distributions of photons emitted by individual molecules to reconstruct super-resolution optical images. Unfortunately, however, the highly valuable spectroscopic information from these photons have been overlooked. Here we report a spectroscopic photon localization microscopy that is capable of capturing the inherent spectroscopic signatures of photons from individual stochastic radiation events. Spectroscopic photon localization microscopy achieved higher spatial resolution than traditional photon localization microscopy through spectral discrimination to identify the photons emitted from individual molecules. As a result, we resolved two fluorescent molecules, which were 15 nm apart, with the corresponding spatial resolution of 10 nm-a four-fold improvement over photon localization microscopy. Using spectroscopic photon localization microscopy, we further demonstrated simultaneous multi-colour super-resolution imaging of microtubules and mitochondria in COS-7 cells and showed that background autofluorescence can be identified through its distinct emission spectra.

  8. Super-resolution spectroscopic microscopy via photon localization

    PubMed Central

    Dong, Biqin; Almassalha, Luay; Urban, Ben E.; Nguyen, The-Quyen; Khuon, Satya; Chew, Teng-Leong; Backman, Vadim; Sun, Cheng; Zhang, Hao F.

    2016-01-01

    Traditional photon localization microscopy analyses only the spatial distributions of photons emitted by individual molecules to reconstruct super-resolution optical images. Unfortunately, however, the highly valuable spectroscopic information from these photons have been overlooked. Here we report a spectroscopic photon localization microscopy that is capable of capturing the inherent spectroscopic signatures of photons from individual stochastic radiation events. Spectroscopic photon localization microscopy achieved higher spatial resolution than traditional photon localization microscopy through spectral discrimination to identify the photons emitted from individual molecules. As a result, we resolved two fluorescent molecules, which were 15 nm apart, with the corresponding spatial resolution of 10 nm—a four-fold improvement over photon localization microscopy. Using spectroscopic photon localization microscopy, we further demonstrated simultaneous multi-colour super-resolution imaging of microtubules and mitochondria in COS-7 cells and showed that background autofluorescence can be identified through its distinct emission spectra. PMID:27452975

  9. Conventional and nonlinear optical microscopy of liquid crystal colloids

    NASA Astrophysics Data System (ADS)

    Lee, Taewoo; Smalyukh, Ivan I.

    The fast-growing field of liquid crystal colloids requires increasingly sophisticated optical microscopy tools for experimental studies. Recent technological advances have resulted in a vast body of new imaging modalities, such as nonlinear optical microscopy techniques, that were developed to achieve high resolution while probing director structures and material composition at length scales ranging from hundreds of nanometers to oscopic. These techniques are ideally suited for experimental exploration of liquid crystal colloids. The goal of this chapter is to introduce a variety of optical microscopy techniques available to researchers in the field, starting from basic principles and finishing with a discussion of the most advanced microscopy systems. We describe traditional imaging tools, such as bright field and polarizing optical microscopy, along with state-of-the-art orientationsensitive three-dimensional imaging techniques, such as various nonlinear optical microscopies. Applications of these different imaging approaches are illustrated by providing specific examples of imaging of liquid crystal colloids and other soft matter systems.

  10. Scanned probe microscopy for thin film superconductor development

    SciTech Connect

    Moreland, J.

    1996-12-31

    Scanned probe microscopy is a general term encompassing the science of imaging based on piezoelectric driven probes for measuring local changes in nanoscale properties of materials and devices. Techniques like scanning tunneling microscopy, atomic force microscopy, and scanning potentiometry are becoming common tools in the production and development labs in the semiconductor industry. The author presents several examples of applications specific to the development of high temperature superconducting thin films and thin-film devices.

  11. Hard x-ray Zernike microscopy reaches 30 nm resolution.

    SciTech Connect

    Chen, Y.; Chen, T.; Yi, J.; Chu, Y.; Lee, W.-K.; Wang, C.; Kempson, I.; Hwu, Y.; Gajdosik, V.; Margaritondo, G.

    2011-03-30

    Since its invention in 1930, Zernike phase contrast has been a pillar in optical microscopy and more recently in x-ray microscopy, in particular for low-absorption-contrast biological specimens. We experimentally demonstrate that hard-x-ray Zernike microscopy now reaches a lateral resolution below 30?nm while strongly enhancing the contrast, thus opening many new research opportunities in biomedicine and materials science.

  12. Hard x-ray Zernike Microscopy Reaches 30 nm Resolution

    SciTech Connect

    Chen, Y.T.; Chu, Y.; Chen, T-Y.; Yi, J.; Lee, W-K.; Wang, C-L.; Kempson, I. M.; Hwu, Y.; Gajdosik, V.; Margaritondo, G.

    2011-03-30

    Since its invention in 1930, Zernike phase contrast has been a pillar in optical microscopy and more recently in x-ray microscopy, in particular for low-absorption-contrast biological specimens. We experimentally demonstrate that hard-x-ray Zernike microscopy now reaches a lateral resolution below 30 nm while strongly enhancing the contrast, thus opening many new research opportunities in biomedicine and materials science.

  13. Modeling Graphene Contrast on Copper Surfaces Using Optical Microscopy

    DTIC Science & Technology

    2014-10-01

    requirement for transfer. Atomic force microscopy was used to determine copper oxide thickness, and a Matlab model based on Fresnel theory was used to...consuming process involving specialized techniques such as atomic force microscopy (AFM), scanning electron microscopy (SEM), transmission electron...laser was used as the excitation source. A Cypher SPM in noncontact mode was used for topography and phase imaging characterization. 2.4 Broadband

  14. Fast and long term lipid droplet tracking with CARS microscopy.

    PubMed

    Jüngst, Christian; Winterhalder, Martin J; Zumbusch, Andreas

    2011-06-01

    Photobleaching of organic fluorophores commonly used in fluorescence microscopy puts a limit to the number of images which can be acquired. Label-free imaging techniques therefore offer advantages both for rapid image acquisition and for long-term observations. CARS microscopy is a label-free imaging technique offering molecule specific contrast. Here we demonstrate that CARS microscopy allows video-rate tracking of intracellular transport of lipid droplets, but also continuous long-term observation of cells over several hours.

  15. A quick guide to light microscopy in cell biology.

    PubMed

    Thorn, Kurt

    2016-01-15

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

  16. Single molecule microscopy and spectroscopy: concluding remarks.

    PubMed

    van Hulst, Niek F

    2015-01-01

    Chemistry is all about molecules: control, synthesis, interaction and reaction of molecules. All too easily on a blackboard, one draws molecules, their structures and dynamics, to create an insightful picture. The dream is to see these molecules in reality. This is exactly what "Single Molecule Detection" provides: a look at molecules in action at ambient conditions; a breakthrough technology in chemistry, physics and biology. Within the realms of the Royal Society of Chemistry, the Faraday Discussion on "Single Molecule Microscopy and Spectroscopy" was a very appropriate topic for presentation, deliberation and debate. Undoubtedly, the Faraday Discussions have a splendid reputation in stimulating scientific debates along the traditions set by Michael Faraday. Interestingly, back in the 1830's, Faraday himself pursued an experiment that led to the idea that atoms in a compound were joined by an electrical component. He placed two opposite electrodes in a solution of water containing a dissolved compound, and observed that one of the elements of the compound accumulated on one electrode, while the other was deposited on the opposite electrode. Although Faraday was deeply opposed to atomism, he had to recognize that electrical forces were responsible for the joining of atoms. Probably a direct view on the atoms or molecules in his experiment would have convinced him. As such, Michael Faraday might have liked the gathering at Burlington House in September 2015 (). Surely, with the questioning eyes of his bust on the 1st floor corridor, the non-believer Michael Faraday has incited each passer-by to enter into discussion and search for deeper answers at the level of single molecules. In these concluding remarks, highlights of the presented papers and discussions are summarized, complemented by a conclusion on future perspectives.

  17. Microelectrophoresis of Silica Rods Using Confocal Microscopy

    PubMed Central

    2017-01-01

    The electrophoretic mobility and the zeta potential (ζ) of fluorescently labeled colloidal silica rods, with an aspect ratio of 3.8 and 6.1, were determined with microelectrophoresis measurements using confocal microscopy. In the case where the colloidal particles all move at the same speed parallel to the direction of the electric field, we record a xyz-stack over the whole depth of the capillary. This method is faster and more robust compared to taking xyt-series at different depths inside the capillary to obtain the parabolic flow profile, as was done in previous work from our group. In some cases, rodlike particles do not move all at the same speed in the electric field, but exhibit a velocity that depends on the angle between the long axis of the rod and the electric field. We measured the orientation-dependent velocity of individual silica rods during electrophoresis as a function of κa, where κ–1 is the double layer thickness and a is the radius of the rod associated with the diameter. Thus, we determined the anisotropic electrophoretic mobility of the silica rods with different sized double layers. The size of the double layer was tuned by suspending silica rods in different solvents at different electrolyte concentrations. We compared these results with theoretical predictions. We show that even at already relatively high κa when the Smoluchowski limiting law is assumed to be valid (κa > 10), an orientation dependent velocity was measured. Furthermore, we observed that at decreasing values of κa the anisotropy in the electrophoretic mobility of the rods increases. However, in low polar solvents with κa < 1, this trend was reversed: the anisotropy in the electrophoretic mobility of the rods decreased. We argue that this decrease is due to end effects, which was already predicted theoretically. When end effects are not taken into account, this will lead to strong underestimation of the experimentally determined zeta potential. PMID:28045541

  18. Quantitative reflection contrast microscopy of living cells

    PubMed Central

    1979-01-01

    Mammalian cells in culture (BHK-21, PtK2, Friend, human flia, and glioma cells) have been observed by reflection contrast microscopy. Images of cells photographed at two different wavelengths (546 and 436 nm) or at two different angles of incidence allowed discrimination between reflected light and light that was both reflected and modulated by interference. Interference is involved when a change in reflected intensity (relative to glass/medium background reflected intensity) occurs on changing either the illumination wavelength or the reflection incidence angle. In cases where interference occurs, refractive indices can be determined at points where the optical path difference is known, by solving the given interference equation. Where cells are at least 50 nm distant from the glass substrate, intensities are also influenced by that distance as well as by the light's angle of incidence and wavelength. The reflected intensity at the glass/medium interface is used as a standard in calculating the refractive index of the cortical cytoplasm. Refractive indices were found to be higher (1.38--1.40) at points of focal contact, where stress fibers terminate, than in areas of close contact (1.354--1.368). In areas of the cortical cytoplasm, between focal contacts, not adherent to the glass substrate, refractive indices between 1.353 and 1.368 were found. This was thought to result from a microfilamentous network within the cortical cytoplasm. Intimate attachment of cells to their substrate is assumed to be characterized by a lack of an intermediate layer of culture medium. PMID:389938

  19. Doppler Encoded Excitation Patterning (DEEP) Microscopy

    NASA Astrophysics Data System (ADS)

    Feldkhun, Daniel

    Traditional optical imaging systems rely on lenses and spatially-resolved detection to probe distinct locations on the object. We develop a novel computational approach to 2D and 3D imaging that instead measures the object's spatial Fourier transform using a single-element detector and without requiring precision optics. This wide-field technique can be used to image biological and synthetic structures in fluoresced or scattered light using coherent or broadband illumination. It employs dynamic structured illumination, acousto-optics, RF electronics, and tomographic algorithms to circumvent several trade-offs in conventional imaging, such as the dependence of the optical transfer function on the imaging lenses and the coupling of resolution and depth of field. We use Fourier optics concepts to derive the dynamic optical transfer function, evaluate different Fourier sampling strategies, and investigate and compare tomographic algorithms for 2D and 3D image synthesis. We also develop conceptual and analytical models to describe imaging of fluorescent as well as amplitude and phase scattering objects, the effects of broadband and spatially-incoherent illumination, and nonlinear wide-field super-resolution imaging. We consider sources of noise, analyze and simulate SNR behavior for several types of noise and Fourier sampling strategies, and compare the sensitivity of the technique to conventional imaging. We describe several experimental proof-of-concept systems and present two-dimensional high-resolution tomographic image reconstructions in both scattered and fluoresced light demonstrating a thousandfold improvement in the depth of field compared to conventional lens-based microscopy. Finally, we explore approaches for high-speed Fourier sampling and propose several related sensing techniques, including wide-field fluorescence imaging in scattering media.

  20. Attosecond photoelectron microscopy of H2+

    NASA Astrophysics Data System (ADS)

    Hu, S. X.; Collins, L. A.; Schneider, B. I.

    2009-08-01

    We present a numerical study of the ultrafast ionization dynamics of H2+ exposed to attosecond extreme ultraviolet (xuv) pulses that goes beyond the Born-Openheimer approximation. The four-dimensional, time-dependent Schrödinger equation was solved using a generalization of the finite-element discrete-variable-representation/real-space-product technique used in our previous calculations to include the dynamical motion of the nuclei. This has enabled us to expose the target to any polarized light at arbitrary angles with respect to the molecular axis. Calculations have been performed at different angles and photon energies ( ℏω=50eV up to 630 eV) to investigate the energy and orientation dependence of the photoionization probability. A strong orientation dependence of the photoionization probability of H2+ was found at a photon energy of ℏω=50eV . At this energy, we found that the ionization probability is three times larger in the perpendicular polarization than in the parallel case. These observations are explained by the different geometric “cross sections” seen by the photoejected electron as it leaves the molecule. This ionization anisotropy vanishes at the higher-photon energy of ℏω≥170eV . When these higher-energy xuv pulses are polarized perpendicular to the internuclear axis, a “double-slit-like” interference pattern is observed. However, we find that the diffraction angle only approaches the classical formula ϕn=sin-1(nλe/R0) , where n is the diffraction order, λe is the released electron wavelength, and R0 is the internuclear distance, when nλe becomes less than 65% of R0 . These results illustrate the possibility of employing attosecond pulses to perform photoelectron microscopy of molecules.