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Sample records for spectroscopic phase microscopy

  1. Dynamic spectroscopic phase microscopy for quantifying hemoglobin concentration and dynamic membrane fluctuation in red blood cells.

    PubMed

    Jang, Yunhun; Jang, Jaeduck; Park, YongKeun

    2012-04-23

    We report a technique for simultaneous label-free quantification of cytoplasmic hemoglobin Hb concentration and dynamic membrane fluctuation in individual red blood cells (RBCs). Spectroscopic phase microscopy equipped with three different coherent laser sources and a color detector records three wavelength-dependent quantitative phase images in a single shot of a color-coded hologram. Using molecular specific dispersion, we demonstrate the extraction of Hb concentration and the dynamic membrane fluctuation from individual RBCs. © 2012 Optical Society of America

  2. Spectroscopic phase microscopy for quantifying hemoglobin concentrations in intact red blood cells.

    PubMed

    Park, YongKeun; Yamauchi, Toyohiko; Choi, Wonshik; Dasari, Ramachandra; Feld, Michael S

    2009-12-01

    We report a practical method for label-free quantification of specific molecules using spectroscopic imaging of sample-induced phase shifts. Diffraction phase microscopy equipped with various wavelengths of light source is used to record wavelength-dependent phase images. We first perform dispersion measurements on pure solutions of single molecular species present in the cells, such as albumin and hemoglobin (Hb). With this prior calibration of molecular specific dispersion, we demonstrate the extraction of Hb concentration from individual human red blood cells. The end point of this study is noninvasive monitoring of physiological states of intact living cells.

  3. 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.

  4. Structural studies on Si:H network before and after solid phase crystallization using spectroscopic ellipsometry: Correlation with Raman spectroscopy and transmission electron microscopy

    NASA Astrophysics Data System (ADS)

    Goswami, Romyani; Ray, Swati

    2013-10-01

    The structure of hydrogenated silicon films (Si:H) before and after solid phase crystallization (SPC) has been investigated by detailed study of spectroscopic ellipsometry (SE). The Si:H films have been deposited by radio frequency plasma enhanced chemical vapor deposition (RF PECVD) system varying deposition power density from 0.03 W/cm2 to 0.46 W/cm2, just below the onset of amorphous to nano-crystalline transition region. Solid phase crystallization of the Si:H network has been done by thermal annealing of the films in a vacuum furnace. Different bulk compositions of the as deposited Si:H network and annealed (polycrystalline) films have been calculated from the fitted parameters obtained from the simulation of the ellipsometry data by Bruggeman effective medium approximation (BEMA) method. More compact and void free structure in the bulk layer of the as deposited films has been observed at low power deposition region. Whereas void fraction in the bulk and surface roughness layer has increased with increase of deposition power density. For the annealed films higher crystallinity at the bulk layer with fewer voids has been observed at the low power region but in the surface roughness layer void fraction dominates in all the low and high power deposited films. The results obtained from the spectroscopic ellipsometry study have been correlated with Raman spectroscopy and transmission electron microscopy for both the as deposited and annealed films.

  5. 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.

  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 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

  8. Infrared spectroscopic imaging microscopy: Applications to biological systems

    SciTech Connect

    Kidder, Linda H.; Levin, Ira W.; Lewis, E. Neil

    1998-06-01

    The coupling of imaging modalities with spectroscopic techniques adds additional dimensions to sample analysis in both the spectroscopic and spatial domains. The particular ability of infrared (IR) imaging to explore the spatial distribution of chemically distinct species on length scales ranging from microns to kilometers demonstrates the versatility and diversity of spectroscopic imaging. In this paper, we focus on the further development of our Fourier-transform (FT) based mid-IR spectroscopic imaging technique which combines the analytical capabilities of mid-IR spectroscopy with the morphological information obtained from optical imaging. The seamless combination of spectroscopy for molecular analysis with the power of visualization represents the future of infrared microscopy. Our spectroscopic imaging instrument integrates several infrared focal-plane arrays with a Michelson step-scan interferometer, generating high-fidelity and high spectral resolution mid-infrared spectroscopic images. The instrumentation produces multidimensional, chemically specific images, while simultaneously obtaining high resolution spectra for each detector pixel. The spatial resolution of the images approaches the diffraction limit for mid-infrared wavelengths, while the spectral resolution is determined by the interferometer, and can be 4 cm{sup -1} or higher. Data derived from a variety of materials, particularly biological samples, illustrate the capabilities of the technique for readily visualizing chemical complexity and for providing statistical data on sample heterogeneity.

  9. Scanning Tunneling Microscopy methods for spectroscopic imaging of subsurface interfaces

    NASA Technical Reports Server (NTRS)

    Bell, L. D.; Kaiser, W. J.

    1988-01-01

    A new method for spatially-resolved, spectroscopic investigation of subsurface interface structure has been developed. The method, Ballistic Electron Emission Microscopy (BEEM), is based on Scanning Tunneling Microscopy (STM) techniques. BEEM combines STM vacuum tunneling with unique ballistic electron spectroscopy capabilities. BEEM enables, for the first time, direct imaging of subsurface interface electronic properties with nanometer spatial resolution. STM topographic images of surface structure and BEEM images of subsurface properties are obtained simultaneously. BEEM capabilities are demonstrated by investigation of important metal-semiconductor interfaces.

  10. Scanning Tunneling Microscopy methods for spectroscopic imaging of subsurface interfaces

    NASA Technical Reports Server (NTRS)

    Bell, L. D.; Kaiser, W. J.

    1988-01-01

    A new method for spatially-resolved, spectroscopic investigation of subsurface interface structure has been developed. The method, Ballistic Electron Emission Microscopy (BEEM), is based on Scanning Tunneling Microscopy (STM) techniques. BEEM combines STM vacuum tunneling with unique ballistic electron spectroscopy capabilities. BEEM enables, for the first time, direct imaging of subsurface interface electronic properties with nanometer spatial resolution. STM topographic images of surface structure and BEEM images of subsurface properties are obtained simultaneously. BEEM capabilities are demonstrated by investigation of important metal-semiconductor interfaces.

  11. 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.

  12. Confocal diffraction phase microscopy of live cells.

    PubMed

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

    2008-09-15

    We present a new quantitative phase microscopy technique, confocal diffraction phase microscopy, which provides quantitative phase measurements from localized sites on a sample with high sensitivity. The technique combines common-path interferometry with confocal microscopy in a transmission geometry. The capability of the technique for static imaging is demonstrated by imaging polystyrene microspheres and live HT29 cells, while dynamic imaging is demonstrated by quantifying the nanometer scale fluctuations of red blood cell membranes.

  13. Spectroscopic analysis of skin intrinsic signals for multiphoton microscopy

    NASA Astrophysics Data System (ADS)

    Pena, Ana-Maria; Strupler, Mathias; Boulesteix, Thierry; Senni, Karim; Godeau, Gaston; Beaurepaire, Emmanuel; Schanne-Klein, Marie-Claire

    2006-02-01

    We recorded multiphoton images of human skin biopsies using endogenous sources of nonlinear optical signals. We detected simultaneously two-photon excited fluorescence (2PEF) from intrinsic fluorophores and second harmonic generation (SHG) from collagen. We observed SHG from fibrillar collagens in the dermis, whereas no SHG was detectable from the non fibrillar type IV collagen in the basal laminae. We compared these distinct behaviours of collagens I and IV in SHG microscopy to polarization-resolved surface SHG experiments on thin films of collagens I and IV molecules. We observed similar signals for both types of molecular films, except for the chiroptical contributions which are present only for collagen I and enhance the signal typically by a factor of 2. We concluded that SHG microscopy is a sensitive probe of the micrometer-scale structural organization of collagen in biological tissues. In order to elucidate the origin of the endogenous fluorescence signals, we recorded 2PEF spectra at various positions in the skin biopsies, and compared these data to in vitro spectroscopic analysis. In particular, we studied the keratin fluorescence and determined its 2PEF action cross section. We observed a good agreement between 2PEF spectra recorded in the keratinized upper layers of the epidermis and in a solution of purified keratin. Finally, to illustrate the capabilities of this technique, we recorded 2PEF/SHG images of skin biopsies obtained from patients of various ages.

  14. Vibrational spectroscopic imaging and multiphoton microscopy of spinal cord injury.

    PubMed

    Galli, Roberta; Uckermann, Ortrud; Winterhalder, Martin J; Sitoci-Ficici, Kerim H; Geiger, Kathrin D; Koch, Edmund; Schackert, Gabriele; Zumbusch, Andreas; Steiner, Gerald; Kirsch, Matthias

    2012-10-16

    Spinal cord injury triggers a series of complex biochemical alterations of nervous tissue. Up to now, such cellular events could not be studied without conventional tissue staining. The development of optical, label-free imaging techniques could provide powerful monitoring tools with the potential to be applied in vivo. In this work, we assess the ability of vibrational spectroscopy to generate contrast at molecular level between normal and altered regions in a rat model of spinal cord injury. Using tissue sections, we demonstrate that Fourier transform infrared (FT-IR) spectroscopy and spontaneous Raman spectroscopy are able to identify the lesion, the surrounding scar, and unharmed normal tissue, delivering insight into the biochemical events induced by the injury and allowing mapping of tissue degeneration. The FT-IR and Raman spectroscopic imaging provides the basis for fast multimodal nonlinear optical microscopy (coherent anti-Stokes Raman scattering, endogenous two-photon fluorescence, and second harmonic generation). The latter proves to be a fast tool for imaging of the lesion on unstained tissue samples, based on the alteration in lipid content, extracellular matrix composition, and microglia/macrophages distribution pattern. The results establish these technologies in the field of regeneration in central nervous system, with the long-term goal to extend them to intravital use, where fast and nonharmful imaging is required.

  15. 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.

  16. 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. © The Author 2015. Published by Oxford University Press on behalf of The Japanese Society of Microscopy. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

  17. 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.

  18. Parallel phase-shifting digital holographic microscopy

    PubMed Central

    Tahara, Tatsuki; Ito, Kenichi; Kakue, Takashi; Fujii, Motofumi; Shimozato, Yuki; Awatsuji, Yasuhiro; Nishio, Kenzo; Ura, Shogo; Kubota, Toshihiro; Matoba, Osamu

    2010-01-01

    We propose parallel phase-shifting digital holographic microscopy (PPSDHM) which has the ability of three-dimensional (3-D) motion measurement using space-division multiplexing technique. By the PPSDHM, instantaneous information of both the 3-D structure and the phase distributions of specimens can be simultaneously acquired with a single-shot exposure. We constructed a parallel phase-shifting digital holographic microscope consisting of an optical interferometer and an image sensor on which micro polarizers are attached pixel by pixel. The validity of the PPSDHM was experimentally verified by demonstrating the single-shot 3-D imaging and phase-imaging ability of the constructed microscope. PMID:21258494

  19. Spectroscopic scanning tunneling microscopy insights into Fe-based superconductors

    NASA Astrophysics Data System (ADS)

    Hoffman, Jennifer E.

    2011-12-01

    In the first three years since the discovery of Fe-based high Tc superconductors, scanning tunneling microscopy (STM) and spectroscopy have shed light on three important questions. First, STM has demonstrated the complexity of the pairing symmetry in Fe-based materials. Phase-sensitive quasiparticle interference (QPI) imaging and low temperature spectroscopy have shown that the pairing order parameter varies from nodal to nodeless s± within a single family, FeTe1-xSex. Second, STM has imaged C4 → C2 symmetry breaking in the electronic states of both parent and superconducting materials. As a local probe, STM is in a strong position to understand the interactions between these broken symmetry states and superconductivity. Finally, STM has been used to image the vortex state, giving insights into the technical problem of vortex pinning, and the fundamental problem of the competing states introduced when superconductivity is locally quenched by a magnetic field. Here we give a pedagogical introduction to STM and QPI imaging, discuss the specific challenges associated with extracting bulk properties from the study of surfaces, and report on progress made in understanding Fe-based superconductors using STM techniques.

  20. 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.

  1. Spectroscopic imaging with prospective motion correction and retrospective phase correction.

    PubMed

    Lange, Thomas; Maclaren, Julian; Buechert, Martin; Zaitsev, Maxim

    2012-06-01

    Motion-induced artifacts are much harder to recognize in magnetic resonance spectroscopic imaging than in imaging experiments and can therefore lead to erroneous interpretation. A method for prospective motion correction based on an optical tracking system has recently been proposed and has already been successfully applied to single voxel spectroscopy. In this work, the utility of prospective motion correction in combination with retrospective phase correction is evaluated for spectroscopic imaging in the human brain. Retrospective phase correction, based on the interleaved reference scan method, is used to correct for motion-induced frequency shifts and ensure correct phasing of the spectra across the whole spectroscopic imaging slice. It is demonstrated that the presented correction methodology can reduce motion-induced degradation of spectroscopic imaging data. Copyright © 2011 Wiley-Liss, Inc.

  2. Spectroscopic analysis of keratin endogenous signal for skin multiphoton microscopy

    NASA Astrophysics Data System (ADS)

    Pena, A.-M.; Strupler, M.; Boulesteix, T.; Schanne-Klein, M.-C.

    2005-08-01

    We recorded one-photon excited fluorescence (1PEF) and two-photon excited fluorescence (2PEF) spectra of purified keratin from human epidermis, and determined the action cross section of this endogenous chromophore. We used this spectroscopic analysis to analyse multiphoton images of skin biopsies and assign the intrinsic fluorescence signals in the epidermis. We observed a good agreement between in situ and in vitro 2PEF spectra of keratin. This study provides a comprehensive characterization of the 2PEF signal of the keratins from the epidermis, and will be of practical interest for multiphoton imaging of the skin.

  3. Ex-vivo holographic microscopy and spectroscopic analysis of head and neck cancer

    NASA Astrophysics Data System (ADS)

    Holler, Stephen; Wurtz, Robert; Auyeung, Kelsey; Auyeung, Kris; Paspaley-Grbavac, Milan; Mulroe, Brigid; Sobrero, Maximiliano; Miles, Brett

    2015-03-01

    Optical probes to identify tumor margins in vivo would greatly reduce the time, effort and complexity in the surgical removal of malignant tissue in head and neck cancers. Current approaches involve visual microscopy of stained tissue samples to determine cancer margins, which results in the excision of excess of tissue to assure complete removal of the cancer. Such surgical procedures and follow-on chemotherapy can adversely affect the patient's recovery and subsequent quality of life. In order to reduce the complexity of the process and minimize adverse effects on the patient, we investigate ex vivo tissue samples (stained and unstained) using digital holographic microscopy in conjunction with spectroscopic analyses (reflectance and transmission spectroscopy) in order to determine label-free, optically identifiable characteristic features that may ultimately be used for in vivo processing of cancerous tissues. The tissue samples studied were squamous cell carcinomas and associated controls from patients of varying age, gender and race. Holographic microscopic imaging scans across both cancerous and non-cancerous tissue samples yielded amplitude and phase reconstructions that were correlated with spectral signatures. Though the holographic reconstructions and measured spectra indicate variations even among the same class of tissue, preliminary results indicate the existence of some discriminating features. Further analyses are presently underway to further this work and extract additional information from the imaging and spectral data that may prove useful for in vivo surgical identification.

  4. 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.

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

    PubMed

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

    2012-08-01

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

  6. 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

  7. 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.

  8. 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.

  9. Near-infrared spectroscopic photoacoustic microscopy using a multi-color fiber laser source

    PubMed Central

    Buma, Takashi; Wilkinson, Benjamin C.; Sheehan, Timothy C.

    2015-01-01

    We demonstrate a simple multi-wavelength optical source suitable for spectroscopic optical resolution photoacoustic microscopy (OR-PAM) of lipid-rich tissue. 1064 nm laser pulses are converted to multiple wavelengths beyond 1300 nm via nonlinear optical propagation in a birefringent optical fiber. OR-PAM experiments with lipid phantoms clearly show the expected absorption peak near 1210 nm. We believe this simple multi-color technique is a promising cost-effective approach to spectroscopic OR-PAM of lipid-rich tissue. PMID:26309746

  10. Spectroscopic studies of cold, gas-phase biomolecular ions

    NASA Astrophysics Data System (ADS)

    Rizzo, Thomas R.; Stearns, Jaime A.; Boyarkin, Oleg V.

    While the marriage of mass spectrometry and laser spectroscopy is not new, developments over the last few years in this relationship have opened up new horizons for the spectroscopic study of biological molecules. The combination of electrospray ionisation for producing large biological molecules in the gas phase together with cooled ion traps and multiple-resonance laser schemes are allowing spectroscopic investigation of individual conformations of peptides with more than a dozen amino acids. Highly resolved infrared spectra of single conformations of such species provide important benchmarks for testing the accuracy of theoretical calculations. This review presents a number of techniques employed in our laboratory and in others for measuring the spectroscopy of cold, gas-phase protonated peptides. We show examples that demonstrate the power of these techniques and evaluate their extension to still larger biological molecules.

  11. Electron microscopy: Phase transition singled out

    SciTech Connect

    Browning, Nigel D.

    2013-04-23

    One of the fundamental challenges within nanotechnology is to understand and control how nanoscale properties are initiated, evolve, and eventually terminate as a system moves from an individual nanostructure towards the meso- and macro-scale ensembles that are used in most applications. The ability to directly observe individual nanostructures and characterize their structure and composition has long been within the purview of transmission electron microscopy (TEM). The almost ubiquitous application of spherical aberration correction in TEM and in scanning-TEM (STEM) that has occurred over the last 10 years, now means it is possible to routinely characterize such nanostructures with both atomic resolution and sensitivity [1,2]. The development of temporal resolution in the TEM and the ability to study fast dynamics, on the other hand, has only recently come to the forefront of instrumentation development and is currently defined by two different approaches in the use of photoemission sources: the single shot s-ns dynamic TEM (DTEM) [3] and the stroboscopic ps-fs 4-D EM [4]. In the case of the DTEM, the goal is to observe the longer timescale irreversible structural changes that occur during nucleation and growth phenomena (here the single shot approach means there are enough electrons in a single pulsed beam to form a complete image). The 4-D EM focuses on a stroboscopic approach with the goal of studying very rapid reversible effects that occur during phase transitions (here an image is composed of thousands of pump-probe events each occurring with exactly the same time signature, with an individual pulse containing only a few electrons).

  12. Analyzing cell structure and dynamics with confocal light scattering and absorption spectroscopic microscopy

    NASA Astrophysics Data System (ADS)

    Qiu, Le; Vitkin, Edward; Fang, Hui; Zaman, Munir M.; Andersson, Charlotte; Salahuddin, Saira; Modell, Mark D.; Freedman, Steven D.; Hanlon, Eugene B.; Itzkan, Irving; Perelman, Lev T.

    2007-02-01

    We recently developed a new microscopic optical technique capable of noninvasive analysis of cell structure and cell dynamics on the submicron scale [1]. It combines confocal microscopy, a well-established high-resolution microscopic technique, with light scattering spectroscopy (LSS) and is called confocal light absorption and scattering spectroscopic (CLASS) microscopy. CLASS microscopy requires no exogenous labels and is capable of imaging and continuously monitoring individual viable cells, enabling the observation of cell and organelle functioning at scales on the order of 100 nm. To test the ability of CLASS microscopy to monitor cellular dynamics in vivo we performed experiments with human bronchial epithelial cells treated with DHA and undergoing apoptosis. The treated and untreated cells show not only clear differences in organelle spatial distribution but time sequencing experiments on a single cell show disappearance of certain types of organelles and change of the nuclear shape and density with the progression of apoptosis. In summary, CLASS microscopy provides an insight into metabolic processes within the cell and opens doors for the noninvasive real-time assessment of cellular dynamics. Noninvasive monitoring of cellular dynamics with CLASS microscopy can be used for a real-time dosimetry in a wide variety of medical and environmental applications that have no immediate observable outcome, such as photodynamic therapy, drug screening, and monitoring of toxins.

  13. 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.

  14. Vibrational Spectroscopic Microscopy: Raman, Near-Infrared and Mid-Infrared Imaging Techniques

    NASA Astrophysics Data System (ADS)

    Lewis, E. Neil; Levin, Ira W.

    1995-02-01

    New instrumental approaches for performing vibrational Raman, near-infrared and mid-infrared spectroscopic imaging microscopy are described. The instruments integrate imaging quality filters such as acousto-optic tunable filters (AOTFs), with visible charge-coupled device (CCD) and infrared focal-plane array detectors. These systems are used in conjunction with infinity-corrected, refractive microscopes for operation in the visible and near-infrared spectral regions and with Cassegrainian reflective optics for operation in the mid-infrared spectral interval. Chemically specific images at moderate spectral resolution (2 nm) and high spatial resolution (1 [mu]m) can be collected rapidly and noninvasively. Image data are presented containing 128 × 128 pixels, although significantly larger format images can be collected in approximately the same time. The instruments can be readily configured for both absorption and reflectance spectroscopies. We present Raman emission images of polystyrene microspheres and a lipid/amino acid mixture and near-infrared images of onion epidermis and a hydrated phospholipid dispersion. Images generated from mid-infrared spectral data are presented for a KBr disk containing nonhomogeneous domains of lipid and for 50-[mu]m slices of monkey cerebellum. These are the first results illustrating the use of infrared focal-plane array detectors as chemically specific spectroscopic imaging devices and demonstrating their application in biomolecular areas. Extensions and future applications of the various vibrational spectroscopic imaging techniques are discussed.

  15. Quantitative phase microscopy with asynchronous digital holography.

    PubMed

    Chalut, Kevin J; Brown, William J; Wax, Adam

    2007-03-19

    We demonstrate a new method of measuring quantitative phase in imaging of biological materials. This method, asynchronous digital holography, employs knowledge of a moving fringe created by acousto-optic modulators to execute phase-shifting interferometry using two near-simultaneous interferograms. The method can be used to obtain quantitative phase images of dynamic biological samples on millisecond time scales. We present results on a standard sample, and on live cell samples.

  16. Quantitative phase microscopy with asynchronous digital holography

    PubMed Central

    Brown, William J.; Wax, Adam

    2009-01-01

    We demonstrate a new method of measuring quantitative phase in imaging of biological materials. This method, asynchronous digital holography, employs knowledge of a moving fringe created by acousto-optic modulators to execute phase-shifting interferometry using two near-simultaneous interferograms. The method can be used to obtain quantitative phase images of dynamic biological samples on millisecond time scales. We present results on a standard sample, and on live cell samples. PMID:19532543

  17. Phase sensitive demodulation in multiphoton microscopy.

    PubMed

    Fisher, Walt G; Piston, David W; Wachter, Eric A

    2002-06-01

    Multiphoton laser scanning microscopy offers advantages in depth of penetration into intact samples over other optical sectioning techniques. To achieve these advantages it is necessary to detect the emitted light without spatial filtering. In this nondescanned (nonconfocal) approach, ambient room light can easily contaminate the signal, forcing experiments to be performed in absolute darkness. For multiphoton microscope systems employing mode-locked lasers, signal processing can be used to reduce such problems by taking advantage of the pulsed characteristics of such lasers. Specifically, by recovering fluorescence generated at the mode-locked frequency, interference from stray light and other ambient noise sources can be significantly reduced. This technology can be adapted to existing microscopes by inserting demodulation circuitry between the detector and data collection system. The improvement in signal-to-noise ratio afforded by this approach yields a more robust microscope system and opens the possibility of moving multiphoton microscopy from the research lab to more demanding settings, such as the clinic.

  18. 3D differential phase contrast microscopy

    NASA Astrophysics Data System (ADS)

    Chen, Michael; Tian, Lei; Waller, Laura

    2016-03-01

    We demonstrate three-dimensional (3D) optical phase and amplitude reconstruction based on coded source illumination using a programmable LED array. Multiple stacks of images along the optical axis are computed from recorded intensities captured by multiple images under off-axis illumination. Based on the first Born approximation, a linear differential phase contrast (DPC) model is built between 3D complex index of refraction and the intensity stacks. Therefore, 3D volume reconstruction can be achieved via a fast inversion method, without the intermediate 2D phase retrieval step. Our system employs spatially partially coherent illumination, so the transverse resolution achieves twice the NA of coherent systems, while axial resolution is also improved 2× as compared to holographic imaging.

  19. Phase contrast in high resolution electron microscopy

    DOEpatents

    Rose, H.H.

    1975-09-23

    This patent relates to a device for developing a phase contrast signal for a scanning transmission electron microscope. The lens system of the microscope is operated in a condition of defocus so that predictable alternate concentric regions of high and low electron density exist in the cone of illumination. Two phase detectors are placed beneath the object inside the cone of illumination, with the first detector having the form of a zone plate, each of its rings covering alternate regions of either higher or lower electron density. The second detector is so configured that it covers the regions of electron density not covered by the first detector. Each detector measures the number of electrons incident thereon and the signal developed by the first detector is subtracted from the signal developed by the record detector to provide a phase contrast signal. (auth)

  20. 3D differential phase contrast microscopy

    PubMed Central

    Chen, Michael; Tian, Lei; Waller, Laura

    2016-01-01

    We demonstrate 3D phase and absorption recovery from partially coherent intensity images captured with a programmable LED array source. Images are captured through-focus with four different illumination patterns. Using first Born and weak object approximations (WOA), a linear 3D differential phase contrast (DPC) model is derived. The partially coherent transfer functions relate the sample’s complex refractive index distribution to intensity measurements at varying defocus. Volumetric reconstruction is achieved by a global FFT-based method, without an intermediate 2D phase retrieval step. Because the illumination is spatially partially coherent, the transverse resolution of the reconstructed field achieves twice the NA of coherent systems and improved axial resolution. PMID:27867705

  1. Growth of vacuum evaporated ultraporous silicon studied with spectroscopic ellipsometry and scanning electron microscopy

    NASA Astrophysics Data System (ADS)

    Kaminska, Kate; Amassian, Aram; Martinu, Ludvik; Robbie, Kevin

    2005-01-01

    Using a combination of variable-angle spectroscopic ellipsometry and scanning electron microscopy, we investigated the scaling behavior of uniaxially anisotropic, ultraporous silicon manufactured with glancing angle deposition. We found that both the diameter of the nanocolumns and the spacing between them increase with film thickness according to a power-law relationship consistent with self-affine fractal growth. An ellipsometric model is proposed to fit the optical properties of the anisotropic silicon films employing an effective medium approximation mixture of Tauc-Lorentz oscillator and void. This study shows that the optical response of silicon films made at glancing incidence differs significantly from that of amorphous silicon prepared by other methods due to highly oriented nanocolumn formation and power-law scaling.

  2. Operando Spectroscopic Microscopy of LiCoO 2 Cathodes Outside Standard Operating Potentials

    DOE PAGES

    Nelson Weker, Johanna; Wise, Anna M.; Lim, Kipil; ...

    2017-07-14

    LiCoO2 can experience over-lithiation (over-discharge) in an electrochemical cell due to poor battery management, failure such as a short circuit, or when LiCoO2 is utilized as a negative electrode conversion material. Furthermore, in order to understand the chemical and morphological changes which occur during over-lithiation, LiCoO2 electrodes were studied during deep discharge to 0.8 V with operando X-ray absorption spectroscopy and spectroscopic X-ray microscopy. During over-lithiation, micron-sized LiCoO2 particles suffer significant cracking, pulverization and an incomplete conversion to Co metal. These irreversible morphological and structural changes then inflict permanent damage on the cathode even during a single over-lithiation event andmore » highlight the need for more sophisticated battery management systems.« less

  3. Tissue refractometry using Hilbert phase microscopy.

    PubMed

    Lue, Niyom; Bewersdorf, Joerg; Lessard, Mark D; Badizadegan, Kamran; Dasari, Ramachandra R; Feld, Michael S; Popescu, Gabriel

    2007-12-15

    We present, for the first time to our knowledge, quantitative phase images associated with unstained 5 mum thick tissue slices of mouse brain, spleen, and liver. The refractive properties of the tissue are retrieved in terms of the average refractive index and its spatial variation. We find that the average refractive index varies significantly with tissue type, such that the brain is characterized by the lowest value and the liver by the highest. The spatial power spectra of the phase images reveal power law behavior with different exponents for each tissue type. This approach opens a new possibility for stain-free characterization of tissues, where the diagnostic power is provided by the intrinsic refractive properties of the biological structure. We present results obtained for liver tissue affected by a lysosomal storage disease and show that our technique can quantify structural changes during this disease development.

  4. Tissue refractometry using Hilbert phase microscopy

    PubMed Central

    Lue, Niyom; Bewersdorf, Joerg; Lessard, Mark D.; Badizadegan, Kamran; Dasari, Ramachandra R.; Feld, Michael S.; Popescu, Gabriel

    2009-01-01

    We present, for the first time to our knowledge, quantitative phase images associated with unstained 5 μm thick tissue slices of mouse brain, spleen, and liver. The refractive properties of the tissue are retrieved in terms of the average refractive index and its spatial variation. We find that the average refractive index varies significantly with tissue type, such that the brain is characterized by the lowest value and the liver by the highest. The spatial power spectra of the phase images reveal power law behavior with different exponents for each tissue type. This approach opens a new possibility for stain-free characterization of tissues, where the diagnostic power is provided by the intrinsic refractive properties of the biological structure. We present results obtained for liver tissue affected by a lysosomal storage disease and show that our technique can quantify structural changes during this disease development. PMID:18087529

  5. Partial wave spectroscopic microscopy can predict prostate cancer progression and mitigate over-treatment (Conference Presentation)

    NASA Astrophysics Data System (ADS)

    Zhang, Di; Graff, Taylor; Crawford, Susan; Subramanian, Hariharan; Thompson, Sebastian; Derbas, Justin R.; Lyengar, Radha; Roy, Hemant K.; Brendler, Charles B.; Backman, Vadim

    2016-02-01

    Prostate Cancer (PC) is the second leading cause of cancer deaths in American men. While prostate specific antigen (PSA) test has been widely used for screening PC, >60% of the PSA detected cancers are indolent, leading to unnecessary clinical interventions. An alternative approach, active surveillance (AS), also suffer from high expense, discomfort and complications associated with repeat biopsies (every 1-3 years), limiting its acceptance. Hence, a technique that can differentiate indolent from aggressive PC would attenuate the harms from over-treatment. Combining microscopy with spectroscopy, our group has developed partial wave spectroscopic (PWS) microscopy, which can quantify intracellular nanoscale organizations (e.g. chromatin structures) that are not accessible by conventional microscopy. PWS microscopy has previously been shown to predict the risk of cancer in seven different organs (N ~ 800 patients). Herein we use PWS measurement of label-free histologically-normal prostatic epithelium to distinguish indolent from aggressive PC and predict PC risk. Our results from 38 men with low-grade PC indicated that there is a significant increase in progressors compared to non-progressors (p=0.002, effect size=110%, AUC=0.80, sensitivity=88% and specificity=72%), while the baseline clinical characteristics were not significantly different. We further improved the diagnostic power by performing nuclei-specific measurements using an automated system that separates in real-time the cell nuclei from the remaining prostate epithelium. In the long term, we envision that the PWS based prognostication can be coupled with AS without any change to the current procedure to mitigate the harms caused by over-treatment.

  6. Single-shot quantitative dispersion phase microscopy

    NASA Astrophysics Data System (ADS)

    Lue, Niyom; Kang, Jeon Woong; Hillman, Timothy R.; Dasari, Ramachandra R.; Yaqoob, Zahid

    2012-08-01

    We present an imaging modality capable of providing high-speed optical dispersion measurements of live cells. The technique permits wide-field measurement of the optical phase shifts introduced by a sample for multiple discrete wavelengths in a single image capture. Utilizing spatial modulation and the wavelength dependence of the interference-fringe spacing, average refractive index as a function of wavelength is obtained, yielding optical dispersion measurements of the sample under observation. Because of its simple and low-cost design, the technique can be readily integrated into a standard microscope to collect additional diagnostic information about biological cells.

  7. High-speed quantitative interferometric microscopy based phase imaging cytometer

    NASA Astrophysics Data System (ADS)

    Xue, Liang; Sun, Nan; Yan, Keding; Liu, Fei; Wang, Shouyu

    2014-11-01

    The paper proposed a simple large scale bio-sample phase detecting equipment called gravity driven phase detecting cytometer, which is based on quantitative interferometric microscopy to realize flowing red blood cells phase distribution detection. The method has advantages on high throughput phase detecting and statistical analysis with high detecting speed and in real-time. The statistical characteristics of red blood cells are useful for biological analysis and disease detection. We believe this method is shedding more light on quantitatively measurement of the phase distribution of bio-samples.

  8. Scanning Electron and Phase-Contrast Microscopy of Bacterial Spores

    PubMed Central

    Bulla, L. A.; Julian, G. St.; Rhodes, R. A.; Hesseltine, C. W.

    1969-01-01

    The three-dimensional immages of free and intrasporangial spores produced by scanning electron microscopy show surface structures not visible by phase-contrast microscopy. Although fine surface detail is not elucidated by scanning electron microscopy, this technique does afford a definitive picture of the general shape of spores. Spores of Bacillus popilliae, B. lentimorbus, B. thuringiensis, B. alvei, B. cereus, and Sarcina ureae have varying patterns of surface ridge formation, whereas spores of B. larvae, B. subtilis, and B. licheniformis have relatively smooth surfaces. Images PMID:4907010

  9. Nonlinear dynamic phase contrast microscopy for microfluidic and microbiological applications

    NASA Astrophysics Data System (ADS)

    Denz, C.; Holtmann, F.; Woerdemann, M.; Oevermann, M.

    2008-08-01

    In live sciences, the observation and analysis of moving living cells, molecular motors or motion of micro- and nano-objects is a current field of research. At the same time, microfluidic innovations are needed for biological and medical applications on a micro- and nano-scale. Conventional microscopy techniques are reaching considerable limits with respect to these issues. A promising approach for this challenge is nonlinear dynamic phase contrast microscopy. It is an alternative full field approach that allows to detect motion as well as phase changes of living unstained micro-objects in real-time, thereby being marker free, without contact and non destructive, i.e. fully biocompatible. The generality of this system allows it to be combined with several other microscope techniques such as conventional bright field or fluorescence microscopy. In this article we will present the dynamic phase contrast technique and its applications in analysis of micro organismic dynamics, micro flow velocimetry and micro-mixing analysis.

  10. Drive frequency dependent phase imaging in piezoresponse force microscopy

    NASA Astrophysics Data System (ADS)

    Bo, Huifeng; Kan, Yi; Lu, Xiaomei; Liu, Yunfei; Peng, Song; Wang, Xiaofei; Cai, Wei; Xue, Ruoshi; Zhu, Jinsong

    2010-08-01

    The drive frequency dependent piezoresponse (PR) phase signal in near-stoichiometric lithium niobate crystals is studied by piezoresponse force microscopy. It is clearly shown that the local and nonlocal electrostatic forces have a great contribution to the PR phase signal. The significant PR phase difference of the antiparallel domains are observed at the contact resonances, which is related to the electrostatic dominated electromechanical interactions of the cantilever and tip-sample system. Moreover, the modulation voltage induced frequency shift at higher eigenmodes could be attributed to the change of indention force depending on the modulation amplitude with a piezoelectric origin. The PR phase of the silicon wafer is also measured for comparison. It is certificated that the electrostatic interactions are universal in voltage modulated scanning probe microscopy and could be extended to other phase imaging techniques.

  11. Three-dimensional microscopy with phase-shifting digital holography.

    PubMed

    Zhang, T; Yamaguchi, I

    1998-08-01

    We applied phase-shifting digital holography to microscopy by deriving the complex amplitude of light scattered from microscopic three-dimensional objects through a microscope objective by video camera recording, phase-shifting analysis, and computer reconstruction. This method requires no mechanical movement and provides a flexible display and quantitative evaluation of the reconstructed images. A theory of image formation and experimental verification with specimens are described.

  12. Spectroscopic microscopy can quantify the statistics of subdiffractional refractive-index fluctuations in media with random rough surfaces.

    PubMed

    Zhang, Di; Cherkezyan, Lusik; Capoglu, Ilker; Subramanian, Hariharan; Chandler, John; Thompson, Sebastian; Taflove, Allen; Backman, Vadim

    2015-11-01

    We previously established that spectroscopic microscopy can quantify subdiffraction-scale refractive index (RI) fluctuations in a label-free dielectric medium with a smooth surface. However, to study more realistic samples, such as biological cells, the effect of rough surface should be considered. In this Letter, we first report an analytical theory to synthesize microscopic images of a rough surface, validate this theory by finite-difference time-domain (FDTD) solutions of Maxwell's equations, and characterize the spectral properties of light reflected from a rough surface. Then, we report a technique to quantify the RI fluctuations beneath a rough surface and demonstrate its efficacy on FDTD-synthesized spectroscopic microscopy images, as well as experimental data obtained from biological cells.

  13. Spectroscopic microscopy can quantify the statistics of subdiffractional refractive-index fluctuations in media with random rough surfaces

    PubMed Central

    Zhang, Di; Cherkezyan, Lusik; Capoglu, Ilker; Subramanian, Hariharan; Chandler, John; Thompson, Sebastian; Taflove, Allen; Backman, Vadim

    2016-01-01

    We previously established that spectroscopic microscopy can quantify subdiffraction-scale refractive index (RI) fluctuations in a label-free dielectric medium with a smooth surface. However, to study more realistic samples, such as biological cells, the effect of rough surface should be considered. In this Letter, we first report an analytical theory to synthesize microscopic images of a rough surface, validate this theory by finite-difference time-domain (FDTD) solutions of Maxwell’s equations, and characterize the spectral properties of light reflected from a rough surface. Then, we report a technique to quantify the RI fluctuations beneath a rough surface and demonstrate its efficacy on FDTD-synthesized spectroscopic microscopy images, as well as experimental data obtained from biological cells. PMID:26512486

  14. Interference microscopy in cell biophysics. 1. Principles and methodological aspects of coherent phase microscopy.

    PubMed

    Tychinsky, Vladimir P; Tikhonov, Alexander N

    2010-12-01

    The purpose of our tandem publications is to review the applications of the coherent phase microscopy to cell biophysics. In this article, we briefly consider the fundamentals and methodological aspects of the coherent phase microscopy (CPM). One of important advantages of this method is a high sensitivity of CPM images to changes in physical-chemical properties of biological samples. The optical path length difference (OPLD), measured with CPM instruments in different domains of an object, serves as the informative optical parameter used for visualization of individual cells or intracellular organelles. Metabolically dependent variations of CPM images reflect changes in the functional state of biological objects. In the next article, we review results of CPM studies of different cells and intracellular energy-transducing organelles.

  15. Label-free imaging of the native, living cellular nanoarchitecture using partial-wave spectroscopic microscopy

    PubMed Central

    Almassalha, Luay M.; Bauer, Greta M.; Chandler, John E.; Gladstein, Scott; Cherkezyan, Lusik; Stypula-Cyrus, Yolanda; Weinberg, Samuel; Zhang, Di; Thusgaard Ruhoff, Peder; Roy, Hemant K.; Subramanian, Hariharan; Chandel, Navdeep S.; Szleifer, Igal; Backman, Vadim

    2016-01-01

    The organization of chromatin is a regulator of molecular processes including transcription, replication, and DNA repair. The structures within chromatin that regulate these processes span from the nucleosomal (10-nm) to the chromosomal (>200-nm) levels, with little known about the dynamics of chromatin structure between these scales due to a lack of quantitative imaging technique in live cells. Previous work using partial-wave spectroscopic (PWS) microscopy, a quantitative imaging technique with sensitivity to macromolecular organization between 20 and 200 nm, has shown that transformation of chromatin at these length scales is a fundamental event during carcinogenesis. As the dynamics of chromatin likely play a critical regulatory role in cellular function, it is critical to develop live-cell imaging techniques that can probe the real-time temporal behavior of the chromatin nanoarchitecture. Therefore, we developed a live-cell PWS technique that allows high-throughput, label-free study of the causal relationship between nanoscale organization and molecular function in real time. In this work, we use live-cell PWS to study the change in chromatin structure due to DNA damage and expand on the link between metabolic function and the structure of higher-order chromatin. In particular, we studied the temporal changes to chromatin during UV light exposure, show that live-cell DNA-binding dyes induce damage to chromatin within seconds, and demonstrate a direct link between higher-order chromatin structure and mitochondrial membrane potential. Because biological function is tightly paired with structure, live-cell PWS is a powerful tool to study the nanoscale structure–function relationship in live cells. PMID:27702891

  16. Label-free imaging of the native, living cellular nanoarchitecture using partial-wave spectroscopic microscopy.

    PubMed

    Almassalha, Luay M; Bauer, Greta M; Chandler, John E; Gladstein, Scott; Cherkezyan, Lusik; Stypula-Cyrus, Yolanda; Weinberg, Samuel; Zhang, Di; Thusgaard Ruhoff, Peder; Roy, Hemant K; Subramanian, Hariharan; Chandel, Navdeep S; Szleifer, Igal; Backman, Vadim

    2016-10-18

    The organization of chromatin is a regulator of molecular processes including transcription, replication, and DNA repair. The structures within chromatin that regulate these processes span from the nucleosomal (10-nm) to the chromosomal (>200-nm) levels, with little known about the dynamics of chromatin structure between these scales due to a lack of quantitative imaging technique in live cells. Previous work using partial-wave spectroscopic (PWS) microscopy, a quantitative imaging technique with sensitivity to macromolecular organization between 20 and 200 nm, has shown that transformation of chromatin at these length scales is a fundamental event during carcinogenesis. As the dynamics of chromatin likely play a critical regulatory role in cellular function, it is critical to develop live-cell imaging techniques that can probe the real-time temporal behavior of the chromatin nanoarchitecture. Therefore, we developed a live-cell PWS technique that allows high-throughput, label-free study of the causal relationship between nanoscale organization and molecular function in real time. In this work, we use live-cell PWS to study the change in chromatin structure due to DNA damage and expand on the link between metabolic function and the structure of higher-order chromatin. In particular, we studied the temporal changes to chromatin during UV light exposure, show that live-cell DNA-binding dyes induce damage to chromatin within seconds, and demonstrate a direct link between higher-order chromatin structure and mitochondrial membrane potential. Because biological function is tightly paired with structure, live-cell PWS is a powerful tool to study the nanoscale structure-function relationship in live cells.

  17. Newly designed, simple relief phase contrast for microscopy of microorganisms.

    PubMed

    Zižka, Z

    2010-11-01

    A new method providing a relief phase contrast for investigation of microorganisms by optical microscopy used a neutral filter Zeiss NG 10/1 that could be controllably slid at a certain azimuthal angle below the aperture condenser diaphragm of the microscope phase contrast. Two ways of application are described depending on the type of the microscope: (1) in a special holder, and (2) fixed on a rubber ring. The device enabled us to obtain excellent results in the area of both optical microscopy and microphotography. With the microorganisms visualized, a better resolution, higher contrast and a significant 3D effect were obtained; outer morphology and organelles (chloroplasts, nuclei, granules, oil reserve vacuoles, etc.) could also be investigated.

  18. Common-path phase-shifting lensless holographic microscopy.

    PubMed

    Micó, Vicente; García, Javier

    2010-12-01

    We present an approach capable of high-NA imaging in a lensless digital in-line holographic microscopy layout even outside the Gabor's regime. The method is based on spatial multiplexing at the sample plane, allowing a common-path interferometric architecture, where two interferometric beams are generated by a spatial light modulator (SLM) prior to illuminating the sample. The SLM allows phase-shifting interferometry by phase modulation of the SLM diffracted beam. After proper digital processing, the complex amplitude distribution of the diffracted object wavefront is recovered and numerically propagated to image the sample. Experimental results are reported that validate the proposed method.

  19. Movable aperture lensless transmission microscopy: a novel phase retrieval algorithm.

    PubMed

    Faulkner, H M L; Rodenburg, J M

    2004-07-09

    We propose an iterative phase retrieval method that uses a series of diffraction patterns, measured only in intensity, to solve for both amplitude and phase of the image wave function over a wide field of view and at wavelength-limited resolution. The new technique requires an aperture that is scanned to two or more positions over the object wave function. A simple implementation of the method is modeled and demonstrated, showing how the algorithm uses overlapping data in real space to resolve ambiguities in the solution. The technique opens up the possibility of practical transmission lensless microscopy at subatomic resolution using electrons, x rays, or nuclear particles.

  20. Phase modulation mode of scanning ion conductance microscopy

    SciTech Connect

    Li, Peng; Zhang, Changlin; Liu, Lianqing E-mail: gli@engr.pitt.edu; Wang, Yuechao; Yang, Yang; Li, Guangyong E-mail: gli@engr.pitt.edu

    2014-08-04

    This Letter reports a phase modulation (PM) mode of scanning ion conductance microscopy. In this mode, an AC current is directly generated by an AC voltage between the electrodes. The portion of the AC current in phase with the AC voltage, which is the current through the resistance path, is modulated by the tip-sample distance. It can be used as the input of feedback control to drive the scanner in Z direction. The PM mode, taking the advantages of both DC mode and traditional AC mode, is less prone to electronic noise and DC drift but maintains high scanning speed. The effectiveness of the PM mode has been proven by experiments.

  1. Single beam Fourier transform digital holographic quantitative phase microscopy

    SciTech Connect

    Anand, A. Chhaniwal, V. K.; Mahajan, S.; Trivedi, V.; Faridian, A.; Pedrini, G.; Osten, W.; Dubey, S. K.; Javidi, B.

    2014-03-10

    Quantitative phase contrast microscopy reveals thickness or height information of a biological or technical micro-object under investigation. The information obtained from this process provides a means to study their dynamics. Digital holographic (DH) microscopy is one of the most used, state of the art single-shot quantitative techniques for three dimensional imaging of living cells. Conventional off axis DH microscopy directly provides phase contrast images of the objects. However, this process requires two separate beams and their ratio adjustment for high contrast interference fringes. Also the use of two separate beams may make the system more vulnerable to vibrations. Single beam techniques can overcome these hurdles while remaining compact as well. Here, we describe the development of a single beam DH microscope providing whole field imaging of micro-objects. A hologram of the magnified object projected on to a diffuser co-located with a pinhole is recorded with the use of a commercially available diode laser and an arrayed sensor. A Fourier transform of the recorded hologram directly yields the complex amplitude at the image plane. The method proposed was investigated using various phase objects. It was also used to image the dynamics of human red blood cells in which sub-micrometer level thickness variation were measurable.

  2. Laser-scanning photoacoustic microscopy with ultrasonic phased array transducer.

    PubMed

    Zheng, Fan; Zhang, Xiangyang; Chiu, Chi Tat; Zhou, Bill L; Shung, K Kirk; Zhang, Hao F; Jiao, Shuliang

    2012-11-01

    In this paper, we report our latest progress on proving the concept that ultrasonic phased array can improve the detection sensitivity and field of view (FOV) in laser-scanning photoacoustic microscopy (LS-PAM). A LS-PAM system with a one-dimensional (1D) ultrasonic phased array was built for the experiments. The 1D phased array transducer consists of 64 active elements with an overall active dimension of 3.2 mm × 2 mm. The system was tested on imaging phantom and mouse ear in vivo. Experiments showed a 15 dB increase of the signal-to-noise ratio (SNR) when beamforming was employed compared to the images acquired with each single element. The experimental results demonstrated that ultrasonic phased array can be a better candidate for LS-PAM in high sensitivity applications like ophthalmic imaging.

  3. Laser-scanning photoacoustic microscopy with ultrasonic phased array transducer

    PubMed Central

    Zheng, Fan; Zhang, Xiangyang; Chiu, Chi Tat; Zhou, Bill L.; Shung, K. Kirk; Zhang, Hao F.; Jiao, Shuliang

    2012-01-01

    In this paper, we report our latest progress on proving the concept that ultrasonic phased array can improve the detection sensitivity and field of view (FOV) in laser-scanning photoacoustic microscopy (LS-PAM). A LS-PAM system with a one-dimensional (1D) ultrasonic phased array was built for the experiments. The 1D phased array transducer consists of 64 active elements with an overall active dimension of 3.2 mm × 2 mm. The system was tested on imaging phantom and mouse ear in vivo. Experiments showed a 15 dB increase of the signal-to-noise ratio (SNR) when beamforming was employed compared to the images acquired with each single element. The experimental results demonstrated that ultrasonic phased array can be a better candidate for LS-PAM in high sensitivity applications like ophthalmic imaging. PMID:23162708

  4. Improved Zernike-type phase contrast for transmission electron microscopy.

    PubMed

    Koeck, P J B

    2015-07-01

    Zernike phase contrast has been recognized as a means of recording high-resolution images with high contrast using a transmission electron microscope. This imaging mode can be used to image typical phase objects such as unstained biological molecules or cryosections of biological tissue. According to the original proposal discussed in Danev and Nagayama (2001) and references therein, the Zernike phase plate applies a phase shift of π/2 to all scattered electron beams outside a given scattering angle and an image is recorded at Gaussian focus or slight underfocus (below Scherzer defocus). Alternatively, a phase shift of -π/2 is applied to the central beam using the Boersch phase plate. The resulting image will have an almost perfect contrast transfer function (close to 1) from a given lowest spatial frequency up to a maximum resolution determined by the wave length, the amount of defocus and the spherical aberration of the microscope. In this paper, I present theory and simulations showing that this maximum spatial frequency can be increased considerably without loss of contrast by using a Zernike or Boersch phase plate that leads to a phase shift between scattered and unscattered electrons of only π /4, and recording images at Scherzer defocus. The maximum resolution can be improved even more by imaging at extended Scherzer defocus, though at the cost of contrast loss at lower spatial frequencies. © 2015 The Authors Journal of Microscopy © 2015 Royal Microscopical Society.

  5. Fourier transform infrared vibrational spectroscopic imaging: integrating microscopy and molecular recognition.

    PubMed

    Levin, Ira W; Bhargava, Rohit

    2005-01-01

    The recent development of Fourier transform infrared (FTIR) spectroscopic imaging has enhanced our capability to examine, on a microscopic scale, the spatial distribution of vibrational spectroscopic signatures of materials spanning the physical and biomedical disciplines. Recent activity in this emerging area has concentrated on instrumentation development, theoretical analyses to provide guidelines for imaging practice, novel data processing algorithms, and the introduction of the technique to new fields. To illustrate the impact and promise of this spectroscopic imaging methodology, we present fundamental principles of the technique in the context of FTIR spectroscopy and review new applications in various venues ranging from the physical chemistry of macromolecular systems to the detection of human disease.

  6. Atomic force microscopy images of lyotropic lamellar phases.

    PubMed

    Garza, C; Thieghi, L T; Castillo, R

    2007-02-07

    For the very first time, atomic force microscope images of lamellar phases were observed combined with a freeze fracture technique that does not involve the use of replicas. Samples are rapidly frozen, fractured, and scanned directly with atomic force microscopy, at liquid nitrogen temperature and in high vacuum. This procedure can be used to investigate micro-structured liquids. The lamellar phases in Sodium bis(2-ethylhexyl) sulfosuccinate (AOT)/water and in C12E5/water systems were used to asses this new technique. Our observations were compared with x-ray diffraction measurements and with other freeze fracture methods reported in the literature. Our results show that this technique is useful to image lyotropic lamellar phases and the estimated repeat distances for lamellar periodicity are consistent with those obtained by x-ray diffraction.

  7. Physical spherical phase compensation in reflection digital holographic microscopy

    NASA Astrophysics Data System (ADS)

    Weijuan, Qu; Oi Choo, Chee; Tan Rongwei, Lewis; Qiangsheng, Xu; Zhaomin, Wang; Zhenzhong, Xiao; Asundi, Anand

    2012-04-01

    Reflection configured digital holographic microscopy (DHM) can perform accurate optical topography measurements of reflecting objects, such as MEMs, MOEMs, and semiconductor wafer. It can provide non-destructive quantitative measurements of surface roughness and geometric pattern characterization with nanometric axial resolution in real-time. However, the measurement results may be affected by an additional phase curvature introduced by the microscope objective (MO) used in DHM. It needs to be removed either by numerical compensation or by physical compensation. We present a method of physical spherical phase compensation for reflection DHM in the Michelson configuration. In the object arm, collimated light is used for illumination. Due to the use of the MO, the object wavefront may have a spherical phase curvature. In the reference arm, a lens and mirror combination is used to generate a spherical recording reference wave in order to physically compensate the spherical phase curvature of the object wavefront. By controlling the position of the mirror and the sample stage, the compensation process has been demonstrated. The relative positions of the test specimen and the reference mirror must be fixed for the physical spherical phase to be totally compensated. A numerical plane reference wave is preferred for the numerical reconstruction of the phase introduced by the test specimen. Experimental results on wafer pattern recognition are also given.

  8. Halo-free phase contrast microscopy (Conference Presentation)

    NASA Astrophysics Data System (ADS)

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

    2017-02-01

    The phase contrast (PC) method is one of the most impactful developments in the four-century long history of microscopy. It allows for intrinsic, nondestructive contrast of transparent specimens, such as live cells. However, PC is plagued by the halo artifact, a result of insufficient spatial coherence in the illumination field, which limits its applicability. 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. Measuring 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.

  9. Zernike phase contrast in scanning microscopy with X-rays

    PubMed Central

    Holzner, Christian; Feser, Michael; Vogt, Stefan; Hornberger, Benjamin; Baines, Stephen B.; Jacobsen, Chris

    2011-01-01

    Scanning X-ray microscopy focuses radiation to a small spot and probes the sample by raster scanning. It allows information to be obtained from secondary signals such as X-ray fluorescence, which yields an elemental mapping of the sample not available in full-field imaging. The analysis and interpretation from these secondary signals can be considerably enhanced if these data are coupled with structural information from transmission imaging. However, absorption often is negligible and phase contrast has not been easily available. Originally introduced with visible light, Zernike phase contrast1 is a well-established technique in full-field X-ray microscopes for visualization of weakly absorbing samples2–7. On the basis of reciprocity, we demonstrate the implementation of Zernike phase contrast in scanning X-ray microscopy, revealing structural detail simultaneously with hard-X-ray trace-element measurements. The method is straightforward to implement without significant influence on the resolution of the fluorescence images and delivers complementary information. We show images of biological specimens that clearly demonstrate the advantage of correlating morphology with elemental information. PMID:21544232

  10. Phase stretch transform for super-resolution localization microscopy

    PubMed Central

    Ilovitsh, Tali; Jalali, Bahram; Asghari, Mohammad H.; Zalevsky, Zeev

    2016-01-01

    Super-resolution localization microscopy has revolutionized the observation of living structures at the cellular scale, by achieving a spatial resolution that is improved by more than an order of magnitude compared to the diffraction limit. These methods localize single events from isolated sources in repeated cycles in order to achieve super-resolution. The requirement for sparse distribution of simultaneously activated sources in the field of view dictates the acquisition of thousands of frames in order to construct the full super-resolution image. As a result, these methods have slow temporal resolution which is a major limitation when investigating live-cell dynamics. In this paper we present the use of a phase stretch transform for high-density super-resolution localization microscopy. This is a nonlinear frequency dependent transform that emulates the propagation of light through a physical medium with a specific warped diffractive property and applies a 2D phase function to the image in the frequency domain. By choosing properly the transform parameters and the phase kernel profile, the point spread function of each emitter can be sharpened and narrowed. This enables the localization of overlapping emitters, thus allowing a higher density of activated emitters as well as shorter data collection acquisition rates. The method is validated by numerical simulations and by experimental data obtained using a microtubule sample. PMID:27867725

  11. Zernike phase contrast in scanning microscopy with X-rays.

    PubMed

    Holzner, Christian; Feser, Michael; Vogt, Stefan; Hornberger, Benjamin; Baines, Stephen B; Jacobsen, Chris

    2010-11-01

    Scanning X-ray microscopy focuses radiation to a small spot and probes the sample by raster scanning. It allows information to be obtained from secondary signals such as X-ray fluorescence, which yields an elemental mapping of the sample not available in full-field imaging. The analysis and interpretation from these secondary signals can be considerably enhanced if these data are coupled with structural information from transmission imaging. However, absorption often is negligible and phase contrast has not been easily available. Originally introduced with visible light, Zernike phase contrast(1) is a well-established technique in full-field X-ray microscopes for visualization of weakly absorbing samples(2-7). On the basis of reciprocity, we demonstrate the implementation of Zernike phase contrast in scanning X-ray microscopy, revealing structural detail simultaneously with hard-X-ray trace-element measurements. The method is straightforward to implement without significant influence on the resolution of the fluorescence images and delivers complementary information. We show images of biological specimens that clearly demonstrate the advantage of correlating morphology with elemental information.

  12. Phase mask optimization for 3D parallax EDF microscopy

    NASA Astrophysics Data System (ADS)

    Beckers, Ingeborg E.; Gierlack, Michael; Höppel, Robert; Landskron, Jürgen

    2014-03-01

    Extended depth-of-field (EDF) microscopy is a well-investigated and very simple method to obtain projection images with an extended depth of focus. Despite its advantages of being a real-time method applicable to any microscopic mode with high lateral resolution that can be simply realized by extending a commercial microscope, the lack of z-correlation is still a problem. In this work we present a combined technique of EDF and stereomicroscopy. By cross-correlation depth information is obtained. Finally, 3D images are reconstructed for best phase masks and simulation results are evaluated experimentally.

  13. Fresnel particle tracing in three dimensions using diffraction phase microscopy.

    PubMed

    Park, Yongkeun; Popescu, Gabriel; Badizadegan, Kamran; Dasari, Ramachandra R; Feld, Michael S

    2007-04-01

    We have developed a novel experimental technique for tracking small particles in three dimensions with nanometer accuracy. The longitudinal positioning of a micrometer-sized particle is determined by using the Fresnel approximation to describe the transverse distribution of the wavefront that originated in the particle. The method utilizes the high-sensitivity quantitative phase imaging capability of diffraction phase microscopy recently developed in our laboratory. We demonstrate the principle of the technique with experiments on Brownian particles jittering in water both in bulk and in the vicinity of a boundary. The particles are localized in space within an error cube of 20 nm x 20 nm x 20 nm for a 33 Hz acquisition rate and 20s recording time.

  14. Efficient Phase Unwrapping Architecture for Digital Holographic Microscopy

    PubMed Central

    Hwang, Wen-Jyi; Cheng, Shih-Chang; Cheng, Chau-Jern

    2011-01-01

    This paper presents a novel phase unwrapping architecture for accelerating the computational speed of digital holographic microscopy (DHM). A fast Fourier transform (FFT) based phase unwrapping algorithm providing a minimum squared error solution is adopted for hardware implementation because of its simplicity and robustness to noise. The proposed architecture is realized in a pipeline fashion to maximize throughput of the computation. Moreover, the number of hardware multipliers and dividers are minimized to reduce the hardware costs. The proposed architecture is used as a custom user logic in a system on programmable chip (SOPC) for physical performance measurement. Experimental results reveal that the proposed architecture is effective for expediting the computational speed while consuming low hardware resources for designing an embedded DHM system. PMID:22163688

  15. Differential phase contrast in scanning x-ray microscopy with half-wavelength phase shifter

    SciTech Connect

    Suzuki, Yoshio Takeuchi, Akihisa

    2016-01-28

    A method for differential-phase-contrast imaging in scanning x-ray microscopy is proposed. The microfocus beam is produced with an x-ray focusing optics, and a half of the optical aperture is masked with a λ/2 phase shifter. This generates a pair of focused beam at the focal plane, with π phase difference. Combining with a diaphragm in front of the transmission beam detector, differential phase contrast (contrast proportional to the phase-difference between two foci) can be obtained. Preliminary results with a Fresnel zone plate focusing optics at 12.4 keV x-ray energy are shown.

  16. Multi-pore carbon phase plate for phase-contrast transmission electron microscopy.

    PubMed

    Sannomiya, Takumi; Junesch, Juliane; Hosokawa, Fumio; Nagayama, Kuniaki; Arai, Yoshihiro; Kayama, Yoko

    2014-11-01

    A new fabrication method of carbon based phase plates for phase-contrast transmission electron microscopy is presented. This method utilizes colloidal masks to produce pores as well as disks on thin carbon membranes for phase modulation. Since no serial process is involved, carbon phase plate membranes containing hundreds of pores can be mass-produced on a large scale, which allows "disposal" of contaminated or degraded phase modulating objects after use. Due to the spherical shape of the mask colloid particles, the produced pores are perfectly circular. The pore size and distribution can be easily tuned by the mask colloid size and deposition condition. By using the stencil method, disk type phase plates can also be fabricated on a pore type phase plate. Both pore and disk type phase plates were tested by measuring amorphous samples and confirmed to convert the sinus phase contrast transfer function to the cosine shape.

  17. Digital holographic microscopy of phase separation in multicomponent lipid membranes

    NASA Astrophysics Data System (ADS)

    Farzam Rad, Vahideh; Moradi, Ali-Reza; Darudi, Ahmad; Tayebi, Lobat

    2016-12-01

    Lateral in-homogeneities in lipid compositions cause microdomains formation and change in the physical properties of biological membranes. With the presence of cholesterol and mixed species of lipids, phospholipid membranes segregate into lateral domains of liquid-ordered and liquid-disordered phases. Coupling of two-dimensional intralayer phase separations and interlayer liquid-crystalline ordering in multicomponent membranes has been previously demonstrated. By the use of digital holographic microscopy (DHMicroscopy), we quantitatively analyzed the volumetric dynamical behavior of such membranes. The specimens are lipid mixtures composed of sphingomyelin, cholesterol, and unsaturated phospholipid, 1,2-dioleoyl-sn-glycero-3-phosphocholine. DHMicroscopy in a transmission mode is an effective tool for quantitative visualization of phase objects. By deriving the associated phase changes, three-dimensional information on the morphology variation of lipid stacks at arbitrary time scales is obtained. Moreover, the thickness distribution of the object at demanded axial planes can be obtained by numerical focusing. Our results show that the volume evolution of lipid domains follows approximately the same universal growth law of previously reported area evolution. However, the thickness of the domains does not alter significantly by time; therefore, the volume evolution is mostly attributed to the changes in area dynamics. These results might be useful in the field of membrane-based functional materials.

  18. Combined atomic force microscopy and spectroscopic ellipsometry applied to the analysis of lipid-protein thin films.

    PubMed

    Finot, Eric; Markey, Laurent; Hane, Francis; Amrein, Mathias; Leonenko, Zoya

    2013-04-01

    Pulmonary surfactant is a complex mixture of phospholipids and proteins and forms a thin film at the lung alveolar interface separating air from liquid environment. The film reduces the work of breathing during repeatable compressions of the alveoli which form a characteristic multilayer upon compression. In this work, we investigated the structure of bovine lipid extract surfactant (BLES). We analysed the BLES films by atomic force microscopy (AFM) and spectroscopic ellipsometry (SE) in order to provide combined characterization of both morphology and thickness of surfactant films. We show how the spectroscopic ellipsometry can be used to supplement the data obtained by AFM. We demonstrate that indium tin oxide (ITO) substrate used for spectroscopic ellipsometry is preferable over glass substrate to enhance the optical contrast. An optical model was proposed to account for non-uniform film morphology. We obtained good correlations between the multilayer surface coverage, determined by both AFM and SE. SE measures the thickness of the first uniform monolayer as 2.6 nm that cannot be achieved by AFM imaging alone.

  19. Femtosecond optical spectroscopy and microscopy with phase coherent pulses

    NASA Astrophysics Data System (ADS)

    Li, Chunqiang

    This dissertation focuses on femtosecond optical spectroscopy and microscopy by using ultrashort laser pulse shaping technology. Collinear phase coherent two-dimensional spectroscopy on the organic dye Coumarin 102 was performed. The collinear phase coherent three-pulse sequences in the near infrared wavelength (800 nm) were generated by an acousto-optic pulse shaper. Coumarin 102 has a broad absorption band of around 400 nm. Two-photon absorption of the 800 nm light in Coumarin 102 is essential to perform the experiment. For comparison another set of experiments with the light frequency doubled to 400 nm was also performed. The results obtained show similar features with different details. The energy level information, e.g. linewidth, can be retrieved from the two-dimensional spectral peaks. With pulse shaping technology the possibility to perform ultrafast laser microscopy aiming at potential biomedical application was also explored. The information utilized to differentiate chemicals is two-photon absorption and the structural information can also be retrieved from self-phase modulation. The nonlinear signal from these experiments is very small compared with input laser power. A heterodyne detection method was performed by creating a spectral hole in the laser spectrum with the pulse shaper. The signal generated is mixed with the local oscillator. The results from Rhodamine 6G, oxyHemoglobin and pheomelanin show the difference and imply promising further development with this method. The last part of this dissertation is a previous work on a novel structure wavelength tunable semiconductor laser based on asymmetric twin waveguide technology. To integrate several optoelectronic devices together, several schemes are available. Asymmetric twin waveguide technology is the only integration platform which does not require material regrowth and quantum well intermixing. The tunable laser based on this technology can be used as a local oscillator in a coherent optical

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

    PubMed

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

    2009-11-26

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

  1. Imaging red blood cell dynamics by quantitative phase microscopy.

    PubMed

    Popescu, Gabriel; Park, YoungKeun; Choi, Wonshik; Dasari, Ramachandra R; Feld, Michael S; Badizadegan, Kamran

    2008-01-01

    Red blood cells (RBCs) play a crucial role in health and disease, and structural and mechanical abnormalities of these cells have been associated with important disorders such as Sickle cell disease and hereditary cytoskeletal abnormalities. Although several experimental methods exist for analysis of RBC mechanical properties, optical methods stand out as they enable collecting mechanical and dynamic data from live cells without physical contact and without the need for exogenous contrast agents. In this report, we present quantitative phase microscopy techniques that enable imaging RBC membrane fluctuations with nanometer sensitivity at arbitrary time scales from milliseconds to hours. We further provide a theoretical framework for extraction of membrane mechanical and dynamical properties using time series of quantitative phase images. Finally, we present an experimental approach to extend quantitative phase imaging to 3-dimensional space using tomographic methods. By providing non-invasive methods for imaging mechanics of live cells, these novel techniques provide an opportunity for high-throughput analysis and study of RBC mechanical properties in health and disease.

  2. Single-shot quantitative phase microscopy with color-multiplexed differential phase contrast (cDPC)

    PubMed Central

    2017-01-01

    We present a new technique for quantitative phase and amplitude microscopy from a single color image with coded illumination. Our system consists of a commercial brightfield microscope with one hardware modification—an inexpensive 3D printed condenser insert. The method, color-multiplexed Differential Phase Contrast (cDPC), is a single-shot variant of Differential Phase Contrast (DPC), which recovers the phase of a sample from images with asymmetric illumination. We employ partially coherent illumination to achieve resolution corresponding to 2× the objective NA. Quantitative phase can then be used to synthesize DIC and phase contrast images or extract shape and density. We demonstrate amplitude and phase recovery at camera-limited frame rates (50 fps) for various in vitro cell samples and c. elegans in a micro-fluidic channel. PMID:28152023

  3. Single-shot quantitative phase microscopy with color-multiplexed differential phase contrast (cDPC).

    PubMed

    Phillips, Zachary F; Chen, Michael; Waller, Laura

    2017-01-01

    We present a new technique for quantitative phase and amplitude microscopy from a single color image with coded illumination. Our system consists of a commercial brightfield microscope with one hardware modification-an inexpensive 3D printed condenser insert. The method, color-multiplexed Differential Phase Contrast (cDPC), is a single-shot variant of Differential Phase Contrast (DPC), which recovers the phase of a sample from images with asymmetric illumination. We employ partially coherent illumination to achieve resolution corresponding to 2× the objective NA. Quantitative phase can then be used to synthesize DIC and phase contrast images or extract shape and density. We demonstrate amplitude and phase recovery at camera-limited frame rates (50 fps) for various in vitro cell samples and c. elegans in a micro-fluidic channel.

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

    PubMed

    Dubois, Arnaud

    2004-03-01

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

  5. Microsecond Scale Vibrational Spectroscopic Imaging by Multiplex Stimulated Raman Scattering Microscopy

    PubMed Central

    Liao, Chien-Sheng; Slipchenko, Mikhail N.; Wang, Ping; Li, Junjie; Lee, Seung-Young; Oglesbee, Robert A.; Cheng, Ji-Xin

    2015-01-01

    Real-time vibrational spectroscopic imaging is desired for monitoring cellular states and cellular processes in a label-free manner. Raman spectroscopic imaging of highly dynamic systems is inhibited by relatively slow spectral acquisition on millisecond to second scale. Here, we report microsecond scale vibrational spectroscopic imaging by lock-in free parallel detection of spectrally dispersed stimulated Raman scattering signal. Using a homebuilt tuned amplifier array, our method enables Raman spectral acquisition, within the window defined by the broadband pulse, at the speed of 32 microseconds and with close to shot-noise limited detection sensitivity. Incorporated with multivariate curve resolution analysis, our platform allows compositional mapping of lipid droplets in single live cells, observation of intracellular retinoid metabolism, discrimination of fat droplets from protein-rich organelles in Caenorhabditis elegans, spectral detection of fast flowing tumor cells, and monitoring drug diffusion through skin tissue in vivo. The reported technique opens new opportunities for compositional analysis of cellular compartment in a microscope setting and high-throughput spectral profiling of single cells in a flow cytometer setting. PMID:26167336

  6. Microsecond Scale Vibrational Spectroscopic Imaging by Multiplex Stimulated Raman Scattering Microscopy.

    PubMed

    Liao, Chien-Sheng; Slipchenko, Mikhail N; Wang, Ping; Li, Junjie; Lee, Seung-Young; Oglesbee, Robert A; Cheng, Ji-Xin

    Real-time vibrational spectroscopic imaging is desired for monitoring cellular states and cellular processes in a label-free manner. Raman spectroscopic imaging of highly dynamic systems is inhibited by relatively slow spectral acquisition on millisecond to second scale. Here, we report microsecond scale vibrational spectroscopic imaging by lock-in free parallel detection of spectrally dispersed stimulated Raman scattering signal. Using a homebuilt tuned amplifier array, our method enables Raman spectral acquisition, within the window defined by the broadband pulse, at the speed of 32 microseconds and with close to shot-noise limited detection sensitivity. Incorporated with multivariate curve resolution analysis, our platform allows compositional mapping of lipid droplets in single live cells, observation of intracellular retinoid metabolism, discrimination of fat droplets from protein-rich organelles in Caenorhabditis elegans, spectral detection of fast flowing tumor cells, and monitoring drug diffusion through skin tissue in vivo. The reported technique opens new opportunities for compositional analysis of cellular compartment in a microscope setting and high-throughput spectral profiling of single cells in a flow cytometer setting.

  7. Phase-measurement interferometric microscopy of microlithographically fabricated platinum electrodes

    SciTech Connect

    Smith, C.P.; Kennedy, H.L.; Kragt, H.J.; White, H.S. ); Biegen, J.F. )

    1990-06-01

    Phase-measurement interferometric microscopy (PMIM) was used to measure the topography of Pt microelectrodes fabricated on silicon wafers. Disk (10 {mu}m radius) and band (10 {mu}m {times} 1,000 {mu}m) electrodes were prepared by deposition and lithographic patterning of a 0.2 {mu}m thick SiO{sub 2} layer on a 0.05 {mu}m thick Pt film. Quantitative topographical images of the resulting electrodes are obtained from PMIM by accounting for the complex refractive index of Pt and for multiple reflections of light which occur within the transparent SiO{sub 2} layer. In situ, quantitative measurement of microelectrode topography in water is also demonstrated.

  8. Histological staining can enhance the performance of spectroscopic microscopy on sensing nanoarchitectural alterations of biological cells (Conference Presentation)

    NASA Astrophysics Data System (ADS)

    Zhang, Di; Cherkezyan, Lusik; Li, Yue; Capoglu, Ilker; Subramanian, Hariharan; Taflove, Allen; Backman, Vadim

    2017-02-01

    Our group had previously established that nanoscale three-dimensional refractive index (RI) fluctuations of a linear, dielectric, label-free medium can be sensed in the far field through spectroscopic microscopy, regardless of the diffraction limit of optical microscopy. Adopting this technique, Partial Wave Spectroscopic (PWS) Microscopy was able to sense nanoarchitectural alterations in early-stage cancers. With the success of PWS on detecting cancer from healthy clinical samples, we further investigated whether and how histological staining can enhance the performance of PWS by both finite difference time domain (FDTD) simulations and experiments. In this investigation, the dispersion models of hematoxylin and eosin were extracted from the absorption spectra of H and E stained cells. Using these models, the effect of staining were studied via FDTD simulations of unstained versus stained samples with various nanostructures. We observed that, the spectral variance was increased and the spectral variance difference between two samples with distinct nanostructures was enhanced in stained samples by over 200%. Furthermore, we investigated with FDTD whether molecule-specific staining can be used to enhance signals from a medium composing of the desired molecule. Samples with two mixed random media were created and the desired medium was either stained or unstained. Our results showed that the difference between the nanostructures of only the desired medium was enhanced in stained samples. We concluded that, with molecule-specific staining, PWS can selectively target the nanoarchitecture of a desired molecule. Lastly, these results were validated by experiments using human buccal cells from healthy or lung cancer patients. This study has significant impact in improving the performance of PWS on quantifying nanoarchitectural alterations during cancer.

  9. Three-dimensional quantitative phase imaging via tomographic deconvolution phase microscopy.

    PubMed

    Jenkins, Micah H; Gaylord, Thomas K

    2015-11-01

    The field of three-dimensional quantitative phase imaging (3D QPI) is expanding rapidly with applications in biological, medical, and industrial research, development, diagnostics, and metrology. Much of this research has centered on developing optical diffraction tomography (ODT) for biomedical applications. In addition to technical difficulties associated with coherent noise, ODT is not congruous with optical microscopy utilizing partially coherent light, which is used in most biomedical laboratories. Thus, ODT solutions have, for the most part, been limited to customized optomechanical systems which would be relatively expensive to implement on a wide scale. In the present work, a new phase reconstruction method, called tomographic deconvolution phase microscopy (TDPM), is described which makes use of commercial microscopy hardware in realizing 3D QPI. TDPM is analogous to methods used in deconvolution microscopy which improve spatial resolution and 3D-localization accuracy of fluorescence micrographs by combining multiple through-focal scans which are deconvolved by the system point spread function. TDPM is based on the 3D weak object transfer function theory which is shown here to be capable of imaging "nonweak" phase objects with large phase excursions. TDPM requires no phase unwrapping and recovers the entire object spectrum via object rotation, mitigating the need to fill in the "missing cone" of spatial frequencies algorithmically as in limited-angle ODT. In the present work, TDPM is demonstrated using optical fibers, including single-mode, polarization-maintaining, and photonic-crystal fibers as well as an azimuthally varying CO2-laser-induced long-period fiber grating period as test phase objects.

  10. Polymorphic phases of galactocerebrosides: spectroscopic evidence of lamellar crystalline structures.

    PubMed

    Bou Khalil, M; Carrier, D; Wong, P T; Tanphaichitr, N

    2001-06-06

    Fourier transform infrared spectroscopy was applied to study the structural and thermal properties of bovine brain galactocerebroside (GalCer) containing amide linked non-hydroxylated or alpha-hydroxy fatty acids (NFA- and HFA-GalCer, respectively). Over the temperature range 0-90 degrees C, both GalCer displayed complex thermal transitions, characteristic of polymorphic phase behavior. Upon heating, aqueous dispersions of NFA- and HFA-GalCer exhibited high order-disorder transition temperatures near 80 and 72 degrees C, respectively. En route to the chain melting transition, the patterns of the amide I band of NFA-GalCer were indicative of two different lamellar crystalline phases, whereas those of HFA-GalCer were suggestive of lamellar gel and crystalline bilayers. Cooling from the liquid-crystalline phase resulted in the formation of another crystalline phase of NFA-GalCer and a gel phase of HFA-GalCer, with a phase transition near 62 and 66 degrees C, respectively. Prolonged incubation of GalCer bilayers at 38 degrees C revealed conversions among lamellar crystalline phases (NFA-GalCer) or between lamellar gel and crystalline bilayer structures (HFA-GalCer). Spectral changes indicated that the temperature and/or time induced formation of the lamellar crystalline structures of NFA- and HFA-GalCer was accompanied by partial dehydration and by rearrangements of the hydrogen bonding network and bilayer packing mode of GalCer.

  11. Spectroscopic fingerprint of phase-incoherent superconductivity in the cuprate pseudogap state [corrected].

    PubMed

    Lee, Jhinhwan; Fujita, K; Schmidt, A R; Kim, Chung Koo; Eisaki, H; Uchida, S; Davis, J C

    2009-08-28

    A possible explanation for the existence of the cuprate "pseudogap" state is that it is a d-wave superconductor without quantum phase rigidity. Transport and thermodynamic studies provide compelling evidence that supports this proposal, but few spectroscopic explorations of it have been made. One spectroscopic signature of d-wave superconductivity is the particle-hole symmetric "octet" of dispersive Bogoliubov quasiparticle interference modulations. Here we report on this octet's evolution from low temperatures to well into the underdoped pseudogap regime. No pronounced changes occur in the octet phenomenology at the superconductor's critical temperature Tc, and it survives up to at least temperature T approximately 1.5 Tc. In this pseudogap regime, we observe the detailed phenomenology that was theoretically predicted for quasiparticle interference in a phase-incoherent d-wave superconductor. Thus, our results not only provide spectroscopic evidence to confirm and extend the transport and thermodynamics studies, but they also open the way for spectroscopic explorations of phase fluctuation rates, their effects on the Fermi arc, and the fundamental source of the phase fluctuations that suppress superconductivity in underdoped cuprates.

  12. Electronic Structure of the Cuprate Superconducting and Pseudogap Phases from Spectroscopic Imaging STM

    SciTech Connect

    Davis, J.C.; Schmidt, A.R.; Fujita, K.; Kim, E.-A.; Lawler, M.J.; Eisaki, H.; Uchida, S.; Lee, D.-H.

    2011-06-21

    We survey the use of spectroscopic imaging scanning tunneling microscopy (SI-STM) to probe the electronic structure of underdoped cuprates. Two distinct classes of electronic states are observed in both the d-wave superconducting (dSC) and the pseudogap (PG) phases. The first class consists of the dispersive Bogoliubov quasiparticle excitations of a homogeneous d-wave superconductor, existing below a lower energy scale E = {Delta}{sub 0}. We find that the Bogoliubov quasiparticle interference (QPI) signatures of delocalized Cooper pairing are restricted to a k-space arc, which terminates near the lines connecting k = {+-}({pi}/a{sub 0},0) to k = {+-}(0,{pi}/a{sub 0}). This arc shrinks continuously with decreasing hole density such that Luttinger's theorem could be satisfied if it represents the front side of a hole-pocket that is bounded behind by the lines between k = {+-}({pi}/a{sub 0},0) and k = {+-}(0,{pi}/a{sub 0}). In both phases, the only broken symmetries detected for the |E| < {Delta}{sub 0} states are those of a d-wave superconductor. The second class of states occurs proximate to the PG energy scale E = {Delta}{sub 1}. Here the non-dispersive electronic structure breaks the expected 90{sup o}-rotational symmetry of electronic structure within each unit cell, at least down to 180{sup o}-rotational symmetry. This electronic symmetry breaking was first detected as an electronic inequivalence at the two oxygen sites within each unit cell by using a measure of nematic (C{sub 2}) symmetry. Incommensurate non-dispersive conductance modulations, locally breaking both rotational and translational symmetries, coexist with this intra-unit-cell electronic symmetry breaking at E = {Delta}{sub 1}. Their characteristic wavevector Q is determined by the k-space points where Bogoliubov QPI terminates and therefore changes continuously with doping. The distinct broken electronic symmetry states (intra-unit-cell and finite Q) coexisting at E {approx} {Delta}{sub 1} are

  13. Silver nanoparticle-induced degranulation observed with quantitative phase microscopy

    NASA Astrophysics Data System (ADS)

    Yang, Wenzhong; Lee, Seungrag; Lee, Jiyong; Bae, Yoonsung; Kim, Dugyoung

    2010-02-01

    The use of AgNP is becoming more and more widespread in biomedical field. But compared with the promising bactericidal function, other physiological effects of AgNP on cells are relatively scant. In this research, we propose quantitative phase microscopy (QPM) as a new method to study the degranulation, and AgNP-induced RBL-2H3 cell degranulation is studied as well. Firstly, HeLa cells as the cell control and PBS as the solvent control, we measured the cell volume and cross section profile (x-z plane) with QPM. The results showed that the volume and cross section profile changed only the RBL-2H3 cells exposed to calcium ionophore A23187, which demonstrates the validity of QPM in degranulation research. Secondly, 50μg/mL of AgNP was used instead of A23187, and the measurement of cell volume and cross section profile was carried out again. RBL-2H3 cell volume increased immediately after AgNP was added, and cross section profile showed that the cell surface became granulated, but HeLa cell was lack of that effect. Phase images obviously indicated the RBL-2H3 cell deformation. Thirdly, stained with Fluo-3/AM, intracellular calcium Ca2+]i of single RBL-2H3 cell treated with AgNP was observed with fluorescent microscopy; incubated with AgNP for 20min, the supernatant of RBL-2H3 cells was collected and reacted with o-phthalaldehyde (OPA), then the fluorescent intensity of histamine-OPA complex was assayed with spectrofluorometer. The results of Ca2+]i and histamine increase showed that degranulation of AgNP-induced RBL-2H3 cell occurred. So, the cell volume was used as a parameter of degranulation in our study and AgNP-induced RBL-2H3 cells degranulation was confirmed by the cell volume increment, cross section profile change, and [Ca2+]i and histamine in supernatant increase.

  14. CONFOCAL MICROSCOPY SYSTEM PERFORMANCE: FOUNDATIONS FOR QUANTIFYING CYTOMETRIC APPLICATIONS WITH SPECTROSCOPIC INSTRUMENTS

    EPA Science Inventory

    The confocal laser-scanning microscopy (CLSM) has enormous potential in many biological fields. The goal of a CLSM is to acquire and quantify fluorescence and in some instruments acquire spectral characterization of the emitted signal. The accuracy of these measurements demands t...

  15. CONFOCAL MICROSCOPY SYSTEM PERFORMANCE: FOUNDATIONS FOR QUANTIFYING CYTOMETRIC APPLICATIONS WITH SPECTROSCOPIC INSTRUMENTS

    EPA Science Inventory

    The confocal laser-scanning microscopy (CLSM) has enormous potential in many biological fields. The goal of a CLSM is to acquire and quantify fluorescence and in some instruments acquire spectral characterization of the emitted signal. The accuracy of these measurements demands t...

  16. Global Abundance and Temperature Constraints via Joint Spectroscopic Phase Curve Retrievals

    NASA Astrophysics Data System (ADS)

    Line, Michael R.; Stevenson, Kevin B.; Bean, Jacob; Kreidberg, Laura; Fortney, Jonathan J.

    2016-01-01

    Spectroscopic thermal emission phase curves can provide us with a global view of an exoplanet's atmosphere. Different wavelengths probe different atmospheric depths whereas different phases probe different planetary longitudes. This in essence allows us to reconstruct the "3D" thermal and compositional structure of these atmospheres. In this contribution I will discuss the application of powerful atmospheric retrieval approaches to spectroscopic phase curve data, specifically the WFC3+Spitzer IRAC observations of the hot-Jupiter WASP-43b. First I will show the variation in thermal structures and molecular abundances with phase, assuming each phase is independent. Secondly, I will present a new framework for performing a joint retrieval on multiple phases simultaneously. In such a framework, I will test, via Bayesian hypothesis testing, a variety of assumptions. For instance, can the absorption features across all phases be explained with a global metallicity and C-to-O ratio under the assumption of thermochemical equilibrium? Can chemical quenching perturb the abundances on the cooler phases more than the hotter phases? Can we tell the difference? Can a global thermal structure "shape" explain all phases or is there structure variation with phase? Answering such questions are critical to understanding the complex interactions of atmospheric dynamics, chemical processes, and radiative energy balance in exoplanet atmospheres.

  17. Photoelectrochemical fabrication of spectroscopic diffraction gratings, phase 2

    NASA Technical Reports Server (NTRS)

    Rauh, R. David; Carrabba, Michael M.; Li, Jianguo; Cartland, Robert F.; Hachey, John P.; Mathew, Sam

    1990-01-01

    This program was directed toward the production of Echelle diffraction gratings by a light-driven, electrochemical etching technique (photoelectrochemical etching). Etching is carried out in single crystal materials, and the differential rate of etching of the different crystallographic planes used to define the groove profiles. Etching of V-groove profiles was first discovered by us during the first phase of this project, which was initially conceived as a general exploration of photoelectrochemical etching techniques for grating fabrication. This highly controllable V-groove etching process was considered to be of high significance for producing low pitch Echelles, and provided the basis for a more extensive Phase 2 investigation.

  18. Study of spatial homogeneity and nitridation of an Al nanopattern template with spectroscopic photoemission and low energy electron microscopy

    NASA Astrophysics Data System (ADS)

    Qi, B.; Ólafsson, S.; Zakharov, A. A.; Göthelid, M.; Agnarsson, B.; Gislason, H. P.

    2013-01-01

    We report a study on the spatial homogeneity and nitridation of a nanopattern template using a spectroscopic photoemission and low energy electron microscopy. The template was composed of Al nanodots which were patterned into a SiO2/Si(1 1 1) surface using e-beam lithography and reactive ion etching. The template exhibited a global inhomogeneity in terms of the local topography, Al composition and structure of the individual nanopatterns. After nitridation, the individual nanopatterns were diminished, more corrugated and faceted. The nitridated nanopatterns were structurally ordered but differently orientated. The nitridation effectively removed the fluorine contaminants by decomposition of the fluorocarbon sidewalls, resulting in the AlN nanopatterns and partially nitridated Si substrate surface outside the nanopattern domains.

  19. Atomic oxygen effects on thin film space coatings studied by spectroscopic ellipsometry, atomic force microscopy, and laser light scattering

    NASA Technical Reports Server (NTRS)

    Synowicki, R. A.; Hale, Jeffrey S.; Woollam, John A.

    1992-01-01

    The University of Nebraska is currently evaluating Low Earth Orbit (LEO) simulation techniques as well as a variety of thin film protective coatings to withstand atomic oxygen (AO) degradation. Both oxygen plasma ashers and an electron cyclotron resonance (ECR) source are being used for LEO simulation. Thin film coatings are characterized by optical techniques including Variable Angle Spectroscopic Ellipsometry, Optical spectrophotometry, and laser light scatterometry. Atomic Force Microscopy (AFM) is also used to characterize surface morphology. Results on diamondlike carbon (DLC) films show that DLC degrades with simulated AO exposure at a rate comparable to Kapton polyimide. Since DLC is not as susceptible to environmental factors such as moisture absorption, it could potentially provide more accurate measurements of AO fluence on short space flights.

  20. Condensed phase photoacoustic spectroscopic detection of porphyrins and dyes

    SciTech Connect

    Voigtman, E.; Jurgensen, A.; Winefordner, J.

    1981-08-01

    A simple piezoelectric detection system suitable for performing highly sensitive liquid-phase photoacoustic spectroscopy is presented. Performance of the system with respect to linearity, solvent effects, and excitation pulse characteristics is experimentally evaluated and compared with theoretical figures of merit. Limits of detection for various porphyrins, laser dyes, and drugs are presented.

  1. Method and apparatus for differential spectroscopic atomic-imaging using scanning tunneling microscopy

    DOEpatents

    Kazmerski, Lawrence L.

    1990-01-01

    A Method and apparatus for differential spectroscopic atomic-imaging is disclosed for spatial resolution and imaging for display not only individual atoms on a sample surface, but also bonding and the specific atomic species in such bond. The apparatus includes a scanning tunneling microscope (STM) that is modified to include photon biasing, preferably a tuneable laser, modulating electronic surface biasing for the sample, and temperature biasing, preferably a vibration-free refrigerated sample mounting stage. Computer control and data processing and visual display components are also included. The method includes modulating the electronic bias voltage with and without selected photon wavelengths and frequency biasing under a stabilizing (usually cold) bias temperature to detect bonding and specific atomic species in the bonds as the STM rasters the sample. This data is processed along with atomic spatial topography data obtained from the STM raster scan to create a real-time visual image of the atoms on the sample surface.

  2. Silver nanoparticle-induced degranulation observed with quantitative phase microscopy

    NASA Astrophysics Data System (ADS)

    Yang, Wenzhong; Lee, Seungrag; Lee, Jiyong; Bae, Yoonsung; Kim, Dugyoung

    2010-07-01

    Monitoring a degranulation process in a live mast cell is a quite important issue in immunology and pharmacology. Because the size of a granule is normally much smaller than the resolution limit of an optical microscope system, there is no direct real-time live cell imaging technique for observing degranulation processes except for fluorescence imaging techniques. In this research, we propose optical quantitative phase microscopy (QPM) as a new observation tool to study degranulation processes in a live mast cell without any fluorescence labeling. We measure the cell volumes and the cross sectional profiles (x-z plane) of an RBL-2H3 cell and a HeLa cell, before and after they are exposed to calcium ionophore A23187 and silver nanoparticles (AgNPs). We verify that the volume and the cross sectional line profile of the RBL-2H3 cell were changed significantly when it was exposed to A23187. When 50 μg/mL of AgNP is used instead of A23187, the measurements of cell volume and cross sectional profiles indicate that RBL-2H3 cells also follow degranulation processes. Degranulation processes for these cells are verified by monitoring the increase of intracellular calcium ([Ca2+]i) and histamine with fluorescent methods.

  3. Superresolution microscopy using common-path phase-shifting interferometry

    NASA Astrophysics Data System (ADS)

    Micó, Vicente; Zalevsky, Zeev; García, Javier

    2008-04-01

    In this contribution, we present a new approach aimed to achieve superresolution in digital holographic microscopy that overcomes the limitations imposed by the Abbe's diffraction limit. Based on time multiplexing and off-axis illumination, a common-path interferometric architecture allows the synthesis of an enlarged aperture that expands the cutoff frequency of the conventional imaging system. Complex object amplitude distribution is recovered using an extremely simple phase-shifting configuration instead of image plane off-axis holographic recording. As a consequence, the setup becomes easy-to-configure (less requirements and lower number of optical elements) and it is useful for practical implementation in microscopes while only simple modifications are required (no higher magnifications are needed to resolve the interference pattern at the CCD plane). Experimental results validating the capabilities of the presented approach when a low numerical aperture commercial microscope objective is used as imaging lens are included using both a synthetic object (high resolution USAF test target) and a biosample (red blood cells) as objects under test.

  4. On measuring cell confluence in phase contrast microscopy

    NASA Astrophysics Data System (ADS)

    Dempsey, K. P.; Richardson, J. B.; Lam, K. P.

    2014-03-01

    A principal focus highlighting recent advances in cell based therapies concerns the development of effective treatments for osteoarthritis. Earlier clinicaltrials have shown that 80% of patients receiving mesenchymal stem cell(MSC) based treatment have improved their quality of life by alleviating pain whilst extending the life of their natural joints. The current challenge facing researchers is to identify the biological differences between the treatments that have worked and those which have shown little improvement. One possible candidate for the difference in treatment prognosis is an examination of the proliferation of the ( type) cells as they grow. To further understanding of the proliferation and differentiation of MSC, non-invasive live cell imaging techniques have been developed which capture important cell events and dynamics in cell divisions over an extended period of time. An automated image analysis procedure capable of tracking cell confluence over time has also been implemented, providing an objective and realistic estimation of cell growth within continuous live cell cultures. The proposed algorithm accounts for the halo artefacts that occur in phase microscopy. In addition to a favourable run-time performance, the method was also validated using continuous live MSC cultures, with consistent and meaningful results.

  5. Scanning electron microscopy of the endometrium during the secretory phase.

    PubMed Central

    Motta, P M; Andrews, P M

    1976-01-01

    Scanning electron microscopy was used to study the surface morphology of the rabbit endometrium during the secretory phase of the oestrous cycle. The free surfaces of ciliated and of inactive active secretory cells are described. Changes in secretory cell surface morphology resulting from accumulation and secretion of material involve the apparent retraction of microvilli and the formation of one or more bulbous protrusions of the cell's apical surface. These protrusions may be relatively smooth surfaced or exhibit long slender micro-extensions. The protrusions grow in size and are eventually pinched off. Loss of the bulbous protrusions often leaves behind crater-like invaginations of the cell's surface. Secretory cells adjacent to the endometrial glands are the first to exhibit signs of mucin accumulation and secretion. The single cilium of a secretory cell is not apparently affected by the secretory process. Signs of ciliated and secretory cell degeneration, and possible sloughing, are also described. Images Fig. 1 Fig. 2 Fig. 3 Fig. 4 Fig. 5 Fig. 6 Fig. 7 PMID:1033932

  6. Analytical electron microscopy in mineralogy; exsolved phases in pyroxenes

    USGS Publications Warehouse

    Nord, G.L.

    1982-01-01

    Analytical scanning transmission electron microscopy has been successfully used to characterize the structure and composition of lamellar exsolution products in pyroxenes. At operating voltages of 100 and 200 keV, microanalytical techniques of x-ray energy analysis, convergent-beam electron diffraction, and lattice imaging have been used to chemically and structurally characterize exsolution lamellae only a few unit cells wide. Quantitative X-ray energy analysis using ratios of peak intensities has been adopted for the U.S. Geological Survey AEM in order to study the compositions of exsolved phases and changes in compositional profiles as a function of time and temperature. The quantitative analysis procedure involves 1) removal of instrument-induced background, 2) reduction of contamination, and 3) measurement of correction factors obtained from a wide range of standard compositions. The peak-ratio technique requires that the specimen thickness at the point of analysis be thin enough to make absorption corrections unnecessary (i.e., to satisfy the "thin-foil criteria"). In pyroxenes, the calculated "maximum thicknesses" range from 130 to 1400 nm for the ratios Mg/Si, Fe/Si, and Ca/Si; these "maximum thicknesses" have been contoured in pyroxene composition space as a guide during analysis. Analytical spatial resolutions of 50-100 nm have been achieved in AEM at 200 keV from the composition-profile studies, and analytical reproducibility in AEM from homogeneous pyroxene standards is ?? 1.5 mol% endmember. ?? 1982.

  7. Study of fossil bones by synchrotron radiation micro-spectroscopic techniques and scanning electron microscopy.

    PubMed

    Zougrou, I M; Katsikini, M; Pinakidou, F; Paloura, E C; Papadopoulou, L; Tsoukala, E

    2014-01-01

    Earlymost Villafranchian fossil bones of an artiodactyl and a perissodactyl from the Milia excavation site in Grevena, Greece, were studied in order to evaluate diagenetic effects. Optical microscopy revealed the different bone types (fibro-lamellar and Haversian, respectively) of the two fragments and their good preservation state. The spatial distribution of bone apatite and soil-originating elements was studied using micro-X-ray fluorescence (µ-XRF) mapping and scanning electron microscopy. The approximate value of the Ca/P ratio was 2.2, as determined from scanning electron microscopy measurements. Bacterial boring was detected close to the periosteal region and Fe bearing oxides were found to fill bone cavities, e.g. Haversian canals and osteocyte lacunae. In the perissodactyl bone considerable amounts of Mn were detected close to cracks (the Mn/Fe weight ratio takes values up to 3.5). Goethite and pyrite were detected in both samples by means of metallographic microscopy. The local Ca/P ratio determined with µ-XRF varied significantly in metal-poor spots indicating spatial inhomogeneities in the ionic substitutions. XRF line scans that span the bone cross sections revealed that Fe and Mn contaminate the bones from both the periosteum and medullar cavity and aggregate around local maxima. The formation of goethite, irrespective of the local Fe concentration, was verified by the Fe K-edge X-ray absorption fine structure (XAFS) spectra. Finally, Sr K-edge extended XAFS (EXAFS) revealed that Sr substitutes for Ca in bone apatite without obvious preference to the Ca1 or Ca2 unit-cell site occupation.

  8. Reconstruction of explicit structural properties at the nanoscale via spectroscopic microscopy

    NASA Astrophysics Data System (ADS)

    Cherkezyan, Lusik; Zhang, Di; Subramanian, Hariharan; Taflove, Allen; Backman, Vadim

    2016-02-01

    The spectrum registered by a reflected-light bright-field spectroscopic microscope (SM) can quantify the microscopically indiscernible, deeply subdiffractional length scales within samples such as biological cells and tissues. Nevertheless, quantification of biological specimens via any optical measures most often reveals ambiguous information about the specific structural properties within the studied samples. Thus, optical quantification remains nonintuitive to users from the diverse fields of technique application. In this work, we demonstrate that the SM signal can be analyzed to reconstruct explicit physical measures of internal structure within label-free, weakly scattering samples: characteristic length scale and the amplitude of spatial refractive-index (RI) fluctuations. We present and validate the reconstruction algorithm via finite-difference time-domain solutions of Maxwell's equations on an example of exponential spatial correlation of RI. We apply the validated algorithm to experimentally measure structural properties within isolated cells from two genetic variants of HT29 colon cancer cell line as well as within a prostate tissue biopsy section. The presented methodology can lead to the development of novel biophotonics techniques that create two-dimensional maps of explicit structural properties within biomaterials: the characteristic size of macromolecular complexes and the variance of local mass density.

  9. Reconstruction of explicit structural properties at the nanoscale via spectroscopic microscopy.

    PubMed

    Cherkezyan, Lusik; Zhang, Di; Subramanian, Hariharan; Taflove, Allen; Backman, Vadim

    2016-02-01

    The spectrum registered by a reflected-light bright-field spectroscopic microscope (SM) can quantify the microscopically indiscernible, deeply subdiffractional length scales within samples such as biological cells and tissues. Nevertheless, quantification of biological specimens via any optical measures most often reveals ambiguous information about the specific structural properties within the studied samples. Thus, optical quantification remains nonintuitive to users from the diverse fields of technique application. In this work, we demonstrate that the SM signal can be analyzed to reconstruct explicit physical measures of internal structure within label-free, weakly scattering samples: characteristic length scale and the amplitude of spatial refractive-index (RI) fluctuations. We present and validate the reconstruction algorithm via finite-difference time-domain solutions of Maxwell's equations on an example of exponential spatial correlation of RI. We apply the validated algorithm to experimentally measure structural properties within isolated cells from two genetic variants of HT29 colon cancer cell line as well as within a prostate tissue biopsy section. The presented methodology can lead to the development of novel biophotonics techniques that create two-dimensional maps of explicit structural properties within biomaterials: the characteristic size of macromolecular complexes and the variance of local mass density.

  10. Reconstruction of explicit structural properties at the nanoscale via spectroscopic microscopy

    PubMed Central

    Cherkezyan, Lusik; Zhang, Di; Subramanian, Hariharan; Taflove, Allen; Backman, Vadim

    2016-01-01

    Abstract. The spectrum registered by a reflected-light bright-field spectroscopic microscope (SM) can quantify the microscopically indiscernible, deeply subdiffractional length scales within samples such as biological cells and tissues. Nevertheless, quantification of biological specimens via any optical measures most often reveals ambiguous information about the specific structural properties within the studied samples. Thus, optical quantification remains nonintuitive to users from the diverse fields of technique application. In this work, we demonstrate that the SM signal can be analyzed to reconstruct explicit physical measures of internal structure within label-free, weakly scattering samples: characteristic length scale and the amplitude of spatial refractive-index (RI) fluctuations. We present and validate the reconstruction algorithm via finite-difference time-domain solutions of Maxwell’s equations on an example of exponential spatial correlation of RI. We apply the validated algorithm to experimentally measure structural properties within isolated cells from two genetic variants of HT29 colon cancer cell line as well as within a prostate tissue biopsy section. The presented methodology can lead to the development of novel biophotonics techniques that create two-dimensional maps of explicit structural properties within biomaterials: the characteristic size of macromolecular complexes and the variance of local mass density. PMID:26886803

  11. A spectroscopic and catalytic investigation of active phase-support interactions

    SciTech Connect

    Haller, G.L.

    1991-01-01

    Active catalytic phases (metal, mixed metals, oxide or mixed oxides) interacting with oxide support on which the active phase is dispersed can affect the percentage exposed, the morphology of supported particles, the degree of reducibility of cations, etc., in a variety of ways. Our objective is to characterize the physical chemistry of the active phase-oxide support by spectroscopic methods and to correlate this structure with catalytic function. The three systems discussed in this progress report are Ag/TiO{sub 2}, Ru-Cu/SiO{sub 2} and SiO{sub 2}/Al{sub 2}O{sub 3}. 24 refs., 3 figs., 2 tabs.

  12. Volta potential phase plate for in-focus phase contrast transmission electron microscopy

    PubMed Central

    Danev, Radostin; Buijsse, Bart; Khoshouei, Maryam; Plitzko, Jürgen M.; Baumeister, Wolfgang

    2014-01-01

    We describe a phase plate for transmission electron microscopy taking advantage of a hitherto-unknown phenomenon, namely a beam-induced Volta potential on the surface of a continuous thin film. The Volta potential is negative, indicating that it is not caused by beam-induced electrostatic charging. The film must be heated to ∼200 °C to prevent contamination and enable the Volta potential effect. The phase shift is created “on the fly” by the central diffraction beam eliminating the need for precise phase plate alignment. Images acquired with the Volta phase plate (VPP) show higher contrast and unlike Zernike phase plate images no fringing artifacts. Following installation into the microscope, the VPP has an initial settling time of about a week after which the phase shift behavior becomes stable. The VPP has a long service life and has been used for more than 6 mo without noticeable degradation in performance. The mechanism underlying the VPP is the same as the one responsible for the degradation over time of the performance of thin-film Zernike phase plates, but in the VPP it is used in a constructive way. The exact physics and/or chemistry behind the process causing the Volta potential are not fully understood, but experimental evidence suggests that radiation-induced surface modification combined with a chemical equilibrium between the surface and residual gases in the vacuum play an important role. PMID:25331897

  13. GAS PHASE MOLECULAR DYNAMICS: HIGH-RESOLUTION SPECTROSCOPIC PROBES OF CHEMICAL DYNAMICS.

    SciTech Connect

    HALL, G.E.

    2006-05-30

    This research is carried out as part of the Gas Phase Molecular Dynamics group program in the Chemistry Department at Brookhaven National Laboratory. High-resolution spectroscopic tools are developed and applied to problems in chemical dynamics. Recent topics have included the state-resolved studies of collision-induced electronic energy transfer, dynamics of barrierless unimolecular reactions, and the kinetics and spectroscopy of transient species.

  14. Detection of secondary phases in duplex stainless steel by magnetic force microscopy and scanning Kelvin probe force microscopy

    SciTech Connect

    Ramírez-Salgado, J.; Domínguez-Aguilar, M.A.; Castro-Domínguez, B.; Hernández-Hernández, P.; Newman, R.C.

    2013-12-15

    The secondary phase transformations in a commercial super duplex stainless steel were investigated by micro-chemical analyses and high resolution scanning probe microscopy. Energy dispersive X-ray and electron probe detected ferrite and austenite as well as secondary phases in unetched aged duplex stainless steel type 25Cr-7Ni-3Mo. Volta potential indicated that nitride and sigma appeared more active than ferrite, while secondary austenite and austenite presented a nobler potential. Reversal order in nobility is thought to be attributable to the potential ranking provided by oxide nature diversity as a result of secondary phase surface compositions on steel. After eutectoid transformation, secondary austenite was detected by electron probe microanalysis, whereas atomic force microscopy distinguished this phase from former austenite by image contrast. Magnetic force microscopy revealed a “ghosted” effect on the latter microstructure probably derived from metal memory reminiscence of mechanical polishing at passivity and long range magnetic forces of ferrite phase. - Highlights: • Nobility detection of secondary phases by SKPFM in DSS particles is not a straightforward procedure. • As Volta potential and contrast are not always consistent SKPFM surface oxides is thought played an important role in detection. • AFM distinguished secondary austenite from former austenite by image contrast though SEM required EPMA.

  15. Intensity and phase fields behind Phase Shifting Masks studied with High Resolution Interference Microscopy

    NASA Astrophysics Data System (ADS)

    Puthankovilakam, Krishnaparvathy; Scharf, Toralf; Herzig, Hans Peter; Weichelt, Tina; Zeitner, Uwe; Vogler, Uwe; Voelkel, Reinhard

    2015-03-01

    The proximity printing industry is in real need of high resolution results and it can be done using Phase Shift Mask (PSM) or by applying Optical Proximity Correction (OPC). In our research we are trying to find out details of how light fields behind the structures of photo masks develop in order to determine the best conditions and designs for proximity printing. We focus here on parameters that are used in real situation with gaps up to 50 μm and structure sizes down to 2 μm. The light field evolution behind the structures is studied and delivers insight in to precisions and tolerances that need to be respected. It is the first time that an experimental analysis of light propagation through mask is presented in detail, which includes information on intensity and phase. The instrument we use is known as High Resolution Interference Microscopy (HRIM). HRIM is a Mach-Zehnder interferometer which is capable of recording three dimensional distributions of intensity and phase with diffraction limited resolution. Our characterization technique allows plotting the evolution of the desired light field and therefore printable structure till the desired proximity gap. In this paper we discuss in detail the evolution of intensity and phase fields of elbow or corner structure at different position behind a phase mask and interpret the main parameters. Of particular interest are tolerances against proximity gap variation and the resolution in printed structures.

  16. Development of in-situ full-field spectroscopic imaging analysis and application on Li-ion battery using transmission x-ray microscopy

    NASA Astrophysics Data System (ADS)

    Chen-Wiegart, Yu-chen K.; Wang, Jiajun; Wang, Jun

    2013-09-01

    This paper presents the advance in spectroscopic imaging technique and analysis method from the newly developed transmission x-ray microscopy (TXM) at the beamline X8C of National Synchrotron Light Source. Through leastsquares linear combination fitting we developed on the in situ spectroscopic images, a time-dependent and spatially resolved chemical composition mapping can be obtained and quantitatively analyzed undergone chemical/electrochemical reactions. A correlation of morphological evolution, chemical state distribution changes and reaction conditions can be revealed. We successfully applied this method to study the electrochemical evolution of CuO, an anode material of Li-ion battery, during the lithiation-delitiation cycling.

  17. A view of the implanted SiC damage by Rutherford backscattering spectroscopy, spectroscopic ellipsometry, and transmission electron microscopy

    SciTech Connect

    Battistig, G.; Khanh, N. Q.; Petrik, P.; Lohner, T.; Dobos, L.; Pecz, B.; Garcia Lopez, J.; Morilla, Y.

    2006-11-01

    4H-SiC single crystalline substrates were implanted at room temperature with 150 keV Al{sup +} ions using fluences of 4x10{sup 14}, 1x10{sup 15}, and 2x10{sup 15} cm{sup -2} with current density of 2.5 {mu}A cm{sup -2}. The samples were subsequently annealed at 1100 deg. C in N{sub 2} for 1 h in order to analyze their structural recovery. The disorder induced in both sublattices by the Al{sup +} ions was studied by backscattering spectrometry in channeling geometry with a 3.5 MeV He{sup 2+} beam. The results were compared with the optical properties of the samples measured by spectroscopic ellipsometry. In a previous work, we concluded that during the postimplantation annealing of a highly damaged SiC crystalline material the short distance order can be recovered, while the long distance disorder remains. We also presented the possibility to have grains of different polytypes oriented faraway from the original direction. Now, this alternative is confirmed by the cross-sectional transmission and high resolution electron microscopy studies, carried out to obtain information about the crystal structure.

  18. Laser scanning saturated structured illumination microscopy based on phase modulation

    NASA Astrophysics Data System (ADS)

    Huang, Yujia; Zhu, Dazhao; Jin, Luhong; Kuang, Cuifang; Xu, Yingke; Liu, Xu

    2017-08-01

    Wide-field saturated structured illumination microscopy has not been widely used due to the requirement of high laser power. We propose a novel method called laser scanning saturated structured illumination microscopy (LS-SSIM), which introduces high order of harmonics frequency and greatly reduces the required laser power for SSIM imaging. To accomplish that, an excitation PSF with two peaks is generated and scanned along different directions on the sample. Raw images are recorded cumulatively by a CCD detector and then reconstructed to form a high-resolution image with extended optical transfer function (OTF). Our theoretical analysis and simulation results show that LS-SSIM method reaches a resolution of 0.16 λ, equivalent to 2.7-fold resolution than conventional wide-field microscopy. In addition, LS-SSIM greatly improves the optical sectioning capability of conventional wide-field illumination system by diminishing our-of-focus light. Furthermore, this modality has the advantage of implementation in multi-photon microscopy with point scanning excitation to image samples in greater depths.

  19. Phase-shifting by means of an electronically tunable lens: quantitative phase imaging of biological specimens with digital holographic microscopy.

    PubMed

    Trujillo, Carlos; Doblas, Ana; Saavedra, Genaro; Martínez-Corral, Manuel; García-Sucerquia, Jorge

    2016-04-01

    The use of an electronically tunable lens (ETL) to produce controlled phase shifts in interferometric arrangements is shown. The performance of the ETL as a phase-shifting device is experimentally validated in phase-shifting digital holographic microscopy. Quantitative phase maps of a section of the thorax of a Drosophila melanogaster fly and of human red blood cells have been obtained using our proposal. The experimental results validate the possibility of using the ETL as a reliable phase-shifter device.

  20. Intensity and phase fields behind phase-shifting masks studied with high-resolution interference microscopy

    NASA Astrophysics Data System (ADS)

    Puthankovilakam, Krishnaparvathy; Scharf, Toralf; Kim, Myun Sik; Naqavi, Ali; Herzig, Hans Peter; Weichelt, Tina; Zeitner, Uwe; Vogler, Uwe; Voelkel, Reinhard

    2016-04-01

    We try to find out the details of how light fields behind the structures of photomasks develop in order to determine the best conditions and designs for proximity printing. The parameters that we use approach real situations like structure printing at proximity gaps of 20 to 50 μm and structure sizes down to 2 μm. This is the first time that an experimental analysis of light propagation through a mask is presented in detail, which includes information on intensity and phase. We use high-resolution interference microscopy (HRIM) for the measurement. HRIM is a Mach-Zehnder interferometer, which is capable of recording three-dimensional distributions of intensity and phase with diffraction-limited resolution. Our characterization technique allows plotting the evolution of the desired light field, usually called the aerial image, and therefore gives access to the printable structure until the desired proximity gap. Here, we discuss in detail the evolution of intensity and phase fields of elbow or corner structures at different positions behind a phase mask and interpret the main parameters. Of particular interest are tolerances against proximity gap variation and the theoretical explanation of the resolution in printed structures.

  1. Super-resolution Microscopy of Lipid Bilayer Phases

    PubMed Central

    Kuo, Chinkuei; Hochstrasser, Robin M.

    2011-01-01

    Sub-diffraction optical imaging with nanometer resolution of lipid phase separated regions is reported. Merocyanine 540, a probe whose fluorescence is sensitive to the lipid phase, is combined with super-resolution imaging to distinguish the liquid and gel phase nanoscale domains of lipid bilayers supported on glass. The monomer-dimer equilibrium of MC 540 in membranes is deemed responsible for the population difference of single molecule fluorescence bursts in the different phase regions. The extension of this method to other binary or ternary lipid models or natural systems provides a promising new super-resolution strategy. PMID:21405121

  2. Lensless phase contrast microscopy based on multiwavelength Fresnel diffraction.

    PubMed

    Noom, Daniel W E; Eikema, Kjeld S E; Witte, Stefan

    2014-01-15

    We demonstrate a compact, wide-field, quantitative phase contrast microscope that does not require lenses for image formation. High-resolution images are retrieved from Fresnel diffraction patterns recorded at multiple wavelengths, combined with a robust iterative phase retrieval algorithm. Quantitative phase contrast images of living cultured neurons are obtained with a transverse resolution of <2 μm. Our system is well suited for high-resolution live cell imaging and provides a compact, cost-effective alternative to full-sized phase-contrast microscopes.

  3. Synthetic aperture common-path digital holographic microscopy with spiral phase filter

    NASA Astrophysics Data System (ADS)

    Tu, Han-Yen; Hsiao, Wei-Jen; Lai, Xin-Ji; Lin, Yu-Chih; Cheng, Chau-Jern

    2017-06-01

    This paper describes a synthetic aperture common-path digital holographic microscopy through phase-shifting in a rotating spiral phase filter (SPF) to strengthen axial phase accuracy and fully utilize frequency coverage to improve of the spatial resolution. A phase-type liquid crystal on a silicon spatial light modulator was employed to generate an SPF on the Fourier plane of the digital holographic microscopy system. Subsequently, the scattering wave passing through the spiral phase plate and the reference wave generated through the center of phase plate as a spatial filter were used for the interference process, thereby completing the recording of the on-axis digital hologram. The common-path digital holographic microscopy system integrated with synthetic aperture imaging can be applied for Fresnel hologram recording and numerical focusing on objects without being affected by environmental vibration and coherence, thereby improving the spatial resolution and axial phase accuracy.

  4. Microscopy

    Treesearch

    Patricia A. Moss; Les Groom

    2001-01-01

    Microscopy is the study and interpretation of images produced by a microscope. "Interpretation" is the keyword, because the microscope enables one to see structures that are too small or too close together to be resolved by the unaided eye. (The human eye cannot separate two points or lines that are closer together than 0.1 mm.) it is important to...

  5. LDRD final report : raman spectroscopic measurements to monitor the HMX beta-delta phase transition.

    SciTech Connect

    Renlund, Anita Mariana; Tappan, Alexander Smith; Miller, Jill C.

    2000-11-01

    The HMX {beta}-{delta} solid-solid phase transition, which occurs as HMX is heated near 170 C, is linked to increased reactivity and sensitivity to initiation. Thermally damaged energetic materials (EMs) containing HMX therefore may present a safety concern. Information about the phase transition is vital to predictive safety models for HMX and HMX-containing EMs. We report work on monitoring the phase transition with real-time Raman spectroscopy aimed towards obtaining a better understanding of physical properties of HMX through the phase transition. HMX samples were confined in a cell of minimal free volume in a displacement-controlled or load-controlled arrangement. The cell was heated and then cooled at controlled rates while real-time Raman spectroscopic measurements were performed. Raman spectroscopy provides a clear distinction between the phases of HMX because the vibrational transitions of the molecule change with conformational changes associated with the phase transition. Temperature of phase transition versus load data are presented for both the heating and cooling cycles in the load-controlled apparatus, and general trends are discussed. A weak dependence of the temperature of phase transition on load was discovered during the heating cycle, with higher loads causing the phase transition to occur at a higher temperature. This was especially true in the temperature of completion of phase transition data as opposed to the temperature of onset of phase transition data. A stronger dependence on load was observed in the cooling cycle, with higher loads causing the reverse phase transitions to occur at a higher cooling temperature. Also, higher loads tended to cause the phase transition to occur over a longer period of time in the heating cycle and over a shorter period of time in the cooling cycle. All three of the pure HMX phases ({alpha}, {beta} and {delta}) were detected on cooling of the heated samples, either in pure form or as a mixture.

  6. X-ray Phase Imaging Microscopy using a Fresnel Zone Plate and a Transmission Grating

    SciTech Connect

    Yashiro, Wataru; Momose, Atsushi; Takeuchi, Akihisa; Suzuki, Yoshio

    2010-06-23

    We report on a hard X-ray phase imaging microscopy (a phase-difference microscopy) that consists of an objective and a transmission grating. The simple optical system provides a quantitative phase image, and does not need a wave field mostly coherent on the objective. Our method has a spatial resolution almost same as that of the absorption contrast microscope image obtained by removing the grating. We demonstrate how our approach provides a phase image from experimentally obtained images. Our approach is attractive for easily appending a quantitative phase-sensitive mode to normal X-ray microscopes, and has potentially broad applications in biology and material sciences.

  7. In-vivo analysis of human cancer cells by means of laser phase microscopy

    NASA Astrophysics Data System (ADS)

    Perevedentseva, Elena V.; Graschev, G.; Balanos, Evangelos; Dressler, Cathrin; Beuthan, Juergen; Schlag, Peter M.

    1999-12-01

    Phase images of human colon carcinoma cells CX1, HCT15, HCT116 and human urinary bladder carcinoma cells EJ28 were obtained with laser phase microscope and have been analyzed. The phase shift images are compared with conventional light microscopy and confocal laser scanning microscopy data in order to detect the correspondence between morphological peculiarities and phase characteristics of measured cell. Influence of interactions between human carcinoma cells and specific antibodies (CC18 monoclonal antibody to cytokeratin and MRK 16 monoclonal antibody) on the cells phase images was studied.

  8. Dynamic phase imaging of host cells attacked by Vibrio vulnificus using quantitative phase microscopy

    NASA Astrophysics Data System (ADS)

    Lee, Seungrag; Yang, Wenzhong; Lee, Ji Yong; Cha, Mi Hye; Kim, Young Ran; Kim, Dug Young

    2010-02-01

    We present the real time quantitative analysis of Vibrio vulnificus-infected host cells using high stability quantitative phase microscopy (HSQPM). It provides the ability to retrieve the phase or optical path length distribution over the cell from a single interferogram image, which has been measured with nanometer path length sensitivity for long periods of time. We have applied HSQPM to study dynamic cell morphologic changes and to quantify noninvasively cell volumes of rat basophilic leukemia RBL-2H3 cells infected with pathogenic bacteria V. vulnificus strains, wild type (MO6-24/O) and RTX toxin mutant (CMM770). During the process of V. vulnificus wild type infection to RBL-2H3 cells, the dynamic changes of quantitative phase images, cell volumes and areas were observed in real time using HSQPM. In contrast, the dramatic changes were not detected in RBL-2H3 cells infected with RTX toxin mutant. The results showed the good correlation between HSQPM analysis and biochemical assays such as lactate dehydrogenase (LDH) assay and β-hexosaminidase release assay. We suggest that HSQPM is useful real time quantitative method to study the dynamic process of host cells infected with pathogen in a noninvasive manner.

  9. Theoretical study of ferroelectric nanoparticles using phase reconstructed electron microscopy

    SciTech Connect

    Phatak, C.; Petford-Long, A. K.; Beleggia, M.; De Graef, M.

    2014-06-01

    Ferroelectric nanostructures are important for a variety of applications in electronic and electro-optical devices, including nonvolatile memories and thin-film capacitors. These applications involve stability and switching of polarization using external stimuli, such as electric fields. We present a theoretical model describing how the shape of a nanoparticle affects its polarization in the absence of screening charges, and quantify the electron-optical phase shift for detecting ferroelectric signals with phase-sensitive techniques in a transmission electron microscope. We provide an example phase shift computation for a uniformly polarized prolate ellipsoid with varying aspect ratio in the absence of screening charges.

  10. Transfer characteristics of rectangular phase gratings in interference microscopy.

    PubMed

    Lehmann, Peter; Xie, Weichang; Niehues, Jan

    2012-02-15

    In this Letter, the transfer characteristics of rectangular periodic phase objects are studied. It turns out that there are significant differences compared to amplitude objects. The imaging of an amplitude object can be understood as a linear process, whereas phase objects behave nonlinearly. It is shown that under certain conditions the correct shape of a rectangular phase grating can be obtained by an interference microscope as long as the first order diffraction component passes the optical imaging system. This result is in a good agreement with experimental observations and computer simulation results.

  11. Raman Spectroscopic and Ultrasonic Measurements to Monitor the HMX ( ) Phase Transition

    SciTech Connect

    GIESKE,JOHN H.; MILLER,JILL C.; RENLUND,ANITA M.; TAPPAN,ALEXANDER S.

    1999-10-14

    The HMX {beta}-{delta} solid-solid phase transition, which occurs as HMX is heated near 170 C, is clearly linked to increased reactivity and sensitivity to initiation. Thermally damaged energetic materials (EMs) containing HMX therefore may present a safety concern. Information about the phase transition is vital to a predictive safety model for HMX and HMX-containing EMs. We report work in progress on monitoring the phase transition with real-time Raman spectroscopy and ultrasonic measurements aimed towards a better understanding of physical properties through the phase transition. HMX samples were confined with minimal free volume.in a cell with constant volume. The cell was heated at a controlled rate and real-time Raman spectroscopic or ultrasonic measurements were performed. Raman spectroscopy provides a clear distinction between the two phases because the vibrational transitions of the molecule change with confirmational changes associated with the phase transition. Ultrasonic time-of-flight measurements provide an additional method of distinguishing the two phases because the sound speed through the material changes with the phase transition. Ultrasonic attenuation measurements also provide information about microstructural changes such as increased porosity due to evolution of gaseous decomposition products.

  12. 4D phase-space multiplexing for fluorescent microscopy

    NASA Astrophysics Data System (ADS)

    Liu, Hsiou-Yuan; Zhong, Jingshan; Waller, Laura

    2016-03-01

    Phase-space measurements enable characterization of second-order spatial coherence properties and can be used for digital aberration removal or 3D position reconstruction. Previous methods use a scanning aperture to measure the phase space spectrogram, which is slow and light inefficient, while also attenuating information about higher-order correlations. We demonstrate a significant improvement of speed and light throughput by incorporating multiplexing techniques into our phase-space imaging system. The scheme implements 2D coded aperture patterning in the Fourier (pupil) plane of a microscope using a Spatial Light Modulator (SLM), while capturing multiple intensity images in real space. We compare various multiplexing schemes to scanning apertures and show that our phase-space reconstructions are accurate for experimental data with biological samples containing many 3D fluorophores.

  13. Phase, amplitude, and polarization microscopy with a sampling field sensor.

    PubMed

    Tumbar, Remy

    2008-07-01

    I describe an improved implementation of a previously reported interferometric device, the sampling field sensor (SFS) [Appl. Opt.47, B32-B43 (2008)]. It provides X, Y, and XY shearing interferometric information simultaneously (space multiplexed) with amplitude and polarization information while using time-multiplexed phase shifting. Its simple common-path configuration makes it compact and vibration insensitive, as demonstrated by the ~lambda/125 phase estimation repeatability that was below the coherent noise floor (estimated at ~lambda/50). The SFS may be viewed as an efficient, robust and accurate full-field optical-digital interface, easy to integrate with traditional imaging systems. This is demonstrated by using the sensor as the focal plane array of a transmitted-light microscope in a straightforward setup using an illumination path polarization phase shifter. This work is focused on a qualitative demonstration and presents phase, amplitude, and polarization images of different types of human cheek cells and Caenorhabditiselegans larvae.

  14. Quantitative phase microscopy: a new tool for investigating the structure and function of unstained live cells.

    PubMed

    Curl, Claire L; Bellair, Catherine J; Harris, Peter J; Allman, Brendan E; Roberts, Ann; Nugent, Keith A; Delbridge, Lea M D

    2004-12-01

    1. The optical transparency of unstained live cell specimens limits the extent to which information can be recovered from bright-field microscopic images because these specimens generally lack visible amplitude-modulating components. However, visualization of the phase modulation that occurs when light traverses these specimens can provide additional information. 2. Optical phase microscopy and derivatives of this technique, such as differential interference contrast (DIC) and Hoffman modulation contrast (HMC), have been used widely in the study of cellular materials. With these techniques, enhanced contrast is achieved, which is useful in viewing specimens, but does not allow quantitative information to be extracted from the phase content available in the images. 3. An innovative computational approach to phase microscopy, which provides mathematically derived information about specimen phase-modulating characteristics, has been described recently. Known as quantitative phase microscopy (QPM), this method derives quantitative phase measurements from images captured using a bright-field microscope without phase- or interference-contrast optics. 4. The phase map generated from the bright-field images by the QPM method can be used to emulate other contrast image modes (including DIC and HMC) for qualitative viewing. Quantitative phase microscopy achieves improved discrimination of cellular detail, which permits more rigorous image analysis procedures to be undertaken compared with conventional optical methods. 5. The phase map contains information about cell thickness and refractive index and can allow quantification of cellular morphology under experimental conditions. As an example, the proliferative properties of smooth muscle cells have been evaluated using QPM to track growth and confluency of cell cultures. Quantitative phase microscopy has also been used to investigate erythrocyte cell volume and morphology in different osmotic environments. 6. Quantitative

  15. Artifact characterization and reduction in scanning X-ray Zernike phase contrast microscopy.

    PubMed

    Vartiainen, Ismo; Holzner, Christian; Mohacsi, Istvan; Karvinen, Petri; Diaz, Ana; Pigino, Gaia; David, Christian

    2015-05-18

    Zernike phase contrast microscopy is a well-established method for imaging specimens with low absorption contrast. It has been successfully implemented in full-field microscopy using visible light and X-rays. In microscopy Cowley's reciprocity principle connects scanning and full-field imaging. Even though the reciprocity in Zernike phase contrast has been discussed by several authors over the past thirty years, only recently it was experimentally verified using scanning X-ray microscopy. In this paper, we investigate the image and contrast formation in scanning Zernike phase contrast microscopy with a particular and detailed focus on the origin of imaging artifacts that are typically associated with Zernike phase contrast. We demonstrate experimentally with X-rays the effect of the phase mask design on the contrast and halo artifacts and present an optimized design of the phase mask with respect to photon efficiency and artifact reduction. Similarly, due to the principle of reciprocity the observations and conclusions of this work have direct applicability to Zernike phase contrast in full-field microscopy as well.

  16. Thermomorphic phase separation in ionic liquid-organic liquid systems--conductivity and spectroscopic characterization.

    PubMed

    Riisager, Anders; Fehrmann, Rasmus; Berg, Rolf W; van Hal, Roy; Wasserscheid, Peter

    2005-08-21

    Electrical conductivity, FT-Raman and NMR measurements are demonstrated as useful tools to probe and determine phase behavior of thermomorphic ionic liquid-organic liquid systems. To illustrate the methods, consecutive conductivity measurements of a thermomorphic methoxyethoxyethyl-imidazolium ionic liquid/1-hexanol system are performed in the temperature interval 25-80 degrees C using a specially constructed double-electrode cell. In addition, FT-Raman and 1H-NMR spectroscopic studies performed on the phase-separable system in the same temperature interval confirm the mutual solubility of the components in the system, the liquid-liquid equilibrium phase diagram of the binary mixture, and signify the importance of hydrogen bonding between the ionic liquid and the hydroxyl group of the alcohol.

  17. Spectroscopic and Structural Investigations of alpha-beta-, and gamma-AIH3 Phases

    SciTech Connect

    Manciu, F.S.; Graetz, J.; Reza, L.; Durrer, W.G.; Bronson, A.; Lacina, D.

    2010-07-01

    With its reputation as a high-energy density fuel, aluminum hydride (AlH{sub 3}) has received renewed attention as a material that is particularly suitable, not only for hydrogen storage but also for rocket propulsion. While the various phases of AlH{sub 3} have been investigated theoretically, there is a shortage of experimental studies corroborating the theoretical findings. In response to this, we present here an investigation of these compounds based primarily on two research areas in which there is the greatest scarcity of information in the literature, namely Raman and infrared (IR) absorption analysis. To the authors knowledge, this is the first report of experimental far-IR absorption results on these compounds. Two different samples prepared by broadly similar ethereal reactions of AlCl{sub 3} with LiAlH{sub 4} were analyzed. Both Raman and IR absorption measurements indicate that one sample is purely {gamma}-AlH{sub 3} and that the other is a mixture of {alpha}-, {beta}-, and {gamma}-AlH{sub 3} phases. X-ray diffraction confirms the spectroscopic findings, most notably for the {beta}-AlH{sub 3} phase, for which optical spectroscopic data are reported here for the first time.

  18. Simultaneous microscopic measurements of thermal and spectroscopic fields of a phase change material

    NASA Astrophysics Data System (ADS)

    Romano, M.; Ryu, M.; Morikawa, J.; Batsale, J. C.; Pradere, C.

    2016-05-01

    In this paper, simultaneous microscopic measurements of thermal and spectroscopic fields of a paraffin wax n-alkane phase change material are reported. Measurements collected using an original set-up are presented and discussed with emphasis on the ability to perform simultaneous characterization of the system when the proposed imaging process is used. Finally, this work reveals that the infrared wavelength contains two sets of important information. Furthermore, this versatile and flexible technique is well adapted to characterize many systems in which the mass and heat transfers effects are coupled.

  19. Spectroscopic evidence of α,α-dimethylbenzyl radicals in the gas phase

    NASA Astrophysics Data System (ADS)

    Chae, Sang Youl; Lim, Manho; Lee, Sang Kuk

    2016-11-01

    We report the spectroscopic evidence of the jet-cooled α,α-dimethylbenzyl radical, which was generated in a technique of corona excited supersonic expansion using a pinhole-type glass nozzle. For identification of the α,α-dimethylbenzyl radical, precursors that form a common molecular radical, as a result of dissociation of either a Csbnd H bond or Csbnd C bond of alkyl group in a corona discharge, were employed. Based on comparisons of the spectra observed from the corona discharges of isopropylbenzene and tert-butylbenzene, evidence was obtained of the existence of the α,α-dimethylbenzyl radical in the gas phase.

  20. Compact diffraction phase microscopy for quantitative visualization of cells in biomedical applications

    NASA Astrophysics Data System (ADS)

    Talaikova, N. A.; Ryabukho, V. P.

    2016-08-01

    We consider a simplified and compact scheme of interference phase microscopy using a diffraction grating and spatial filtering of the diffracted field, i.e., diffraction phase microscopy. The scheme and the parameters of the device with the possibility of using the optical system of a smartphone and its software are analysed. The results of experimental determination of the spatial structure parameters of erythrocytes are presented.

  1. Enlightening intracellular complexity of living cells with quantitative phase microscopy

    NASA Astrophysics Data System (ADS)

    Martinez Torres, C.; Laperrousaz, B.; Berguiga, L.; Boyer Provera, E.; Elezgaray, J.; Nicolini, F. E.; Maguer-Satta, V.; Arneodo, A.; Argoul, F.

    2016-03-01

    The internal distribution of refractive indices (RIs) of a living cell is much more complex than usually admitted in multi-shell models. The reconstruction of RI maps from single phase images has rarely been achieved for several reasons: (i) we still have very little knowledge of the impact of internal macromolecular complexes on the local RI and (ii) phase changes produced by light propagation through the sample are mixed with diffraction effects by internal cell bodies. We propose the implementation a 2D wavelet-based contour chain detection method to distinguish internal boundaries thanks to their greatest optical path difference gradients. These contour chains correspond to the highest image phase contrast and follow the local RI inhomogeneities linked to the intracellular structural intricacy. Their statistics and spatial distribution are morphological indicators for distinguishing cells of different origins and to follow their transformation in pathologic situations. We use this method to compare non adherent blood cells from primary and laboratory culture origins, in healthy and pathological situations (chronic myelogenous leukaemia). In a second part of this presentation, we concentrate on the temporal dynamics of the phase contour chains and we discuss the spectral decomposition of their dynamics in both health and disease.

  2. Quantitative phase imaging of human red blood cells using phase-shifting white light interference microscopy with colour fringe analysis

    NASA Astrophysics Data System (ADS)

    Singh Mehta, Dalip; Srivastava, Vishal

    2012-11-01

    We report quantitative phase imaging of human red blood cells (RBCs) using phase-shifting interference microscopy. Five phase-shifted white light interferograms are recorded using colour charge coupled device camera. White light interferograms were decomposed into red, green, and blue colour components. The phase-shifted interferograms of each colour were then processed by phase-shifting analysis and phase maps for red, green, and blue colours were reconstructed. Wavelength dependent refractive index profiles of RBCs were computed from the single set of white light interferogram. The present technique has great potential for non-invasive determination of refractive index variation and morphological features of cells and tissues.

  3. Spectroscopic Studies of Cyclopentane under Extreme Conditions and Implications for the P-T Phase Diagram

    NASA Astrophysics Data System (ADS)

    Ma, Chunli; Liu, Zhenxian; Cui, Qiliang; Hemley, Russell

    2013-06-01

    Infrared (IR) spectroscopy and Raman scattering combined with diamond anvil cell (DAC) and cryogenic techniques have been employed to investigate cyclopentane up to 4 GPa in the temperature range of 100-350 K and isothermal compression up to 84 GPa at room temperature. Four phases including liquid, plastic phases I and II, and truly crystalline phase III are clearly identified in the P-T range studied based on the changes of the ring breathing mode and CH2 rocking modes. The phase diagram is extended to the pressure and temperature range of 0 - 4.0 GPa and 105 - 350 K. Further compression at room temperature up to 84 GPa, another high-pressure phase (IV) is observed based on the appearance of low frequency peaks related to the lattice vibrational modes in the synchrotron far-IR spectra. The spectroscopic results indicate that cyclopentane persists the orientation ordered crystalline phase up to 84 GPa at room temperature. This work was supported by NSF (DMR-0805056; EAR 06-49658, COMPRES) and DOE/ NNSA (DE-FC03-03N00144, CDAC). NSLS is supported by the DOE/BES (DE-AC02-98CH10886).

  4. Time Resolved Phase Transitions via Dynamic Transmission Electron Microscopy

    SciTech Connect

    Reed, B W; Armstrong, M R; Blobaum, K J; Browning, N D; Burnham, A K; Campbell, G H; Gee, R; Kim, J S; King, W E; Maiti, A; Piggott, W T; Torralva, B R

    2007-02-22

    The Dynamic Transmission Electron Microscope (DTEM) project is developing an in situ electron microscope with nanometer- and nanosecond-scale resolution for the study of rapid laser-driven processes in materials. We report on the results obtained in a year-long LDRD-supported effort to develop DTEM techniques and results for phase transitions in molecular crystals, reactive multilayer foils, and melting and resolidification of bismuth. We report the first in situ TEM observation of the HMX {beta}-{delta} phase transformation in sub-{micro}m crystals, computational results suggesting the importance of voids and free surfaces in the HMX transformation kinetics, and the first electron diffraction patterns of intermediate states in fast multilayer foil reactions. This project developed techniques which are applicable to many materials systems and will continue to be employed within the larger DTEM effort.

  5. A phase space model of Fourier ptychographic microscopy

    PubMed Central

    Horstmeyer, Roarke; Yang, Changhuei

    2014-01-01

    A new computational imaging technique, termed Fourier ptychographic microscopy (FPM), uses a sequence of low-resolution images captured under varied illumination to iteratively converge upon a high-resolution complex sample estimate. Here, we propose a mathematical model of FPM that explicitly connects its operation to conventional ptychography, a common procedure applied to electron and X-ray diffractive imaging. Our mathematical framework demonstrates that under ideal illumination conditions, conventional ptychography and FPM both produce datasets that are mathematically linked by a linear transformation. We hope this finding encourages the future cross-pollination of ideas between two otherwise unconnected experimental imaging procedures. In addition, the coherence state of the illumination source used by each imaging platform is critical to successful operation, yet currently not well understood. We apply our mathematical framework to demonstrate that partial coherence uniquely alters both conventional ptychography’s and FPM’s captured data, but up to a certain threshold can still lead to accurate resolution-enhanced imaging through appropriate computational post-processing. We verify this theoretical finding through simulation and experiment. PMID:24514995

  6. Two-step phase-shifting fluorescence incoherent holographic microscopy

    PubMed Central

    Qin, Wan; Yang, Xiaoqi; Li, Yingying; Peng, Xiang; Yao, Hai; Qu, Xinghua; Gao, Bruce Z.

    2014-01-01

    Abstract. Fluorescence holographic microscope (FINCHSCOPE) is a motionless fluorescence holographic imaging technique based on Fresnel incoherent correlation holography (FINCH) that shows promise in reconstructing three-dimensional fluorescence images of biological specimens with three holograms. We report a developing two-step phase-shifting method that reduces the required number of holograms from three to two. Using this method, we resolved microscopic fluorescent beads that were three-dimensionally distributed at different depths with two interferograms captured by a CCD camera. The method enables the FINCHSCOPE to work in conjunction with the frame-straddling technique and significantly enhance imaging speed. PMID:24972355

  7. Two-step phase-shifting fluorescence incoherent holographic microscopy.

    PubMed

    Qin, Wan; Yang, Xiaoqi; Li, Yingying; Peng, Xiang; Yao, Hai; Qu, Xinghua; Gao, Bruce Z

    2014-06-01

    Fluorescence holographic microscope (FINCHSCOPE) is a motionless fluorescence holographic imaging technique based on Fresnel incoherent correlation holography (FINCH) that shows promise in reconstructing three-dimensional fluorescence images of biological specimens with three holograms. We report a developing two-step phase-shifting method that reduces the required number of holograms from three to two. Using this method, we resolved microscopic fluorescent beads that were three-dimensionally distributed at different depths with two interferograms captured by a CCD camera. The method enables the FINCHSCOPE to work in conjunction with the frame-straddling technique and significantly enhance imaging speed.

  8. Phase contrast hard x-ray microscopy with submicron resolution

    SciTech Connect

    Lagomarsino, S.; Cedola, A.; Cloetens, P.; Di Fonzo, S.; Jark, W.; Soullie, G.; Riekel, C.

    1997-11-01

    In this letter we present a hard x-ray phase contrast microscope based on the divergent and coherent beam exiting an x-ray waveguide. It uses lensless geometrical projection to magnify spatial variations in optical path length more than 700 times. Images of a nylon fiber and a gold test pattern were obtained with a resolution of 0.14 {mu}m in one direction. Exposure times as short as 0.1 s gave already visible contrast, opening the way to high resolution, real time studies. {copyright} {ital 1997 American Institute of Physics.}

  9. Understanding the Phase Contrast Optics to Restore Artifact-free Microscopy Images for Segmentation

    PubMed Central

    Yin, Zhaozheng; Kanade, Takeo; Chen, Mei

    2012-01-01

    Phase contrast, a noninvasive microscopy imaging technique, is widely used to capture time-lapse images to monitor the behavior of transparent cells without staining or altering them. Due to the optical principle, phase contrast microscopy images contain artifacts such as the halo and shade-off that hinder image segmentation, a critical step in automated microscopy image analysis. Rather than treating phase contrast microscopy images as general natural images and applying generic image processing techniques on them, we propose to study the optical properties of the phase contrast microscope to model its image formation process. The phase contrast imaging system can be approximated by a linear imaging model. Based on this model and input image properties, we formulate a regularized quadratic cost function to restore artifact-free phase contrast images that directly correspond to the specimen's optical path length. With artifacts removed, high quality segmentation can be achieved by simply thresholding the restored images. The imaging model and restoration method are quantitatively evaluated on microscopy image sequences with thousands of cells captured over several days. We also demonstrate that accurate restoration lays the foundation for high performance in cell detection and tracking. PMID:22386070

  10. Parallel on-axis holographic phase microscopy of biological cells and unicellular microorganism dynamics.

    PubMed

    Shaked, Natan T; Newpher, Thomas M; Ehlers, Michael D; Wax, Adam

    2010-05-20

    We apply a wide-field quantitative phase microscopy technique based on parallel two-step phase-shifting on-axis interferometry to visualize live biological cells and microorganism dynamics. The parallel on-axis holographic approach is more efficient with camera spatial bandwidth consumption compared to previous off-axis approaches and thus can capture finer sample spatial details, given a limited spatial bandwidth of a specific digital camera. Additionally, due to the parallel acquisition mechanism, the approach is suitable for visualizing rapid dynamic processes, permitting an interferometric acquisition rate equal to the camera frame rate. The method is demonstrated experimentally through phase microscopy of neurons and unicellular microorganisms.

  11. Phase reconstruction in annular bright-field scanning transmission electron microscopy.

    PubMed

    Ishida, Takafumi; Kawasaki, Tadahiro; Tanji, Takayoshi; Kodama, Tetsuji; Matsutani, Takaomi; Ogai, Keiko; Ikuta, Takashi

    2015-04-01

    A novel technique for reconstructing the phase shifts of electron waves was applied to Cs-corrected scanning transmission electron microscopy (STEM). To realize this method, a new STEM system equipped with an annular aperture, annularly arrayed detectors and an arrayed image processor has been developed and evaluated in experiments. We show a reconstructed phase image of graphite particles and demonstrate that this new method works effectively for high-resolution phase imaging. © The Author 2014. Published by Oxford University Press on behalf of The Japanese Society of Microscopy. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

  12. Controlling stimulated coherent spectroscopy and microscopy by a position-dependent phase

    NASA Astrophysics Data System (ADS)

    Chung, Chao-Yu; Hsu, Julie; Mukamel, Shaul; Potma, Eric O.

    2013-03-01

    We study the role of geometry-dependent phase shifts of the optical electric field in stimulated coherent spectroscopy, a special class of heterodyne optical spectroscopy techniques. We generalize the theoretical description of stimulated spectroscopy to include spatial phase effects, and study the measured material response for several representative excitation and detection configurations. Using stimulated Raman scattering microscopy as an example, we show that different components of the material response are measured by varying the position of the object in focus. We discuss the implications of the position-dependent phase in stimulated coherent microscopy and point out a detection configuration in which its effects are minimized.

  13. Quantitative interferometric microscopy with improved full-field phase aberration compensation

    NASA Astrophysics Data System (ADS)

    Xue, Liang; Wang, Shouyu; Yan, Keding; Sun, Nan; Li, Zhenhua; Liu, Fei

    2014-11-01

    Single-shot quantitative interferometric microscopy (QIM) needs a high-accuracy and rapid phase retrieval algorithm. Retrieved phase distributions are often influenced by phase aberration background caused by both imaging system and phase retrieval algorithms. Here, we propose an improved phase aberration compensation (PAC) approach in order to eliminate the phase aberrations inherent in the data. With this method, sample-free parts are identified and used to calculate the background phase, reducing phase errors induced in samples and providing high-quality phase images. We now demonstrate that QIM based on this PAC approach realizes high-quality phase imaging from a single interferogram. This is of great potential for a real-time speedy diagnosis.

  14. Unstained viable cell recognition in phase-contrast microscopy

    NASA Astrophysics Data System (ADS)

    Skoczylas, M.; Rakowski, W.; Cherubini, R.; Gerardi, S.

    2011-09-01

    Individual cell recognition is a relevant task to be accomplished when single-ion microbeam irradiations are performed. At INFN-LNL facility cell visualization system is based on a phase-contrast optical microscope, without the use of any cell dye. Unstained cells are seeded in the special designed Petri dish, between two mylar foils, and at present the cell recognition is achieved manually by an expert operator. Nevertheless, this procedure is time consuming and sometimes it could be not practical if the amount of living cells to be irradiated is large. To reduce the time needed to recognize unstained cells on the Petri dish, it has been designed and implemented an automated, parallel algorithm. Overlapping ROIs sliding in steps over the captured grayscale image are firstly pre-classified and potential cell markers for the segmentation are obtained. Segmented objects are additionally classified to categorize cell bodies from other structures considered as sample dirt or background. As a result, cell coordinates are passed to the dedicated CELLView program that controls all the LNL single-ion microbeam irradiation protocol, including the positioning of individual cells in front of the ion beam. Unstained cell recognition system was successfully tested in experimental conditions with two different mylar surfaces. The recognition time and accuracy was acceptable, however, improvements in speed would be useful.

  15. Spectroscopic Identification of p-CHLORO-α-METHYLBENZYL Radical in the Gas Phase

    NASA Astrophysics Data System (ADS)

    Lee, Seung Woon; Lee, Gi Woo; Lee, Sang Kuk

    2009-06-01

    We report the first spectroscopic identification of the p-fluoro-α-methylbenzyl radical in the gas phase. Precursor p-fluoro-ethylbenzene seeded in a large amount of inert carrier gas helium was electrically discharged to produce the benzyl-type radicals in a corona excited supersonic expansion using a pinhole-type glass nozzle, from which the vibronic emission spectrum was recorded in the visible region using a long path monochromator. From an analysis of the spectrum observed, we found the formation of p-fluoro-α-methylbenzyl radical as well as p-fluorobenzyl radical in the jet from the precursor. After eliminating the bands belonging to p-fluorobenzyl radical using the known data, we identified spectroscopically the formation of the p-fluoro-α-methylbenzyl radical, in which the energy of the D_1 → D_0 electronic transition and a few vibrational mode frequencies in the ground electronic state were determined by comparison with those from an ab initio calculation and with those from the known data of the precursor. S. K. Lee and D. Y. Baek Chem. Phys. Lett. 301(3-4), 407-412 (1999).

  16. Phase-difference and spectroscopic imaging for monitoring of human brain temperature during cooling.

    PubMed

    Weis, Jan; Covaciu, Lucian; Rubertsson, Sten; Allers, Mats; Lunderquist, Anders; Ortiz-Nieto, Francisco; Ahlström, Håkan

    2012-12-01

    Decrease of the human brain temperature was induced by intranasal cooling. The main purpose of this study was to compare the two magnetic resonance methods for monitoring brain temperature changes during cooling: phase-difference and magnetic resonance spectroscopic imaging (MRSI) with high spatial resolution. Ten healthy volunteers were measured. Selective brain cooling was performed through nasal cavities using saline-cooled balloon catheters. MRSI was based on a radiofrequency spoiled gradient echo sequence. The spectral information was encoded by incrementing the echo time of the subsequent eight image records. Reconstructed voxel size was 1×1×5 mm(3). Relative brain temperature was computed from the positions of water spectral lines. Phase maps were obtained from the first image record of the MRSI sequence. Mild hypothermia was achieved in 15-20 min. Mean brain temperature reduction varied in the interval <-3.0; -0.6>°C and <-2.7; -0.7>°C as measured by the MRSI and phase-difference methods, respectively. Very good correlation was found in all locations between the temperatures measured by both techniques except in the frontal lobe. Measurements in the transversal slices were more robust to the movement artifacts than those in the sagittal planes. Good agreement was found between the MRSI and phase-difference techniques. Copyright © 2012 Elsevier Inc. All rights reserved.

  17. Combined confocal Raman and quantitative phase microscopy system for biomedical diagnosis.

    PubMed

    Kang, Jeon Woong; Lue, Niyom; Kong, Chae-Ryon; Barman, Ishan; Dingari, Narahara C; Goldfless, Stephen J; Niles, Jacquin C; Dasari, Ramachandra R; Feld, Michael S

    2011-09-01

    We have developed a novel multimodal microscopy system that incorporates confocal Raman, confocal reflectance, and quantitative phase microscopy (QPM) into a single imaging entity. Confocal Raman microscopy provides detailed chemical information from the sample, while confocal reflectance and quantitative phase microscopy show detailed morphology. Combining these intrinsic contrast imaging modalities makes it possible to obtain quantitative morphological and chemical information without exogenous staining. For validation and characterization, we have used this multi-modal system to investigate healthy and diseased blood samples. We first show that the thickness of a healthy red blood cell (RBC) shows good correlation with its hemoglobin distribution. Further, in malaria infected RBCs, we successfully image the distribution of hemozoin (malaria pigment) inside the cell. Our observations lead us to propose morphological screening by QPM and subsequent chemical imaging by Raman for investigating blood disorders. This new approach allows monitoring cell development and cell-drug interactions with minimal perturbation of the biological system of interest.

  18. Automatic full compensation of quantitative phase imaging in off-axis digital holographic microscopy.

    PubMed

    Trujillo, Carlos; Castañeda, Raúl; Piedrahita-Quintero, Pablo; Garcia-Sucerquia, Jorge

    2016-12-20

    An automatic method that fully compensates the quantitative phase measurements in off-axis digital holographic microscopy (DHM) is presented. The two main perturbations of the quantitative phase measurements in off-axis DHM are automatically removed. While the curvature phase flaw introduced by the microscope objective is avoided by the use of an optimized telecentric imaging system for the recording of the holograms, the remaining phase perturbation due to the tilt of the reference wave is removed by the automatic computation of a digital compensating reference wave. The method has been tested on both nonbiological and biological samples with and improving on the quality of the recovered phase maps.

  19. Quantitative interferometric microscopy with two dimensional Hilbert transform based phase retrieval method

    NASA Astrophysics Data System (ADS)

    Wang, Shouyu; Yan, Keding; Xue, Liang

    2017-01-01

    In order to obtain high contrast images and detailed descriptions of label free samples, quantitative interferometric microscopy combining with phase retrieval is designed to obtain sample phase distributions from fringes. As accuracy and efficiency of recovered phases are affected by phase retrieval methods, thus approaches owning higher precision and faster processing speed are still in demand. Here, two dimensional Hilbert transform based phase retrieval method is adopted in cellular phase imaging, it not only reserves more sample specifics compared to classical fast Fourier transform based method, but also overcomes disadvantages of traditional algorithm according to Hilbert transform which is a one dimensional processing causing phase ambiguities. Both simulations and experiments are provided, proving the proposed phase retrieval approach can acquire quantitative sample phases with high accuracy and fast speed.

  20. Single-exposure quantitative phase imaging in color-coded LED microscopy (Conference Presentation)

    NASA Astrophysics Data System (ADS)

    Lee, Wonchan; Jung, Daeseong; Joo, Chulmin

    2017-02-01

    Quantitative phase-gradient or phase imaging in LED microscopy has been recently demonstrated. The methods enable measurement of phase distribution of transparent specimens in a simple and cost-effective manner, but require multiple image acquisitions with different source or pupil configurations to improve phase accuracy. Here, we demonstrate a strategy for single-shot quantitative phase imaging in color-coded LED microscopy. We employ a circular LED illumination pattern that is trisected into subregions with equal area, assigned to red, green and blue colors, respectively. Additional color filter is also employed to mitigate the color leakage of light into different color channels of the image sensor. Image acquisition with a color image sensor and subsequent computation based on the weak object transfer function allow for quantitative amplitude and phase measurements of a specimen. We describe computational model and single-shot quantitative phase imaging capability of our method by presenting phase images of calibrated phase sample and dynamics of cells. Phase measurement accuracy is validated with pre-characterized phase plate, and single-shot phase imaging capability is demonstrated with time-lapse imaging of cells acquired at 30 Hz.

  1. Correlative Energy-Dispersive X-Ray Spectroscopic Tomography and Atom Probe Tomography of the Phase Separation in an Alnico 8 Alloy

    SciTech Connect

    Guo, Wei; Sneed, Brian T.; Zhou, Lin; Tang, Wei; Kramer, Matthew J.; Cullen, David A.; Poplawsky, Jonathan D.

    2016-12-21

    Alnico alloys have long been used as strong permanent magnets because of their ferromagnetism and high coercivity. Understanding their structural details allows for better prediction of the resulting magnetic properties. However, quantitative three-dimensional characterization of the phase separation in these alloys is still challenged by the spatial quantification of nanoscale phases. Herein, we apply a dual tomography approach, where correlative scanning transmission electron microscopy (STEM) energy-dispersive X-ray spectroscopic (EDS) tomography and atom probe tomography (APT) are used to investigate the initial phase separation process of an alnico 8 alloy upon non-magnetic annealing. STEM-EDS tomography provides information on the morphology and volume fractions of Fe–Co-rich and Νi–Al-rich phases after spinodal decomposition in addition to quantitative information of the composition of a nanoscale volume. Subsequent analysis of a portion of the same specimen by APT offers quantitative chemical information of each phase at the sub-nanometer scale. Furthermore, APT reveals small, 2–4 nm Fe-rich α1 phases that are nucleated in the Ni-rich α2 matrix. From this information, we show that phase separation of the alnico 8 alloy consists of both spinodal decomposition and nucleation and growth processes. Lastly, we discuss the complementary benefits and challenges associated with correlative STEM-EDS and APT.

  2. Correlative Energy-Dispersive X-Ray Spectroscopic Tomography and Atom Probe Tomography of the Phase Separation in an Alnico 8 Alloy

    DOE PAGES

    Guo, Wei; Sneed, Brian T.; Zhou, Lin; ...

    2016-12-21

    Alnico alloys have long been used as strong permanent magnets because of their ferromagnetism and high coercivity. Understanding their structural details allows for better prediction of the resulting magnetic properties. However, quantitative three-dimensional characterization of the phase separation in these alloys is still challenged by the spatial quantification of nanoscale phases. Herein, we apply a dual tomography approach, where correlative scanning transmission electron microscopy (STEM) energy-dispersive X-ray spectroscopic (EDS) tomography and atom probe tomography (APT) are used to investigate the initial phase separation process of an alnico 8 alloy upon non-magnetic annealing. STEM-EDS tomography provides information on the morphology andmore » volume fractions of Fe–Co-rich and Νi–Al-rich phases after spinodal decomposition in addition to quantitative information of the composition of a nanoscale volume. Subsequent analysis of a portion of the same specimen by APT offers quantitative chemical information of each phase at the sub-nanometer scale. Furthermore, APT reveals small, 2–4 nm Fe-rich α1 phases that are nucleated in the Ni-rich α2 matrix. From this information, we show that phase separation of the alnico 8 alloy consists of both spinodal decomposition and nucleation and growth processes. Lastly, we discuss the complementary benefits and challenges associated with correlative STEM-EDS and APT.« less

  3. Spectroscopic-ellipsometric study of native oxide removal by liquid phase HF process

    NASA Astrophysics Data System (ADS)

    Kurhekar, Anil Sudhakar; Apte, Prakash R.

    2013-02-01

    Ex situ spectroscopic ellipsometry (SE) measurements have been employed to investigate the effect of liquid-phase hydrofluoric acid (HF) cleaning on Si<100> surfaces for microelectromechanical systems application. The hydrogen terminated (H-terminated) Si surface was realized as an equivalent dielectric layer, and SE measurements are performed. The SE analyses indicate that after a 20-s 100:5 HF dip with rinse, the Si (100) surface was passivated by the hydrogen termination and remained chemically stable. Roughness of the HF-etched bare Si (100) surface was observed and analyzed by the ex-situ SE. Evidence for desorption of the H-terminated Si surface layer is studied using Fourier transform infrared spectroscopy and ellipsometry, and discussed. This piece of work explains the usage of an ex situ, non-destructive technique capable of showing state of passivation, the H-termination of Si<100> surfaces.

  4. Spectroscopic-ellipsometric study of native oxide removal by liquid phase HF process

    PubMed Central

    Kurhekar, Anil Sudhakar; Apte, Prakash R

    2014-01-01

    Ex situ spectroscopic ellipsometry (SE) measurements have been employed to investigate the effect of liquid-phase hydrofluoric acid (HF) cleaning on Si<100> surfaces for microelectromechanical systems application. The hydrogen terminated (H-terminated) Si surface was realized as an equivalent dielectric layer, and SE measurements are performed. The SE analyses indicate that after a 20-s 100:5 HF dip with rinse, the Si (100) surface was passivated by the hydrogen termination and remained chemically stable. Roughness of the HF-etched bare Si (100) surface was observed and analyzed by the ex-situ SE. Evidence for desorption of the H-terminated Si surface layer is studied using Fourier transform infrared spectroscopy and ellipsometry, and discussed. This piece of work explains the usage of an ex situ, non-destructive technique capable of showing state of passivation, the H-termination of Si<100> surfaces. PMID:24619506

  5. Multimodal imaging of human cerebellum - merging X-ray phase microtomography, magnetic resonance microscopy and histology

    PubMed Central

    Schulz, Georg; Waschkies, Conny; Pfeiffer, Franz; Zanette, Irene; Weitkamp, Timm; David, Christian; Müller, Bert

    2012-01-01

    Imaging modalities including magnetic resonance imaging and X-ray computed tomography are established methods in daily clinical diagnosis of human brain. Clinical equipment does not provide sufficient spatial resolution to obtain morphological information on the cellular level, essential for applying minimally or non-invasive surgical interventions. Therefore, generic data with lateral sub-micrometer resolution have been generated from histological slices post mortem. Sub-cellular spatial resolution, lost in the third dimension as a result of sectioning, is obtained using magnetic resonance microscopy and micro computed tomography. We demonstrate that for human cerebellum grating-based X-ray phase tomography shows complementary contrast to magnetic resonance microscopy and histology. In this study, the contrast-to-noise values of magnetic resonance microscopy and phase tomography were comparable whereas the spatial resolution in phase tomography is an order of magnitude better. The registered data with their complementary information permit the distinct segmentation of tissues within the human cerebellum. PMID:23145319

  6. Multimodal imaging of human cerebellum - merging X-ray phase microtomography, magnetic resonance microscopy and histology

    NASA Astrophysics Data System (ADS)

    Schulz, Georg; Waschkies, Conny; Pfeiffer, Franz; Zanette, Irene; Weitkamp, Timm; David, Christian; Müller, Bert

    2012-11-01

    Imaging modalities including magnetic resonance imaging and X-ray computed tomography are established methods in daily clinical diagnosis of human brain. Clinical equipment does not provide sufficient spatial resolution to obtain morphological information on the cellular level, essential for applying minimally or non-invasive surgical interventions. Therefore, generic data with lateral sub-micrometer resolution have been generated from histological slices post mortem. Sub-cellular spatial resolution, lost in the third dimension as a result of sectioning, is obtained using magnetic resonance microscopy and micro computed tomography. We demonstrate that for human cerebellum grating-based X-ray phase tomography shows complementary contrast to magnetic resonance microscopy and histology. In this study, the contrast-to-noise values of magnetic resonance microscopy and phase tomography were comparable whereas the spatial resolution in phase tomography is an order of magnitude better. The registered data with their complementary information permit the distinct segmentation of tissues within the human cerebellum.

  7. Direct Imaging of Phase Objects Enables Conventional Deconvolution in Bright Field Light Microscopy

    PubMed Central

    Hernández Candia, Carmen Noemí; Gutiérrez-Medina, Braulio

    2014-01-01

    In transmitted optical microscopy, absorption structure and phase structure of the specimen determine the three-dimensional intensity distribution of the image. The elementary impulse responses of the bright field microscope therefore consist of separate absorptive and phase components, precluding general application of linear, conventional deconvolution processing methods to improve image contrast and resolution. However, conventional deconvolution can be applied in the case of pure phase (or pure absorptive) objects if the corresponding phase (or absorptive) impulse responses of the microscope are known. In this work, we present direct measurements of the phase point- and line-spread functions of a high-aperture microscope operating in transmitted bright field. Polystyrene nanoparticles and microtubules (biological polymer filaments) serve as the pure phase point and line objects, respectively, that are imaged with high contrast and low noise using standard microscopy plus digital image processing. Our experimental results agree with a proposed model for the response functions, and confirm previous theoretical predictions. Finally, we use the measured phase point-spread function to apply conventional deconvolution on the bright field images of living, unstained bacteria, resulting in improved definition of cell boundaries and sub-cellular features. These developments demonstrate practical application of standard restoration methods to improve imaging of phase objects such as cells in transmitted light microscopy. PMID:24558478

  8. Infrared spectroscopic studies of myeloid leukemia (ML-1) cells at different phases of the cell cycle

    NASA Astrophysics Data System (ADS)

    Boydston-White, Susie; Diem, Max

    1999-06-01

    Advances in infrared spectroscopic methodology permit excellent infrared spectra to be collected from objects as small as single human cells. These advances have lead to an increased interest of the use of infrared spectroscopy as a medical diagnostic tool. Infrared spectra of myeloid leukemia (ML-1) cells are reported for cells derived from an asynchronous, exponentially-growing culture, as well as for cells that were fractionated according to their stage within the cell division cycle. The observed results suggest that the cells' DNA is detectable by infrared spectroscopy mainly when the cell is in the S phase, during the replication of DNA. In the G1 and G2 phases, the DNA is so tightly packed in the nucleus that it appears opaque to infrared radiation. Consequently, the nucleic acid spectral contributions in the G1 and G2 phases would be mostly that of cytoplasmic RNA. These results suggest that infrared spectral changes observed earlier between normal and abnormal cells may have been due to different distributions of cells within the stages of the cell division cycle.

  9. Irreversible reactions studied with nanosecond transmission electron microscopy movies: Laser crystallization of phase change materials

    NASA Astrophysics Data System (ADS)

    Santala, M. K.; Reed, B. W.; Raoux, S.; Topuria, T.; LaGrange, T.; Campbell, G. H.

    2013-04-01

    We use multi-frame, nanosecond-scale photo-emission transmission electron microscopy to create movies of irreversible reactions that occur too rapidly to capture with conventional microscopy. The technique is applied to the crystallization of phase change materials used for optical and resistive memory. For those applications, laser- or current-induced crystallization is orders of magnitude too fast to capture with other imaging techniques. We recorded movies of laser-induced crystallization and measured crystal growth rates at temperatures close to where the maximum growth rate occurs. This paves the way for studying crystallization kinetics of phase change materials over the whole range of technologically relevant temperatures.

  10. The striped phases of ethylthiolate monolayers on the Au(111) surface: a scanning tunneling microscopy study.

    PubMed

    Li, Fangsen; Tang, Lin; Voznyy, Oleksandr; Gao, Jianzhi; Guo, Quanmin

    2013-05-21

    Striped phases of ethylthiolate monolayers, corresponding to surface coverage in between 0.2 ML and 0.27 ML, were studied using high-resolution scanning tunneling microscopy. Striped phases consist of rows of Au-adatom-diethythiolate (AAD) aligned along the [112] direction. In the perpendicular [110] direction, the AAD rows adjust their spacing according to the surface coverage. A (5√3 × √3)-R30° striped phase with 0.27 ML thiolate and a (6√3 × √3)-R30° striped phase with 0.23 ML thiolate, both with long-range order, are found. A localized (5 × √3)-rect. phase is also found as a minority phase embedded in the 5√3 × √3)-R30° phase. This (5 × √3)-rect. phase can be constructed using di-Au-adatom-tri-thiolate species.

  11. Phase stabilized homodyne of infrared scattering type scanning near-field optical microscopy

    SciTech Connect

    Xu, Xiaoji G.; Gilburd, Leonid; Walker, Gilbert C.

    2014-12-29

    Scattering type scanning near-field optical microscopy (s-SNOM) allows sub diffraction limited spatial resolution. Interferometric homodyne detection in s-SNOM can amplify the signal and extract vibrational responses based on sample absorption. A stable reference phase is required for a high quality homodyne-detected near-field signal. This work presents the development of a phase stabilization mechanism for s-SNOM to provide stable homodyne conditions. The phase stability is found to be better than 0.05 rad for the mid infrared light source. Phase stabilization results in improved near field images and vibrational spectroscopies. Spatial inhomogeneities of the boron nitride nanotubes are measured and compared.

  12. Phase shifting technique for extended inline holographic microscopy with a pinhole array.

    PubMed

    Graulig, Christian; Kanka, Mario; Riesenberg, Rainer

    2012-09-24

    Digital inline holographic microscopy using a pinhole for sample illumination allows lensless imaging. To overcome restrictions of the sample size and density in the setup additional reference waves are generated by extending the single pinhole to a regular 2D pinhole array illumination. A technique is presented that uses phase shifting between the pinhole waves. Multiple foci with stable phase differences and a phase error (rms) of 0.027 rad generate pinhole waves which illuminate an undiluted, dense blood smear sample. Amplitude and phase images of the blood sample were successfully reconstructed.

  13. Phase unwrapping method based on multiple recording distances for digital holographic microscopy

    NASA Astrophysics Data System (ADS)

    Li, Yan; Xiao, Wen; Pan, Feng; Rong, Lu

    2015-07-01

    We present a phase unwrapping approach based on multiple recording distances for digital holographic microscopy. It unwrappes the ambiguous phase image by synthesizing the extracted continuous components from a set of multiple reconstructed phase images obtained from a series of holograms by slightly shifting the specimen longitudinally with a step more than the longitudinal correlation length of the coherent noise field. The experimental results demonstrate that the proposed method provides a more accurate calculation and better counteraction of phase noise than the methods proposed in previous research.

  14. Phase aberration compensation of digital holographic microscopy based on least squares surface fitting

    NASA Astrophysics Data System (ADS)

    Di, Jianglei; Zhao, Jianlin; Sun, Weiwei; Jiang, Hongzhen; Yan, Xiaobo

    2009-10-01

    Digital holographic microscopy allows the numerical reconstruction of the complex wavefront of samples, especially biological samples such as living cells. In digital holographic microscopy, a microscope objective is introduced to improve the transverse resolution of the sample; however a phase aberration in the object wavefront is also brought along, which will affect the phase distribution of the reconstructed image. We propose here a numerical method to compensate for the phase aberration of thin transparent objects with a single hologram. The least squares surface fitting with points number less than the matrix of the original hologram is performed on the unwrapped phase distribution to remove the unwanted wavefront curvature. The proposed method is demonstrated with the samples of the cicada wings and epidermal cells of garlic, and the experimental results are consistent with that of the double exposure method.

  15. Variable multimodal light microscopy with interference contrast and phase contrast; dark or bright field.

    PubMed

    Piper, T; Piper, J

    2014-07-01

    Using the optical methods described, specimens can be observed with modified multimodal light microscopes based on interference contrast combined with phase contrast, dark- or bright-field illumination. Thus, the particular visual information associated with interference and phase contrast, dark- and bright-field illumination is joined in real-time composite images appearing in enhanced clarity and purified from typical artefacts, which are apparent in standard phase contrast and dark-field illumination. In particular, haloing and shade-off are absent or significantly reduced as well as marginal blooming and scattering. The background brightness and thus the range of contrast can be continuously modulated and variable transitions can be achieved between interference contrast and complementary illumination techniques. The methods reported should be of general interest for all disciplines using phase and interference contrast microscopy, especially in biology and medicine, and also in material sciences when implemented in vertical illuminators. © 2014 The Authors Journal of Microscopy © 2014 Royal Microscopical Society.

  16. Carbon contamination in scanning transmission electron microscopy and its impact on phase-plate applications.

    PubMed

    Hettler, Simon; Dries, Manuel; Hermann, Peter; Obermair, Martin; Gerthsen, Dagmar; Malac, Marek

    2017-05-01

    We analyze electron-beam induced carbon contamination in a transmission electron microscope. The study is performed on thin films potentially suitable as phase plates for phase-contrast transmission electron microscopy. Electron energy-loss spectroscopy and phase-plate imaging is utilized to analyze the contamination. The deposited contamination layer is identified as a graphitic carbon layer which is not prone to electrostatic charging whereas a non-conductive underlying substrate charges. Several methods that inhibit contamination are evaluated and the impact of carbon contamination on phase-plate imaging is discussed. The findings are in general interesting for scanning transmission electron microscopy applications. Crown Copyright © 2017. Published by Elsevier Ltd. All rights reserved.

  17. Wideband phase-locked loop circuit with real-time phase correction for frequency modulation atomic force microscopy

    NASA Astrophysics Data System (ADS)

    Fukuma, Takeshi; Yoshioka, Shunsuke; Asakawa, Hitoshi

    2011-07-01

    We have developed a wideband phase-locked loop (PLL) circuit with real-time phase correction for high-speed and accurate force measurements by frequency modulation atomic force microscopy (FM-AFM) in liquid. A high-speed operation of FM-AFM requires the use of a high frequency cantilever which, however, increases frequency-dependent phase delay caused by the signal delay within the cantilever excitation loop. Such phase delay leads to an error in the force measurements by FM-AFM especially with a low Q factor. Here, we present a method to compensate this phase delay in real time. Combined with a wideband PLL using a subtraction-based phase comparator, the method allows to perform an accurate and high-speed force measurement by FM-AFM. We demonstrate the improved performance by applying the developed PLL to three-dimensional force measurements at a mica/water interface.

  18. Wideband phase-locked loop circuit with real-time phase correction for frequency modulation atomic force microscopy.

    PubMed

    Fukuma, Takeshi; Yoshioka, Shunsuke; Asakawa, Hitoshi

    2011-07-01

    We have developed a wideband phase-locked loop (PLL) circuit with real-time phase correction for high-speed and accurate force measurements by frequency modulation atomic force microscopy (FM-AFM) in liquid. A high-speed operation of FM-AFM requires the use of a high frequency cantilever which, however, increases frequency-dependent phase delay caused by the signal delay within the cantilever excitation loop. Such phase delay leads to an error in the force measurements by FM-AFM especially with a low Q factor. Here, we present a method to compensate this phase delay in real time. Combined with a wideband PLL using a subtraction-based phase comparator, the method allows to perform an accurate and high-speed force measurement by FM-AFM. We demonstrate the improved performance by applying the developed PLL to three-dimensional force measurements at a mica/water interface.

  19. Instantaneous Spatial Light Interference Microscopy.

    PubMed

    Ding, Huafeng; Popescu, Gabriel

    2010-01-18

    We present Instantaneous Spatial Light Interference Microscopy (iSLIM) as a new quantitative phase method that combines the benefits of white light illumination in Zernike's phase contrast microscopy and phase stability associated diffraction phase microscopy. iSLIM is implemented as an add-on module to a commercial phase contrast microscope, and enables new features to quantitative phase imaging: diminished speckle effects due to white light illumination, multimodal investigation potential due to overlaying with other modalities of the microscope (e.g. fluorescence, DIC, phase contrast), and spectroscopic potential due to the broad band light. We show proof of principle results by multicolor phase imaging of microsphere and red blood cells, and dynamic imaging of nanoscale cell membrane fluctuations.

  20. Parallel-quadrature phase-shifting digital holographic microscopy using polarization beam splitter

    NASA Astrophysics Data System (ADS)

    Das, Bhargab; Yelleswarapu, Chandra S.; Rao, D. V. G. L. N.

    2012-11-01

    We present a digital holography microscopy technique based on a parallel-quadrature phase-shifting method. Two π/2 phase-shifted holograms are recorded simultaneously using polarization phase-shifting principle, slightly off-axis recording geometry, and two identical CCD sensors. The parallel phase-shifting is realized by combining circularly polarized object beam with a 45° degree polarized reference beam through a polarizing beam splitter. DC term is eliminated by subtracting the two holograms from each other and the object information is reconstructed after selecting the frequency spectrum of the real image. Both amplitude and phase object reconstruction results are presented. Simultaneous recording eliminates phase errors caused by mechanical vibrations and air turbulences. The slightly off-axis recording geometry with phase-shifting allows a much larger dimension of the spatial filter for reconstruction of the object information. This leads to better reconstruction capability than traditional off-axis holography.

  1. Parallel-quadrature phase-shifting digital holographic microscopy using polarization beam splitter.

    PubMed

    Das, Bhargab; Yelleswarapu, Chandra S; Rao, Dvgln

    2012-11-01

    We present a digital holography microscopy technique based on parallel-quadrature phase-shifting method. Two π/2 phase-shifted holograms are recorded simultaneously using polarization phase-shifting principle, slightly off-axis recording geometry, and two identical CCD sensors. The parallel phase-shifting is realized by combining circularly polarized object beam with a 45° degree polarized reference beam through a polarizing beam splitter. DC term is eliminated by subtracting the two holograms from each other and the object information is reconstructed after selecting the frequency spectrum of the real image. Both amplitude and phase object reconstruction results are presented. Simultaneous recording eliminates phase errors caused by mechanical vibrations and air turbulences. The slightly off-axis recording geometry with phase-shifting allows a much larger dimension of the spatial filter for reconstruction of the object information. This leads to better reconstruction capability than traditional off-axis holography.

  2. Parallel-quadrature phase-shifting digital holographic microscopy using polarization beam splitter

    PubMed Central

    Das, Bhargab; Yelleswarapu, Chandra S; Rao, DVGLN

    2012-01-01

    We present a digital holography microscopy technique based on parallel-quadrature phase-shifting method. Two π/2 phase-shifted holograms are recorded simultaneously using polarization phase-shifting principle, slightly off-axis recording geometry, and two identical CCD sensors. The parallel phase-shifting is realized by combining circularly polarized object beam with a 45° degree polarized reference beam through a polarizing beam splitter. DC term is eliminated by subtracting the two holograms from each other and the object information is reconstructed after selecting the frequency spectrum of the real image. Both amplitude and phase object reconstruction results are presented. Simultaneous recording eliminates phase errors caused by mechanical vibrations and air turbulences. The slightly off-axis recording geometry with phase-shifting allows a much larger dimension of the spatial filter for reconstruction of the object information. This leads to better reconstruction capability than traditional off-axis holography. PMID:23109732

  3. Multi-modal digital holographic microscopy for wide-field fluorescence and 3D phase imaging

    NASA Astrophysics Data System (ADS)

    Quan, Xiangyu; Xia, Peng; Matoba, Osamu; Nitta, Koichi; Awatsuji, Yasuhiro

    2016-03-01

    Multi-modal digital holographic microscopy is a combination of epifluorescence microscopy and digital holographic microscopy, the main function of which is to obtain images from fluorescence intensity and quantified phase contrasts, simultaneously. The proposed system is mostly beneficial to biological studies, with the reason that often the studies are depending on fluorescent labeling techniques to detect certain intracellular molecules, while phase information reflecting properties of unstained transparent elements. This paper is presenting our latest researches on applications such as randomly moving micro-fluorescent beads and living cells of Physcomitrella patens. The experiments are succeeded on obtaining a succession of wide-field fluorescent images and holograms from micro-beads, and different depths focusing is realized via numerical reconstruction. Living cells of Physcomitrella patens are recorded in the static manner, the reconstruction distance indicates thickness of cellular structure. These results are implementing practical applications toward many biomedical science researches.

  4. High Resolution Phase-Sensitive Magnetomotive Optical Coherence Microscopy for Tracking Magnetic Microbeads and Cellular Mechanics

    PubMed Central

    Crecea, Vasilica; Graf, Benedikt W.; Kim, Taewoo; Popescu, Gabriel; Boppart, Stephen A.

    2014-01-01

    We present a real-time multimodal near-infrared imaging technology that tracks externally induced axial motion of magnetic microbeads in single cells in culture. The integrated multimodal imaging technique consists of phase-sensitive magnetomotive optical coherence microscopy (MM-OCM) and multiphoton microscopy (MPM).MPMis utilized for the visualization of multifunctional fluorescent and magnetic microbeads, while MM-OCM detects, with nanometer-scale sensitivity, periodic displacements of the microbeads induced by the modulation of an external magnetic field. Magnetomotive signals are measured from mouse macrophages, human breast primary ductal carcinoma cells, and human breast epithelial cells in culture, and validated with full-field phase-sensitive microscopy. This methodology demonstrates the capability for imaging controlled cell dynamics and has the potential for measuring cell biomechanical properties, which are important in assessing the health and pathological state of cells. PMID:25400496

  5. Total internal reflection holographic microscopy (TIRHM) for quantitative phase characterization of cell-substrate adhesion

    NASA Astrophysics Data System (ADS)

    Ash, William Mason, III

    Total Internal Reflection Holographic Microscopy (TIRHM) combines near-field microscopy with digital holography to produce a new form of near-field phase microscopy. Using a prism in TIR as a near-field imager, the presence of microscopic organisms, cell-substrate interfaces, and adhesions, causes relative refractive index (RRI) and frustrated TIR (f-TIR) to modulate the object beam's evanescent wave phase front. Quantitative phase images of test specimens such as Amoeba proteus, Dictyostelium Discoideum and cells such as SKOV-3 ovarian cancer and 3T3 fibroblasts are produced without the need to introduce stains or fluorophores. The angular spectrum method of digital holography to compensate for tilt anamorphism due to the inclined TIR plane is also discussed. The results of this work conclusively demonstrate, for the first time, the integration of near-field microscopy with digital holography. The cellular images presented show a correlation between the physical extent of the Amoeba proteus plasma membrane and the adhesions that are quantitatively profiled by phase cross-sectioning of the holographic images obtained by digital holography. With its ability to quantitatively characterise cellular adhesion and motility, it is anticipated that TIRHM can be a tool for characterizing and combating cancer metastasis, as well as improving our understanding of morphogenesis and embryogenesis itself.

  6. Pre-phase A: Development of a far-ultraviolet photometric- and spectroscopic-survey small-explorer experiment

    NASA Technical Reports Server (NTRS)

    Martin, Christopher

    1993-01-01

    We propose to perform a far ultraviolet photometric and spectroscopic survey covering the lambda lambda 1300-2000 band with a sensitivity comparable to that of the Palomar Sky Survey. This survey will proceed in three phases: an all-sky survey in three bands to 18-19.5(sup m), deep surveys of selected targets of interest in the same bands to 21-22(sup m), and a spectroscopic survey of 2 percent of the sky to 18(sup m) with a resolution of 3-20A. This mission, the Joint Ultraviolet Nightsky Observer (JUNO), can be performed by a Small-Explorer-class satellite.

  7. Single-shot and phase-shifting digital holographic microscopy using a 2-D grating.

    PubMed

    Yang, Taeseok Daniel; Kim, Hyung-Jin; Lee, Kyoung J; Kim, Beop-Min; Choi, Youngwoon

    2016-05-02

    We demonstrate digital holographic microscopy that, while being based on phase-shifting interferometry, is capable of single-shot measurements. A two-dimensional (2-D) diffraction grating placed in a Fourier plane of a standard in-line holographic phase microscope generates multiple copies of a sample image on a camera sensor. The identical image copies are spatially separated with different overall phase shifts according to the diffraction orders. The overall phase shifts are adjusted by controlling the lateral position of the grating. These phase shifts are then set to be multiples of π/2. Interferograms composed of four image copies combined with a parallel reference beam are acquired in a single shot. The interferograms are processed through a phase-shifting algorithm to produce a single complex image. By taking advantage of the higher sampling capacity of the in-line holography, we can increase the imaging information density by a factor of 3 without compromising the imaging acquisition speed.

  8. Gravity driven high throughput phase detecting cytometer based on quantitative interferometric microscopy

    NASA Astrophysics Data System (ADS)

    Xue, Liang; Wang, Shouyu; Yan, Keding; Sun, Nan; Ferraro, Pietro; Li, Zhenhua; Liu, Fei

    2014-04-01

    Phase distribution detection of cells and tissues is concerned since it is an important auxiliary method for observing biological samples. High speed and large amount cell detection is needed for its high detecting efficiency. In this paper, we have proposed a simple large scale biological sample phase detection device called gravity driven high throughput phase detecting cytometer based on quantitative interferometric microscopy to obtain flowing red blood cells phase. The system could realize high throughput phase detecting and statistical analysis with high detecting speed and in real time. The statistical characteristics of red blood cells could be obtained which might be helpful for biological analysis and disease detection. We believe this method is a powerful tool to quantitatively measure the phase distribution of biological samples.

  9. Incommensurate phases in barium sodium niobate: Transmission-electron-microscopy study

    SciTech Connect

    Barre, S.; Mutka, H.; Roucau, C.

    1988-11-01

    A transmission-electron-microscopy study of barium sodium niobate has confirmed new features of the incommensurate phase. We present here images showing the nucleation of the incommensurate phase when the temperature increases after thermal cycling the sample or annealing in the incommensurate phase. In agreement with previous inferences, in situ annealing either at 230 /sup 0/C near the lock-in transition or at 270 /sup 0/C close to the tetragonal normal phase transition can favor the stability of a unidirectional 1q or a doubly modulated 2q incommensurate phase, respectively. The difference in the orientation of discommensurations, either perpendicular to the orthorhombic a axis or perpendicular to the tetragonal a and b axes, agree with an assignment of the two phases based on theoretical arguments and on x-ray and optical data.

  10. Integral refractive index determination of living suspension cells by multifocus digital holographic phase contrast microscopy.

    PubMed

    Kemper, Björn; Kosmeier, Sebastian; Langehanenberg, Patrik; von Bally, Gert; Bredebusch, Ilona; Domschke, Wolfram; Schnekenburger, Jürgen

    2007-01-01

    A method for the determination of the integral refractive index of living cells in suspension by digital holographic microscopy is described. Digital holographic phase contrast images of spherical cells in suspension are recorded, and the radius as well as the integral refractive index are determined by fitting the relation between cell thickness and phase distribution to the measured phase data. The algorithm only requires information about the refractive index of the suspension medium and the image scale of the microscope system. The specific digital holographic microscopy advantage of subsequent focus correction allows a simultaneous investigation of cells in different focus planes. Results obtained from human pancreas and liver tumor cells show that the integral cellular refractive index decreases with increasing cell radius.

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

    NASA Astrophysics Data System (ADS)

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

    2016-02-01

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

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

    PubMed Central

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

    2016-01-01

    The ability to image light elements in soft matter at atomic resolution enables unprecedented insight into the structure and properties of molecular heterostructures and beam-sensitive nanomaterials. In this study, we introduce a scanning transmission electron microscopy technique combining a pre-specimen phase plate designed to produce a probe with structured phase with a high-speed direct electron detector to generate nearly linear contrast images with high efficiency. We demonstrate this method by using both experiment and simulation to simultaneously image the atomic-scale structure of weakly scattering amorphous carbon and strongly scattering gold nanoparticles. Our method demonstrates strong contrast for both materials, making it a promising candidate for structural determination of heterogeneous soft/hard matter samples even at low electron doses comparable to traditional phase-contrast transmission electron microscopy. Simulated images demonstrate the extension of this technique to the challenging problem of structural determination of biological material at the surface of inorganic crystals. PMID:26923483

  13. Superresolved common-path phase-shifting digital inline holographic microscopy using a spatial light modulator.

    PubMed

    Micó, Vicente; Zalevsky, Zeev; Garcia, Javier

    2012-12-01

    Common-path phase-shifting lensless holographic microscopy has been recently proposed as a novel approach capable of high numerical aperture imaging in a lensless digital inline holographic microscopy layout [Opt. Lett.35, 3919 (2010)]. Here we present proof-of-concept validation for improving the resolution limit imposed by diffraction in such a setup. This is accomplished by shifting the phase lens displayed at the spatial light modulator, which moves the illumination point source to different off-axis positions. For each off-axis position, a set of inline phase-shifted holograms are recorded by the digital sensor and stored at the computer's memory for later digital postprocessing. As a consequence, each recording allows the recovery of different spatial frequency content of the object's diffracted wavefront meaning a superresolved image of the input object. Experimental results are reported validating the proposed method.

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

    PubMed

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

    2016-02-29

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

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

    SciTech Connect

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

    2016-02-29

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

  16. One-particle spectroscopic intensities as a signature of shape phase transition: The {gamma}-unstable case

    SciTech Connect

    Alonso, C. E.; Arias, J. M.; Vitturi, A.

    2006-08-15

    We investigate the evolution of one-particle spectroscopic intensities as a possible signature of shape phase transitions. The study describes the odd systems in terms of the interacting boson-fermion model. We consider the particular case of an odd j=3/2 particle coupled to an even-even boson core that undergoes a phase transition from spherical U(5) to {gamma}-unstable O(6) situation. At the critical point, our findings are compared with the one-particle spectroscopic intensities that can be obtained within the E(5/4) model proposed by[F. Iachello, Phys. Rev. Lett. 95, 052503 (2005); F. Iachello, in Symmetries and Low-Energy Phase Transitions in Nuclear Structure Physics, edited by G. Lo Bianco (University of Camerino Press, Camerino, Italy, in press)].

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

    NASA Astrophysics Data System (ADS)

    Waller, Laura

    2016-03-01

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

  18. Phase measurements of erythrocytes affected by metal ions with quantitative interferometric microscopy

    NASA Astrophysics Data System (ADS)

    Wang, Shouyu; Yan, Keding; Shan, Yanke; Xu, Mingfei; Liu, Fei; Xue, Liang

    2015-12-01

    Erythrocyte morphology is an important factor in disease diagnosis, however, traditional setups as microscopes and cytometers cannot provide enough quantitative information of cellular morphology for in-depth statistics and analysis. In order to capture variations of erythrocytes affected by metal ions, quantitative interferometric microscopy (QIM) is applied to monitor their morphology changes. Combined with phase retrieval and cell recognition, erythrocyte phase images, as well as phase area and volume, can be accurately and automatically obtained. The research proves that QIM is an effective tool in cellular observation and measurement.

  19. Time-resolved imaging refractometry of microbicidal films using quantitative phase microscopy

    PubMed Central

    Rinehart, Matthew T.; Drake, Tyler K.; Robles, Francisco E.; Rohan, Lisa C.; Katz, David; Wax, Adam

    2011-01-01

    Quantitative phase microscopy is applied to image temporal changes in the refractive index (RI) distributions of solutions created by microbicidal films undergoing hydration. We present a novel method of using an engineered polydimethylsiloxane structure as a static phase reference to facilitate calibration of the absolute RI across the entire field. We present a study of dynamic structural changes in microbicidal films during hydration and subsequent dissolution. With assumptions about the smoothness of the phase changes induced by these films, we calculate absolute changes in the percentage of film in regions across the field of view. PMID:22191912

  20. Time-resolved imaging refractometry of microbicidal films using quantitative phase microscopy.

    PubMed

    Rinehart, Matthew T; Drake, Tyler K; Robles, Francisco E; Rohan, Lisa C; Katz, David; Wax, Adam

    2011-12-01

    Quantitative phase microscopy is applied to image temporal changes in the refractive index (RI) distributions of solutions created by microbicidal films undergoing hydration. We present a novel method of using an engineered polydimethylsiloxane structure as a static phase reference to facilitate calibration of the absolute RI across the entire field. We present a study of dynamic structural changes in microbicidal films during hydration and subsequent dissolution. With assumptions about the smoothness of the phase changes induced by these films, we calculate absolute changes in the percentage of film in regions across the field of view.

  1. Using quantitative interference phase microscopy for sperm acrosome evaluation (Conference Presentation)

    NASA Astrophysics Data System (ADS)

    Balberg, Michal; Kalinowski, Ksawery; Levi, Mattan; Shaked, Natan T.

    2016-03-01

    We demonstrate quantitative assessment of sperm cell morphology, primarily acrosomal volume, using quantitative interference phase microscopy (IPM). Normally, the area of the acrosome is assessed using dyes that stain the acrosomal part of the cell. We have imaged fixed individual sperm cells using IPM. Following, the sample was stained and the same cells were imaged using bright field microscopy (BFM). We identified the acrosome using the stained BFM image, and used it to define a quantitative corresponding area in the IPM image and determine a quantitative threshold for evaluating the volume of the acrosome.

  2. In vivo endoscopic tissue diagnostics based on spectroscopic absorption, scattering, and phase function properties.

    PubMed

    Thueler, Philippe; Charvet, Igor; Bevilacqua, Frederic; St Ghislain, M; Ory, G; Marquet, Pierre; Meda, Paolo; Vermeulen, Ben; Depeursinge, Christian

    2003-07-01

    A fast spectroscopic system for superficial and local determination of the absorption and scattering properties of tissue (480 to 950 nm) is described. The probe can be used in the working channel of an endoscope. The scattering properties include the reduced scattering coefficient and a parameter of the phase function called gamma, which depends on its first two moments. The inverse problem algorithm is based on the fit of absolute reflectance measurements to cubic B-spline functions derived from the interpolation of a set of Monte Carlo simulations. The algorithm's robustness was tested with simulations altered with various amounts of noise. The method was also assessed on tissue phantoms of known optical properties. Finally, clinical measurements performed endoscopically in vivo in the stomach of human subjects are presented. The absorption and scattering properties were found to be significantly different in the antrum and in the fundus and are correlated with histopathologic observations. The method and the instrument show promise for noninvasive tissue diagnostics of various epithelia.

  3. LORENTZ PHASE IMAGING AND IN-SITU LORENTZ MICROSCOPY OF PATTERNED CO-ARRAYS.

    SciTech Connect

    VOLKOV,V.V.ZHU,Y.

    2003-08-03

    Understanding magnetic structures and properties of patterned and ordinary magnetic films at nanometer length-scale is the area of immense technological and fundamental scientific importance. The key feature to such success is the ability to achieve visual quantitative information on domain configurations with a maximum ''magnetic'' resolution. Several methods have been developed to meet these demands (Kerr and Faraday effects, differential phase contrast microscopy, magnetic force microscopy, SEMPA etc.). In particular, the modern off-axis electron holography allows retrieval of the electron-wave phase shifts down to 2{pi}/N (with typical N = 10-20, approaching in the limit N {approx} 100) in TEM equipped with field emission gun, which is already successfully employed for studies of magnetic materials at nanometer scale. However, it remains technically demanding, sensitive to noise and needs highly coherent electron sources. As possible alternative we developed a new method of Lorentz phase microscopy [1,2] based on the Fourier solution [3] of magnetic transport-of-intensity (MTIE) equation. This approach has certain advantages, since it is less sensitive to noise and does not need high coherence of the source required by the holography. In addition, it can be realized in any TEM without basic hardware changes. Our approach considers the electron-wave refraction in magnetic materials (magnetic refraction) and became possible due to general progress in understanding of noninterferometric phase retrieval [4-6] dealing with optical refraction. This approach can also be treated as further development of Fresnel microscopy, used so far for imaging of in-situ magnetization process in magnetic materials studied by TEM. Figs. 1-3 show some examples of what kind information can be retrieved from the conventional Fresnel images using the new approach. Most of these results can be compared with electron-holographic data. Using this approach we can shed more light on fine

  4. Coherent noise reduction in digital holographic phase contrast microscopy by slightly shifting object.

    PubMed

    Pan, Feng; Xiao, Wen; Liu, Shuo; Wang, FanJing; Rong, Lu; Li, Rui

    2011-02-28

    A method to reduce coherent noise in digital holographic phase contrast microscopy is proposed. By slightly shifting the specimen, a series of digital holograms with different coherent noise patterns is recorded. Each hologram is reconstructed individually, while the different phase tilts of the reconstructed complex amplitudes due to the specimen shifts are corrected in the hologram plane by using numerical parametric lens method. Afterward, the lateral displacements of the phase maps from different holograms are compensated in the image plane by using digital image registration method. Thus, all phase images have same distribution, but uncorrelated coherent noise patterns. By a proper averaging procedure, the coherent noise of phase contrast image is reduced significantly. The experimental results are given to confirm the proposed method.

  5. 3D motion picture recording by parallel phase-shifting digital holographic microscopy

    NASA Astrophysics Data System (ADS)

    Tahara, Tatsuki; Xia, Peng; Kakue, Takashi; Awatsuji, Yasuhiro; Nishio, Kenzo; Ura, Shogo; Kubota, Toshihiro; Matoba, Osamu

    2013-12-01

    Three-dimensional (3-D) motion-picture recording by parallel phase-shifting digital holographic microscopy that has the ability of instantaneous 3-D recording of dynamic phenomena in the microscopic field of view is presented. Parallel phase-shifting digital holography is a scheme to record multiple phase-shifted holograms with a single-shot exposure, and to achieve 3-D motion-picture recording of objects with high accuracy and wide 3-D area, based on space-division multiplexing of phase-shifted holograms. Parallel phase-shifting digital holographic microscopy is implemented by an optical interferometer and an image sensor on which polarization-detection function is introduced pixel by pixel. This time, we constructed a parallel phase-shifting digital holographic microscope for recording high-speed dynamic phenomena, and then motions of biological objects in water were recorded at more than 10,000 frames per second, which is the fastest among the previous reports on 3-D imaging of biological objects.

  6. Spectroscopic characterization of charged defects in polycrystalline pentacene by time- and wavelength-resolved electric force microscopy.

    PubMed

    Luria, Justin L; Schwarz, Kathleen A; Jaquith, Michael J; Hennig, Richard G; Marohn, John A

    2011-02-01

    Spatial maps of topography and trapped charge are acquired for polycrystalline pentacene thin-film transistors using electric and atomic force microscopy. In regions of trapped charge, the rate of trap clearing is studied as a function of the wavelength of incident radiation.

  7. TV-regularized phase reconstruction in differential-interference-contrast (DIC) microscopy

    NASA Astrophysics Data System (ADS)

    Rebegoldi, Simone; Bautista, Lola; Blanc-Féraud, Laure; Prato, Marco; Zanni, Luca; Plata, Arturo

    2016-10-01

    In this paper we address the problem of reconstructing the phase from color images acquired with differential-interference-contrast (DIC) microscopy. In particular, we reformulate the problem as the minimization of a least-squares fidelity function regularized with a total variation term, and we address the solution by exploiting a recently proposed inexact forward-backward approach. The effectiveness of this method is assessed on a realistic synthetic test.

  8. Reflective-type digital holographic microscopy using induced self-pumped phase conjugation technique

    NASA Astrophysics Data System (ADS)

    Chew, Yang Kun; Chan, Huang Tian; Shiu, Min Tzung; Wu, Chia Hao; Chang, Chi Ching

    2017-05-01

    In digital holography, techniques of noise suppression from the perspective of reference arm have been maturely developed. The object counterpart, however, is still in its infancy. Self-pumped phase conjugation technique which involves a BaTiO3 crystal was introduced unto reflective-type digital holographic microscopy to suppress scattering noise derived from the object arm, prior to recording stage. A phase distorter was introduced as scattering source, and signal-to-noise ratio was calculated. Furthermore, induced method was proposed to speed up the response time.

  9. Digital microscopy using phase-shifting digital holography with two reference waves.

    PubMed

    Guo, Pengyi; Devaney, Anthony J

    2004-04-15

    A lensless, coherent optical microscope is described that uses a version of phase-shifting digital holography (PSDH) in conjunction with a field backpropagation algorithm to form coherent images of transmission-type objects. The PSDH is implemented by use of only two reference waves, in contrast with the usual implementation that requires four quadrature phase-shifting reference waves. Therefore only two digital holograms need to be recorded, and the complexity of the microscopic system is reduced. Experimental results are presented that compare images generated from conventional Gabor digital holography, two-reference-wave PSDH, and conventional white-light microscopy.

  10. Phase contrast x-ray microscopy study of rabbit primo vessels

    NASA Astrophysics Data System (ADS)

    Kim, M.-S.; Oh, S.-W.; Lim, J.-H.; Han, S.-W.

    2010-11-01

    The microstructural properties of the primo-vascular systems of rabbits were examined by phase contrast x-ray microscopy. The primo-vascular systems with an average diameter of 32 μm were extracted from the surfaces of the rabbit internal organs. Phase contrast x-ray imaging showed that the primo-vascular systems were tubules with random holes on their lateral surfaces. The size of the holes on the vascular surface was 2-5 μm. The holes might act as size selective channels for microcells coming in and out and assist in the unidirectional flow of the primo-vascular systems.

  11. Two-channel interferometric detection for the compensation of phase fluctuation noise in nonlinear infrared microscopy

    NASA Astrophysics Data System (ADS)

    Lee, E. S.; Lee, S. W.; Park, J. H.; Lee, J. Y.

    2013-01-01

    We demonstrate an interferometric implementation of nonlinear optical infrared microscopy in which a probe beam interrogates the refractive index change of samples that is induced by molecular vibrational absorption of an infrared pump beam. To compensate for the phase noise caused by the ambient airflow and external vibration, we configure two coupled Mach-Zehnder interferometers sharing a single microscope setup in the sample arm. Proper control over the phase between the two interference channels allows two simultaneous quadrature mode outputs. The validity of our method is proven by imaging polystyrene bead aggregates and comparing the images with the conventional single-channel results.

  12. Frequency, amplitude, and phase measurements in contact resonance atomic force microscopies.

    PubMed

    Stan, Gheorghe; Solares, Santiago D

    2014-01-01

    The resonance frequency, amplitude, and phase response of the first two eigenmodes of two contact-resonance atomic force microscopy (CR-AFM) configurations, which differ in the method used to excite the system (cantilever base vs sample excitation), are analyzed in this work. Similarities and differences in the observables of the cantilever dynamics, as well as the different effect of the tip-sample contact properties on those observables in each configuration are discussed. Finally, the expected accuracy of CR-AFM using phase-locked loop detection is investigated and quantification of the typical errors incurred during measurements is provided.

  13. Projection phase contrast microscopy with a hard x-ray nanofocused beam: Defocus and contrast transfer

    SciTech Connect

    Salditt, T.; Giewekemeyer, K.; Fuhse, C.; Krueger, S. P.; Tucoulou, R.; Cloetens, P.

    2009-05-01

    We report a projection phase contrast microscopy experiment using hard x-ray pink beam undulator radiation focused by an adaptive mirror system to 100-200 nm spot size. This source is used to illuminate a lithographic test pattern with a well-controlled range of spatial frequencies. The oscillatory nature of the contrast transfer function with source-to-sample distance in this holographic imaging scheme is quantified and the validity of the weak phase object approximation is confirmed for the experimental conditions.

  14. [Phase contrast microscopy demonstration of glomerular erythrocytes in urine: practicable in ambulatory practice?].

    PubMed

    Conzelmann, M; Conen, D; Besch, W; Dubach, U C; Thiel, G

    1988-04-16

    The use of phase-contrast microscopy in microhematuria, as proposed in 1979 by Birch and Fairley, renders morphological changes in red cells easily detectable and makes it possible to distinguish glomerular from non-glomerular bleeding. The aim of this study was to evaluate the practicability of this method as a routine laboratory test in ambulatory care. 60 patients with asymptomatic microhematuria (greater than or equal to 2 erythrocytes per high power field) were followed up over a one-year period. All patients were investigated by intravenous pyelography, ultrasound of urinary tract and three cytological examinations of the urine. The description of urine samples was done with phase-contrast microscopy by a first investigator at the beginning of the study and by a second after 12.8 months, blinded to clinical results and previous examinations. In 21 patients a definitive diagnosis was possible. In 18 patients the morphologic descriptions of the two investigators correlated with the clinical results. Only in two patients with established diagnosis there were differences between the urine description of the two investigators, and in one patient the interpretations of both investigators were wrong. These incorrect descriptions concerned patients with low-grade microhematuria. Thus, phase-contrast microscopy is a practicable method for the practitioner's use as a routine laboratory investigation. In low-grade microhematuria the method seems to be of minor value.

  15. Phase imaging and nanoscale energy dissipation of supported graphene using amplitude modulation atomic force microscopy.

    PubMed

    Vasic, Borislav; Matkovic, Aleksandar; Gajic, Rados

    2017-09-21

    We investigate phase imaging of supported graphene using amplitude modulation atomic force microscopy (AFM), so called, tapping mode. The phase contrast between graphene and neighbouring substrate grows at hard tapping conditions and the contrast is enhanced compared to the topographic one. Therefore, the phase measurements could enable a high contrast imaging of graphene and related two dimensional materials and heterostructures, not achievable with conventional AFM based topographic measurements. Obtained phase maps are then transformed into energy dissipation maps which are important for graphene applications in various mechanical systems. From fundamental point of view, the energy dissipation gives further insights into mechanical properties. Reliable measurements, obtained in the repulsive regime, show that the energy dissipation on graphene covered substrate is lower than on bare one, so graphene provides a certain shielding in tip-substrate interaction. According to obtained phase curves and their derivatives, as well as according to correlation measurements based on AFM nanoindentation and force modulation microscopy, we conclude that the main dissipation channels in graphene-substrate systems are short range hysteresis and long range interfacial forces. © 2017 IOP Publishing Ltd.

  16. Interferometric phase microscopy using slightly-off-axis reflective point diffraction interferometer

    NASA Astrophysics Data System (ADS)

    Bai, Hongyi; Zhong, Zhi; Shan, Mingguang; Liu, Lei; Guo, Lili; Zhang, Yabin

    2017-03-01

    An interferometric phase microscopy (IPM) is proposed using slightly-off-axis reflective point diffraction interferometry for quantitative phase imaging. A retro-reflector consisting two mirrors is used to generate an angle between the object beam and reference beam, and a 45° tilted polarizing beam splitter is used to split the horizontal and vertical components of the both beams. Two carrier interferograms with π/2 phase-shift can be acquired in one shot, and the phase distribution of a thin specimen can be retrieved using a fast reconstruction method. The new IPM without loss in the utilization of the input-plane field of view combines the real time and optimizing detector bandwidth measurement benefit associated with slightly-off-axis method, high stability associated with common path geometry, and simplicity in terms of procedure and setup. Experiments are carried out on both static and dynamic specimens to demonstrate the validity and stability of the proposed method.

  17. Single-shot slightly-off-axis interferometry based Hilbert phase microscopy of red blood cells.

    PubMed

    Xue, Liang; Lai, Jiancheng; Wang, Shouyu; Li, Zhenhua

    2011-03-29

    A slightly-off-axis interferometry based Hilbert phase microscopy (HPM) method is developed to quantitatively obtain the phase distribution. Owing to its single-shot nature and details detection ability, HPM can be used to investigate rapid phenomena that take place in transparent structures such as biological cells. Moreover, the slightly-off-axis interferometry owns higher effective bandwidth and more sensitivity than traditional off-axis interferometry. The proposed method takes advantages of the above techniques to obtain the phase image of the red blood cells and compared with the traditional off-axis interferometry and phase retrieval algorithm based on the FFT. The experimental results show that the proposed method owns fine spatial details and real-time imaging ability. We are sure that the proposed method provides a breakthrough for real-time observing and quantitative analyzing of cells in vivo.

  18. Online quantitative phase imaging of vascular endothelial cells under fluid shear stress utilizing digital holographic microscopy

    NASA Astrophysics Data System (ADS)

    Odenthal-Schnittler, Maria; Schnittler, Hans Joachim; Kemper, Björn

    2016-03-01

    We have explored the utilization of quantitative phase imaging with digital holographic microscopy (DHM) as a novel tool for quantifying the dynamics of morphologic parameters (morphodynamics) of confluent endothelial cell layers under fluid shear stress conditions. Human umbilical vein endothelial cells (HUVECs) were exposed to fluid shear stress in a transparent cone/plate flow device (BioTech-Flow-System) and imaged with a modular setup for quantitative DHM phase imaging for up to 48 h. The resulting series of quantitative phase image sequences were analyzed for the average surface roughness of the cell layers and cell alignment. Our results demonstrate that quantitative phase imaging is a powerful and reliable tool to quantify the dynamics of morphological adaptation of endothelial cells to fluid shear stress.

  19. Analysis of wavelength error in spectral phase shifting of digital holographic microscopy

    NASA Astrophysics Data System (ADS)

    Wang, Jie; Zhang, Xiangchao; Zhang, Xiaolei; Xiao, Hong; Xu, Min

    2016-10-01

    Digital holographic microscopy is an attractive technology of precision measurement. Phase shifting is required to correctly reconstruct the measured surfaces from interferograms. Spectral phase shifting scheme, as an alternative approach of phase shifting, has drawn intensive attention in recent years. However, the wavelength modulated by the acousto-optic tunable filter (AOTF) is not sufficiently precise. As a consequence, severe measurement errors will be caused. In this paper, an iterative calibration algorithm is proposed. It estimates the unknown wavelength errors in the 3-step spectral phase shifting interferometry and then reconstructs the complex object wave. The actual wavelength is obtained by minimizing the difference between the measured and calculated intensities. Numerical examples have demonstrated that this algorithm can achieve very high accuracy over a wide range of wavelengths.

  20. From images to interactions: high-resolution phase imaging in tapping-mode atomic force microscopy.

    PubMed Central

    Stark, M; Möller, C; Müller, D J; Guckenberger, R

    2001-01-01

    In tapping-mode atomic force microscopy, the phase shift between excitation and response of the cantilever is used as a material-dependent signal complementary to topography. The localization of information in the phase signal is demonstrated with 1.4-nm lateral resolution on purple membrane of Halobacterium salinarum in buffer solution. In a first-order approximation, the phase signal is found to correlate with modulations of the tip oscillation amplitude, induced by topography. Extending the analysis to contributions of the tip-sample interaction area as a second-order approximation, a method is proposed to extract information about the interaction from the phase signal for surfaces with a roughness in the order of the tip radius. PMID:11371473

  1. Quantitative phase microscopy using defocusing by means of a spatial light modulator.

    PubMed

    Camacho, Luis; Micó, Vicente; Zalevsky, Zeev; García, Javier

    2010-03-29

    A new method for recovery the quantitative phase information of microscopic samples is presented. It is based on a spatial light modulator (SLM) and digital image processing as key elements to extract the sample's phase distribution. By displaying a set of lenses with different focal power, the SLM produces a set of defocused images of the input sample at the CCD plane. Such recorded images are then numerically processed to retrieve phase information. This iterative process is based on the wave propagation equation and leads on a complex amplitude image containing information of both amplitude and phase distributions of the input sample diffracted wave front. The proposed configuration is a non-interferometric architecture (conventional transmission imaging mode) where no moving elements are included. Experimental results perfectly correlate with the results obtained by conventional digital holographic microscopy (DHM).

  2. A Method for accurate in vivo micro-Raman spectroscopic measurements under guidance of advanced microscopy imaging

    PubMed Central

    Wang, Hequn; Lee, Anthony M. D.; Lui, Harvey; McLean, David I.; Zeng, Haishan

    2013-01-01

    The movement from the subjects during in vivo confocal Raman spectral measurements could change the measurement volume, leading to non-specific signals and inaccurate interpretation of the acquired spectrum. Here we introduce a generally applicable method that includes (1) developing a multimodal system to achieve real-time monitoring of every spectral measurement with reflectance confocal microscopy (RCM) and multiphoton microscopy (MPM) imaging; (2) performing region-of-interest measurement by scanning an area of the tissue during spectral acquisition. The developed method has been validated by measuring different micro-structures of in vivo human skin. Our results demonstrated great consistency between RCM images and confocal Raman spectra. The superior quality of the images and spectra allows us to derive blood flow velocity and blood glucose level. We believe this method is valuable for realizing accurate microscopic spectral measurement and have great potential to be adapted into clinic to achieve non-invasive measurement of important biological parameters. PMID:23712517

  3. Improved cancer risk stratification and diagnosis via quantitative phase microscopy (Conference Presentation)

    NASA Astrophysics Data System (ADS)

    Liu, Yang; Uttam, Shikhar; Pham, Hoa V.; Hartman, Douglas J.

    2017-02-01

    Pathology remains the gold standard for cancer diagnosis and in some cases prognosis, in which trained pathologists examine abnormality in tissue architecture and cell morphology characteristic of cancer cells with a bright-field microscope. The limited resolution of conventional microscope can result in intra-observer variation, missed early-stage cancers, and indeterminate cases that often result in unnecessary invasive procedures in the absence of cancer. Assessment of nanoscale structural characteristics via quantitative phase represents a promising strategy for identifying pre-cancerous or cancerous cells, due to its nanoscale sensitivity to optical path length, simple sample preparation (i.e., label-free) and low cost. I will present the development of quantitative phase microscopy system in transmission and reflection configuration to detect the structural changes in nuclear architecture, not be easily identifiable by conventional pathology. Specifically, we will present the use of transmission-mode quantitative phase imaging to improve diagnostic accuracy of urine cytology and the nuclear dry mass is progressively correlate with negative, atypical, suspicious and positive cytological diagnosis. In a second application, we will present the use of reflection-mode quantitative phase microscopy for depth-resolved nanoscale nuclear architecture mapping (nanoNAM) of clinically prepared formalin-fixed, paraffin-embedded tissue sections. We demonstrated that the quantitative phase microscopy system detects a gradual increase in the density alteration of nuclear architecture during malignant transformation in animal models of colon carcinogenesis and in human patients with ulcerative colitis, even in tissue that appears histologically normal according to pathologists. We evaluated the ability of nanoNAM to predict "future" cancer progression in patients with ulcerative colitis.

  4. Magneto-optical spectroscopic studies of solid and solution-phase tetra-phenyl porphyrin

    NASA Astrophysics Data System (ADS)

    Wahlen-Strothman, Jacob; Pan, Zhen Wen; Lamarche, Cody; Manning, Lane; Rawat, Naveen; Tokumoto, Takahisa; McGill, Stephen; Furis, Madalina; Chu, Kelvin

    2012-02-01

    Tetraphenylporphyrin (TPP) is a heterocyclic model system for porphyrins found in heme proteins, cytochromes and photosynthetic cofactors. TPP can accommodate a metal ion in the center; D-shell ion porphyrin complexes with a crystalline solid phase are of interest for magnetic studies because of the possibility of macroscopic long-range magnetic order of the ion spins. We have investigated the 5K magnetic properties of poly-crystalline thin films of TPP complexed with Zn, Mn and Cu and deposited through a room temperature capillary pen technique that produces grain size in the 100 micron to 1mm range. Our novel setup measures the UV/VIS, linear dichroism and MCD simultaneously and incorporates a photoelastic modulator and a microscopy superconducting magnet for high-field (5T) measurements. In addition, we present 25T data on samples from the new split magnet at NHMFL. We present solution and crystalline data on metal-complexed TPP; data are analyzed in terms of A and B-type MCD using a perimeter model. We find good agreement with previous solution data, and novel crystalline phase spectra that are correlated to the long range ordering of the solid state.

  5. Magneto-optical spectroscopic studies of solid and solution-phase tetra-phenyl porphyrin

    NASA Astrophysics Data System (ADS)

    Wahlen-Strothman, Jacob; Pan, Zhen Wen; Manning, Lane; Furis, Madalina; Chu, Kelvin

    2011-03-01

    Tetraphenylporphyrin (TPP) is a synthetic heterocyclic compound that serves as a model system for heme proteins and cytochromes. TPP can accomodate a metal ion in the center; D-shell ion porphyrin complexes with a crystalline solid phase are of interest for magnetic studies because of the possibility of macroscopic long range magnetic order of the ion spins. We have investigated the 5K magnetic properties of poly-crystalline thin films of the heme protoporphyrin IX analogue tetra-phenyl porphyrin, complexed with Zn and Mn, deposited through a capillary pen technique that produces 100um to 1 mm sized grains. Our novel experimental setup measures the UV/VIS, linear dichroism and magnetic circular dichorism simultaneously, incorporates a photoelastic modulator and a microscopy superconducting magnet for high-field (5T) measurements. We present solution and crystalline data on metal-complexed TPP; data are analyzed in terms of A and B-type MCD using a perimeter model. We find good agreement with previous solution data, and novel crystalline phase spectra that are correlated to the long range ordering. This work supported by NSF DMR-0821268, DUE-0942562 and EPS-0701410.

  6. Spectroscopic studies of molecular iodine emitted into the gas phase by seaweed

    NASA Astrophysics Data System (ADS)

    Ball, S. M.; Hollingsworth, A. M.; Humbles, J.; Leblanc, C.; Potin, P.; McFiggans, G.

    2009-12-01

    Time profiles of molecular iodine emissions from seven species of seaweed have been measured at high time resolution (7.5 s) by direct spectroscopic quantification of the gas phase I2 using broadband cavity enhanced absorption spectroscopy. Substantial differences were found between species, both in the amounts of I2 emitted when the plants were exposed to air and in the shapes of their emission time profiles. Two species of kelp, Laminaria digitata and Laminaria hyperborea, were found to be the most potent emitters, producing an intense burst of I2 when first exposed to air. I2 was also observed from Saccharina latissima and Ascophyllum nodosum but in lower amounts and with broader time profiles. I2 mixing ratios from two Fucus species and Dictyopteris membranacea were at or below the detection limit of the present instrument (25 pptv). A further set of experiments investigated the time dependence of I2 emissions and aerosol particle formation when fragments of L. digitata were exposed to desiccation in air, to ozone and to oligoguluronate stress factors. Particle formation occurred in all L. digitata stress experiments where ozone and light were present, subject to the I2 mixing ratios being above certain threshold amounts. Moreover, the particle number concentrations closely tracked variations in the I2 mixing ratios, confirming the results of previous studies that the condensable particle-forming gases derive from the photochemical oxidation of the plant's I2 emissions. This work also supports the theory that particle nucleation in the coastal atmosphere occurs in "hot-spot" regions of locally elevated concentrations of condensable gases: the greatest atmospheric concentrations of I2 and hence of condensable iodine oxides are likely to be above plants of the most efficiently emitting kelp species and localised in time to shortly after these seaweeds are uncovered by a receding tide.

  7. Spectroscopic studies of molecular iodine emitted into the gas phase by seaweed

    NASA Astrophysics Data System (ADS)

    Ball, S. M.; Hollingsworth, A. M.; Humbles, J.; Leblanc, C.; Potin, P.; McFiggans, G.

    2010-07-01

    Time profiles of molecular iodine emissions from seven species of seaweed have been measured at high time resolution (7.5 s) by direct spectroscopic quantification of the gas phase I2 using broadband cavity enhanced absorption spectroscopy. Substantial differences were found between species, both in the amounts of I2 emitted when the plants were exposed to air and in the shapes of their emission time profiles. Two species of kelp, Laminaria digitata and Laminaria hyperborea, were found to be the most potent emitters, producing an intense burst of I2 when first exposed to air. I2 was also observed from Saccharina latissima and Ascophyllum nodosum but in lower amounts and with broader time profiles. I2 mixing ratios from two Fucus species and Dictyopteris membranacea were at or below the detection limit of the present instrument (25 pptv). A further set of experiments investigated the time dependence of I2 emissions and aerosol particle formation when fragments of L. digitata were exposed to desiccation in air, to ozone and to oligoguluronate stress factors. Particle formation occurred in all L. digitata stress experiments where ozone and light were present, subject to the I2 mixing ratios being above certain threshold amounts. Moreover, the particle number concentrations closely tracked variations in the I2 mixing ratios, confirming the results of previous studies that the condensable particle-forming gases derive from the photochemical oxidation of the plant's I2 emissions. This work also supports the theory that particle nucleation in the coastal atmosphere occurs in "hot-spot" regions of locally elevated concentrations of condensable gases: the greatest atmospheric concentrations of I2 and hence of condensable iodine oxides are likely to be above plants of the most efficiently emitting kelp species and localised in time to shortly after these seaweeds are uncovered by a receding tide.

  8. High-temperature phase transitions, spectroscopic properties, and dimensionality reduction in rubidium thorium molybdate family.

    PubMed

    Xiao, Bin; Gesing, Thorsten M; Kegler, Philip; Modolo, Giuseppe; Bosbach, Dirk; Schlenz, Hartmut; Suleimanov, Evgeny V; Alekseev, Evgeny V

    2014-03-17

    Four new rubidium thorium molybdates have been synthesized by high-temperature solid-state reactions. The crystal structures of Rb8Th(MoO4)6, Rb2Th(MoO4)3, Rb4Th(MoO4)4, and Rb4Th5(MoO4)12 were determined using single-crystal X-ray diffraction. All these compounds construct from MoO4 tetrahedra and ThO8 square antiprisms. The studied compounds adopt the whole range of possible structure dimensionalities from zero-dimensional (0D) to three-dimensional (3D): finite clusters, chains, sheets, and frameworks. Rb8Th(MoO4)6 crystallizes in 0D containing clusters of [Th(MoO4)6](8-). The crystal structure of Rb2Th(MoO4)3 is based upon one-dimensional chains with configuration units of [Th(MoO4)3](2-). Two-dimensional sheets occur in compound Rb4Th(MoO4)4, and a 3D framework with channels formed by thorium and molybdate polyhedra has been observed in Rb4Th5(MoO4)12. The Raman and IR spectroscopic properties of these compounds are reported. Temperature-depended phase transition effects were observed in Rb2Th(MoO4)3 and Rb4Th(MoO4)4 using thermogravimetry-differential scanning calorimetry analysis and high-temperature powder diffraction methods.

  9. In situ spectroscopic studies on vapor phase catalytic decomposition of dimethyl oxalate.

    PubMed

    Hegde, Shweta; Tharpa, Kalsang; Akuri, Satyanarayana Reddy; K, Rakesh; Kumar, Ajay; Deshpande, Raj; Nair, Sreejit A

    2017-03-15

    Dimethyl Oxalate (DMO) has recently gained prominence as a valuable intermediate for the production of compounds of commercial importance. The stability of DMO is poor and hence this can result in the decomposition of DMO under reaction conditions. The mechanism of DMO decomposition is however not reported and more so on catalytic surfaces. Insights into the mechanism of decomposition would help in designing catalysts for its effective molecular transformation. It is well known that DMO is sensitive to moisture, which can also be a factor contributing to its decomposition. The present work reports the results of decomposition of DMO on various catalytic materials. The materials studied consist of acidic (γ-Al2O3), basic (MgO), weakly acidic (ZnAl2O4) and neutral surfaces such as α-Al2O3 and mesoporous precipitated SiO2. Infrared spectroscopy is used to identify the nature of adsorption of the molecule on the various surfaces. The spectroscopy study is done at a temperature of 200 °C, which is the onset of gas phase decomposition of DMO. The results indicate that the stability of DMO is lower than the corresponding acid, i.e. oxalic acid. It is also one of the products of decomposition. Spectroscopic data suggest that DMO decomposition is related to surface acidity and the extent of decomposition depends on the number of surface hydroxyl groups. Decomposition was also observed on α-Al2O3, which was attributed to the residual surface hydroxyl groups. DMO decomposition to oxalic acid was not observed on the basic surface (MgO).

  10. Laser Spectroscopic Study of Cold Gas-Phase Host-Guest Complexes of Crown Ethers.

    PubMed

    Ebata, Takayuki; Inokuchi, Yoshiya

    2016-06-01

    The structure, molecular recognition, and inclusion effect on the photophysics of guest species are investigated for neutral and ionic cold host-guest complexes of crown ethers (CEs) in the gas phase. Here, the cold neutral host-guest complexes are produced by a supersonic expansion technique and the cold ionic complexes are generated by the combination of electrospray ionization (ESI) and a cryogenically cooled ion trap. The host species are 3n-crown-n (3nCn; n = 4, 5, 6, 8) and (di)benzo-3n-crown-n ((D)B3nCn; n = 4, 5, 6, 8). For neutral guests, we have chosen water and aromatic molecules, such as phenol and benzenediols, and as ionic species we have chosen alkali-metal ions (M(+) ). The electronic spectra and isomer-specific vibrational spectra for the complexes are observed with various laser spectroscopic methods: laser-induced fluorescence (LIF); ultraviolet-ultraviolet hole-burning (UV-UV HB); and IR-UV double resonance (IR-UV DR) spectroscopy. The obtained spectra are analyzed with the aid of quantum chemical calculations. We will discuss how the host and guest species change their flexible structures for forming best-fit stable complexes (induced fitting) and what kinds of interactions are operating for the stabilization of the complexes. For the alkali metal ion•CE complexes, we investigate the solvation effect by attaching water molecules. In addition to the ground-state stabilization problem, we will show that the complexation leads to a drastic effect on the excited-state electronic structure and dynamics of the guest species, which we call a "cage-like effect".

  11. Invited Review Article: Methods for imaging weak-phase objects in electron microscopy

    SciTech Connect

    Glaeser, Robert M.

    2013-11-15

    Contrast has traditionally been produced in electron-microscopy of weak phase objects by simply defocusing the objective lens. There now is renewed interest, however, in using devices that apply a uniform quarter-wave phase shift to the scattered electrons relative to the unscattered beam, or that generate in-focus image contrast in some other way. Renewed activity in making an electron-optical equivalent of the familiar “phase-contrast” light microscope is based in part on the improved possibilities that are now available for device microfabrication. There is also a better understanding that it is important to take full advantage of contrast that can be had at low spatial frequency when imaging large, macromolecular objects. In addition, a number of conceptually new phase-plate designs have been proposed, thus increasing the number of options that are available for development. The advantages, disadvantages, and current status of each of these options is now compared and contrasted. Experimental results that are, indeed, superior to what can be accomplished with defocus-based phase contrast have been obtained recently with two different designs of phase-contrast aperture. Nevertheless, extensive work also has shown that fabrication of such devices is inconsistent, and that their working lifetime is short. The main limitation, in fact, appears to be electrostatic charging of any device that is placed into the electron diffraction pattern. The challenge in fabricating phase plates that are practical to use for routine work in electron microscopy thus may be more in the area of materials science than in the area of electron optics.

  12. Invited Review Article: Methods for imaging weak-phase objects in electron microscopy

    PubMed Central

    Glaeser, Robert M.

    2013-01-01

    Contrast has traditionally been produced in electron-microscopy of weak phase objects by simply defocusing the objective lens. There now is renewed interest, however, in using devices that apply a uniform quarter-wave phase shift to the scattered electrons relative to the unscattered beam, or that generate in-focus image contrast in some other way. Renewed activity in making an electron-optical equivalent of the familiar “phase-contrast” light microscope is based in part on the improved possibilities that are now available for device microfabrication. There is also a better understanding that it is important to take full advantage of contrast that can be had at low spatial frequency when imaging large, macromolecular objects. In addition, a number of conceptually new phase-plate designs have been proposed, thus increasing the number of options that are available for development. The advantages, disadvantages, and current status of each of these options is now compared and contrasted. Experimental results that are, indeed, superior to what can be accomplished with defocus-based phase contrast have been obtained recently with two different designs of phase-contrast aperture. Nevertheless, extensive work also has shown that fabrication of such devices is inconsistent, and that their working lifetime is short. The main limitation, in fact, appears to be electrostatic charging of any device that is placed into the electron diffraction pattern. The challenge in fabricating phase plates that are practical to use for routine work in electron microscopy thus may be more in the area of materials science than in the area of electron optics. PMID:24289381

  13. Spectral interferometric microscopy reveals absorption by individual optical nanoantennas from extinction phase

    PubMed Central

    Gennaro, Sylvain D.; Sonnefraud, Yannick; Verellen, Niels; Van Dorpe, Pol; Moshchalkov, Victor V.; Maier, Stefan A.; Oulton, Rupert F.

    2014-01-01

    Optical antennas transform light from freely propagating waves into highly localized excitations that interact strongly with matter. Unlike their radio frequency counterparts, optical antennas are nanoscopic and high frequency, making amplitude and phase measurements challenging and leaving some information hidden. Here we report a novel spectral interferometric microscopy technique to expose the amplitude and phase response of individual optical antennas across an octave of the visible to near-infrared spectrum. Although it is a far-field technique, we show that knowledge of the extinction phase allows quantitative estimation of nanoantenna absorption, which is a near-field quantity. To verify our method we characterize gold ring-disk dimers exhibiting Fano interference. Our results reveal that Fano interference only cancels a bright mode’s scattering, leaving residual extinction dominated by absorption. Spectral interference microscopy has the potential for real-time and single-shot phase and amplitude investigations of isolated quantum and classical antennas with applications across the physical and life sciences. PMID:24781663

  14. Phase contrast microscopy analysis of breast tissue: differences in benign vs. malignant epithelium and stroma.

    PubMed

    Wells, Wendy A; Wang, Xin; Daghlian, Charles P; Paulsen, Keith D; Pogue, Brian W

    2009-08-01

    To assess how optical scatter properties in breast tissue, as measured by phase contrast microscopy and interpreted pathophysiologically, might be exploited as a diagnostic tool to differentiate cancer from benign tissue. We evaluated frozen human breast tissue sections of adipose tissue, normal breast parenchyma, benign fibroadenoma tumors and noninvasive and invasive malignant cancers by phase contrast microscopy through quantification of grayscale values, using multiple regions of interest (ROI). Student's t tests were performed on phase contrast measures across diagnostic categories testing data from individual cases; all ROI data were used as separate measures. Stroma demonstrated significantly higher scatter intensity than did epithelium, with lower scattering in tumor-associated stroma as compared with normal or benign-associated stroma. Measures were comparable for invasive and noninvasive malignant tumors but were higher than those found in benign tumors and were lowest in adipose tissue. Significant differences were found in scatter coefficient properties of epithelium and stroma across diagnostic categories of breast tissue, particularly between benign and malignant-associated stroma. Improved understanding of how scatter properties correlate with morphologic criteria used in routine pathologic diagnoses could have a significant clinical impact as developing optical technology allows macroscopic in situ phase contrast imaging.

  15. Characterization of Homopolymer and Polymer Blend Films by Phase Sensitive Acoustic Microscopy

    NASA Astrophysics Data System (ADS)

    Ngwa, Wilfred; Wannemacher, Reinhold; Grill, Wolfgang

    2003-03-01

    CHARACTERIZATION OF HOMOPOLYMER AND POLYMER BLEND FILMS BY PHASE SENSITIVE ACOUSTIC MICROSCOPY W Ngwa, R Wannemacher, W Grill Institute of Experimental Physics II, University of Leipzig, 04103 Leipzig, Germany Abstract We have used phase sensitive acoustic microscopy (PSAM) to study homopolymer thin films of polystyrene (PS) and poly (methyl methacrylate) (PMMA), as well as PS/PMMA blend films. We show from our results that PSAM can be used as a complementary and highly valuable technique for elucidating the three-dimensional (3D) morphology and micromechanical properties of thin films. Three-dimensional image acquisition with vector contrast provides the basis for: complex V(z) analysis (per image pixel), 3D image processing, height profiling, and subsurface image analysis of the polymer films. Results show good agreement with previous studies. In addition, important new information on the three dimensional structure and properties of polymer films is obtained. Homopolymer film structure analysis reveals (pseudo-) dewetting by retraction of droplets, resulting in a morphology that can serve as a starting point for the analysis of polymer blend thin films. The outcome of confocal laser scanning microscopy studies, performed on the same samples are correlated with the obtained results. Advantages and limitations of PSAM are discussed.

  16. Phase microscopy using light-field reconstruction method for cell observation.

    PubMed

    Xiu, Peng; Zhou, Xin; Kuang, Cuifang; Xu, Yingke; Liu, Xu

    2015-08-01

    The refractive index (RI) distribution can serve as a natural label for undyed cell imaging. However, the majority of images obtained through quantitative phase microscopy is integrated along the illumination angle and cannot reflect additional information about the refractive map on a certain plane. Herein, a light-field reconstruction method to image the RI map within a depth of 0.2 μm is proposed. It records quantitative phase-delay images using a four-step phase shifting method in different directions and then reconstructs a similar scattered light field for the refractive sample on the focus plane. It can image the RI of samples, transparent cell samples in particular, in a manner similar to the observation of scattering characteristics. The light-field reconstruction method is therefore a powerful tool for use in cytobiology studies.

  17. Microscopy of non-birefringent transmissive phase samples using Sagnac laser interferometer.

    PubMed

    Sarkar, Sanjukta; Bhattacharya, K

    2014-11-01

    A cyclic interferometer, appropriately combined with a long working distance microscope objective, is adapted for quantitative phase microscopy. In such an arrangement, the sample information, in terms of the diffracted orders emerging from the sample, is carried by both the counter propagating beams within the cyclic interferometer. However, positioning the sample close to the input/output cube beam splitter and use of a suitably converging laser beam of light as the input to the interferometer ensure that only one of the counter propagating beams carries the object information to the objective while the other beam, which serves as the reference, allows only the undiffracted component to contribute to the process of image formation. Use of suitable polarization optics renders the interferometer its polarization phase shifting property. Using the proposed arrangement, the experimental results showing the quantitative 3D phase rendering of polystyrene microspheres and micro-wells etched in glass are presented.

  18. 2D motility tracking of Pseudomonas putida KT2440 in growth phases using video microscopy

    PubMed Central

    Davis, Michael L.; Mounteer, Leslie C.; Stevens, Lindsey K.; Miller, Charles D.; Zhou, Anhong

    2011-01-01

    Pseudomonas putida KT2440 is a gram negative motile soil bacterium important in bioremediation and biotechnology. Thus, it is important to understand its motility characteristics as individuals and in populations. Population characteristics were determined using a modified Gompertz model. Video microscopy and imaging software were utilized to analyze two dimensional (2D) bacteria movement tracks to quantify individual bacteria behavior. It was determined that inoculum density increased the lag time as seeding densities decreased, and that the maximum specific growth rate decreased as seeding densities increased. Average bacterial velocity remained relatively similar throughout exponential growth phase (~20.9 µm/sec), while maximum velocities peak early in exponential growth phase at a velocity of 51.2 µm/sec. Pseudomonas putida KT2440 also favor smaller turn angles indicating they often continue in the same direction after a change in flagella rotation throughout the exponential growth phase. PMID:21334971

  19. Measurement of nonlinear characteristics of silver-halide holographic materials by phase-contrast microscopy

    NASA Astrophysics Data System (ADS)

    Banyasz, Istvan

    2003-05-01

    Lin-curves of plane-wave phase holograms recorded in Agfa-Gevaert 8E75HD emulsions were measured for the combinations of AAC developer and the R9 bleaching agent. Then each holographic grating was studied by phase-contrast microscopy, using both medium-power (40 X) and high-power immersion (100 X) objectives. Thus, besides of the Lin-curves, the modulation of the refractive index as a function of the bias exposure and the visibility of the recording interference pattern can also been determined. This latter characteristics is used in coupled wave theory to calculate the diffraction efficiency of holographic gratings, thus the measured diffraction efficiencies can be compared to those predicted by the theory. Moreover, this direct study of the phase profile of the gratings can be used for optimising processing.

  20. Spectral-domain optical coherence phase microscopy for label-free multiplexed protein microarray assay.

    PubMed

    Joo, Chulmin; Ozkumur, Emre; Unlü, M Selim; Boer, Johannes F de

    2009-10-15

    Quantitative measurement of affinities and kinetics of various biomolecular interactions such as protein-protein, protein-DNA and receptor-ligand is central to our understanding of basic molecular and cellular functions and is useful for therapeutic evaluation. Here, we describe a laser-scanning quantitative imaging method, referred to as spectral-domain optical coherence phase microscopy, as an optical platform for label-free detection of biomolecular interactions. The instrument is based on a confocal interferometric microscope that enables depth-resolved quantitative phase measurements on sensor surface with high spatial resolution and phase stability. We demonstrate picogram per square millimeter surface mass sensitivity, and show its sensing capability by presenting static and dynamic detection of multiplexed protein microarray as immobilized antigens capture their corresponding antibodies.

  1. Phase-contrast microscopy at high x-ray energy with a laboratory setup.

    PubMed

    Endrizzi, Marco; Vittoria, Fabio A; Diemoz, Paul C; Lorenzo, Rodolfo; Speller, Robert D; Wagner, Ulrich H; Rau, Christoph; Robinson, Ian K; Olivo, Alessandro

    2014-06-01

    We report on the design and realization of an x-ray imaging system for quantitative phase-contrast microscopy at high x-ray energy with laboratory-scale instrumentation. Phase and amplitude were separated quantitatively at x-ray energies up to 80 keV with micrometric spatial resolution. The accuracy of the results was tested against numerical simulations, and the spatial resolution was experimentally quantified by measuring a Siemens star phase object. This simple setup should find broad application in those areas of x-ray imaging where high energy and spatial resolution are simultaneously required and in those difficult cases where the sample contains materials with similar x-ray absorption.

  2. Compensation of phase aberration by using a virtual confocal scheme in digital holographic microscopy.

    PubMed

    Chew, Yang-Kun; Shiu, Min-Tzung; Wang, Je-Chung; Chang, Chi-Ching

    2014-09-20

    This work presents cost-effective, simple arbitrary phase-step digital holographic microscopy to suppress both zero-order and twin-image terms. A virtual confocal offset lens under in-line configuration is also used to compensate for the introduced quadratic phase by using a microscope objective lens. In addition to reducing the difficulties of physical confocal configurations, the proposed method significantly increases the magnification power, ultimately achieving the purposes of an optical zoom. An attempt is also made to reduce the noise interference of a high magnification system by developing a long focal lens to reduce light detection size, subsequently gaining an approximately plane wave light source to illuminate the object within the effective depth of focus. Experimental results indicate that the proposed high magnification system can be elevated with low noise interference, and image reconstruction without quadratic phase terms.

  3. Superresolution Microscopy of the Volume Phase Transition of pNIPAM Microgels

    NASA Astrophysics Data System (ADS)

    Conley, Gaurasundar Marc; Nöjd, Sofi; Braibanti, Marco; Schurtenberger, Peter; Scheffold, Frank

    Hierarchical polymer structures such as pNIPAM microgels have been extensively studied for their ability to undergo significant structural and physical transformations that can be controlled by external stimuli such as temperature, pH or solvent composition. Here we discuss in-situ three-dimensional superresolution microscopy of dye-labeled submicron sized PNiPAM microgels. We use direct STochastic Optical Reconstruction Microscopy (dSTORM) to study the internal density distribution and the particle-to-particle variability of the volume phase transition. Moreover we discuss the potential of this technique towards future applications to more complex architectures for example microgel with anisotropic shape or ones that are doped or decorated with nanoparticles.

  4. Total three-dimensional imaging of phase objects using defocusing microscopy: Application to red blood cells

    NASA Astrophysics Data System (ADS)

    Roma, P. M. S.; Siman, L.; Amaral, F. T.; Agero, U.; Mesquita, O. N.

    2014-06-01

    We introduce Defocusing Microscopy (DM), a bright-field optical microscopy technique able to perform total three-dimensional (3D) imaging of transparent objects. By total 3D imaging, we mean the determination of the actual shapes of the upper and lower surfaces of a phase object. We propose a methodology using DM and apply it to red blood cells subject to different osmolality conditions: hypotonic, isotonic, and hypertonic solutions. For each situation, the shapes of the upper and lower cell surface-membranes (lipid bilayer/cytoskeleton) are completely recovered, displaying the deformation of red blood cell (RBC) surfaces due to adhesion on the glass-substrate. The axial resolution of our technique allowed us to image surface-membranes separated by distances as small as 300 nm. Finally, we determine the volume, surface area, sphericity index, and RBC refractive index for each osmotic condition.

  5. Rigid gas permeable contact lenses surface roughness examined by interferential shifting phase and scanning electron microscopies.

    PubMed

    Merindano, M D; Canals, M; Saona, C; Costa, J

    1998-01-01

    The anterior surface roughness of seven factory new rigid gas permeable (RGP) contact lenses has been studied by interferential shifting phase microscopy (ISPM) and scanning electron microscopy (SEM). Five lenses were fluorsilicone acrylate and two lenses were silicone acrylate. Their material Dk ranged from 14 to 210. ISPM is shown to be a reliable and non-destructive method to observe and measure the relief of the contact lens surface. Moreover, profile and contour data are easily stored for further quantitative studies. ISPM contour patterns of the studied lenses are qualitatively compared with those obtained by SEM for the same lenses. Results point out that ISPM gives similar accuracy but it is non-destructive and cheaper than SEM. Moreover, the quantitative study of surface roughness suggests that there is a relationship between surface roughness and Dk of the lens material: surface roughness increases with Dk and allows to distinguish between lenses with low, medium and high Dk.

  6. Use of Fluorescence Lifetime Imaging Microscopy (FLIM) as a Timer of Cell Cycle S Phase.

    PubMed

    Okkelman, Irina A; Dmitriev, Ruslan I; Foley, Tara; Papkovsky, Dmitri B

    2016-01-01

    Incorporation of thymidine analogues in replicating DNA, coupled with antibody and fluorophore staining, allows analysis of cell proliferation, but is currently limited to monolayer cultures, fixed cells and end-point assays. We describe a simple microscopy imaging method for live real-time analysis of cell proliferation, S phase progression over several division cycles, effects of anti-proliferative drugs and other applications. It is based on the prominent (~ 1.7-fold) quenching of fluorescence lifetime of a common cell-permeable nuclear stain, Hoechst 33342 upon the incorporation of 5-bromo-2'-deoxyuridine (BrdU) in genomic DNA and detection by fluorescence lifetime imaging microscopy (FLIM). We show that quantitative and accurate FLIM technique allows high-content, multi-parametric dynamic analyses, far superior to the intensity-based imaging. We demonstrate its uses with monolayer cell cultures, complex 3D tissue models of tumor cell spheroids and intestinal organoids, and in physiological study with metformin treatment.

  7. Deconstructing the Late Phase of Vimentin Assembly by Total Internal Reflection Fluorescence Microscopy (TIRFM)

    PubMed Central

    Winheim, Stefan; Hieb, Aaron R.; Silbermann, Marleen; Surmann, Eva-Maria; Wedig, Tatjana; Herrmann, Harald; Langowski, Jörg; Mücke, Norbert

    2011-01-01

    Quantitative imaging of intermediate filaments (IF) during the advanced phase of the assembly process is technically difficult, since the structures are several µm long and therefore they exceed the field of view of many electron (EM) or atomic force microscopy (AFM) techniques. Thereby quantitative studies become extremely laborious and time-consuming. To overcome these difficulties, we prepared fluorescently labeled vimentin for visualization by total internal reflection fluorescence microscopy (TIRFM). In order to investigate if the labeling influences the assembly properties of the protein, we first determined the association state of unlabeled vimentin mixed with increasing amounts of labeled vimentin under low ionic conditions by analytical ultracentrifugation. We found that bona fide tetrameric complexes were formed even when half of the vimentin was labeled. Moreover, we demonstrate by quantitative atomic force microscopy and electron microscopy that the morphology and the assembly properties of filaments were not affected when the fraction of labeled vimentin was below 10%. Using fast frame rates we observed the rapid deposition of fluorescently labeled IFs on glass supports by TIRFM in real time. By tracing their contours, we have calculated the persistence length of long immobilized vimentin IFs to 1 µm, a value that is identical to those determined for shorter unlabeled vimentin. These results indicate that the structural properties of the filaments were not affected significantly by the dye. Furthermore, in order to analyze the late elongation phase, we mixed long filaments containing either Alexa 488- or Alexa 647-labeled vimentin. The ‘patchy’ structure of the filaments obtained unambiguously showed the elongation of long IFs through direct end-to-end annealing of individual filaments. PMID:21544245

  8. Real time spectroscopic ellipsometry of the growth and phase evolution of thin film silicon-germanium:hydrogen

    NASA Astrophysics Data System (ADS)

    Podraza, Nikolas J.

    Spectroscopic ellipsometry (SE) is one of the most important characterization techniques available for accurately and precisely measuring thin films including their structural parameters such as thickness and their optical properties such as the index of refraction and extinction coefficient. The importance of SE derives from the fact that it can provide useful physical and optical characteristics of thin films and how they vary in the fabrication process. These characteristics serve as key inputs for optical modeling of complex structures in order to better understand and predict the performance of multilayer devices. In this thesis, the growth of hydrogenated silicon (Si:H) and silicon germanium alloy (Si1-xGex:H) thin films by plasma enhanced chemical vapor deposition (PECVD) has been studied by real time spectroscopic ellipsometry (RTSE). The motivation is to develop deposition phase diagrams that can provide greater insights into the optimization of amorphous silicon germanium alloys (a-Si1-xGex:H) for multijunction photovoltaics. Real time spectroscopic ellipsometry (RTSE), performed both in situ and in real time, was applied to deduce the microstructural evolution and phase diagrams of Si1-xGex:H thin films suitable for the middle and bottom intrinsic layers of triple junction photovoltaic devices. These thin film depositions were performed under several different deposition conditions involving variations in hydrogen dilution ratio (which generally forms the abscissa of the deposition phase diagram), germane flow ratio, substrate temperature, electrode configuration, and helium dilution ratio. These phase diagrams depict the bulk layer thicknesses at which structural and phase transitions occur as functions of hydrogen dilution ratio, germane flow ratio, or substrate temperature. These transitions include the amorphous-to-amorphous roughening transition [a → a], the amorphous-to-(mixed-phase amorphous-plus-microcrystalline) phase transition [a → (a

  9. Self interference digital holographic microscopy approach for inspection of technical and biological phase specimens

    NASA Astrophysics Data System (ADS)

    Kemper, Björn; Schlichthaber, Frank; Vollmer, Angelika; Ketelhut, Steffi; Przibilla, Sabine; von Bally, Gert

    2011-05-01

    Quantitative holographic phase contrast imaging enables high-resolution inspection of reflective surfaces and technical phase specimen as well as the minimally invasive analysis of living cells. However, a drawback of many experimental arrangements is the requirement for a separate reference wave which results in a phase stability decrease and the demand for a precise adjustment of the intensity ratio between object and reference wave. Thus, a self interference digital holographic microscopy (DHM) approach was explored which only requires a single object illumination wave. Due to the Michelson interferometer design of the proposed setup two wave fronts with an almost identical curvature are superimposed. This results in a nearly ideal pattern of spatial off-axis carrier fringes and a constant interferogram contrast in the hologram plane. Moreover, the hologram evaluation with spatial phase shifting reconstruction algorithms and Fourier transformation-based spatial filtering methods as well as the integration of DHM in common research microscopes is simplified. Furthermore, the use of laser light sources with a short coherence length is enabled. The applicability of the proposed self interference principle is illustrated by data from the analysis of technical and biological phase specimens. The obtained results demonstrate that the method prospects to be a versatile tool for quantitative phase contrast imaging.

  10. An improved phase shift reconstruction algorithm of fringe scanning technique for X-ray microscopy

    SciTech Connect

    Lian, S.; Yang, H.; Kudo, H.; Momose, A.; Yashiro, W.

    2015-02-15

    The X-ray phase imaging method has been applied to observe soft biological tissues, and it is possible to image the soft tissues by using the benefit of the so-called “Talbot effect” by an X-ray grating. One type of the X-ray phase imaging method was reported by combining an X-ray imaging microscope equipped by a Fresnel zone plate with a phase grating. Using the fringe scanning technique, a high-precision phase shift image could be obtained by displacing the grating step by step and measuring dozens of sample images. The number of the images was selected to reduce the error caused by the non-sinusoidal component of the Talbot self-image at the imaging plane. A larger number suppressed the error more but increased radiation exposure and required higher mechanical stability of equipment. In this paper, we analyze the approximation error of fringe scanning technique for the X-ray microscopy which uses just one grating and proposes an improved algorithm. We compute the approximation error by iteration and substitute that into the process of reconstruction of phase shift. This procedure will suppress the error even with few sample images. The results of simulation experiments show that the precision of phase shift image reconstructed by the proposed algorithm with 4 sample images is almost the same as that reconstructed by the conventional algorithm with 40 sample images. We also have succeeded in the experiment with real data.

  11. Nanoimaging and spectroscopic analysis of rubber/ZnO interfaces by energy-filtering transmission electron microscopy.

    PubMed

    Horiuchi, Shin; Dohi, Hidehiko

    2006-05-09

    Energy-filtering transmission electron microscopy (EFTEM) was employed for investigating interactions between rubber and ZnO particles in the accelerated vulcanization process. Combining elemental mapping and electron energy loss spectroscopy (EELS) by EFTEM enabled the characterization of the interfaces with spatial resolutions of less than 10 nm and with high elemental detection sensitivity. We found that a sulfur- and zinc-rich compound was generated around ZnO particles, and that product was then revealed to be ZnS-generated as a byproduct in the accelerated vulcanization process. Through this study, it is indicated that the accelerated vulcanization with ZnO does not occur uniformly in the rubber matrix; it occurs locally around ZnO particles at a higher reaction rate, implying that the rubber network structure is not uniform on the nanoscale.

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

    PubMed

    Piper, T; Piper, J

    2012-09-01

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

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

    DOE PAGES

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

    2016-02-29

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

  14. Accurate phase measurements for thick spherical objects using optical quadrature microscopy

    NASA Astrophysics Data System (ADS)

    Warger, William C., II; DiMarzio, Charles A.

    2009-02-01

    In vitro fertilization (IVF) procedures have resulted in the birth of over three million babies since 1978. Yet the live birth rate in the United States was only 34% in 2005, with 32% of the successful pregnancies resulting in multiple births. These multiple pregnancies were directly attributed to the transfer of multiple embryos to increase the probability that a single, healthy embryo was included. Current viability markers used for IVF, such as the cell number, symmetry, size, and fragmentation, are analyzed qualitatively with differential interference contrast (DIC) microscopy. However, this method is not ideal for quantitative measures beyond the 8-cell stage of development because the cells overlap and obstruct the view within and below the cluster of cells. We have developed the phase-subtraction cell-counting method that uses the combination of DIC and optical quadrature microscopy (OQM) to count the number of cells accurately in live mouse embryos beyond the 8-cell stage. We have also created a preliminary analysis to measure the cell symmetry, size, and fragmentation quantitatively by analyzing the relative dry mass from the OQM image in conjunction with the phase-subtraction count. In this paper, we will discuss the characterization of OQM with respect to measuring the phase accurately for spherical samples that are much larger than the depth of field. Once fully characterized and verified with human embryos, this methodology could provide the means for a more accurate method to score embryo viability.

  15. Temporal phase-unwrapping algorithm for dynamic interference pattern analysis in interference-contrast microscopy.

    PubMed

    van den Doel, L R; van Vliet, L J

    2001-09-01

    A temporal phase-unwrapping algorithm has been developed for the analysis of dynamic interference patterns generated with interference-contrast microscopy in micromachined picoliter vials. These vials are etched in silicon dioxide, have a typical depth of 6 mum, and are filled with a liquid sample. In this kind of microscopy, fringe patterns are observed at the air-liquid interface. These fringe patterns are caused by interference between the directly reflected part of an incident plane wave and the part of that wave that is reflected on the bottom of the vial. The optical path difference (OPD) between both parts is proportional to the distance to the reflecting bottom of the vial. Evaporation decreases the OPD at the meniscus level and causes alternating constructive and destructive interference of the incident light, resulting in an interferogram. Imaging of the space-varying OPD yields a fringe pattern in which the isophotes correspond to isoheight curves of the meniscus. When the bottom is flat, the interference pattern allows for monitoring of the meniscus as a function of time during evaporation. However, when there are objects on the bottom of the vial, the heights of these objects are observed as phase jumps in the fringes proportional to their heights. First, we present a classical electromagnetic description of interference-contrast microscopy. Second, a temporal phase-unwrapping algorithm is described that retrieves the meniscus profile from the interference pattern. Finally, this algorithm is applied to measure height differences of objects on the bottom in other micromachined vials with a precision of ~5 nm.

  16. Phase II of the Small Main-Belt Asteroid Spectroscopic Survey. A Feature-Based Taxonomy

    NASA Astrophysics Data System (ADS)

    Bus, Schelte J.; Binzel, Richard P.

    2002-07-01

    The second phase of the Small Main-belt Asteroid Spectroscopic Survey (SMASSII) produced an internally consistent set of visible-wavelength charge-coupled device (CCD) spectra for 1447 asteroids (Bus and Binzel 2002, Icarus, ). These data provide a basis for developing a new asteroid taxonomy that utilizes more of the information contained in CCD spectra. Here we construct a classification system that builds on the robust framework provided by existing asteroid taxonomies. In particular, we define three major groupings (the S-, C-, and X-complexes) that adhere to the classical definitions of the S-, C-, and X-type asteroids. A total of 26 classes are defined, based on the presence or absence of specific spectral features. Definitions and boundary parameters are provided for each class, allowing new spectral observations to be placed in this system. Of these 26 classes, 12 bear familiar single-letter designations that follow previous conventions: A, B, C, D, K, O, Q, R, S, T, V, and X. A new L-class is introduced to describe 35 objects with spectra having a steep UV slope shortward of 0.75 μm, but which are relatively flat longward of 0.75 μm. Asteroids with intermediate spectral characteristics are assigned multiletter designations: Cb, Cg, Cgh, Ch, Ld, Sa, Sk, Sl, Sq, Sr, Xc, Xe, and Xk. Members of the Cgh- and Ch-classes have spectra containing a 0.7-μm feature that is generally attributed to hydration. Although previously considered featureless, CCD observations reveal distinct features of varying strengths in the spectra of asteroids in the X-complex, thus allowing the Xc-, Xe-, and Xk-classes to be established. Most notably, the spectra of Xe-type asteroids contain an absorption feature centered near 0.49 μm that may be associated with troilite. Several new members are identified for previously unique or sparsely populated classes: 12 A-types, 3 O-types, and 3 R-types. Q-types are common within the near-Earth asteroid population but remain unobserved in

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

    PubMed Central

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

    2010-01-01

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

  18. Nanoscale nuclear architecture for cancer diagnosis by spatial-domain low-coherence quantitative phase microscopy

    NASA Astrophysics Data System (ADS)

    Wang, Pin; Bista, Rajan K.; Khalbuss, Walid E.; Qiu, Wei; Staton, Kevin D.; Zhang, Lin; Brentnall, Teresa A.; Brand, Randall E.; Liu, Yang

    2011-03-01

    Alterations in nuclear architecture are the hallmark diagnostic characteristic of cancer cells. In this work, we show that the nuclear architectural characteristics quantified by spatial-domain low-coherence quantitative phase microscopy (SL-QPM), is more sensitive for the identification of cancer cells than conventional cytopathology. We demonstrated the importance of nuclear architectural characteristics in both an animal model of intestinal carcinogenesis - APC/Min mouse model and human cytology specimens with colorectal cancer by identifying cancer from cytologically noncancerous appearing cells. The determination of nanoscale nuclear architecture using this simple and practical optical instrument is a significant advance towards cancer diagnosis.

  19. Space-bandwidth conditions for efficient phase-shifting digital holographic microscopy.

    PubMed

    Stern, Adrian; Javidi, Bahram

    2008-03-01

    Microscopy by holographic means is attractive because it permits true three-dimensional (3D) visualization and 3D display of the objects. We investigate the necessary condition on the object size and spatial bandwidth for complete 3D microscopic imaging with phase-shifting digital holography with various common arrangements. The cases for which a Fresnel holographic arrangement is sufficient and those for which object magnification is necessary are defined. Limitations set by digital sensors are analyzed in the Wigner domain. The trade-offs between the various holographic arrangements in terms of conditions on the object size and bandwidth, recording conditions required for complete representation, and complexity are discussed.

  20. Dual mode diffraction phase microscopy for quantitative functional assessment of biological cells

    NASA Astrophysics Data System (ADS)

    Talaikova, N. A.; Popov, A. P.; Kalyanov, A. L.; Ryabukho, V. P.; Meglinski, I. V.

    2017-10-01

    A diffraction phase microscopy approach with a combined use of transmission and reflection imaging modes has been developed and applied for non-invasive quantitative assessment of the refractive index of red blood cells (RBCs). We present the theoretical background of signal formation for both imaging modes, accompanied by the results of experimental studies. We demonstrate that simultaneous use of the two modes has great potential for accurate assessment of the refractive index of biological cells, and we perform a reconstruction of spatial distribution of the refractive index of RBC in 3D.

  1. Spatially resolved quantitative mapping of thermomechanical properties and phase transition temperatures using scanning probe microscopy

    DOEpatents

    Jesse, Stephen; Kalinin, Sergei V; Nikiforov, Maxim P

    2013-07-09

    An approach for the thermomechanical characterization of phase transitions in polymeric materials (polyethyleneterephthalate) by band excitation acoustic force microscopy is developed. This methodology allows the independent measurement of resonance frequency, Q factor, and oscillation amplitude of a tip-surface contact area as a function of tip temperature, from which the thermal evolution of tip-surface spring constant and mechanical dissipation can be extracted. A heating protocol maintained a constant tip-surface contact area and constant contact force, thereby allowing for reproducible measurements and quantitative extraction of material properties including temperature dependence of indentation-based elastic and loss moduli.

  2. Nanosecond switching in GeSe phase change memory films by atomic force microscopy

    SciTech Connect

    Bosse, James L.; Huey, Bryan D.; Grishin, Ilya; Kolosov, Oleg V.; Gyu Choi, Yong; Cheong, Byung-ki; Lee, Suyoun

    2014-02-03

    Nanosecond scale threshold switching is investigated with conducting atomic force microscopy (AFM) for an amorphous GeSe film. Switched bits exhibit 2–3 orders of magnitude variations in conductivity, as demonstrated in phase change based memory devices. Through the nm-scale AFM probe, this crystallization was achieved with pulse durations of as low as 15 ns, the fastest reported with scanning probe based methods. Conductance AFM imaging of the switched bits further reveals correlations between the switched volume, pulse amplitude, and pulse duration. The influence of film heterogeneities on switching is also directly detected, which is of tremendous importance for optimal device performance.

  3. Characterization of phases in ‘crud’ from boiling-water reactors by transmission electron microscopy

    NASA Astrophysics Data System (ADS)

    Janney, Dawn E.; Porter, Douglas L.

    2007-05-01

    This paper reports phases identified in samples of crud (activated corrosion products) from two commercial boiling-water reactors using transmission and analytical electron microscopy and selected-area electron diffraction. Franklinite (ZnFe 2O 4) was observed in both samples. Hematite (α-Fe 2O 3), crystalline silica (SiO 2), a fine-grained mixture of iron oxides probably including magnetite (Fe 3O 4), hematite (α-Fe 2O 3), and goethite (α-FeOOH), and an unidentified high-Ba, high-S phase were observed in one of the samples. Willemite (Zn 2SiO 4), amorphous silica, and an unidentified iron-chromium phase were observed in the other. Chloride-bearing phases were found in both samples, and are assumed to represent sample contaminants. Because of the small sample volumes and numbers of particles studied and the possibility of contamination, it is not clear whether the differences between the phases observed in the two crud samples represent actual differences in the assemblages formed in the reactors.

  4. Characterization of Phases in “Crud” from Boiling Water Reactors by Transmission Electron Microscopy

    SciTech Connect

    Dawn E. Janney; Douglas L. Porter

    2007-05-01

    This paper reports phases identified in samples of crud (activated corrosion products) from two commercial boilingwater reactors using transmission and analytical electron microscopy and selected-area electron diffraction. Franklinite (ZnFe2O4) was observed in both samples. Hematite (a-Fe2O3), crystalline silica (SiO2), a fine-grained mixture of iron oxides probably including magnetite (Fe3O4), hematite (a-Fe2O3), and goethite (a-FeOOH), and an unidentified high-Ba, high-S phase were observed in one of the samples. Willemite (Zn2SiO4), amorphous silica, and an unidentified iron– chromium phase were observed in the other. Chloride-bearing phases were found in both samples, and are assumed to represent sample contaminants. Because of the small sample volumes and numbers of particles studied and the possibility of contamination, it is not clear whether the differences between the phases observed in the two crud samples represent actual differences in the assemblages formed in the reactors.

  5. Au Colloids Formed by Ion Implantation in Muscovite Mica Studied by Vibrational and Electronic Spectroscopes and Atomic Force Microscopy

    NASA Technical Reports Server (NTRS)

    Tung, Y. S.; Henderson, D. O.; Mu, R.; Ueda, A.; Collins, W. E.; White, C. W.; Zuhr, R. A.; Zhu, Jane G.

    1997-01-01

    Au was implanted into the (001) surface of Muscovite mica at an energy of 1.1 MeV and at doses of 1, 3, 6, and 10 x 10(exp 16) ions/cu cm. Optical spectra of the as-implanted samples revealed a peak at 2.28 eV (545 nm) which is attributed to the surface plasmon absorption of Au colloids. The infrared reflectance measurements show a decreasing reflectivity with increasing ion dose in the Si-O stretching region (900-1200 /cm). A new peak observed at 967 /cm increases with the ion dose and is assigned to an Si-O dangling bond. Atomic force microscopy images of freshly cleaved samples implanted with 6 and 10 x 10(exp 16) ions/sq cm indicated metal colloids with diameters between 0.9- 1.5 nm. AFM images of the annealed samples showed irregularly shaped structures with a topology that results from the fusion of smaller colloids.

  6. Studies on the interaction of heparin with lysozyme by multi-spectroscopic techniques and atomic force microscopy

    NASA Astrophysics Data System (ADS)

    Tian, Lunfu; Hu, Xiaoli; Liu, Zhongfang; Liu, Shaopu

    2016-02-01

    The interaction between heparin (Hep) and lysozyme (Lyso) in vitro was studied by fluorescence, UV-vis, circular dichroism (CD), resonance Rayleigh scattering (RRS) spectroscopy and atomic force microscopy (AFM) under normal physiological conditions. UV-vis spectra of Lyso showed the absorbance was significantly increased with the addition of Hep. Fluorescence studies revealed that the emission quenching of Lyso with Hep was initiated by static quenching mechanism. CD spectral studies showed that Hep induced conformational changes in the secondary structure of Lyso. RRS spectra of Lyso showed the intensity of scattering was significantly increased with the addition of Hep and the enhanced RRS intensities were proportional to the concentration of Hep in a certain range. Thus, a new RRS method using Lyso as a probe could be used for the determination of Hep. The detection limit for Hep was 3.9 ng mL- 1. In addition, the shape of the complex was characterized by AFM. The possible reaction mechanism and the reasons for the enhancement of RRS intensity had been discussed through experimental results.

  7. Coherence-controlled holographic microscopy for live-cell quantitative phase imaging

    NASA Astrophysics Data System (ADS)

    Slabý, TomáÅ.¡; Křížová, Aneta; Lošt'ák, Martin; Čolláková, Jana; Jůzová, Veronika; Veselý, Pavel; Chmelík, Radim

    2015-03-01

    In this paper we present coherence-controlled holographic microscopy (CCHM) and various examples of observations of living cells including combination of CCHM with fluorescence microscopy. CCHM is a novel technique of quantitative phase imaging (QPI). It is based on grating off-axis interferometer, which is fully adapted for the use of incoherent illumination. This enables high-quality QPI free from speckles and parasitic interferences and lateral resolution of classical widefield microscopes. Label-free nature of QPI makes CCHM a useful tool for long-term observations of living cells. Moreover, coherence-gating effect induced by the use of incoherent illumination enables QPI of cells even in scattering media. Combination of CCHM with common imaging techniques brings the possibility to exploit advantages of QPI while simultaneously identifying the observed structures or processes by well-established imaging methods. We used CCHM for investigation of general parameters of cell life cycles and for research of cells reactions to different treatment. Cells were also visualized in 3D collagen gel with the use of CCHM. It was found that both the cell activity and movement of the collagen fibers can be registered. The method of CCHM in combination with fluorescence microscopy was used in order to obtain complementary information about cell morphology and identify typical morphological changes associated with different types of cell death. This combination of CCHM with common imaging technique has a potential to provide new knowledge about various processes and simultaneously their confirmation by comparison with known imaging method.

  8. Label-free imaging of developing vasculature in zebrafish with phase variance optical coherence microscopy

    NASA Astrophysics Data System (ADS)

    Chen, Yu; Fingler, Jeff; Trinh, Le A.; Fraser, Scott E.

    2016-03-01

    A phase variance optical coherence microscope (pvOCM) has been created to visualize blood flow in the vasculature of zebrafish embryos, without using exogenous labels. The pvOCM imaging system has axial and lateral resolutions of 2 μm in tissue, and imaging depth of more than 100 μm. Imaging of 2-5 days post-fertilization zebrafish embryos identified the detailed structures of somites, spinal cord, gut and notochord based on intensity contrast. Visualization of the blood flow in the aorta, veins and intersegmental vessels was achieved with phase variance contrast. The pvOCM vasculature images were confirmed with corresponding fluorescence microscopy of a zebrafish transgene that labels the vasculature with green fluorescent protein. The pvOCM images also revealed functional information of the blood flow activities that is crucial for the study of vascular development.

  9. In situ high temperature microscopy study of the surface oxidation and phase transformations in titanium alloys.

    PubMed

    Malinov, S; Sha, W; Voon, C S

    2002-09-01

    Two popular commercial titanium alloys, Ti-6Al-4V and Ti-6Al-2Sn-4Zr-2Mo-0.08Si, were used for in situ high temperature microscopy study. The experiments were performed on an optical microscope equipped with high temperature stage using both normal and florescence lights. Two kinds of experiments were performed, at continuous heating/cooling with different rates and in isothermal conditions at different temperatures. The changes taking place on the sample surface during the experiments were monitored. The morphology of the alpha ==> beta ==> alpha phase transformation was recorded at different heat treatment conditions using the effect of thermal etching. An effect of sample surface oxidation and deoxidation was observed during continuous heating. The appearance and disappearance of ordered titanium oxides Ti3O and Ti2O are discussed based on the phase equilibrium diagram. The kinetics of the surface oxidation was monitored in both isothermal and continuous cooling conditions.

  10. Lensless transport-of-intensity phase microscopy and tomography with a color LED matrix

    NASA Astrophysics Data System (ADS)

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

    2015-07-01

    We demonstrate lens-less quantitative phase microscopy and diffraction tomography based on a compact on-chip platform, using only a CMOS image sensor and a programmable color LED array. Based on multi-wavelength transport-of- intensity phase retrieval and multi-angle illumination diffraction tomography, this platform offers high quality, depth resolved images with a lateral resolution of ˜3.7μm and an axial resolution of ˜5μm, over wide large imaging FOV of 24mm2. The resolution and FOV can be further improved by using a larger image sensors with small pixels straightforwardly. This compact, low-cost, robust, portable platform with a decent imaging performance may offer a cost-effective tool for telemedicine needs, or for reducing health care costs for point-of-care diagnostics in resource-limited environments.

  11. Spectral-domain phase microscopy with improved sensitivity using two-dimensional detector arrays

    SciTech Connect

    Singh, K.; Dion, C.; Ozaki, T.; Lesk, M. R.; Costantino, S.

    2011-02-15

    In this work we demonstrate the use of two-dimensional detectors to improve the signal-to-noise ratio (SNR) and sensitivity in spectral-domain phase microscopy for subnanometer accuracy measurements. We show that an increase in SNR can be obtained, from 82 dB to 105 dB, using 150 pixel lines of a low-cost CCD camera as compared to a single line, to compute an averaged axial scan. In optimal mechanical conditions, phase stability as small as 92 {mu}rad, corresponding to 6 pm displacement accuracy, could be obtained. We also experimentally demonstrate the benefit of spatial-averaging in terms of the reduction of signal fading due to an axially moving sample. The applications of the improved system are illustrated by imaging live cells in culture.

  12. New approaches for the analysis of confluent cell layers with quantitative phase digital holographic microscopy

    NASA Astrophysics Data System (ADS)

    Pohl, L.; Kaiser, M.; Ketelhut, S.; Pereira, S.; Goycoolea, F.; Kemper, Björn

    2016-03-01

    Digital holographic microscopy (DHM) enables high resolution non-destructive inspection of technical surfaces and minimally-invasive label-free live cell imaging. However, the analysis of confluent cell layers represents a challenge as quantitative DHM phase images in this case do not provide sufficient information for image segmentation, determination of the cellular dry mass or calculation of the cell thickness. We present novel strategies for the analysis of confluent cell layers with quantitative DHM phase contrast utilizing a histogram based-evaluation procedure. The applicability of our approach is illustrated by quantification of drug induced cell morphology changes and it is shown that the method is capable to quantify reliable global morphology changes of confluent cell layers.

  13. Quantitative phase microscopy using dual-plane in-line digital holography.

    PubMed

    Das, Bhargab; Yelleswarapu, Chandra S; Rao, D V G L N

    2012-03-20

    We present detailed theoretical evaluation and thorough experimental investigation of quantitative phase imaging using our previously demonstrated dual-plane in-line digital holographic microscopy technique [Opt. Lett. 35, 3426 (2010)]. This evaluation is based on the recording of two interferograms at slightly different planes and numerically reconstructing the object information. The zero-order diffracted wave is eliminated by using the method of subtraction of average intensity of the entire hologram, and the twin-image diffracted wave is removed by Fourier domain processing of the two recorded holograms. Experiments are performed using controlled amplitude and phase objects and human muscle cells to demonstrate the potential of this technique. © 2012 Optical Society of America

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

    PubMed

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

    2013-08-10

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

  15. Dual-modality wide-field photothermal quantitative phase microscopy and depletion of cell populations

    NASA Astrophysics Data System (ADS)

    Turko, Nir A.; Barnea, Itay; Blum, Omry; Korenstein, Rafi; Shaked, Natan T.

    2015-03-01

    We review our dual-modality technique for quantitative imaging and selective depletion of populations of cells based on wide-field photothermal (PT) quantitative phase imaging and simultaneous PT cell extermination. The cells are first labeled by plasmonic gold nanoparticles, which evoke local plasmonic resonance when illuminated by light in a wavelength corresponding to their specific plasmonic resonance peak. This reaction creates changes of temperature, resulting in changes of phase. This phase changes are recorded by a quantitative phase microscope (QPM), producing specific imaging contrast, and enabling bio-labeling in phase microscopy. Using this technique, we have shown discrimination of EGFR over-expressing (EGFR+) cancer cells from EGFR under-expressing (EGFR-) cancer cells. Then, we have increased the excitation power in order to evoke greater temperatures, which caused specific cell death, all under real-time phase acquisition using QPM. Close to 100% of all EGFR+ cells were immediately exterminated when illuminated with the strong excitation beam, while all EGFR- cells survived. For the second experiment, in order to simulate a condition where circulating tumor cells (CTCs) are present in blood, we have mixed the EGFR+ cancer cells with white blood cells (WBCs) from a healthy donor. Here too, we have used QPM to observe and record the phase of the cells as they were excited for selective visualization and then exterminated. The WBCs survival rate was over 95%, while the EGFR+ survival rate was under 5%. The technique may be the basis for real-time detection and controlled treatment of CTCs.

  16. Deciphering complex, functional structures with synchrotron-based absorption and phase contrast tomographic microscopy

    NASA Astrophysics Data System (ADS)

    Stampanoni, M.; Reichold, J.; Weber, B.; Haberthür, D.; Schittny, J.; Eller, J.; Büchi, F. N.; Marone, F.

    2010-09-01

    Nowadays, thanks to the high brilliance available at modern, third generation synchrotron facilities and recent developments in detector technology, it is possible to record volumetric information at the micrometer scale within few minutes. High signal-to-noise ratio, quantitative information on very complex structures like the brain micro vessel architecture, lung airways or fuel cells can be obtained thanks to the combination of dedicated sample preparation protocols, in-situ acquisition schemes and cutting-edge imaging analysis instruments. In this work we report on recent experiments carried out at the TOMCAT beamline of the Swiss Light Source [1] where synchrotron-based tomographic microscopy has been successfully used to obtain fundamental information on preliminary models for cerebral fluid flow [2], to provide an accurate mesh for 3D finite-element simulation of the alveolar structure of the pulmonary acinus [3] and to investigate the complex functional mechanism of fuel cells [4]. Further, we introduce preliminary results on the combination of absorption and phase contrast microscopy for the visualization of high-Z nanoparticles in soft tissues, a fundamental information when designing modern drug delivery systems [5]. As an outlook we briefly discuss the new possibilities offered by high sensitivity, high resolution grating interferomtery as well as Zernike Phase contrast nanotomography [6].

  17. Broadband quantitative phase microscopy with extended field of view using off-axis interferometric multiplexing

    NASA Astrophysics Data System (ADS)

    Girshovitz, Pinhas; Frenklach, Irena; Shaked, Natan T.

    2015-11-01

    We propose a new portable imaging configuration that can double the field of view (FOV) of existing off-axis interferometric imaging setups, including broadband off-axis interferometers. This configuration is attached at the output port of the off-axis interferometer and optically creates a multiplexed interferogram on the digital camera, which is composed of two off-axis interferograms with straight fringes at orthogonal directions. Each of these interferograms contains a different FOV of the imaged sample. Due to the separation of these two FOVs in the spatial-frequency domain, they can be fully reconstructed separately, while obtaining two complex wavefronts from the sample at once. Since the optically multiplexed off-axis interferogram is recorded by the camera in a single exposure, fast dynamics can be recorded with a doubled imaging area. We used this technique for quantitative phase microscopy of biological samples with extended FOV. We demonstrate attaching the proposed module to a diffractive phase microscopy interferometer, illuminated by a broadband light source. The biological samples used for the experimental demonstrations include microscopic diatom shells, cancer cells, and flowing blood cells.

  18. Improved Method for Analysis of Airborne Asbestos Fibers Using Phase Contrast Microscopy and FTIR Spectrometry

    PubMed Central

    Azari, Mansour R.; Yazdian, Asil; Souri, Hamid; Khodakarim, Soheila; Peirovi, Habibalalah; Panahi, Davod; Kazempour, Marzieh

    2014-01-01

    Background In recent years, some studies have tried to improve Phase Contrast Microscopy (PCM) for counting asbestos fibers. Due to the lack of a universally accepted alternative method, this study aimed to improve PCM for better counting of asbestos fibers. Materials and Methods Confirmed asbestos standards were applied using a dust generator for sampling. Sampling from the dust generator was carried out according to the Occupational Safety and Health Administration (OSHA) ID-160 method and 95 samples with diverse densities were prepared to be counted using conventional and modern PCM. All samples were counted single blindly by a co-author of this study and the obtained data were analyzed by paired t-test, correlation coefficient and Bland-Altman analysis. Duplicate samples were prepared for qualitative analysis by Fourier transform infrared spectroscopy (FT-IR), scanning electron microscopy (SEM) and X ray. Results Asbestos densities on filters were in the range of less than 100 to 600 fibers/mm2. Statistically, significant differences were observed for the count density of the 95 samples counted by the two phase contrast microscopes (P<0.001). Nikon microscope demonstrated higher counts compared to conventional microscope and had a lower coefficient of variation. Samples were analyzed qualitatively using FT-IR and SEM, and the presence of asbestos fibers was confirmed. Conclusion The improved PCM and FT-IR methods presented in this study demonstrated more precise and accurate determination of personal exposure to airborne asbestos fibers and subsequent risk assessment. PMID:25713590

  19. Improved Method for Analysis of Airborne Asbestos Fibers Using Phase Contrast Microscopy and FTIR Spectrometry.

    PubMed

    Azari, Mansour R; Yazdian, Asil; Zendehdel, Rezvan; Souri, Hamid; Khodakarim, Soheila; Peirovi, Habibalalah; Panahi, Davod; Kazempour, Marzieh

    2014-01-01

    In recent years, some studies have tried to improve Phase Contrast Microscopy (PCM) for counting asbestos fibers. Due to the lack of a universally accepted alternative method, this study aimed to improve PCM for better counting of asbestos fibers. Confirmed asbestos standards were applied using a dust generator for sampling. Sampling from the dust generator was carried out according to the Occupational Safety and Health Administration (OSHA) ID-160 method and 95 samples with diverse densities were prepared to be counted using conventional and modern PCM. All samples were counted single blindly by a co-author of this study and the obtained data were analyzed by paired t-test, correlation coefficient and Bland-Altman analysis. Duplicate samples were prepared for qualitative analysis by Fourier transform infrared spectroscopy (FT-IR), scanning electron microscopy (SEM) and X ray. Asbestos densities on filters were in the range of less than 100 to 600 fibers/mm(2). Statistically, significant differences were observed for the count density of the 95 samples counted by the two phase contrast microscopes (P<0.001). Nikon microscope demonstrated higher counts compared to conventional microscope and had a lower coefficient of variation. Samples were analyzed qualitatively using FT-IR and SEM, and the presence of asbestos fibers was confirmed. The improved PCM and FT-IR methods presented in this study demonstrated more precise and accurate determination of personal exposure to airborne asbestos fibers and subsequent risk assessment.

  20. Label-free characterization of living human induced pluripotent stem cells by subcellular topographic imaging technique using full-field quantitative phase microscopy coupled with interference reflection microscopy.

    PubMed

    Sugiyama, Norikazu; Asai, Yasuyuki; Yamauchi, Toyohiko; Kataoka, Takuji; Ikeda, Takahiro; Iwai, Hidenao; Sakurai, Takashi; Mizuguchi, Yoshinori

    2012-09-01

    There is a need for a noninvasive technique to monitor living pluripotent stem cell condition without any labeling. We present an optical imaging technique that is able to capture information about optical path difference through the cell and cell adhesion properties simultaneously using a combination of quantitative phase microscopy (QPM) and interference reflection microscopy (IRM) techniques. As a novel application of QPM and IRM, this multimodal imaging technique demonstrated its ability to distinguish the undifferentiated status of human induced pluripotent stem (hiPS) cells quantitatively based on the variation of optical path difference between the nucleus and cytoplasm as well as hiPS cell-specific cell adhesion properties.

  1. Label-free characterization of living human induced pluripotent stem cells by subcellular topographic imaging technique using full-field quantitative phase microscopy coupled with interference reflection microscopy

    PubMed Central

    Sugiyama, Norikazu; Asai, Yasuyuki; Yamauchi, Toyohiko; Kataoka, Takuji; Ikeda, Takahiro; Iwai, Hidenao; Sakurai, Takashi; Mizuguchi, Yoshinori

    2012-01-01

    There is a need for a noninvasive technique to monitor living pluripotent stem cell condition without any labeling. We present an optical imaging technique that is able to capture information about optical path difference through the cell and cell adhesion properties simultaneously using a combination of quantitative phase microscopy (QPM) and interference reflection microscopy (IRM) techniques. As a novel application of QPM and IRM, this multimodal imaging technique demonstrated its ability to distinguish the undifferentiated status of human induced pluripotent stem (hiPS) cells quantitatively based on the variation of optical path difference between the nucleus and cytoplasm as well as hiPS cell-specific cell adhesion properties. PMID:23024911

  2. Sparsity-based multi-height phase recovery in holographic microscopy

    NASA Astrophysics Data System (ADS)

    Rivenson, Yair; Wu, Yichen; Wang, Hongda; Zhang, Yibo; Feizi, Alborz; Ozcan, Aydogan

    2016-11-01

    High-resolution imaging of densely connected samples such as pathology slides using digital in-line holographic microscopy requires the acquisition of several holograms, e.g., at >6–8 different sample-to-sensor distances, to achieve robust phase recovery and coherent imaging of specimen. Reducing the number of these holographic measurements would normally result in reconstruction artifacts and loss of image quality, which would be detrimental especially for biomedical and diagnostics-related applications. Inspired by the fact that most natural images are sparse in some domain, here we introduce a sparsity-based phase reconstruction technique implemented in wavelet domain to achieve at least 2-fold reduction in the number of holographic measurements for coherent imaging of densely connected samples with minimal impact on the reconstructed image quality, quantified using a structural similarity index. We demonstrated the success of this approach by imaging Papanicolaou smears and breast cancer tissue slides over a large field-of-view of ~20 mm2 using 2 in-line holograms that are acquired at different sample-to-sensor distances and processed using sparsity-based multi-height phase recovery. This new phase recovery approach that makes use of sparsity can also be extended to other coherent imaging schemes, involving e.g., multiple illumination angles or wavelengths to increase the throughput and speed of coherent imaging.

  3. Sparsity-based multi-height phase recovery in holographic microscopy

    PubMed Central

    Rivenson, Yair; Wu, Yichen; Wang, Hongda; Zhang, Yibo; Feizi, Alborz; Ozcan, Aydogan

    2016-01-01

    High-resolution imaging of densely connected samples such as pathology slides using digital in-line holographic microscopy requires the acquisition of several holograms, e.g., at >6–8 different sample-to-sensor distances, to achieve robust phase recovery and coherent imaging of specimen. Reducing the number of these holographic measurements would normally result in reconstruction artifacts and loss of image quality, which would be detrimental especially for biomedical and diagnostics-related applications. Inspired by the fact that most natural images are sparse in some domain, here we introduce a sparsity-based phase reconstruction technique implemented in wavelet domain to achieve at least 2-fold reduction in the number of holographic measurements for coherent imaging of densely connected samples with minimal impact on the reconstructed image quality, quantified using a structural similarity index. We demonstrated the success of this approach by imaging Papanicolaou smears and breast cancer tissue slides over a large field-of-view of ~20 mm2 using 2 in-line holograms that are acquired at different sample-to-sensor distances and processed using sparsity-based multi-height phase recovery. This new phase recovery approach that makes use of sparsity can also be extended to other coherent imaging schemes, involving e.g., multiple illumination angles or wavelengths to increase the throughput and speed of coherent imaging. PMID:27901048

  4. Automatic phase aberration compensation for digital holographic microscopy based on deep learning background detection.

    PubMed

    Nguyen, Thanh; Bui, Vy; Lam, Van; Raub, Christopher B; Chang, Lin-Ching; Nehmetallah, George

    2017-06-26

    We propose a fully automatic technique to obtain aberration free quantitative phase imaging in digital holographic microscopy (DHM) based on deep learning. The traditional DHM solves the phase aberration compensation problem by manually detecting the background for quantitative measurement. This would be a drawback in real time implementation and for dynamic processes such as cell migration phenomena. A recent automatic aberration compensation approach using principle component analysis (PCA) in DHM avoids human intervention regardless of the cells' motion. However, it corrects spherical/elliptical aberration only and disregards the higher order aberrations. Traditional image segmentation techniques can be employed to spatially detect cell locations. Ideally, automatic image segmentation techniques make real time measurement possible. However, existing automatic unsupervised segmentation techniques have poor performance when applied to DHM phase images because of aberrations and speckle noise. In this paper, we propose a novel method that combines a supervised deep learning technique with convolutional neural network (CNN) and Zernike polynomial fitting (ZPF). The deep learning CNN is implemented to perform automatic background region detection that allows for ZPF to compute the self-conjugated phase to compensate for most aberrations.

  5. Early responses of human cancer cells upon photodynamic treatment monitored by laser phase microscopy

    NASA Astrophysics Data System (ADS)

    Roelofs, Theo A.; Graschew, Georgi; Perevedentseva, Elena V.; Rakowsky, Stefan; Dressler, Cathrin; Beuthan, Juergen; Schlag, Peter M.

    2001-04-01

    Photodynamic treatment of cancer cells is known to eventually cause cell death in most cases. The precise pathways and the time course seem to vary among different cell types and modes of photodynamic treatment. In this contribution, the focus was put on the responses of human colon carcinoma cells HCT-116 within the first 15 minutes after laser irradiation in the presence of Photofrin« II (PII). To monitor the cell response in this early time period laser phase microscopic imaging was used, a method sensitive to changes in overall cell shape and intracellular structures, mediated by changes in the local refractive index. Laser irradiation of cells loaded with PII induced a significant reduction of the phase shifts, which probably reflects the induced damage to the different cellular membrane structures. The data suggest that even within the first 30 s after the onset of laser illumination, a significant reduction of the phase shifts can be detected. These results underline that laser phase microscopy is a suitable diagnostic tool for cellular research, also in the early time domain.

  6. Sinter-free phase conversion and scanning transmission electron microscopy of FePt nanoparticle monolayers.

    PubMed

    Johnston-Peck, Aaron C; Scarel, Giovanna; Wang, Junwei; Parsons, Gregory N; Tracy, Joseph B

    2011-10-05

    Thermally robust monolayers of 4-6 nm diameter FePt nanoparticles (NPs) were fabricated by combining chemical synthesis and atomic layer deposition. Spin-cast monolayers of FePt NPs were coated with thin, 11 nm-thick layers of amorphous Al(2)O(3), followed by annealing to convert the FePt NPs from an alloy (A1) into intermetallic FePt (L1(0)) and FePt(3) (L1(2)) phases. The Al(2)O(3) layer serves as a barrier that prevents sintering between NPs during annealing at temperatures up to 730 °C. Electron and X-ray diffraction in conjunction with high-angle annular dark-field scanning transmission electron microscopy (HAADF-STEM) show that as-synthesized A1 FePt NPs convert into L1(0) and L1(2) phase NPs through annealing. HAADF-STEM measurements of individual NPs reveal imperfect ordering and show that the NP composition determines which intermetallic phase is obtained. Mixed-phase NPs with L1(0) cores and FePt(3) L1(2) shells were also observed, as well as a smaller number of unconverted A1 NPs. These results highlight the need for improved control over the compositional uniformity of FePt NPs for their use in bit-patterned magnetic recording.

  7. Keto-polymethines: a versatile class of dyes with outstanding spectroscopic properties for in cellulo and in vivo two-photon microscopy imaging.

    PubMed

    Pascal, Simon; Denis-Quanquin, Sandrine; Appaix, Florence; Duperray, Alain; Grichine, Alexei; Le Guennic, Boris; Jacquemin, Denis; Cuny, Jérôme; Chi, San-Hui; Perry, Joseph W; van der Sanden, Boudewijn; Monnereau, Cyrille; Andraud, Chantal; Maury, Olivier

    2017-01-01

    The synthesis of keto-heptamethine derivatives has been expanded to various new symmetrical and asymmetrical structures, including an unprecedented di-anionic keto-polymethine. The spectroscopic behavior of these new dyes has been systematically and thoroughly investigated, revealing that the formation of hydrogen bond interactions with protic solvents is responsible for a dramatic enhancement of the fluorescence quantum yield in the far-red spectral region. The existence of these strong hydrogen-bond interactions was further confirmed by molecular dynamics simulations. These bis-dipolar polymethines exhibit large two-photon absorption (TPA) cross-sections (σ(2) in GM) in the near-infrared, making them ideal candidates for NIR-to-NIR two-photon microscopy imaging applications. We demonstrate that the molecular engineering of the hydrophilic/hydrophobic balance enables targeting of different cellular components, such as cytoplasm or cell membranes. Addition of appropriate substituents provides the molecule with high-water-solubility, affording efficient two-photon probes for angiography.

  8. Study on the interaction between pelargonidin-3-O-glucoside and bovine serum albumin using spectroscopic, transmission electron microscopy and molecular modeling techniques.

    PubMed

    Li, Shu; Tang, Lin; Bi, Hongna

    2016-03-01

    The aim of this study is to evaluate the binding behavior between pelargonidin-3-O-glucoside (P3G) and bovine serum albumin (BSA) using multi-spectroscopic, transmission electron microscopy (TEM) and molecular docking methods under physiological conditions. Fluorescence spectroscopy and time-resolved fluorescence showed that the fluorescence of BSA could be quenched remarkably by P3G via a static quenching mechanism, and there is a single class of binding site on BSA. In addition, the thermodynamic functions ΔH and ΔS were -21.69 kJ/mol and 24.46 J/mol/K, indicating that an electrostatic interaction was a main acting force. The distance between BSA and P3G was 2.74 nm according to Förster's theory, illustrating that energy transfer occurred. In addition, the secondary structure of BSA changed with a decrease in the α-helix content from 66.2% to 64.0% as seen using synchronous fluorescence, UV/vis, circular dichroism and Fourier transform infrared spectroscopies, whereas TEM images showed that P3G led to BSA aggregation and fibrillation. Furthermore, site marker competitive experiments and molecular docking indicated that P3G could bind with subdomain IIA of BSA. The calculated results of the equilibrium fraction showed that the concentration of free P3G in plasma was high enough to be stored and transported from the circulatory system to its target sites to provide therapeutic effects.

  9. Spectroscopic evidence of α-methylbenzyl radical in the gas phase

    NASA Astrophysics Data System (ADS)

    Lee, Gi Woo; Ahn, Hyeon Geun; Kim, Tae Kyu; Lee, Sang Kuk

    2008-11-01

    We report the observation of the spectroscopic evidence of the α-methylbenzyl radical in a corona excited supersonic expansion using a pinhole-type glass nozzle for the first time. The precursors, toluene, ethylbenzene, and isopropylbenzene, seeded in a large amount of inert carrier gas helium, were electrically discharged to produce benzyl-type radicals as a result of the breaking off of a C-H or a C-C bond from the alkyl chain. The vibronic emission spectra, obtained in the visible region from the precursors, were compared to identify the species generated in the corona discharge of the precursors, from which we found the spectroscopic evidence of the α-methylbenzyl radical.

  10. Phase states of water near the surface of a polymer membrane. Phase microscopy and luminescence spectroscopy experiments

    SciTech Connect

    Bunkin, N. F.; Gorelik, V. S.; Kozlov, V. A. Shkirin, A. V. Suyazov, N. V.

    2014-11-15

    Phase microscopy is used to show that the refractive index in the near-surface layer of water at the surface of a polymer Nafion membrane increases by a factor of 1.1 as compared to bulk water. Moreover, this layer exhibits birefringence. Experiments on UV irradiation of dry (anhydrous) and water-soaked Nafion are performed in grazing-incidence geometry to study their stimulated luminescence spectra. These spectra are found to be identical in both cases. For dry Nafion, luminescence can only be excited if probing radiation illuminates the polymer surface. The luminescence of water-soaked Nafion can also be excited if the distance between the optical axis and the surface is several hundred micrometers.

  11. Spectroscopic descriptors for dynamic changes of soluble microbial products from activated sludge at different biomass growth phases under prolonged starvation.

    PubMed

    Maqbool, Tahir; Cho, Jinwoo; Hur, Jin

    2017-10-15

    In this study, the spectroscopic indices of soluble microbial products (SMP) were explored using absorption and fluorescence spectroscopy to identify different distinctive biomass growth phases (i.e., exponential phase, pseudo-endogenous phase, and endogenous phase) and to describe the microbial activity of activated sludge in a batch type bioreactor under prolonged starvation. The optical descriptors, including UV absorption at 254 nm (UVA254), spectral slope, absorbance slope index (ASI), biological index (BIX), humification index (HIX), and the ratio of tryptophan-like to humic-like components (C1/C2), were examined to describe the dynamic changes in SMP. These indices were mostly associated with dissolved organic carbon (DOC) of SMPs and specific oxygen uptake rate (SOUR). Among those, ASI was the most strongly correlated with the SOUR data for the pseudo-endogenous and the endogenous periods. Although the three microbial phases were well discriminated using the spectral slope, BIX, and the C1/C2 ratio, the C1/C2 ratio can be suggested as the most preferable indicator as it can also trace the changes of the relative abundance of proteins to humic-like substances in SMPs. The suggested spectroscopic descriptors were reasonably explained by the general trends of decreased large-sized biopolymer fractions (e.g., proteins) and increased humic substrates (HS) with starvation time, which were detected by size exclusion chromatography. This study provides a novel insight into the strong potential of using optical descriptors to easily probe microbial status in biological treatment systems. Copyright © 2017 Elsevier Ltd. All rights reserved.

  12. Hexatic and blue phases in a chiral liquid crystal: optical polarizing microscopy, synchrotron radiation and dielectric study

    NASA Astrophysics Data System (ADS)

    Sinha, Debashis; Debnath, Asim; Mandal, Pradip Kumar

    2014-09-01

    Phase behavior, structure and molecular dynamics of a chiral liquid crystalline compound, which exhibits SmG*, SmJ*, SmF*, SmI*, SmC*, SmA*, N* and BP*, have been investigated. Observed optical textures, synchrotron radiation diffraction data and frequency dependent dielectric spectroscopic study clearly depict the temperature evolution of the different hexatic smectic phases along with cholesteric and blue phase in a single compound. In hexatic phases dielectric absorption spectra show one low frequency relaxation process, related to the phase fluctuation of the bond orientational order, and one high frequency process related to amplitude fluctuation of the bond orientational order coupled with the polarization and tilt of the molecules. Goldstone and soft mode relaxation processes are detected, respectively, in SmC* and SmA* phases.

  13. Towards non-invasive 3D hepatotoxicity assays with optical coherence phase microscopy

    NASA Astrophysics Data System (ADS)

    Nelson, Leonard J.; Koulovasilopoulos, Andreas; Treskes, Philipp; Hayes, Peter C.; Plevris, John N.; Bagnaninchi, Pierre O.

    2015-03-01

    Three-dimensional tissue-engineered models are increasingly recognised as more physiologically-relevant than standard 2D cell culture for pre-clinical drug toxicity testing. However, many types of conventional toxicity assays are incompatible with dense 3D tissues. This study investigated the use of optical coherence phase microscopy (OCPM) as a novel approach to assess cell death in 3D tissue culture. For 3D micro-spheroid formation Human hepatic C3A cells were encapsulated in hyaluronic acid gels and cultured in 100μl MEME/10%FBS in 96-well plates. After spheroid formation the 3D liver constructs were exposed to acetaminophen on culture day 8. Acetaminophen hepatotoxicity in 3D cultures was evaluated using standard biochemical assays. An inverted OCPM in common path configuration was developed with a Callisto OCT engine (Thorlabs), centred at 930nm and a custom scanning head. Intensity data were used to perform in-depth microstructural imaging. In addition, phase fluctuations were measured by collecting several successive B scans at the same location, and statistics on the first time derivative of the phase, i.e. time fluctuations, were analysed over the acquisition time interval to retrieve overall cell viability. OCPM intensity (cell cluster size) and phase fluctuation statistics were directly compared with biochemical assays. In this study, we investigated optical coherence phase tomography to assess cell death in a 3d liver model after exposure to a prototypical hepatotoxin, acetaminophen. We showed that OCPM has the potential to assess noninvasively and label-free drug toxicity in 3D tissue models.

  14. Video-rate processing in tomographic phase microscopy of biological cells using CUDA.

    PubMed

    Dardikman, Gili; Habaza, Mor; Waller, Laura; Shaked, Natan T

    2016-05-30

    We suggest a new implementation for rapid reconstruction of three-dimensional (3-D) refractive index (RI) maps of biological cells acquired by tomographic phase microscopy (TPM). The TPM computational reconstruction process is extremely time consuming, making the analysis of large data sets unreasonably slow and the real-time 3-D visualization of the results impossible. Our implementation uses new phase extraction, phase unwrapping and Fourier slice algorithms, suitable for efficient CPU or GPU implementations. The experimental setup includes an external off-axis interferometric module connected to an inverted microscope illuminated coherently. We used single cell rotation by micro-manipulation to obtain interferometric projections from 73 viewing angles over a 180° angular range. Our parallel algorithms were implemented using Nvidia's CUDA C platform, running on Nvidia's Tesla K20c GPU. This implementation yields, for the first time to our knowledge, a 3-D reconstruction rate higher than video rate of 25 frames per second for 256 × 256-pixel interferograms with 73 different projection angles (64 × 64 × 64 output). This allows us to calculate additional cellular parameters, while still processing faster than video rate. This technique is expected to find uses for real-time 3-D cell visualization and processing, while yielding fast feedback for medical diagnosis and cell sorting.

  15. High throughput imaging of blood smears using white light diffraction phase microscopy

    NASA Astrophysics Data System (ADS)

    Majeed, Hassaan; Kandel, Mikhail E.; Bhaduri, Basanta; Han, Kevin; Luo, Zelun; Tangella, Krishnarao; Popescu, Gabriel

    2015-03-01

    While automated blood cell counters have made great progress in detecting abnormalities in blood, the lack of specificity for a particular disease, limited information on single cell morphology and intrinsic uncertainly due to high throughput in these instruments often necessitates detailed inspection in the form of a peripheral blood smear. Such tests are relatively time consuming and frequently rely on medical professionals tally counting specific cell types. These assays rely on the contrast generated by chemical stains, with the signal intensity strongly related to staining and preparation techniques, frustrating machine learning algorithms that require consistent quantities to denote the features in question. Instead we opt to use quantitative phase imaging, understanding that the resulting image is entirely due to the structure (intrinsic contrast) rather than the complex interplay of stain and sample. We present here our first steps to automate peripheral blood smear scanning, in particular a method to generate the quantitative phase image of an entire blood smear at high throughput using white light diffraction phase microscopy (wDPM), a single shot and common path interferometric imaging technique.

  16. Dynamic analysis of pathogen-infected host cells using quantitative phase microscopy

    NASA Astrophysics Data System (ADS)

    Lee, Seungrag; Kim, Young Ran; Lee, Ji Yong; Rhee, Joon Haeng; Park, Chang-Soo; Kim, Dug Young

    2011-03-01

    We present the real-time quantitative analysis of Vibrio vulnificus-infected host cells using quantitative phase microscopy (QPM) based on interferometric techniques. This provides the ability to retrieve the phase or optical path-length distribution over the cell with nanometer path-length sensitivity from a single interferogram image. We have used QPM to study dynamic cell morphologic changes and to noninvasively quantify the cell volumes of rat basophilic leukemia RBL-2H3 cells infected with V. vulnificus strains: wild type (MO6-24/O) and RtxA1 toxin mutant (CMM770). During the process of V. vulnificus infection in RBL-2H3 cells, the dynamic changes of quantitative phase images, cell volumes, and areas were observed in real time using QPM. In contrast, dramatic changes were not detected in RBL-2H3 cells infected with the noncytotoxic RtxA1 toxin mutant. The results showed good correlation between QPM analysis and biochemical assays, such as lactate dehydrogenase assay or β-hexosaminidase release assay. We suggest that QPM is a powerful quantitative method to study the dynamic process of host cells infected with pathogens in a noninvasive manner.

  17. Thin-Film Phase Plates for Transmission Electron Microscopy Fabricated from Metallic Glasses.

    PubMed

    Dries, Manuel; Hettler, Simon; Schulze, Tina; Send, Winfried; Müller, Erich; Schneider, Reinhard; Gerthsen, Dagmar; Luo, Yuansu; Samwer, Konrad

    2016-10-01

    Thin-film phase plates (PPs) have become an interesting tool to enhance the contrast of weak-phase objects in transmission electron microscopy (TEM). The thin film usually consists of amorphous carbon, which suffers from quick degeneration under the intense electron-beam illumination. Recent investigations have focused on the search for alternative materials with an improved material stability. This work presents thin-film PPs fabricated from metallic glass alloys, which are characterized by a high electrical conductivity and an amorphous structure. Thin films of the zirconium-based alloy Zr65.0Al7.5Cu27.5 (ZAC) were fabricated and their phase-shifting properties were evaluated. The ZAC film was investigated by different TEM techniques, which reveal beneficial properties compared with amorphous carbon PPs. Particularly favorable is the small probability for inelastic plasmon scattering, which results from the combined effect of a moderate inelastic mean free path and a reduced film thickness due to a high mean inner potential. Small probability plasmon scattering improves contrast transfer at high spatial frequencies, which makes the ZAC alloy a promising material for PP fabrication.

  18. Spatial light modulator phase mask implementation of wavefront encoded 3D computational-optical microscopy.

    PubMed

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

    2015-10-10

    Spatial light modulator (SLM) implementation of wavefront encoding enables various types of engineered point-spread functions (PSFs), including the generalized-cubic and squared-cubic phase mask wavefront encoded (WFE) PSFs, shown to reduce the impact of sample-induced spherical aberration in fluorescence microscopy. This investigation validates dynamic experimental parameter variation of these WFE-PSFs. We find that particular design parameter bounds exist, within which the divergence of computed and experimental WFE-PSFs is of the same order of magnitude as that of computed and experimental conventional PSFs, such that model-based approaches for solving the inverse imaging problem can be applied to a wide range of SLM-WFE systems. Interferometric measurements were obtained to evaluate the SLM implementation of the numeric mask. Agreement between experiment and theory in terms of a wrapped phase, 0-2π, validates the phase mask implementation and allows characterization of the SLM response. These measurements substantiate experimental practice of computational-optical microscope imaging with an SLM-engineered PSF.

  19. X-ray microscopy using two phase contrast imaging techniques: two dimensional grating interferometry and speckle tracking

    NASA Astrophysics Data System (ADS)

    Wang, Hongchang; Berujon, Sebastien; Pape, Ian; Sawhney, Kawal

    2013-10-01

    Two phase contrast imaging techniques, namely two dimensional grating interferometry and X-ray speckle tracking (XST), have been combined with the use of a Fresnel Zone Plate (FZP) for application to X-ray microscopy. Both techniques allows the phase shift introduced by a sample on a hard X-ray beam in two dimensions, to be recovered with a high sensitivity and low requirements on transverse and longitudinal coherence. Sub-micron phase imaging of carbon fibres was achieved using the two methods thanks to the high magnification ratio of the FZP. Advantages, drawbacks and differences between these two techniques for X-ray microscopy are discussed.

  20. Transport-of-intensity approach to differential interference contrast (TI-DIC) microscopy for quantitative phase imaging.

    PubMed

    Kou, Shan Shan; Waller, Laura; Barbastathis, George; Sheppard, Colin J R

    2010-02-01

    Differential interference contrast (DIC) microscopy is an inherently qualitative phase-imaging technique. What is obtained is an image with mixed phase-gradient and amplitude information rather than a true linear mapping of actual optical path length (OPL) differences. Here we investigate an approach that combines the transport-of-intensity equation (TIE) with DIC microscopy, thus improving direct visual observation. There is little hardware modification and the computation is noniterative. Numerically solving for the propagation of light in a series of through-focus DIC images allows linear phase information in a single slice to be completely determined and restored from DIC intensity values.

  1. Real-space observation of nanoscale magnetic phase separation in dysprosium by aberration-corrected Lorentz microscopy

    NASA Astrophysics Data System (ADS)

    Nagai, Takuro; Kimoto, Koji; Inoke, Koji; Takeguchi, Masaki

    2017-09-01

    Magnetic phase separation in single-crystal dysprosium at 97-187 K was investigated using aberration-corrected Lorentz microscopy. The high-resolution Lorentz microscopy combined with the transport-of-intensity equation method successfully visualized the in-plane magnetization distribution of the coexisting magnetic phases. The onset of a phase transition from the ferromagnetic (FM) phase to helical antiferromagnetic (HAFM) phase was observed at ˜100 K , and the two nanoscale phases coexisted up to ˜140 K . The volume fraction of the FM phase decreased with increasing temperature, eventually resulting in the formation of static magnetic solitons, which are isolated single domains of the FM phase, at around 130 K. We also performed the in situ observation of the HAFM phase at 142 K by applying an external magnetic field normal to the helical axis. With increasing field, a distorted HAFM phase emerged and the nanoscale phase separation between the HAFM phase and the fan phase subsequently occurred from ˜6 to ˜11 kOe . It was proven that the boundaries between these nanoscale coexisting phases were perpendicular to the z axis, which is the rotation axis common to the modulated magnetic structures.

  2. Common-path phase-shifting digital holographic microscopy: A way to quantitative phase imaging and superresolution

    NASA Astrophysics Data System (ADS)

    Mico, Vicente; Zalevsky, Zeev; García, Javier

    2008-09-01

    We present an experimental setup useful for complex amplitude evaluation and phase image quantification of three-dimensional (3-D) samples in digital holographic microscopy (DHM). It is based on a common-path interferometric configuration performed by dividing the input plane in two contiguous regions and by placing a translation grating near to the Fourier plane. Then, complex amplitude distribution of the sample under test is recovered with phase-shifting standard method obtained by moving the grating using a linear motion stage. Some experimental results of an USAF resolution test are presented for different numerical aperture (NA) microscope lenses. In a second part, the proposed setup is tested under superresolution purposes. Based on the object's spectrum shift produced by off-axis illumination, we use time multiplexing to generate a synthetic aperture enlargement that improves the final image resolution. Experimental results for the case of a biosample (human red blood cells) and a commercial low NA microscope lens validates the suggested superresolution approach.

  3. Evaluation of Chemical Interactions between Small Molecules in the Gas Phase Using Chemical Force Microscopy.

    PubMed

    Lee, Jieun; Ju, Soomi; Kim, In Tae; Jung, Sun-Hwa; Min, Sun-Joon; Kim, Chulki; Sim, Sang Jun; Kim, Sang Kyung

    2015-12-04

    Chemical force microscopy analyzes the interactions between various chemical/biochemical moieties in situ. In this work we examined force-distance curves and lateral force to measure the interaction between modified AFM tips and differently functionalized molecular monolayers. Especially for the measurements in gas phase, we investigated the effect of humidity on the analysis of force-distance curves and the images in lateral force mode. Flat chemical patterns composed of different functional groups were made through micro-contact printing and lateral force mode provided more resolved analysis of the chemical patterns. From the images of 1-octadecanethiol/11-mercapto-1-undecanoic acid patterns, the amine group functionalized tip brought out higher contrast of the patterns than an intact silicon nitride tip owing to the additional chemical interaction between carboxyl and amine groups. For more complex chemical interactions, relative chemical affinities toward specific peptides were assessed on the pattern of 1-octadecanethiol/phenyl-terminated alkanethiol. The lateral image of chemical force microscopy reflected specific preference of a peptide to phenyl group as well as the hydrophobic interaction.

  4. Ultrahigh-speed, phase-sensitive full-field interferometric confocal microscopy for quantitative microscale physiology

    PubMed Central

    Sencan, Ikbal; Huang, Brendan K.; Bian, Yong; Mis, Emily; Khokha, Mustafa K.; Cao, Hui; Choma, Michael

    2016-01-01

    We developed ultra-high-speed, phase-sensitive, full-field reflection interferometric confocal microscopy (FFICM) for the quantitative characterization of in vivo microscale biological motions and flows. We demonstrated 2D frame rates in excess of 1 kHz and pixel throughput rates up to 125 MHz. These fast FFICM frame rates were enabled by the use of a low spatial coherence, high-power laser source. Specifically, we used a dense vertical cavity surface emitting laser (VCSEL) array that synthesized low spatial coherence light through a large number of narrowband, mutually-incoherent emitters. Off-axis interferometry enabled single-shot acquisition of the complex-valued interferometric signal. We characterized the system performance (~2 μm lateral resolution, ~8 μm axial gating depth) with a well-known target. We also demonstrated the use of this highly parallelized confocal microscopy platform for visualization and quantification of cilia-driven surface flows and cilia beat frequency in an important animal model (Xenopus embryos) with >1 kHz frame rate. Such frame rates are needed to see large changes in local flow velocity over small distance (high shear flow), in this case, local flow around a single ciliated cell. More generally, our results are an important demonstration of low-spatial coherence, high-power lasers in high-performance, quantitative biomedical imaging. PMID:27896006

  5. Evaluation of Chemical Interactions between Small Molecules in the Gas Phase Using Chemical Force Microscopy

    PubMed Central

    Lee, Jieun; Ju, Soomi; Kim, In Tae; Jung, Sun-Hwa; Min, Sun-Joon; Kim, Chulki; Sim, Sang Jun; Kim, Sang Kyung

    2015-01-01

    Chemical force microscopy analyzes the interactions between various chemical/biochemical moieties in situ. In this work we examined force-distance curves and lateral force to measure the interaction between modified AFM tips and differently functionalized molecular monolayers. Especially for the measurements in gas phase, we investigated the effect of humidity on the analysis of force-distance curves and the images in lateral force mode. Flat chemical patterns composed of different functional groups were made through micro-contact printing and lateral force mode provided more resolved analysis of the chemical patterns. From the images of 1-octadecanethiol/11-mercapto-1-undecanoic acid patterns, the amine group functionalized tip brought out higher contrast of the patterns than an intact silicon nitride tip owing to the additional chemical interaction between carboxyl and amine groups. For more complex chemical interactions, relative chemical affinities toward specific peptides were assessed on the pattern of 1-octadecanethiol/phenyl-terminated alkanethiol. The lateral image of chemical force microscopy reflected specific preference of a peptide to phenyl group as well as the hydrophobic interaction. PMID:26690165

  6. Use of Fluorescence Lifetime Imaging Microscopy (FLIM) as a Timer of Cell Cycle S Phase

    PubMed Central

    Foley, Tara; Papkovsky, Dmitri B.

    2016-01-01

    Incorporation of thymidine analogues in replicating DNA, coupled with antibody and fluorophore staining, allows analysis of cell proliferation, but is currently limited to monolayer cultures, fixed cells and end-point assays. We describe a simple microscopy imaging method for live real-time analysis of cell proliferation, S phase progression over several division cycles, effects of anti-proliferative drugs and other applications. It is based on the prominent (~ 1.7-fold) quenching of fluorescence lifetime of a common cell-permeable nuclear stain, Hoechst 33342 upon the incorporation of 5-bromo-2’-deoxyuridine (BrdU) in genomic DNA and detection by fluorescence lifetime imaging microscopy (FLIM). We show that quantitative and accurate FLIM technique allows high-content, multi-parametric dynamic analyses, far superior to the intensity-based imaging. We demonstrate its uses with monolayer cell cultures, complex 3D tissue models of tumor cell spheroids and intestinal organoids, and in physiological study with metformin treatment. PMID:27973570

  7. Observation of fresh Bos indicus embryos comparing stereoscopic and phase contrast microscopy.

    PubMed

    Gutiérrez, M E; Galina, C S; Moreno-Mendoza, N; Alarcón, M A; Godínez, B; Lammoglia, M A

    2014-05-01

    Summary The precision of embryo evaluation using stereoscopic microscopy (SM) and inverted phase contrast microscopy (PCM) was compared in 20 Bos indicus cows superovulated at two different times of the year. In total, 118 embryos were collected and classified according to their developmental stage and quality by two independent evaluators using SM and inverted PCM. Cohen's kappa coefficient was used to determine concordance between SM and PCM observations. A good level of agreement (k = 0.616) was found for quality level, and a moderate one (k = 0.464) for developmental stage, particularly at the morula stage. Using the TUNEL (terminal deoxynucleotidyl transferase dUTP nick-end labelling) technique, concordance level was deemed to be low with the SM (k = 0.169), and poor with the PCM (k = 0.217). Differences in concordance levels were also found between observations made at the two times of year, 78 embryos were evaluated in the rainy season when concordance level was good (k = 0.68), in contrast to the 40 embryos evaluated in the dry season when agreement was found to be poor (k = 0.24). In conclusion, inverted PCM was somewhat more effective for evaluating embryos, particularly at the morula stage. However, considering the high cost of an inverted PCM, the differences observed do not justify its purchase for routine embryo evaluation.

  8. Multifocus microscopy with precise color multi-phase diffractive optics applied in functional neuronal imaging

    PubMed Central

    Abrahamsson, Sara; Ilic, Rob; Wisniewski, Jan; Mehl, Brian; Yu, Liya; Chen, Lei; Davanco, Marcelo; Oudjedi, Laura; Fiche, Jean-Bernard; Hajj, Bassam; Jin, Xin; Pulupa, Joan; Cho, Christine; Mir, Mustafa; El Beheiry, Mohamed; Darzacq, Xavier; Nollmann, Marcelo; Dahan, Maxime; Wu, Carl; Lionnet, Timothée; Liddle, J. Alexander; Bargmann, Cornelia I.

    2016-01-01

    Multifocus microscopy (MFM) allows high-resolution instantaneous three-dimensional (3D) imaging and has been applied to study biological specimens ranging from single molecules inside cells nuclei to entire embryos. We here describe pattern designs and nanofabrication methods for diffractive optics that optimize the light-efficiency of the central optical component of MFM: the diffractive multifocus grating (MFG). We also implement a “precise color” MFM layout with MFGs tailored to individual fluorophores in separate optical arms. The reported advancements enable faster and brighter volumetric time-lapse imaging of biological samples. In live microscopy applications, photon budget is a critical parameter and light-efficiency must be optimized to obtain the fastest possible frame rate while minimizing photodamage. We provide comprehensive descriptions and code for designing diffractive optical devices, and a detailed methods description for nanofabrication of devices. Theoretical efficiencies of reported designs is ≈90% and we have obtained efficiencies of > 80% in MFGs of our own manufacture. We demonstrate the performance of a multi-phase MFG in 3D functional neuronal imaging in living C. elegans. PMID:27231594

  9. Image formation in phase-shifting digital holography and applications to microscopy.

    PubMed

    Yamaguchi, I; Kato, J; Ohta, S; Mizuno, J

    2001-12-01

    We discuss image formation in phase-shifting digital holography by developing an analytical formulation based on the Fresnel-Kirchhoff diffraction theory. Image-plane position and imaging magnification are derived for general configurations in which a spherical reference is employed. The influences of discrete sampling of the resulting interference patterns by a CCD and numerical reconstruction on qualities of point images are investigated. Dependence of the point images on the ratio of the minimum fringe spacing to pixel pitch of the CCD is numerically analyzed. Two-point resolution and magnification are also investigated as a function of pixel numbers by a simulation using a one-dimensional model. In experiments magnified images of biological objects and a resolution target were reconstructed with the same quality as by conventional microscopy.

  10. Phasing of the Triatoma virus diffraction data using a cryo-electron microscopy reconstruction

    SciTech Connect

    Estrozi, L.F.; Neumann, E.; Squires, G.; Rozas-Dennis, G.; Costabel, M.; Rey, F.A.; Guerin, D.M.A. Navaza, J.

    2008-05-25

    The blood-sucking reduviid bug Triatoma infestans, one of the most important vector of American human trypanosomiasis (Chagas disease) is infected by the Triatoma virus (TrV). TrV has been classified as a member of the Cripavirus genus (type cricket paralysis virus) in the Dicistroviridae family. This work presents the three-dimensional cryo-electron microscopy (cryo-EM) reconstruction of the TrV capsid at about 25 A resolution and its use as a template for phasing the available crystallographic data by the molecular replacement method. The main structural differences between the cryo-EM reconstruction of TrV and other two viruses, one from the same family, the cricket paralysis virus (CrPV) and the human rhinovirus 16 from the Picornaviridae family are presented and discussed.

  11. Bacteria-clay interactions investigated by light scattering and phase contrast microscopy

    NASA Astrophysics Data System (ADS)

    Alimova, Alexandra; Block, Karin; Rudolph, Elizabeth; Katz, A.; Steiner, J. C.; Gottlieb, P.; Alfano, R. R.

    2006-02-01

    Light scattering experiments and phase contrast microscopy are used to evaluate the aggregate-forming characteristics of simple clay-bacteria mixtures. Colloidal suspensions of negatively charged Pseudomonas syringae (Ps) and Mg 2+-, Li + - or Ca 2+ -exchanged smectite (and non-exchanged smectite) are flocculated in neutral (pH 7) aqueous media. Aggregate formation is monitored using changes in optical transmission. Clustering is observed in all the clay-bacteria preparations. The Li +-substituted clay aggregates average 50-300 microns in diameter, in contrast to the Ca 2+- substituted clay that produces aggregates of 10-50 microns in diameter. Light scattering measurements indicate that aggregates begin forming 3 hours after mixing and that the (larger sized) aggregates exhibit less scattering than a mixture with an equivalent concentration of unattached Ps and clay particles.

  12. Fresnel diffraction correction by phase-considered iteration procedure in soft X-ray projection microscopy

    NASA Astrophysics Data System (ADS)

    Shiina, Tatsuo; Suzuki, Tsuyoshi; Honda, Toshio; Ito, Atsushi; Kinjo, Yasuhito; Yoshimura, Hideyuki; Yada, Keiji; Shinohara, Kunio

    2009-09-01

    In soft X-ray projection microscopy, it is easy to alter the magnification by changing the distance between the pinhole and the specimen, while the image is blurred because the soft X-rays are diffracted through the propagation from specimen to CCD detector. We corrected the blurred image by the iteration procedure of Fresnel to inverse Fresnel transformation taking phase distribution of the specimen into account. The experiments were conducted at the BL-11A of the Photon Factory, KEK, Japan for the specimens such as glass-capillaries, latex-particles, dried mammalian cells and human chromosomes. Many of those blurred images were corrected adequately by the iteration procedure, though some images such as those which have high-contrast or are overlapped by small cells still remain to be improved.

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

    PubMed Central

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

    2010-01-01

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

  14. Polymorphic phase behavior of lysophosphatidylethanolamine dispersions. A thermodynamic and spectroscopic characterization.

    PubMed Central

    Slater, J L; Huang, C H; Adams, R G; Levin, I W

    1989-01-01

    We have investigated the phase behavior of aqueous dispersions of a series of synthetic lysophosphatidylethanolamines as a function of the acyl chain length. Lysophosphatidylethanolamines exhibit phase polymorphism encompassing a well-ordered crystalline phase which may arise either from a metastable interdigitated lamellar gel phase or a metastable micellar phase. The time course of interconversion between these various phases have been outlined by observing the low temperature incubation time dependence of the calorimetric thermograms. We have determined differences in structure of these phases by Raman spectroscopy and 31P nuclear magnetic resonance spectroscopy. It appears that a principal contribution to this polymorphic phase behavior lies in the nature of headgroup hydration and headgroup-headgroup interactions. PMID:2775827

  15. Comparison of fluorescence microscopy and solid-phase cytometry methods for counting bacteria in water

    USGS Publications Warehouse

    Lisle, John T.; Hamilton, Martin A.; Willse, Alan R.; McFeters, Gordon A.

    2004-01-01

    Total direct counts of bacterial abundance are central in assessing the biomass and bacteriological quality of water in ecological and industrial applications. Several factors have been identified that contribute to the variability in bacterial abundance counts when using fluorescent microscopy, the most significant of which is retaining an adequate number of cells per filter to ensure an acceptable level of statistical confidence in the resulting data. Previous studies that have assessed the components of total-direct-count methods that contribute to this variance have attempted to maintain a bacterial cell abundance value per filter of approximately 106 cells filter-1. In this study we have established the lower limit for the number of bacterial cells per filter at which the statistical reliability of the abundance estimate is no longer acceptable. Our results indicate that when the numbers of bacterial cells per filter were progressively reduced below 105, the microscopic methods increasingly overestimated the true bacterial abundance (range, 15.0 to 99.3%). The solid-phase cytometer only slightly overestimated the true bacterial abundances and was more consistent over the same range of bacterial abundances per filter (range, 8.9 to 12.5%). The solid-phase cytometer method for conducting total direct counts of bacteria was less biased and performed significantly better than any of the microscope methods. It was also found that microscopic count data from counting 5 fields on three separate filters were statistically equivalent to data from counting 20 fields on a single filter.

  16. Phase retrieval and 3D imaging in gold nanoparticles based fluorescence microscopy (Conference Presentation)

    NASA Astrophysics Data System (ADS)

    Ilovitsh, Tali; Ilovitsh, Asaf; Weiss, Aryeh M.; Meir, Rinat; Zalevsky, Zeev

    2017-02-01

    Optical sectioning microscopy can provide highly detailed three dimensional (3D) images of biological samples. However, it requires acquisition of many images per volume, and is therefore time consuming, and may not be suitable for live cell 3D imaging. We propose the use of the modified Gerchberg-Saxton phase retrieval algorithm to enable full 3D imaging of gold nanoparticles tagged sample using only two images. The reconstructed field is free space propagated to all other focus planes using post processing, and the 2D z-stack is merged to create a 3D image of the sample with high fidelity. Because we propose to apply the phase retrieving on nano particles, the regular ambiguities typical to the Gerchberg-Saxton algorithm, are eliminated. The proposed concept is then further enhanced also for tracking of single fluorescent particles within a three dimensional (3D) cellular environment based on image processing algorithms that can significantly increases localization accuracy of the 3D point spread function in respect to regular Gaussian fitting. All proposed concepts are validated both on simulated data as well as experimentally.

  17. Atomic-force-microscopy studies of phase separations in macromolecular systems

    NASA Astrophysics Data System (ADS)

    Kuznetsov, Yu. G.; Malkin, A. J.; McPherson, A.

    1998-09-01

    Atomic force microscopy (AFM) has been used to visualize events arising from the formation of intervening metastable phases at the surfaces of macromolecular crystals growing from solution. Crystals investigated were of the proteins canavalin, thaumatin, lipase, xylanase, and catalase, crystals of transfer RNA, and crystals of satellite tobacco mosaic virus. Two types of aggregates were observed. The first were small, linear and branched aggregates, perhaps fractile in structure. These were incorporated into growing crystals as impurities, and they produced defects of various kinds. The second aggregate form we infer to be liquid-protein droplets which were particularly evident in freshly mixed protein-precipitant solutions. Droplets, upon sedimentation, have two possible fates. In some cases they immediately restructured as crystalline multilayer stacks whose development was guided by and contiguous with the underlying lattice. These contributed to the ordered growth of the crystal by serving as sources of growth steps. In other cases, liquid-protein droplets formed distinct microcrystals, somehow discontinuous with the underlying lattice, and these were subsequently incorporated into the growing substrate crystal with the formation of defects. Scarring experiments with the AFM tip indicated that liquid-protein droplets with the potential to rapidly crystallize were a consequence of concentration instabilities near the crystal's surfaces. The AFM study suggests that phase separation and the appearance of aggregates having limited order is a common occurrence in supersaturated macromolecular solutions such as the protein-precipitant solutions used for crystallization.

  18. Identifying ferroelectric phase and domain structure using angle-resolved piezoresponse force microscopy

    SciTech Connect

    Kim, K. L.; Huber, J. E.

    2014-03-24

    We used angle-resolved piezoresponse force microscopy (AR-PFM), vertical PFM (VPFM), and electron backscatter diffraction (EBSD) to provide a systematic interpretation of domain patterns in polycrystalline, near-morphotropic lead zirconate titanate. This material was used to illustrate the power of AR-PFM methods in resolving complex domain patterns where multiple phases may be present. AR-PFM was carried out with a 30° rotation interval, and the resulting data were analysed to identify the orientation of the underlying axis of piezoelectricity. The additional information provided by AR-PFM was studied, comparing its capabilities to those of 3-dimensional PFM, consisting of one VPFM image and two orthogonal lateral PFM (LPFM) images. We show that, in certain conditions, using AR-PFM can identify the phases present at the sub-grain scale. This was confirmed using VPFM and EBSD data. Furthermore, the method can discriminate laminated domain patterns that appear similar in VPFM and can reliably expose domain patterns that may not be seen in LPFM data from a single orientation, or even in 3D PFM data.

  19. Analyzer-based x-ray phase-contrast microscopy combining channel-cut and asymmetrically cut crystals

    SciTech Connect

    Hoennicke, M. G.; Cusatis, C.

    2007-11-15

    An analyzer-based x-ray phase-contrast microscopy (ABM) setup combining a standard analyzer-based x-ray phase-contrast imaging (ABI) setup [nondispersive 4-crystal setup (Bonse-Hart setup)] and diffraction by asymmetrically cut crystals is presented here. An attenuation-contrast microscopy setup with conventional x-ray source and asymmetrically cut crystals is first analyzed. Edge-enhanced effects attributed to phase jumps or refraction/total external reflection on the fiber borders were detected. However, the long exposure times and the possibility to achieve high contrast microscopies by using extremely low attenuation-contrast samples motivated us to assemble the ABM setup using a synchrotron source. This setup was found to be useful for low contrast attenuation samples due to the low exposure time, high contrast, and spatial resolution found. Moreover, thanks to the combination with the nondispersive ABI setup, the diffraction-enhanced x-ray imaging algorithm could be applied.

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

    PubMed

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

    2015-08-10

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

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

    PubMed

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

    2015-01-01

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

  2. Visualization of phase evolution in model organic photovoltaic structures via energy-filtered transmission electron microscopy.

    PubMed

    Herzing, Andrew A; Ro, Hyun Wook; Soles, Christopher L; DeLongchamp, Dean M

    2013-09-24

    The morphology of the active layer in an organic photovoltaic bulk-heterojunction device is controlled by the extent and nature of phase separation during processing. We have studied the effects of fullerene crystallinity during heat treatment in model structures consisting of a layer of poly(3-hexylthiophene) (P3HT) sandwiched between two layers of [6,6]-phenyl-C61-butyric acid methyl ester (PCBM). Utilizing a combination of focused ion-beam milling and energy-filtered transmission electron microscopy, we monitored the local changes in phase distribution as a function of annealing time at 140 °C. In both cases, dissolution of PCBM within the surrounding P3HT was directly visualized and quantitatively described. In the absence of crystalline PCBM, the overall phase distribution remained stable after intermediate annealing times up to 60 s, whereas microscale PCBM aggregates were observed after annealing for 300 s. Aggregate growth proceeded vertically from the substrate interface via uptake of PCBM from the surrounding region, resulting in a large PCBM-depleted region in their vicinity. When precrystallized PCBM was present, amorphous PCBM was observed to segregate from the intermediate P3HT layer and ripen the crystalline PCBM underneath, owing to the far lower solubility of crystalline PCBM within P3HT. This process occurred rapidly, with segregation already evident after annealing for 10 s and with uptake of nearly all of the amorphous PCBM by the crystalline layer after 60 s. No microscale aggregates were observed in the precrystallized system, even after annealing for 300 s.

  3. Theoretical Investigation of OCN(-) Charge Transfer Complexes in Condensed Phase Media: Spectroscopic Properties in Amorphous Ice

    NASA Technical Reports Server (NTRS)

    Park, Jin-Young; Woon, David E.

    2004-01-01

    Density functional theory (DFT) calculations of cyanate (OCN(-)) charge-transfer complexes were performed to model the "XCN" feature observed in interstellar icy grain mantles. OCN(-) charge-transfer complexes were formed from precursor combinations of HNCO or HOCN with either NH3 or H2O. Three different solvation strategies for realistically modeling the ice matrix environment were explored, including (1) continuum solvation, (2) pure DFT cluster calculations, and (3) an ONIOM DFT/PM3 cluster calculation. The model complexes were evaluated by their ability to reproduce seven spectroscopic measurements associated with XCN: the band origin of the OCN(-) asymmetric stretching mode, shifts in that frequency due to isotopic substitutions of C, N, O, and H, plus two weak features. The continuum solvent field method produced results consistent with some of the experimental data but failed to account for other behavior due to its limited capacity to describe molecular interactions with solvent. DFT cluster calculations successfully reproduced the available spectroscopic measurements very well. In particular, the deuterium shift showed excellent agreement in complexes where OCN(-) was fully solvated. Detailed studies of representative complexes including from two to twelve water molecules allowed the exploration of various possible solvation structures and provided insights into solvation trends. Moreover, complexes arising from cyanic or isocyanic acid in pure water suggested an alternative mechanism for the formation of OCN(-) charge-transfer complexes without the need for a strong base such as NH3 to be present. An extended ONIOM (B3LYP/PM3) cluster calculation was also performed to assess the impact of a more realistic environment on HNCO dissociation in pure water.

  4. Theoretical Investigation of OCN(-) Charge Transfer Complexes in Condensed Phase Media: Spectroscopic Properties in Amorphous Ice

    NASA Technical Reports Server (NTRS)

    Park, Jin-Young; Woon, David E.

    2004-01-01

    Density functional theory (DFT) calculations of cyanate (OCN(-)) charge-transfer complexes were performed to model the "XCN" feature observed in interstellar icy grain mantles. OCN(-) charge-transfer complexes were formed from precursor combinations of HNCO or HOCN with either NH3 or H2O. Three different solvation strategies for realistically modeling the ice matrix environment were explored, including (1) continuum solvation, (2) pure DFT cluster calculations, and (3) an ONIOM DFT/PM3 cluster calculation. The model complexes were evaluated by their ability to reproduce seven spectroscopic measurements associated with XCN: the band origin of the OCN(-) asymmetric stretching mode, shifts in that frequency due to isotopic substitutions of C, N, O, and H, plus two weak features. The continuum solvent field method produced results consistent with some of the experimental data but failed to account for other behavior due to its limited capacity to describe molecular interactions with solvent. DFT cluster calculations successfully reproduced the available spectroscopic measurements very well. In particular, the deuterium shift showed excellent agreement in complexes where OCN(-) was fully solvated. Detailed studies of representative complexes including from two to twelve water molecules allowed the exploration of various possible solvation structures and provided insights into solvation trends. Moreover, complexes arising from cyanic or isocyanic acid in pure water suggested an alternative mechanism for the formation of OCN(-) charge-transfer complexes without the need for a strong base such as NH3 to be present. An extended ONIOM (B3LYP/PM3) cluster calculation was also performed to assess the impact of a more realistic environment on HNCO dissociation in pure water.

  5. Kinetic precipitation of solution-phase polyoxomolybdate followed by transmission electron microscopy: a window to solution-phase nanostructure.

    PubMed

    Zhu, Yan; Cammers-Goodwin, Arthur; Zhao, Bin; Dozier, Alan; Dickey, Elizabeth C

    2004-05-17

    This study aimed to elucidate the structural nature of the polydisperse, nanoscopic components in the solution and the solid states of partially reduced polyoxomolybdate derived from the [Mo132] keplerate, [(Mo)Mo5]12-[Mo2 acetate]30. Designer tripodal hexamine-tris-crown ethers and nanoscopic molybdate coprecipitated from aqueous solution. These microcrystalline solids distributed particle radii between 2-30 nm as assayed by transmission electron microscopy (TEM). The solid materials and their particle size distributions were snap shots of the solution phase. The mother liquor of the preparation of the [Mo132] keplerate after three days revealed large species (r=20-30 nm) in the coprecipitate, whereas [Mo132] keplerate redissolved in water revealed small species (3-7 nm) in the coprecipitate. Nanoparticles of coprecipitate were more stable than solids derived solely from partially reduced molybdate. The TEM features of all material analyzed lacked facets on the nanometer length scale; however, the structures diffracted electrons and appeared to be defect-free as evidenced by Moiré patterns in the TEM images. Moiré patterns and size-invariant optical densities of the features in the micrographs suggested that the molybdate nanoparticles were vesicular.

  6. Membrane protein stoichiometry studied in intact mammalian cells using liquid-phase electron microscopy.

    PubMed

    DE Jonge, N

    2017-05-04

    Receptor membrane proteins in the plasma membranes of cells respond to extracellular chemical signals by conformational changes, spatial redistribution, and (re-)assembly into protein complexes, for example, into homodimers (pairs of the same protein type). The functional state of the proteins can be determined from information about how subunits are assembled into protein complexes. Stoichiometric information about the protein complex subunits, however, is generally not obtained from intact cells but from pooled material extracted from many cells, resulting in a lack of fundamental knowledge about the functioning of membrane proteins. First, functional states may dramatically differ from cell to cell on account of cell heterogeneity. Second, extracting the membrane proteins from the plasma membrane may lead to many artefacts. Liquid-phase scanning transmission electron microscopy (STEM), in short liquid STEM, is a new technique capable of determining the locations of individual membrane proteins within the intact plasma membranes of cells in liquid. Many tens of whole cells can readily be imaged. It is possible to analyse the stoichiometry of membrane proteins in single cells while accounting for heterogenic cell populations. Liquid STEM was used to image epidermal growth factor receptors in whole COS7 cells. A study of the dimerisation of the HER2 protein in breast cancer cells revealed the presence of rare cancer cells in which HER2 was in a different functional state than in the bulk cells. Stoichiometric information about receptors is essential not only for basic science but also for biomedical application because they present many important pharmaceutical targets. © 2017 The Authors Journal of Microscopy © 2017 Royal Microscopical Society.

  7. Context based mixture model for cell phase identification in automated fluorescence microscopy

    PubMed Central

    Wang, Meng; Zhou, Xiaobo; King, Randy W; Wong, Stephen TC

    2007-01-01

    Background Automated identification of cell cycle phases of individual live cells in a large population captured via automated fluorescence microscopy technique is important for cancer drug discovery and cell cycle studies. Time-lapse fluorescence microscopy images provide an important method to study the cell cycle process under different conditions of perturbation. Existing methods are limited in dealing with such time-lapse data sets while manual analysis is not feasible. This paper presents statistical data analysis and statistical pattern recognition to perform this task. Results The data is generated from Hela H2B GFP cells imaged during a 2-day period with images acquired 15 minutes apart using an automated time-lapse fluorescence microscopy. The patterns are described with four kinds of features, including twelve general features, Haralick texture features, Zernike moment features, and wavelet features. To generate a new set of features with more discriminate power, the commonly used feature reduction techniques are used, which include Principle Component Analysis (PCA), Linear Discriminant Analysis (LDA), Maximum Margin Criterion (MMC), Stepwise Discriminate Analysis based Feature Selection (SDAFS), and Genetic Algorithm based Feature Selection (GAFS). Then, we propose a Context Based Mixture Model (CBMM) for dealing with the time-series cell sequence information and compare it to other traditional classifiers: Support Vector Machine (SVM), Neural Network (NN), and K-Nearest Neighbor (KNN). Being a standard practice in machine learning, we systematically compare the performance of a number of common feature reduction techniques and classifiers to select an optimal combination of a feature reduction technique and a classifier. A cellular database containing 100 manually labelled subsequence is built for evaluating the performance of the classifiers. The generalization error is estimated using the cross validation technique. The experimental results show

  8. A vibrational spectroscopic study on amorphous phase associated with crystallization and deformation behavior of poly(lactic acid)

    NASA Astrophysics Data System (ADS)

    Kang, Shuhui; Hsu, Shaw Ling

    2002-03-01

    We have used Raman spectroscopy in conjunction with differential scanning calorimetry to characterize the structure of biodegradable poly(lactic acid). It is known that crystallization behavior of this bio-commodity polymer is strongly dependent on the amount and sequence of configurational defects, i. e. D isomer. Relatively little effort has been devoted to the study of the amorphous phase. Based on a number of normal coordinate analyses, specific spectroscopic features have been identified which is capable of identifying defect content. By introducing D defects, which have the opposite chirality to native L-unit s, the number of curved conformational sequences increase, or conversely the number of straight conformational sequences decrease. We found that deformation will increase the latter and decrease the former. In addition, the changes in chain persistance length, nucleation rate with different D defects content, sequence distribution, and draw ratio are discussed.

  9. Fast and single solid phase fluorescence spectroscopic batch procedure for (acetyl) salicylic acid determination in drug formulations.

    PubMed

    Ortega Algar, S; Ramos Martos, N; Molina Díaz, A

    2003-03-10

    A solid phase fluorescence spectroscopic batch procedure for (acetyl) salicylic acid in drug formulations have been developed. The procedure is based on the sorption of salicylic acid (SA) on Sephadex DEAE A-25 anion exchanger gel (100 mg) by equilibration from an aqueous solution (10 or 25 ml) for 5 min; the equilibrated gel is transferred into an 1 mm quartz cell and the native fluorescence of SA sorbed on it is directly measured (lambda(ex)=297 nm; lambda(em)=405 nm). Good linearity was found in the 10-200 and 5-100 microg l(-1) ranges (for 10 and 25 ml sample volume, respectively) with R.S.D. (%) of 2.8 and 1.1. The procedure was successfully applied to the determination of acetyl salicylic acid (ASA) in drug formulations after alkaline hydrolysis to yield SA.

  10. [Linear and nonlinear spectroscopic probing of solute interactions with chemically modified silica surface]. [Stationary phase studies

    SciTech Connect

    Not Available

    1993-01-01

    Objective is to understand the surface science underlying liquid chromatographic separations, enabling improvements in design of chromatographic stationary phases. Progress was made both in use of laser spectroscopy to probe chromatographic surfaces and in developing new stationary phases based on self-assembled monolayers.

  11. Phase diagram of ammonium perchlorate: Raman spectroscopic constrains at high pressures and temperatures

    NASA Astrophysics Data System (ADS)

    Dunuwille, Mihindra; Yoo, Choong-Shik

    2016-06-01

    We present the pressure-temperature (PT) induced physical and chemical transformations in ammonium perchlorates (APs) up to 50 GPa and 450 °C, using diamond anvil cells and confocal micro-Raman spectroscopy, which provide new constraints for the phase diagram of AP. The results show spectral evidences for three new polymorphs (III, IV, and VI) of AP, in addition to two previously known phases (I and II), at various PT conditions with varying degrees of hydrogen bonding and lack of strong spectral evidence for previously known high-temperature cubic phase (phase V). Upon further heating, AP chemically decomposes to N2, N2O, and H2O. The present phase diagram is, therefore, in sharp contrast to the previous one, underscoring a rich polymorphism, a large stability field for solids, and a replacement of the melt with a decomposition line.

  12. Simple and fast spectral domain algorithm for quantitative phase imaging of living cells with digital holographic microscopy.

    PubMed

    Min, Junwei; Yao, Baoli; Ketelhut, Steffi; Engwer, Christian; Greve, Burkhard; Kemper, Björn

    2017-01-15

    We present a simple and fast phase aberration compensation method in digital holographic microscopy (DHM) for quantitative phase imaging of living cells. By analyzing the frequency spectrum of an off-axis hologram, phase aberrations can be compensated for automatically without fitting or pre-knowledge of the setup and/or the object. Simple and effective computation makes the method suitable for quantitative online monitoring with highly variable DHM systems. Results from automated quantitative phase imaging of living NIH-3T3 mouse fibroblasts demonstrate the effectiveness and the feasibility of the method.

  13. Quantitative phase-digital holographic microscopy: a new imaging modality to identify original cellular biomarkers of diseases

    NASA Astrophysics Data System (ADS)

    Marquet, P.; Rothenfusser, K.; Rappaz, B.; Depeursinge, C.; Jourdain, P.; Magistretti, P. J.

    2016-03-01

    Quantitative phase microscopy (QPM) has recently emerged as a powerful label-free technique in the field of living cell imaging allowing to non-invasively measure with a nanometric axial sensitivity cell structure and dynamics. Since the phase retardation of a light wave when transmitted through the observed cells, namely the quantitative phase signal (QPS), is sensitive to both cellular thickness and intracellular refractive index related to the cellular content, its accurate analysis allows to derive various cell parameters and monitor specific cell processes, which are very likely to identify new cell biomarkers. Specifically, quantitative phase-digital holographic microscopy (QP-DHM), thanks to its numerical flexibility facilitating parallelization and automation processes, represents an appealing imaging modality to both identify original cellular biomarkers of diseases as well to explore the underlying pathophysiological processes.

  14. Breath analysis system based on phase-shifting interferometric microscopy readout of microcantilever arrays.

    PubMed

    Kelling, S; Huang, J; Capener, M J; Elliott, S R

    2011-09-01

    Microcantilever (MC) sensors can provide ultrasensitive bio-chemical detection. Monitoring the bending response of large arrays of MC sensors coated with a library of receptors will produce a characteristic 'breath-print' corresponding to the breath composition; this promises to be suitable for non-invasive medical diagnostics. While MC arrays with hundreds of individual sensors can be produced readily, conventional MC readout methods are not suitable for the parallel readout of tens or hundreds of individual sensors. We have developed a MC sensor-array readout method based on phase-shifting interferometric microscopy (PSIM) which allows for the simultaneous monitoring of the response of all MCs within the field of view of the readout microscope optics. Based on this sensor technology, we have built an exhaled breath-analysis research instrument for the Point of Care Diagnostics Development Unit at the University of Leicester, UK. In this paper, we describe the PSIM readout system and the breath-analysis instrument that will be used to test sensor surface coatings and develop sensor sets with response patterns suitable for clear correlation to patients' health condition.

  15. Microwave spectroscopic observation of distinct electron solid phases in wide quantum wells.

    PubMed

    Hatke, A T; Liu, Yang; Magill, B A; Moon, B H; Engel, L W; Shayegan, M; Pfeiffer, L N; West, K W; Baldwin, K W

    2014-06-20

    In high magnetic fields, two-dimensional electron systems can form a number of phases in which interelectron repulsion plays the central role, since the kinetic energy is frozen out by Landau quantization. These phases include the well-known liquids of the fractional quantum Hall effect, as well as solid phases with broken spatial symmetry and crystalline order. Solids can occur at the low Landau-filling termination of the fractional quantum Hall effect series but also within integer quantum Hall effects. Here we present microwave spectroscopy studies of wide quantum wells that clearly reveal two distinct solid phases, hidden within what in d.c. transport would be the zero diagonal conductivity of an integer quantum-Hall-effect state. Explanation of these solids is not possible with the simple picture of a Wigner solid of ordinary (quasi) electrons or holes.

  16. Confocal raman microscopy as a non-invasive tool to investigate the phase composition of frozen complex cryopreservation media.

    PubMed

    Kreiner-Møller, A; Stracke, F; Zimmermann, H

    2013-01-01

    Various cryoprotective agents (CPA) are added to cell media in order to avoid cell injury during cryo preservation. The resulting complex environment of the preserved cell, consisting of crystalline and liquid phases can however not be investigated non-invasively by established methods in cryobiology. This study shows how scanning confocal Raman microscopy can non-invasively extract information on chemical composition, phase domain and distribution at cryogenic temperatures. The formation of the salt hydrate, hydrohalite NaCl∙H2O, in solutions comprised of phosphate buffered saline (PBS) and dimethyl sulphoxide (DMSO) is studied in particular. Scanning confocal Raman microscopy can be used to unambiguously identify hydrohalite in a medium containing DMSO and saline. The confocal Raman microscopy imaging along with differential scanning calorimetric measurements further show that the hydrohalite is formed without eutectic formation. This method also allows for discrimination between closely packed hydrohalite crystals that are oriented differently.

  17. Spectroscopic Fingerprint of Phase-Incoherent Superconductivity in the Underdoped Bi2Sr2CaCu2O8+δ

    SciTech Connect

    Lee, J.; Davis, J.; Fujita, K.; Schmidt, A.R.; Kim, C.K.; Eisaki, H.; Uchida, S.

    2009-08-28

    A possible explanation for the existence of the cuprate 'pseudogap' state is that it is a d-wave superconductor without quantum phase rigidity. Transport and thermodynamic studies provide compelling evidence that supports this proposal, but few spectroscopic explorations of it have been made. One spectroscopic signature of d-wave superconductivity is the particle-hole symmetric 'octet' of dispersive Bogoliubov quasiparticle interference modulations. Here we report on this octet's evolution from low temperatures to well into the underdoped pseudogap regime. No pronounced changes occur in the octet phenomenology at the superconductor's critical temperature T{sub c}, and it survives up to at least temperature T {approx} 1.5 T{sub c}. In this pseudogap regime, we observe the detailed phenomenology that was theoretically predicted for quasiparticle interference in a phase-incoherent d-wave superconductor. Thus, our results not only provide spectroscopic evidence to confirm and extend the transport and thermodynamics studies, but they also open the way for spectroscopic explorations of phase fluctuation rates, their effects on the Fermi arc, and the fundamental source of the phase fluctuations that suppress superconductivity in underdoped cuprates.

  18. Serpentines, talc, chlorites, and their high-pressure phase transitions: a Raman spectroscopic study

    NASA Astrophysics Data System (ADS)

    Reynard, Bruno; Bezacier, Lucile; Caracas, Razvan

    2015-09-01

    Raman spectra of magnesian phyllosilicates belonging to the serpentine, talc, and chlorite groups have been obtained at ambient conditions, and at high pressures and up to 200 °C in order to study high-pressure transformations in the 10 GPa range. The complex and distinct Raman spectra of these minerals allow straightforward identification, which may otherwise be difficult from optical microscopy. High-pressure measurements are in good agreement with DFT calculations for talc and lizardite. Pressure-induced displacive modifications are identified in lizardite and antigorite serpentines, and in chlorite at ~4, 7 and 8 GPa, respectively, while talc shows no transition up to ~11 GPa. At high temperature, the high-pressure distortions of serpentines shift to higher pressures. Given the stability limits of these minerals, and the natural range of P-T conditions, none of the high-pressure distortions observed at high pressure are likely to occur at depth in the Earth.

  19. Using digital inline holographic microscopy and quantitative phase contrast imaging to assess viability of cultured mammalian cells

    NASA Astrophysics Data System (ADS)

    Missan, Sergey; Hrytsenko, Olga

    2015-03-01

    Digital inline holographic microscopy was used to record holograms of mammalian cells (HEK293, B16, and E0771) in culture. The holograms have been reconstructed using Octopus software (4Deep inwater imaging) and phase shift maps were unwrapped using the FFT-based phase unwrapping algorithm. The unwrapped phase shifts were used to determine the maximum phase shifts in individual cells. Addition of 0.5 mM H2O2 to cell media produced rapid rounding of cultured cells, followed by cell membrane rupture. The cell morphology changes and cell membrane ruptures were detected in real time and were apparent in the unwrapped phase shift images. The results indicate that quantitative phase contrast imaging produced by the digital inline holographic microscope can be used for the label-free real time automated determination of cell viability and confluence in mammalian cell cultures.

  20. Mesoscopic structural phase progression in photo-excited VO2 revealed by time-resolved x-ray diffraction microscopy

    DOE PAGES

    Zhu, Yi; Cai, Zhonghou; Chen, Pice; ...

    2016-02-26

    Dynamical phase separation during a solid-solid phase transition poses a challenge for understanding the fundamental processes in correlated materials. Critical information underlying a phase transition, such as localized phase competition, is difficult to reveal by measurements that are spatially averaged over many phase seperated regions. The ability to simultanousely track the spatial and temporal evolution of such systems is essential to understanding mesoscopic processes during a phase transition. Using state-of- the-art time-resolved hard x-ray diffraction microscopy, we directly visualize the structural phase progression in a VO2 film upon photoexcitation. Following a homogenous in-plane optical excitation, the phase transformation is initiatedmore » at discrete sites and completed by the growth of one lattice structure into the other, instead of a simultaneous isotropic lattice symmetry change. The time-dependent x-ray diffraction spatial maps show that the in-plane phase progression in laser-superheated VO2 is via a displacive lattice transformation as a result of relaxation from an excited monoclinic phase into a rutile phase. The speed of the phase front progression is quantitatively measured, which is faster than the process driven by in-plane thermal diffusion but slower than the sound speed in VO2. Lastly, the direct visualization of localized structural changes in the time domain opens a new avenue to study mesoscopic processes in driven systems.« less

  1. Mesoscopic structural phase progression in photo-excited VO2 revealed by time-resolved x-ray diffraction microscopy

    PubMed Central

    Zhu, Yi; Cai, Zhonghou; Chen, Pice; Zhang, Qingteng; Highland, Matthew J.; Jung, Il Woong; Walko, Donald A.; Dufresne, Eric M.; Jeong, Jaewoo; Samant, Mahesh G.; Parkin, Stuart S. P.; Freeland, John W.; Evans, Paul G.; Wen, Haidan

    2016-01-01

    Dynamical phase separation during a solid-solid phase transition poses a challenge for understanding the fundamental processes in correlated materials. Critical information underlying a phase transition, such as localized phase competition, is difficult to reveal by measurements that are spatially averaged over many phase separated regions. The ability to simultaneously track the spatial and temporal evolution of such systems is essential to understanding mesoscopic processes during a phase transition. Using state-of-the-art time-resolved hard x-ray diffraction microscopy, we directly visualize the structural phase progression in a VO2 film upon photoexcitation. Following a homogenous in-plane optical excitation, the phase transformation is initiated at discrete sites and completed by the growth of one lattice structure into the other, instead of a simultaneous isotropic lattice symmetry change. The time-dependent x-ray diffraction spatial maps show that the in-plane phase progression in laser-superheated VO2 is via a displacive lattice transformation as a result of relaxation from an excited monoclinic phase into a rutile phase. The speed of the phase front progression is quantitatively measured, and is faster than the process driven by in-plane thermal diffusion but slower than the sound speed in VO2. The direct visualization of localized structural changes in the time domain opens a new avenue to study mesoscopic processes in driven systems. PMID:26915398

  2. Mesoscopic structural phase progression in photo-excited VO2 revealed by time-resolved x-ray diffraction microscopy.

    PubMed

    Zhu, Yi; Cai, Zhonghou; Chen, Pice; Zhang, Qingteng; Highland, Matthew J; Jung, Il Woong; Walko, Donald A; Dufresne, Eric M; Jeong, Jaewoo; Samant, Mahesh G; Parkin, Stuart S P; Freeland, John W; Evans, Paul G; Wen, Haidan

    2016-02-26

    Dynamical phase separation during a solid-solid phase transition poses a challenge for understanding the fundamental processes in correlated materials. Critical information underlying a phase transition, such as localized phase competition, is difficult to reveal by measurements that are spatially averaged over many phase separated regions. The ability to simultaneously track the spatial and temporal evolution of such systems is essential to understanding mesoscopic processes during a phase transition. Using state-of-the-art time-resolved hard x-ray diffraction microscopy, we directly visualize the structural phase progression in a VO2 film upon photoexcitation. Following a homogenous in-plane optical excitation, the phase transformation is initiated at discrete sites and completed by the growth of one lattice structure into the other, instead of a simultaneous isotropic lattice symmetry change. The time-dependent x-ray diffraction spatial maps show that the in-plane phase progression in laser-superheated VO2 is via a displacive lattice transformation as a result of relaxation from an excited monoclinic phase into a rutile phase. The speed of the phase front progression is quantitatively measured, and is faster than the process driven by in-plane thermal diffusion but slower than the sound speed in VO2. The direct visualization of localized structural changes in the time domain opens a new avenue to study mesoscopic processes in driven systems.

  3. Mesoscopic structural phase progression in photo-excited VO2 revealed by time-resolved x-ray diffraction microscopy

    NASA Astrophysics Data System (ADS)

    Zhu, Yi; Cai, Zhonghou; Chen, Pice; Zhang, Qingteng; Highland, Matthew J.; Jung, Il Woong; Walko, Donald A.; Dufresne, Eric M.; Jeong, Jaewoo; Samant, Mahesh G.; Parkin, Stuart S. P.; Freeland, John W.; Evans, Paul G.; Wen, Haidan

    2016-02-01

    Dynamical phase separation during a solid-solid phase transition poses a challenge for understanding the fundamental processes in correlated materials. Critical information underlying a phase transition, such as localized phase competition, is difficult to reveal by measurements that are spatially averaged over many phase separated regions. The ability to simultaneously track the spatial and temporal evolution of such systems is essential to understanding mesoscopic processes during a phase transition. Using state-of-the-art time-resolved hard x-ray diffraction microscopy, we directly visualize the structural phase progression in a VO2 film upon photoexcitation. Following a homogenous in-plane optical excitation, the phase transformation is initiated at discrete sites and completed by the growth of one lattice structure into the other, instead of a simultaneous isotropic lattice symmetry change. The time-dependent x-ray diffraction spatial maps show that the in-plane phase progression in laser-superheated VO2 is via a displacive lattice transformation as a result of relaxation from an excited monoclinic phase into a rutile phase. The speed of the phase front progression is quantitatively measured, and is faster than the process driven by in-plane thermal diffusion but slower than the sound speed in VO2. The direct visualization of localized structural changes in the time domain opens a new avenue to study mesoscopic processes in driven systems.

  4. Origin of p(2 x 1) phase on Si(001) by noncontact atomic force microscopy at 5 k.

    PubMed

    Li, Yan Jun; Nomura, Hikaru; Ozaki, Naoyuki; Naitoh, Yoshitaka; Kageshima, Masami; Sugawara, Yasuhiro; Hobbs, Chris; Kantorovich, Lev

    2006-03-17

    The controversial issue of the origin of the p(2 x 1) reconstruction of the Si(001) surface observed in recent low temperature scanning tunneling microscopy experiments is clarified here using 5 K noncontact atomic force microscopy. The c(4 x 2) phase is observed at separations corresponding to weak tip-surface interactions, confirming that it is the ground state of the surface. At larger frequency shifts the p(2 x 1) phase of symmetric dimers is observed. By studying the interaction of a reactive Si tip with the c(4 x 2) Si(001) surface using an ab initio method, we find that the observed change in the surface reconstruction is an apparent effect caused by tip induced dimer flipping resulting in a modification of the surface structure and appearance of the p(2 x 1) phase in the image. Using an appropriate scanning protocol, one can manipulate the surface reconstruction at will, which has significance in nanotechnology.

  5. Zernike-type phase contrast X-ray microscopy at 4 keV photon energy with 60 nm resolution

    NASA Astrophysics Data System (ADS)

    Neuhäusler, Ulrich; Schneider, Gerd

    2004-05-01

    X-ray microscopy in the multi-keV photon energy range offers unique possibilities to study thick dense samples with high spatial resolution. When employing a high numerical aperture (N.A.) condenser zone plate sample illumination in combination with a high resolution micro zone plate objective lens, a spatial resolution of currently 60 nm is achieved. Since the absorption becomes smaller with increasing photon energy, phase contrast imaging overcomes the limitation for imaging weakly absorbing structures in amplitude contrast mode. We report here on X-ray microscopy of advanced microelectronic devices imaged in Zernike phase contrast mode. While the amplitude contrast between copper and silicon dioxide in these samples is only 7 %, negative as well as positive phase contrast were demonstrated with a contrast of 40 % and 45 %, respectively.

  6. Graphene Near-Degenerate Four-Wave Mixing for Phase Characterization of Broadband Pulses in Ultrafast Microscopy.

    PubMed

    Ciesielski, Richard; Comin, Alberto; Handloser, Matthias; Donkers, Kevin; Piredda, Giovanni; Lombardo, Antonio; Ferrari, Andrea C; Hartschuh, Achim

    2015-08-12

    We investigate near-degenerate four-wave mixing in graphene using femtosecond laser pulse shaping microscopy. Intense near-degenerate four-wave mixing signals on either side of the exciting laser spectrum are controlled by amplitude and phase shaping. Quantitative signal modeling for the input pulse parameters shows a spectrally flat phase response of the near-degenerate four-wave mixing due to the linear dispersion of the massless Dirac Fermions in graphene. Exploiting these properties we demonstrate that graphene is uniquely suited for the intrafocus phase characterization and compression of broadband laser pulses, circumventing disadvantages of common methods utilizing second or third harmonic light.

  7. FTIR spectroscopic study of the mechanism for the gas-phase reaction between ozone and tetramethylethylene

    SciTech Connect

    Niki, H.; Maker, P.D.; Savage, C.M.; Breitenbach, L.P.; Hurley, M.D.

    1987-02-12

    Long-path FTIR spectroscopic product studies were made of mixtures containing part-per-million concentrations of O/sub 3/ and (CH/sub 3/)/sub 2/C double bonded C(CH/sub 3/)/sub 2/ (TME) in 700 Torr of O/sub 2/-N/sub 2/ to determine the formation and reactions of the dimethyl-substituted Criegee intermediate (CH/sub 3/)/sub 2/COO expected from O/sub 3/ + TME ..-->.. (CH/sub 3/)/sub 2/COO + (CH/sub 3/)/sub 2/C double bonded O. The observed stoichiometry, ..delta..-(TME)/..delta..(O/sub 3/), was typically 1.7, and among the products were CO, CO/sub 2/, HCHO, CH/sub 3/OH, CH/sub 3/OOH, CH/sub 3/C(double bonded O)OH, CH/sub 3/C(double bonded O)OOH, (CH/sub 3/)/sub 2/C double bonded O, CH/sub 3/C(double bonded O)CHO, CH/sub 3/C(double bonded O)OCH/sub 3/, CH/sub 3/C(double bonded O)CH/sub 2/(OH), and products tentatively identified as (CH/sub 2/)/sub 2/C(OH)-C(CH/sub 3/)/sub 2/OOH AND CH/sub 3/C(double bonded O)CH/sub 2/(OOH). In addition, /sup 16/O- and /sup 18/O-labeled products were identified in /sup 18/O/sub 3/-TME-air mixtures, and the effects of added HCHO, CH/sub 3/CHO, and NO/sub 2/ (or /sup 15/NO/sub 2/) on the product distribution were determined. The results indicate that ca. 25% of the (CH/sub 3/)/sub 2/COO moiety formed was thermally stable, and that the remainder largely dissociated to yield the following free radicals, i.e., (CH/sub 3/)/sub 2/COO ..-->.. CH/sub 3/C(OOH)double bonded CH/sub 2/ ..-->.. CH/sub 3/C(double bonded O)CH/sub 2/ + HO. The HO-initiated oxidation of TME was shown to account for the excess consumption of TME over O/sub 3/ as well as for some of the major products observed.

  8. Optomechanical properties of cancer cells revealed by light-induced deformation and quantitative phase microscopy

    NASA Astrophysics Data System (ADS)

    Kastl, Lena; Budde, Björn; Isbach, Michael; Rommel, Christina; Kemper, Björn; Schnekenburger, Jürgen

    2015-05-01

    There is a growing interest in cell biology and clinical diagnostics in label-free, optical techniques as the interaction with the sample is minimized and substances like dyes or fixatives do not affect the investigated cells. Such techniques include digital holographic microscopy (DHM) and the optical stretching by fiber optical two beam traps. DHM enables quantitative phase contrast imaging and thereby the determination of the cellular refractive index, dry mass and the volume, whereas optical cell stretching reveals the deformability of cells. Since optical stretching strongly depends on the optical properties and the shape of the investigated material we combined the usage of fiber optical stretching and DHM for the characterization of pancreatic tumor cells. The risk of tumors is their potential to metastasize, spread through the bloodstream and build distal tumors/metastases. The grade of dedifferentiation in which the cells lose their cell type specific properties is a measure for this metastatic potential. The less differentiated the cells are, the higher is their risk to metastasize. Our results demonstrate that pancreatic tumor cells, which are from the same tumor but vary in their grade of differentiation, show significant differences in their deformability. The retrieved data show that differentiated cells have a higher stiffness than less differentiated cells of the same tumor. Even cells that differ only in the expression of a single tumor suppressor gene which is responsible for cell-cell adhesions can be distinguished by their mechanical properties. Additionally, results from DHM measurements yield that the refractive index shows only few variations, indicating that it does not significantly influence optical cell stretching. The obtained results show a promising new approach for the phenotyping of different cell types, especially in tumor cell characterization and cancer diagnostics.

  9. Spectroscopically characterized cadmium sulfide quantum dots lengthening the lag phase of Escherichia coli growth

    NASA Astrophysics Data System (ADS)

    Jaiganesh, T.; Daisy Vimala Rani, J.; Girigoswami, Agnishwar

    2012-06-01

    The present study reports the effect of cadmium sulfide (CdS) quantum dots on the life cycle of Escherichia coli. CdS quantum dots were synthesized by pH sensitive organochemical route using cadmium chloride and sodium sulfide as precursors and mercaptopropionic acid (MPA) as capping agent. It is observed that varying concentration of MPA leads to the production of different sized quantum dots with inverse proportionality and increment in the fluorescence quantum yield. The investigation also shows that CdS quantum dots have no antibacterial activity except it delays the log phase growth of bacteria in terms of size of the particles. The largest synthesized particles significantly elongate the lag phase growth.

  10. Investigational study of tamoxifen phase I metabolites using chromatographic and spectroscopic analytical techniques.

    PubMed

    Teunissen, S F; Rosing, H; Seoane, M Dominguez; Brunsveld, L; Schellens, J H M; Schinkel, A H; Beijnen, J H

    2011-06-01

    A comprehensive overview is presented of currently known phase I metabolites of tamoxifen consisting of their systematic name and molecular structure. Reference standards are utilized to elucidate the MS(n) fragmentation patterns of these metabolites using a linear ion trap mass spectrometer. UV-absorption spectra are recorded and absorption maxima are defined. Serum extracts from ten breast cancer patients receiving 40mg tamoxifen once daily were qualitatively analyzed for tamoxifen phase I metabolites using a liquid chromatography-tandem mass spectrometry set-up. In total, 19 metabolites have been identified in these serum samples. Additionally a synthetic method for the preparation of the putative metabolite 3',4'-dihydroxytamoxifen is described. Copyright © 2011 Elsevier B.V. All rights reserved.

  11. Scattering-type scanning near-field optical microscopy with low-repetition-rate pulsed light source through phase-domain sampling

    PubMed Central

    Wang, Haomin; Wang, Le; Xu, Xiaoji G.

    2016-01-01

    Scattering-type scanning near-field optical microscopy (s-SNOM) allows spectroscopic imaging with spatial resolution below the diffraction limit. With suitable light sources, s-SNOM is instrumental in numerous discoveries at the nanoscale. So far, the light sources have been limited to continuous wave or high-repetition-rate pulsed lasers. Low-repetition-rate pulsed sources cannot be used, due to the limitation of the lock-in detection mechanism that is required for current s-SNOM techniques. Here, we report a near-field signal extraction method that enables low-repetition-rate pulsed light sources. The method correlates scattering signals from pulses with the mechanical phases of the oscillating s-SNOM probe to obtain near-field signal, by-passing the apparent restriction imposed by the Nyquist–Shannon sampling theorem on the repetition rate. The method shall enable s-SNOM with low-repetition-rate pulses with high-peak-powers, such as femtosecond laser amplifiers, to facilitate investigations of strong light–matter interactions and nonlinear processes at the nanoscale. PMID:27748360

  12. Scattering-type scanning near-field optical microscopy with low-repetition-rate pulsed light source through phase-domain sampling.

    PubMed

    Wang, Haomin; Wang, Le; Xu, Xiaoji G

    2016-10-17

    Scattering-type scanning near-field optical microscopy (s-SNOM) allows spectroscopic imaging with spatial resolution below the diffraction limit. With suitable light sources, s-SNOM is instrumental in numerous discoveries at the nanoscale. So far, the light sources have been limited to continuous wave or high-repetition-rate pulsed lasers. Low-repetition-rate pulsed sources cannot be used, due to the limitation of the lock-in detection mechanism that is required for current s-SNOM techniques. Here, we report a near-field signal extraction method that enables low-repetition-rate pulsed light sources. The method correlates scattering signals from pulses with the mechanical phases of the oscillating s-SNOM probe to obtain near-field signal, by-passing the apparent restriction imposed by the Nyquist-Shannon sampling theorem on the repetition rate. The method shall enable s-SNOM with low-repetition-rate pulses with high-peak-powers, such as femtosecond laser amplifiers, to facilitate investigations of strong light-matter interactions and nonlinear processes at the nanoscale.

  13. Mid-infrared spectroscopic study of crystallization of cubic spinel phase from metakaolin

    SciTech Connect

    Ptacek, Petr

    2011-10-15

    The structural changes during thermal conversion of metakaolinite into cubic spinel phase that was in literature considered as Al-Si spinel or {gamma}-Al{sub 2}O{sub 3} were investigated by mid-infrared spectroscopy (FT-IR) and differential scanning calorimetry (DSC) using medium ordered kaolin with high content of kaolinite. Spectrum features were studied in the mid-infrared region from 4000 to 400 cm{sup -1} as function of fractional conversion that resulted from DSC experiments. The increasing contends of Al-O bonds in the octahedral position (AlO{sub 6}) was observed during thermal transformation. The high ratio of antisymmetric stretching of {identical_to}Si-O-Al= and {identical_to}Si-O-Si{identical_to} bands and bands belonging to stretching of =Al-O bonds in the (AlO{sub 4}) tetrahedra and (AlO{sub 6}) octahedra decreases exponentially with fractional conversion. The antisymmetric stretching band of {identical_to}Si-O-Si{identical_to} bond in the (SiO{sub 4}) tetrahedra becomes more expressive due to formation of amorphous SiO{sub 2} phase. - Graphical abstract: The thermal conversion of metakaolinite into spinel phase was investigated by mid-infrared spectroscopy to found relationship between bands features and fractional convesrion. Highlights: > Formation of spinel phase from metakaolinite was studied by FT-IR. > Relationship between spectrum features and fractional conversion was investigated. > Intensity ratios of (AlO{sub 4}) and (AlO{sub 6}) decreases exponentially. > Antisymmetric stretching of (SiO{sub 4}) becomes more expressive during transformation.

  14. Mineralogical composition and phase-to-phase relationships in natural hydraulic lime and/or natural cement - raw materials and burnt products revealed by scanning electron microscopy

    NASA Astrophysics Data System (ADS)

    Kozlovcev, Petr; Přikryl, Richard; Racek, Martin; Přikrylová, Jiřina

    2016-04-01

    In contrast to modern process of production of cement clinker, traditional burning of natural hydraulic lime below sintering temperature relied on the formation of new phases from ion migration between neighbouring mineral grains composing raw material. The importance of the mineralogical composition and spatial distribution of rock-forming minerals in impure limestones used as a raw material for natural hydraulic lime presents not well explored issue in the scientific literature. To fill this gap, the recent study focuses in detailed analysis of experimentally burnt impure limestones (mostly from Barrandian area, Bohemian Massif). The phase changes were documented by optical microscopy, X-ray diffraction, and scanning electron microscopy with an energy dispersive spectrometer (SEM-EDS) coupled with x-ray elemental mapping. The latest allowed for visualization of distribution of elements within raw materials and burnt products. SEM/EDS study brought valuable data on the presence of transitional and/or minor phases, which were poorly detectable by other methods.

  15. Direct measurement of the refractive index profile of phase gratings, recorded in silver halide holographic materials by phase-contrast microscopy

    NASA Astrophysics Data System (ADS)

    Bányász, I.

    2003-11-01

    Plane-wave phase holograms recorded in Agfa-Gevaert 8E75HD emulsions and processed by the combination of AAC developer and the R-9 bleaching agent were studied by phase-contrast microscopy, using high-power immersion (100×) objective. Thus the modulation of the refractive index as a function of the bias exposure and the visibility of the recording interference pattern can also be determined. Measured diffraction efficiencies were compared to those predicted by coupled wave theory, using the measured refractive index modulations. Direct measurement of the phase profile of the gratings can be used for optimizing processing.

  16. Optical spectroscopic and reverse-phase HPLC analyses of Hg(II) binding to phytochelatins.

    PubMed Central

    Mehra, R K; Miclat, J; Kodati, V R; Abdullah, R; Hunter, T C; Mulchandani, P

    1996-01-01

    Optical spectroscopy and reverse-phase HPLC were used to investigate the binding of Hg(II) to plant metal-binding peptides (phytochelatins) with the structure (gammaGlu-Cys)2Gly, (gammaGlu-Cys)3Gly and (gammaGlu-Cys)4Gly. Glutathione-mediated transfer of Hg(II) into phytochelatins and the transfer of the metal ion from one phytochelatin to another was also studied using reverse-phase HPLC. The saturation of Hg(II)-induced bands in the UV/visible and CD spectra of (gammaGlu-Cys)2Gly suggested the formation of a single Hg(II)-binding species of this peptide with a stoichiometry of one metal ion per peptide molecule. The separation of apo-(gammaGlu-Cys)2Gly from its Hg(II) derivative on a C18 reverse-phase column also indicated the same metal-binding stoichiometry. The UV/visible spectra of both (gammaGlu-Cys)3Gly and (gammaGlu-Cys)4Gly at pH 7.4 showed distinct shoulders in the ligand-to-metal charge-transfer region at 280-290 mm. Two distinct Hg(II)-binding species, occurring at metal-binding stoichiometries of around 1.25 and 2.0 Hg(II) ions per peptide molecule, were observed for (gammaGlu-Cys)3Gly. These species exhibited specific spectral features in the charge-transfer region and were separable by HPLC. Similarly, two main Hg(II)-binding species of (gammaGlu-Cys)4Gly were observed by UV/visible and CD spectroscopy at metal-binding stoichiometries of around 1.25 and 2.5 respectively. Only a single peak of Hg(II)-(gammaGlu-Cys)4Gly complexes was resolved under the conditions used for HPLC. The overall Hg(II)-binding stoichiometries of phytochelatins were similar at pH 2.0 and at pH 7.4, indicating that pH did not influence the final Hg(II)-binding capacity of these peptides. The reverse-phase HPLC assays indicated a rapid transfer of Hg(II) from glutathione to phytochelatins. These assays also demonstrated a facile transfer of the metal ion from shorter- to longer-chain phytochelatins. The strength of Hg(II) binding to glutathione and phytochelatins followed the

  17. Direct observation of dopant distribution in GaAs compound semiconductors using phase-shifting electron holography and Lorentz microscopy.

    PubMed

    Sasaki, Hirokazu; Otomo, Shinya; Minato, Ryuichiro; Yamamoto, Kazuo; Hirayama, Tsukasa

    2014-06-01

    Phase-shifting electron holography and Lorentz microscopy were used to map dopant distributions in GaAs compound semiconductors with step-like dopant concentration. Transmission electron microscope specimens were prepared using a triple beam focused ion beam (FIB) system, which combines a Ga ion beam, a scanning electron microscope, and an Ar ion beam to remove the FIB damaged layers. The p-n junctions were clearly observed in both under-focused and over-focused Lorentz microscopy images. A phase image was obtained by using a phase-shifting reconstruction method to simultaneously achieve high sensitivity and high spatial resolution. Differences in dopant concentrations between 1 × 10(19) cm(-3) and 1 × 10(18) cm(-3) regions were clearly observed by using phase-shifting electron holography. We also interpreted phase profiles quantitatively by considering inactive layers induced by ion implantation during the FIB process. The thickness of an inactive layer at different dopant concentration area can be measured from the phase image.

  18. Infrared spectroscopic study of thermotropic phase behavior of newly developed synthetic biopolymers.

    PubMed

    Bista, Rajan K; Bruch, Reinhard F; Covington, Aaron M

    2011-10-15

    The thermotropic phase behavior of a suite of newly developed self-forming synthetic biopolymers has been investigated by variable-temperature Fourier transform infrared (FT-IR) absorption spectroscopy. The temperature-induced infrared spectra of these artificial biopolymers (lipids) composed of 1,2-dimyristoyl-rac-glycerol-3-dodecaethylene glycol (GDM-12), 1,2-dioleoyl-rac-glycerol-3-dodecaethylene glycol (GDO-12) and 1,2-distearoyl-rac-glycerol-3-triicosaethylene glycol (GDS-23) in the spectral range of 4000-500 cm(-1) have been acquired by using a thin layered FT-IR spectrometer in conjunction with a custom built temperature-controlled demountable liquid cell having a pathlength of ∼15 μm. The lipids under consideration have long hydrophobic acyl chains and contain various units of hydrophilic polyethylene glycol (PEG) headgroups. In contrast to conventional phospholipids, this new kind of lipids forms liposomes or nanovesicles spontaneously upon hydration, without requiring external activation energy. We have found that the thermal stability of the PEGylated lipids differs greatly depending upon the acyl chain-lengths as well as the nature of the associated bonds and the number of PEG headgroup units. In particular, GDM-12 (saturated 14 hydrocarbon chains with 12 units of PEG headgroup) exhibits one sharp order-disorder phase transition over a temperature range increasing from 3°C to 5°C. Similarly, GDS-23 (saturated 18 hydrocarbon chains with 23 units of PEG headgroup) displays comparatively broad order-disorder phase transition profiles between temperature 17°C and 22°C. In contrast, GDO-12 (monounsaturated 18 hydrocarbon chains with 12 units of PEG headgroup) does not reveal any order-disorder transition phenomena demonstrating a highly disordered behavior for the entire temperature range. To confirm these observations, differential scanning calorimetry (DSC) was applied to the samples and revealed good agreement with the infrared spectroscopy results

  19. Comprehensive Size-Determination of Whole Virus Vaccine Particles Using Gas-Phase Electrophoretic Mobility Macromolecular Analyzer, Atomic Force Microscopy, and Transmission Electron Microscopy

    PubMed Central

    Havlik, Marlene; Marchetti-Deschmann, Martina; Friedbacher, Gernot; Winkler, Wolfgang; Messner, Paul; Perez-Burgos, Laura; Tauer, Christa; Allmaier, Günter

    2015-01-01

    Biophysical properties including particle size distribution, integrity, and shape of whole virus vaccine particles at different stages in tick-borne encephalitis (TBE) vaccines formulation were analyzed by a new set of methods. Size-exclusion chromatography (SEC) was used as a conservative sample preparation for vaccine particle fractionation and gas-phase electrophoretic mobility macromolecular analyzer (GEMMA) for analyzing electrophoretic mobility diameters of isolated TBE virions. The derived particle diameter was then correlated with molecular weight. The diameter of the TBE virions determined after SEC by GEMMA instrumentation was 46.8 ± 1.1 nm. Atomic force microscopy (AFM) and transmission electron microscopy (TEM) were implemented for comparison purposes and to gain morphological information on the virion particle. Western blotting (Dot Blot) as an immunological method confirmed biological activity of the particles at various stages of the developed analytical strategy. AFM and TEM measurements revealed higher diameters with much higher SD for a limited number of virions, 60.4 ± 8.5 and 53.5 ± 5.3 nm, respectively. GEMMA instrumentation was also used for fractionation of virions with specifically selected diameters in the gas-phase, which were finally collected by means of an electrostatic sampler. At that point (i.e., after particle collection), AFM and TEM showed that the sampled virions were still intact, exhibiting a narrow size distribution (i.e., 59.8 ± 7.8 nm for AFM and 47.5 ± 5.2 nm for TEM images), and most importantly, dot blotting confirmed immunological activity of the collected samples. Furthermore dimers and virion artifacts were detected, too. PMID:26266988

  20. Comprehensive size-determination of whole virus vaccine particles using gas-phase electrophoretic mobility macromolecular analyzer, atomic force microscopy, and transmission electron microscopy.

    PubMed

    Havlik, Marlene; Marchetti-Deschmann, Martina; Friedbacher, Gernot; Winkler, Wolfgang; Messner, Paul; Perez-Burgos, Laura; Tauer, Christa; Allmaier, Günter

    2015-09-01

    Biophysical properties including particle size distribution, integrity, and shape of whole virus vaccine particles at different stages in tick-borne encephalitis (TBE) vaccines formulation were analyzed by a new set of methods. Size-exclusion chromatography (SEC) was used as a conservative sample preparation for vaccine particle fractionation and gas-phase electrophoretic mobility macromolecular analyzer (GEMMA) for analyzing electrophoretic mobility diameters of isolated TBE virions. The derived particle diameter was then correlated with molecular weight. The diameter of the TBE virions determined after SEC by GEMMA instrumentation was 46.8 ± 1.1 nm. Atomic force microscopy (AFM) and transmission electron microscopy (TEM) were implemented for comparison purposes and to gain morphological information on the virion particle. Western blotting (Dot Blot) as an immunological method confirmed biological activity of the particles at various stages of the developed analytical strategy. AFM and TEM measurements revealed higher diameters with much higher SD for a limited number of virions, 60.4 ± 8.5 and 53.5 ± 5.3 nm, respectively. GEMMA instrumentation was also used for fractionation of virions with specifically selected diameters in the gas-phase, which were finally collected by means of an electrostatic sampler. At that point (i.e., after particle collection), AFM and TEM showed that the sampled virions were still intact, exhibiting a narrow size distribution (i.e., 59.8 ± 7.8 nm for AFM and 47.5 ± 5.2 nm for TEM images), and most importantly, dot blotting confirmed immunological activity of the collected samples. Furthermore dimers and virion artifacts were detected, too.

  1. Dynamic and spectroscopic characteristics of atmospheric gliding arc in gas-liquid two-phase flow

    NASA Astrophysics Data System (ADS)

    Tu, X.; Yu, L.; Yan, J. H.; Cen, K. F.; Chéron, B. G.

    2009-11-01

    In this study, an atmospheric alternating-current gliding arc device in gas-liquid two-phase flow has been developed for the purpose of waste water degradation. The dynamic behavior of the gas-liquid gliding arc is investigated through the oscillations of electrical signals, while the spatial evolution of the arc column is analyzed by high speed photography. Different arc breakdown regimes are reported, and the restrike mode is identified as the typical fluctuation characteristic of the hybrid gliding arc in air-water mixture. Optical emission spectroscopy is employed to investigate the active species generated in the gas-liquid plasma. The axial evolution of the OH (309 nm) intensity is determined, while the rotational and vibrational temperatures of the OH are obtained by a comparison between the experimental and simulated spectra. The significant discrepancy between the rotational and translational temperatures has also been discussed.

  2. Spectroscopic Analysis of Various Phases of Water with an Intrinsic Near-Infrared Optical Sensor System

    NASA Astrophysics Data System (ADS)

    Igbinosun, Osazonamen J.

    Results and analysis are reported for near-infrared spectra obtained using an intrinsic optical sensor system. An experimental facility was designed to integrate fiber optic cables with an FTIR spectrometer. Wavelength-dependent spectral signatures were observed for the solid, liquid, and adsorbed phases of water at 1.94 and 1.45 micrometers. The data obtained can be used to characterize the water abundance of a medium on very small scales and can aid our understanding of the behavior of water and thermal properties of planetary surfaces by observing, for example, ice formation in void spaces between mineral grains. This system will be further developed for use in an environmental simulation facility to analyze Mars analog soils. The implications of this work could impact future space missions in regard to the availability of critical resources for human missions and the search for non-terrestrial life.

  3. Infrared spectroscopic characterization of dehydration and accompanying phase transition behaviors in NAT-topology zeolites

    SciTech Connect

    Wang, Hsiu-Wen; Bishop, David

    2012-01-01

    Relative humidity (PH2O, partial pressure of water)-dependent dehydration and accompanying phase transitions in NAT-topology zeolites (natrolite, scolecite, and mesolite) were studied under controlled temperature and known PH2O conditions by in situ diffuse-reflectance infrared Fourier transform spectroscopy and parallel X-ray powder diffraction. Dehydration was characterized by the disappearance of internal H2O vibrational modes. The loss of H2O molecules caused a sequence of structural transitions in which the host framework transformation path was coupled primarily via the thermal motion of guest Na?/Ca2? cations and H2O molecules. The observation of different interactions of H2O molecules and Na?/Ca2? cations with host aluminosilicate frameworks under highand low-PH2O conditions indicated the development of different local strain fields, arising from cation H2O interactions in NAT-type channels. These strain fields influence the Si O/Al O bond strength and tilting angles within and between tetrahedra as the dehydration temperature is approached. The newly observed infrared bands (at 2,139 cm-1 in natrolite, 2,276 cm-1 in scolecite, and 2,176 and 2,259 cm-1 in mesolite) result from strong cation H2O Al Si framework interactions in NAT-type channels, and these bands can be used to evaluate the energetic evolution of Na?/Ca2? cations before and after phase transitions, especially for scolecite and mesolite. The 2,176 and 2,259 cm-1 absorption bands in mesolite also appear to be related to Na?/Ca2? order disorder that occur when mesolite loses its Ow4 H2O molecules.

  4. Infrared spectroscopic characterization of dehydration and accompanying phase transition behaviors in NAT-topology zeolites

    NASA Astrophysics Data System (ADS)

    Wang, Hsiu-Wen; Bish, David L.

    2012-04-01

    Relative humidity ( P_{{{text{H}}_{ 2} {text{O}}}} , partial pressure of water)-dependent dehydration and accompanying phase transitions in NAT-topology zeolites (natrolite, scolecite, and mesolite) were studied under controlled temperature and known P_{{{text{H}}_{ 2} {text{O}}}} conditions by in situ diffuse-reflectance infrared Fourier transform spectroscopy and parallel X-ray powder diffraction. Dehydration was characterized by the disappearance of internal H2O vibrational modes. The loss of H2O molecules caused a sequence of structural transitions in which the host framework transformation path was coupled primarily via the thermal motion of guest Na+/Ca2+ cations and H2O molecules. The observation of different interactions of H2O molecules and Na+/Ca2+ cations with host aluminosilicate frameworks under high- and low- P_{{{text{H}}_{ 2} {text{O}}}} conditions indicated the development of different local strain fields, arising from cation-H2O interactions in NAT-type channels. These strain fields influence the Si-O/Al-O bond strength and tilting angles within and between tetrahedra as the dehydration temperature is approached. The newly observed infrared bands (at 2,139 cm-1 in natrolite, 2,276 cm-1 in scolecite, and 2,176 and 2,259 cm-1 in mesolite) result from strong cation-H2O-Al-Si framework interactions in NAT-type channels, and these bands can be used to evaluate the energetic evolution of Na+/Ca2+ cations before and after phase transitions, especially for scolecite and mesolite. The 2,176 and 2,259 cm-1 absorption bands in mesolite also appear to be related to Na+/Ca2+ order-disorder that occur when mesolite loses its Ow4 H2O molecules.

  5. In-situ microscopy of the first-order magnetic phase transition in FeRh thin films

    NASA Astrophysics Data System (ADS)

    Baldasseroni, Chloe

    Simple ferromagnetic (FM) and antiferromagnetic (AF) materials such as Fe and Cr become paramagnetic when heated above some critical temperature, in what is known as a second-order phase transition. Less usual magnetic transitions are found in the magnetic world, for example a first-order magnetic phase transition from AF to FM with increasing temperature. Equiatomic FeRh has been known to exhibit such a transition for over 50 years, with a transition temperature slightly above room temperature. Interest in this material has been renewed in the recent years due to its potential application for heat-assisted magnetic recording, as well as a test system for fundamental studies of the physics of magnetic phase transitions. Similarly to crystallization, this AF-FM transition is expected to proceed by nucleation of magnetic domains but the features of the first-order hysteretic transition have been difficult to study with macroscopic measurements and very few microscopic studies have been performed. In this work, FeRh thin films were synthesized by magnetron sputtering and structurally and magnetically characterized. A membrane-based heating device was designed to enable temperature-dependent microscopy measurements, providing a thermally uniform and well-controlled sample area. Synchrotron x-ray magnetic microscopy was used to study the temperature-driven AF-FM phase transition in epitaxial FeRh thin films in zero field. Using magnetic microscopy with x-ray magnetic circular dichroism, the different stages of nucleation, growth and coalescence of FM domains were observed across the transition and details of the nucleation were identified. The FM phase nucleates into single domain islands and the width of the transition of the individual nuclei upon heating is sharper than that of the macroscopic transition. Using magnetic microscopy with x-ray magnetic linear dichroism, the evolution of the AF phase was studied. Differences in the morphology of AF and FM phases were

  6. A Combined Gas-Phase Photoelectron Spectroscopic and Theoretical Study of Zeise's Anion and Its Bromine and Iodine Analogues

    SciTech Connect

    Hou, Gaolei; Wen, Hui; Lopata, Kenneth A.; Zheng, Weijun; Kowalski, Karol; Govind, Niranjan; Wang, Xue B.; Xantheas, Sotiris S.

    2012-06-25

    We report the first photoelectron spectroscopic study of Zeise’s anion, [PtCl3(C2H4)], and its Br- and I- analogs in the gas phase. Well-resolved and rich spectral features are obtained for each species, yielding detailed electronic structure information, which is assigned with the aid of highlevel electronic structure calculations at the Coupled Cluster (CC) level of theory. The electron binding energies of [PtX3(C2H4)] are found to decrease with the size of halogen (4.57, 4.51, and 4.18 eV for X = Cl, Br, and I, respectively). The calculations indicate a synergistic η2 interaction [with interaction strengths of 1.54 (Cl), 1.37 (Br) and 1.10 eV (I)] between the perpendicular C2H4 fragment and the nearly horizontal planar PtX3- anions, resulting in activating the ethylene molecule. The detailed insights of the chemical bonding and underlying electronic structure can be used to benchmark interactions between olefins and transition metal complexes, which are crucial to a wide range of catalytic processes.

  7. Parallel on-axis phase-shifting holographic phase microscopy based on reflective point-diffraction interferometer with long-term stability.

    PubMed

    Guo, Rongli; Yao, Baoli; Gao, Peng; Min, Junwei; Han, Jun; Yu, Xun; Lei, Ming; Yan, Shaohui; Yang, Yanlong; Dan, Dan; Ye, Tong

    2013-05-20

    Parallel on-axis two-step phase-shifting reflective point-diffraction interferometry for holographic phase microscopy based on Michelson architecture is proposed. A cube beamsplitter splits the object wave into two copies within the two arms. The reference wave is rebuilt by low-pass filtering with a pinhole-masked mirror. Both object and reference waves are split into two beams by a grating in a 4f imaging system; thus, two interferograms with quadrature phase-shift can be acquired simultaneously with the aid of polarization elements. The approach has the merit of nanometers-scale phase stability over hours due to its quasi-common-path geometry. It can make full use of camera spatial bandwidth while its temporal resolution is as fast as the camera frame rate. Phase imaging on microscale specimen is implemented, and the experimental results demonstrate that the proposed approach is suitable for investigating dynamic processes.

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

    PubMed Central

    Kazemzadeh, Farnoud; Wong, Alexander

    2016-01-01

    Wide-field lensfree on-chip microscopy, which leverages holography principles to capture interferometric light-field encodings without lenses, is an emerging imaging modality with widespread interest given the large field-of-view compared to lens-based techniques. In this study, we introduce the idea of laser light-field fusion for lensfree on-chip phase contrast microscopy for detecting nanoparticles, where interferometric laser light-field encodings acquired using a lensfree, on-chip setup with laser pulsations at different wavelengths are fused to produce marker-free phase contrast images of particles at the nanometer scale. As a proof of concept, we demonstrate, for the first time, a wide-field lensfree on-chip instrument successfully detecting 300 nm particles across a large field-of-view of ~30 mm2 without any specialized or intricate sample preparation, or the use of synthetic aperture- or shift-based techniques. PMID:27958348

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

    PubMed

    Kazemzadeh, Farnoud; Wong, Alexander

    2016-12-13

    Wide-field lensfree on-chip microscopy, which leverages holography principles to capture interferometric light-field encodings without lenses, is an emerging imaging modality with widespread interest given the large field-of-view compared to lens-based techniques. In this study, we introduce the idea of laser light-field fusion for lensfree on-chip phase contrast microscopy for detecting nanoparticles, where interferometric laser light-field encodings acquired using a lensfree, on-chip setup with laser pulsations at different wavelengths are fused to produce marker-free phase contrast images of particles at the nanometer scale. As a proof of concept, we demonstrate, for the first time, a wide-field lensfree on-chip instrument successfully detecting 300 nm particles across a large field-of-view of ~30 mm(2) without any specialized or intricate sample preparation, or the use of synthetic aperture- or shift-based techniques.

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

    NASA Astrophysics Data System (ADS)

    Kazemzadeh, Farnoud; Wong, Alexander

    2016-12-01

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

  11. Twin-beams digital holography for 3D tracking and quantitative phase-contrast microscopy in microfluidics.

    PubMed

    Memmolo, Pasquale; Finizio, Andrea; Paturzo, Melania; Miccio, Lisa; Ferraro, Pietro

    2011-12-05

    We report on a compact twin-beam interferometer that can be adopted as a flexible diagnostic tool in microfluidic platforms with twofold functionality. The novel configuration allows 3D tracking of micro-particles and, at same time, can simultaneously furnish Quantitative Phase-contrast maps of tracked micro-objects by interference microscopy, without changing the configuration. Experimental demonstration is given on for in vitro cells in a microfluidic environment.

  12. Fractional pressure measurements in the W7-X startup phase with a spectroscopically assisted Penning gauge

    NASA Astrophysics Data System (ADS)

    Kremeyer, Thierry; Schmitz, Oliver; Wenzel, Uwe; Flesch, Kurt; W7-X Team

    2016-10-01

    Studies of helium exhaust from stellarator divertors is important to qualify for maintaining low overall helium concentrations for future reactors. Penning gauges assisted by spectroscopy were used to measure total neutral pressure and to resolve the D and He partial pressures. A similar system was installed on an outboard vacuum flange as a generic feasibility test for W7-X. In this contribution, initial results from the W7-X startup phase are shown. A compact CCD spectrometer with a range from 500 to 1000 nm was used to observe the Penning discharge. With a long integration time of 25s, He lines can be seen down to 10-5 mbar and Hα lines down to 10-6 mbar. The impact of He cleaning discharges on the in-situ neutral gas spectrum was measured and will be discussed in comparison to the mass spectrometers on the device. This initial test is the basis for developing optimized Penning gauges, which will be deployed to measure the D/He pressure ratios inside of the island divertor. The status of this development will be discussed. This work was funded in part by the Department of Energy under Grants DE-SC0012315 and DE-SC0014210 and from EUROfusion under Grant No 633053.

  13. Phase transformation of calcium oxalate dihydrate-monohydrate: Effects of relative humidity and new spectroscopic data

    NASA Astrophysics Data System (ADS)

    Conti, Claudia; Casati, Marco; Colombo, Chiara; Realini, Marco; Brambilla, Luigi; Zerbi, Giuseppe

    2014-07-01

    New data on vibrational properties of calcium oxalates and their controversial transformation mechanism are presented. We have focused on whewellite (CaC2O4·H2O) and weddellite [CaC2O4·(2 + x) H2O], the most common phases of calcium oxalate; these compounds occur in many organisms, in kidney stones and in particular kinds of films found on the surface of many works of art. Low temperature experiments carried out by Fourier transform infrared spectroscopy have highlighted both the high structural order in the crystalline state of whewellite and the disordered distribution of the zeolitic water molecules in weddellite. The synthesised nanocrystals of weddellite have been kept under different hygrometric conditions in order to study, by X-ray powder diffraction, the role of “external” water molecules on their stability. Moreover, in order to identify the different kinds of water molecules, a re-investigation, supported by quantum chemical calculations, of the observed vibrational spectra (IR and Raman) of whewellite has been conducted.

  14. A computational and spectroscopic study of the gas-phase conformers of adrenaline

    NASA Astrophysics Data System (ADS)

    Çarçabal, P.; Snoek, L. C.; van Mourik, T.

    The conformational landscapes of the neurotransmitter l-adrenaline (l-epinephrine) and its diastereoisomer pseudo-adrenaline, isolated in the gas phase and un-protonated, have been investigated by using a combination of mass-selected ultraviolet and infrared holeburn spectroscopy, following laser desorption of the sample into a pulsed supersonic argon jet, and DFT and ab initio computation (at the B3LYP/6-31+G*, MP2/6-31+G* and MP2/aug-cc-pVDZ levels of theory). Both for adrenaline and its diastereoisomer, pseudo-adrenaline, one dominant molecular conformation, very similar to the one seen in noradrenaline, has been observed. It could be assigned to an extended side-chain structure (AG1a) stabilized by an OH → N intramolecular hydrogen bond. An intramolecular hydrogen bond is also formed between the neighbouring hydroxyl groups on the catechol ring. The presence of further conformers for both diastereoisomers could not be excluded, but overlapping electronic spectra and low ion signals prevented further assignments.

  15. Characterization of U(VI)-phases in corroded cement products by micro(μ)-spectroscopic methods

    NASA Astrophysics Data System (ADS)

    Rothe, J.; Brendebach, B.; Bube, C.; Dardenne, K.; Denecke, M. A.; Kienzler, B.; Metz, V.; Prüßmann, T.; Rickers-Appel, K.; Schild, D.; Soballa, E.; Vitova, T.

    2013-04-01

    Cementation is an industrial scale conditioning method applied to fix and solidify liquid low and intermediate level radioactive wastes (LLW/ILW) prior to underground disposal in geological formations.To assist prognosis of the long-term safety of cemented waste, alteration of uranium doped cement productswas studied in chloride-rich solutions relevant for final LLW/ILW disposal in rock salt. After long-time exposure of the full-scale LLW/ILW simulates to concentrated NaCl and MgCl2 brines, solid samples were retrieved for chemical and mineralogical analysis with an emphasis on uranium speciation in the corroded cement matrix.Bulk and recent spatially resolved micro(μ) U L3-XAFS measurements point to the occurrence of a diuranate type U(VI) phase forming throughout the corroded cement monoliths. U-enriched hot spots with dimensions up to several tens of μm turn out to be generally X-ray amorphous.

  16. Molecular resolution of a dioleoyl-Sn-glycero-phosphocholine lipid bilayer in liquid by phase modulation atomic force microscopy

    NASA Astrophysics Data System (ADS)

    Hoel, Antonin; Osaki, Toshihisa; Takeuchi, Shoji; Volz, Sebastian; Kawakatsu, Hideki

    2012-08-01

    We present a molecular resolved image of a supported lipid bilayer using atomic force microscopy in liquid conditions. Due to the well-known molecular diffusion processes with velocities of a few nm/s occurring in free-standing membranes, this type of mapping is difficult to achieve. We have obtained a sub-nanometer scale resolution of a dioleoyl-sn-glycero-phosphocholine membrane because of the high sensitivity of our phase modulated microscope and the slowed down diffusion due to the interaction with the substrate. A pair function analysis has revealed local disordered and gel phases with an order range limited to the first neighbour.

  17. Self-interference fluorescence microscopy with three-phase detection for depth-resolved confocal epi-fluorescence imaging.

    PubMed

    Braaf, Boy; de Boer, Johannes F

    2017-03-20

    Three-dimensional confocal fluorescence imaging of in vivo tissues is challenging due to sample motion and limited imaging speeds. In this paper a novel method is therefore presented for scanning confocal epi-fluorescence microscopy with instantaneous depth-sensing based on self-interference fluorescence microscopy (SIFM). A tabletop epi-fluorescence SIFM setup was constructed with an annular phase plate in the emission path to create a spectral self-interference signal that is phase-dependent on the axial position of a fluorescent sample. A Mach-Zehnder interferometer based on a 3 × 3 fiber-coupler was developed for a sensitive phase analysis of the SIFM signal with three photon-counter detectors instead of a spectrometer. The Mach-Zehnder interferometer created three intensity signals that alternately oscillated as a function of the SIFM spectral phase and therefore encoded directly for the axial sample position. Controlled axial translation of fluorescent microsphere layers showed a linear dependence of the SIFM spectral phase with sample depth over axial image ranges of 500 µm and 80 µm (3.9 × Rayleigh range) for 4 × and 10 × microscope objectives respectively. In addition, SIFM was in good agreement with optical coherence tomography depth measurements on a sample with indocyanine green dye filled capillaries placed at multiple depths. High-resolution SIFM imaging applications are demonstrated for fluorescence angiography on a dye-filled capillary blood vessel phantom and for autofluorescence imaging on an ex vivo fly eye.

  18. Comprehensive analysis of TEM methods for LiFePO4/FePO4 phase mapping: spectroscopic techniques (EFTEM, STEM-EELS) and STEM diffraction techniques (ACOM-TEM).

    PubMed

    Mu, X; Kobler, A; Wang, D; Chakravadhanula, V S K; Schlabach, S; Szabó, D V; Norby, P; Kübel, C

    2016-11-01

    Transmission electron microscopy (TEM) has been used intensively in investigating battery materials, e.g. to obtain phase maps of partially (dis)charged (lithium) iron phosphate (LFP/FP), which is one of the most promising cathode material for next generation lithium ion (Li-ion) batteries. Due to the weak interaction between Li atoms and fast electrons, mapping of the Li distribution is not straightforward. In this work, we revisited the issue of TEM measurements of Li distribution maps for LFP/FP. Different TEM techniques, including spectroscopic techniques (energy filtered (EF)TEM in the energy range from low-loss to core-loss) and a STEM diffraction technique (automated crystal orientation mapping (ACOM)), were applied to map the lithiation of the same location in the same sample. This enabled a direct comparison of the results. The maps obtained by all methods showed excellent agreement with each other. Because of the strong difference in the imaging mechanisms, it proves the reliability of both the spectroscopic and STEM diffraction phase mapping. A comprehensive comparison of all methods is given in terms of information content, dose level, acquisition time and signal quality. The latter three are crucial for the design of in-situ experiments with beam sensitive Li-ion battery materials. Furthermore, we demonstrated the power of STEM diffraction (ACOM-STEM) providing additional crystallographic information, which can be analyzed to gain a deeper understanding of the LFP/FP interface properties such as statistical information on phase boundary orientation and misorientation between domains.

  19. Simple and fast spectral domain algorithm for quantitative phase imaging of living cells with digital holographic microscopy

    NASA Astrophysics Data System (ADS)

    Min, Junwei; Yao, Baoli; Ketelhut, Steffi; Kemper, Björn

    2017-02-01

    The modular combination of optical microscopes with digital holographic microscopy (DHM) has been proven to be a powerful tool for quantitative live cell imaging. The introduction of condenser and different microscope objectives (MO) simplifies the usage of the technique and makes it easier to measure different kinds of specimens with different magnifications. However, the high flexibility of illumination and imaging also causes variable phase aberrations that need to be eliminated for high resolution quantitative phase imaging. The existent phase aberrations compensation methods either require add additional elements into the reference arm or need specimen free reference areas or separate reference holograms to build up suitable digital phase masks. These inherent requirements make them unpractical for usage with highly variable illumination and imaging systems and prevent on-line monitoring of living cells. In this paper, we present a simple numerical method for phase aberration compensation based on the analysis of holograms in spatial frequency domain with capabilities for on-line quantitative phase imaging. From a single shot off-axis hologram, the whole phase aberration can be eliminated automatically without numerical fitting or pre-knowledge of the setup. The capabilities and robustness for quantitative phase imaging of living cancer cells are demonstrated.

  20. X-ray Phase Imaging Microscopy with Two-Dimensional Knife-Edge Filters

    NASA Astrophysics Data System (ADS)

    Choi, Jaeho; Park, Yong-Sung

    2012-04-01

    A novel scheme of X-ray differential phase imaging was implemented with an array source and a two-dimensional Foucault knife-edge (2DFK). A pinhole array lens was employed to manipulate the X-ray beam on the Fourier space. An emerging biaxial scanning procedure was also demonstrated with the periodic 2DFK. The differential phase images (DPIs) of the midrib in a leaf of a rose bush were visualized to verify the phase imaging of biological specimens by the proposed method. It also has features of depicting multiple-stack phase images, and rendering morphological DPIs, because it acquires pure phase information.

  1. Recovering correct phase information in multiwavelength digital holographic microscopy by compensation for chromatic aberrations.

    PubMed

    De Nicola, S; Finizio, A; Pierattini, G; Alfieri, D; Grilli, S; Sansone, L; Ferraro, P

    2005-10-15

    We demonstrate experimentally that correct phase imaging without 2pi ambiguity is obtainable in digital holography by using a multiwavelength approach in the microscope configuration. We describe a general approach for removing chromatic aberrations and for controlling the pixel size of the reconstructed phase image in multiwavelength digital holography when the Fourier transform method is adopted for the numerical reconstruction of digital holograms. The retrieved phase is affected by the unavoidable, unwanted chromatic aberration. The correct phase can be obtained by evaluating the phase from the reference holograms reconstructed at different wavelengths to compensate for the chromatic aberration.

  2. In-focus electron microscopy of frozen-hydrated biological samples with a Boersch phase plate.

    PubMed

    Barton, B; Rhinow, D; Walter, A; Schröder, R; Benner, G; Majorovits, E; Matijevic, M; Niebel, H; Müller, H; Haider, M; Lacher, M; Schmitz, S; Holik, P; Kühlbrandt, W

    2011-12-01

    We report the implementation of an electrostatic Einzel lens (Boersch) phase plate in a prototype transmission electron microscope dedicated to aberration-corrected cryo-EM. The combination of phase plate, C(s) corrector and Diffraction Magnification Unit (DMU) as a new electron-optical element ensures minimal information loss due to obstruction by the phase plate and enables in-focus phase contrast imaging of large macromolecular assemblies. As no defocussing is necessary and the spherical aberration is corrected, maximal, non-oscillating phase contrast transfer can be achieved up to the information limit of the instrument. A microchip produced by a scalable micro-fabrication process has 10 phase plates, which are positioned in a conjugate, magnified diffraction plane generated by the DMU. Phase plates remained fully functional for weeks or months. The large distance between phase plate and the cryo sample permits the use of an effective anti-contaminator, resulting in ice contamination rates of <0.6 nm/h at the specimen. Maximal in-focus phase contrast was obtained by applying voltages between 80 and 700 mV to the phase plate electrode. The phase plate allows for in-focus imaging of biological objects with a signal-to-noise of 5-10 at a resolution of 2-3 nm, as demonstrated for frozen-hydrated virus particles and purple membrane at liquid-nitrogen temperature.

  3. Comparison of mounting methods for the evaluation of fibers by phase contrast microscopy.

    PubMed

    Lee, Eun Gyung; Pang, Thomas W S; Nelson, John; Andrew, Mike; Harper, Martin

    2011-07-01

    The objectives of this study were to evaluate mounting methods for fiber examination of air sample filters by phase contrast microscopy (PCM) and to evaluate differences in fiber counts that might be due to fiber movement. Acetone/triacetin (AT) with various amounts of triacetin and acetone/Euparal (AE) where the mounting medium was placed between the cleared filter wedge and the coverslip were tested as a function of time. Field sample slides collected from a taconite iron-ore processing mill, a tremolitic talc-ore processing mill, and from around a crusher in a meta-basalt stone quarry were prepared with relocatable coverslips to revisit the same field areas on the slides. For each slide, three or four field areas were randomly selected and pictures were taken every 2 weeks to determine any sign of fiber movement over time. For 11 AT slides (named as AT-3.5) prepared with 3.5 μl of the mounting medium according to the NIOSH 7400 method, no fiber movements were detected over 59 weeks. On the other hand, AT slides prepared with larger quantities (10, 15, and 20 μl) of the mounting medium (named as AT-10) and AE slides prepared with ∼10 μl mounting medium showed fiber movement from the eighth day at the earliest. Fiber movement began earlier for the slides mounted with excess triacetin than for those mounted with Euparal. The sample slide storage method, either vertically or horizontally, did not seem to accelerate fiber movement. Additionally, two other modified methods, dimethylformamide solution/Euparal (mDE) and dimethylformamide solution/triacetin (mDT), were also prepared where the mounting medium was placed between the cleared filter wedge and the glass slide. The findings of fiber movements were similar; when 3.5 μl of triacetin was used for the mDT slides, fiber movements were not detected, while fibers on slides prepared with 10 μl triacetin (mDT-10) moved around. No fiber movements were observed for the mDE slides at any time during 59 weeks. Once

  4. Transmission electron microscopy analysis of phase separation in GaInAsSb films grown on GaSb substrate.

    PubMed

    Szczeszek, P; Amariei, A; Schöne, J; Zoulis, G; Vouroutzis, N; Polychroniadis, E K; Stróz, D

    2006-10-01

    The GaSb-based quaternary alloys are a good choice for thermophotovoltaic applications. The thermophotovoltaic cell converts infrared radiation to electricity, using the same principles as photovoltaic devices. The aim of the present work was the microstructural study of such an alloy, namely Ga(0.84)In(0.16)As(0.12)Sb(0.88). A thin film of the material was grown by metal organic vapour phase epitaxy on a (100)alpha-->[111]B (alpha = 2 degrees, 4 degrees, 6 degrees) GaSb substrate. The GaInAsSb alloy has an appropriate band gap, but suffers from a phase separation consisting of GaAs-rich and InSb-rich regions that is disadvantageous for cell efficiency. In this work, we employed a morphological approach to phase separation, with the use of conventional transmission electron microscopy and atomic force microscopy. The phase separation occurs in two different orientations: parallel to the growth direction (vertical) and inclined (lateral). After application of fast Fourier transformation filtering, the vertical periodicity was found to be lambda = 5 nm for the pair (black and white) of layers independently of the cut-off angle, whereas the lateral periodicity was related to it.

  5. Late stages of phase separation/gelation of isotropic solutions of rod-like polymers by video microscopy

    NASA Astrophysics Data System (ADS)

    Chowdhury, Aslam H.; Russo, Paul S.

    1990-05-01

    The late stages of phase separation/gelation in concentrated solutions of poly(γ-benzyl-α, L-glutamate) in N,N-dimethylformamide containing 2% added water have been observed by video optical microscopy. Microscopic phase separation is directly evident on cooling homogeneous, isotropic solutions. Within the phase separating mixture, diffuse structural features are separated from one another by a characteristic distance. Fourier transforms of the real space images, equivalent to scattering patterns, show a radially symmetric ring, which collapses to lower wave number as gelation proceeds. The wave number associated with the maximum intensity, qm, obeys a scaling relationship consistent with the Lifshitz-Slyozov evaporation/condensation model, also consistent with the Binder-Stauffer cluster dynamics model: qm=t-1/3, where t is time. A more general scaling relationship proposed by Furukawa is obeyed very well if the dimensionality of the growth of the new phases is 3. The advantages of video microscopy for such studies and possible implications for the role of spinodal decomposition in rod-like polymer solutions are discussed.

  6. Calorimetric and spectroscopic studies of the thermotropic phase behavior of lipid bilayer model membranes composed of a homologous series of linear saturated phosphatidylserines.

    PubMed Central

    Lewis, R N; McElhaney, R N

    2000-01-01

    The thermotropic phase behavior of lipid bilayer model membranes composed of the even-numbered, N-saturated 1,2-diacyl phosphatidylserines was studied by differential scanning calorimetry and by Fourier-transform infrared and (31)P-nuclear magnetic resonance spectroscopy. At pH 7.0, 0.1 M NaCl and in the absence of divalent cations, aqueous dispersions of these lipids, which have not been incubated at low temperature, exhibit a single calorimetrically detectable phase transition that is fully reversible, highly cooperative, and relatively energetic, and the transition temperatures and enthalpies increase progressively with increases in hydrocarbon chain length. Our spectroscopic observations confirm that this thermal event is a lamellar gel (L(beta))-to-lamellar liquid crystalline (L(alpha)) phase transition. However, after low temperature incubation, the L(beta)/L(alpha) phase transition of dilauroyl phosphatidylserine is replaced by a higher temperature, more enthalpic, and less cooperative phase transition, and an additional lower temperature, less enthalpic, and less cooperative phase transition appears in the longer chain phosphatidylserines. Our spectroscopic results indicate that this change in thermotropic phase behavior when incubated at low temperatures results from the conversion of the L(beta) phase to a highly ordered lamellar crystalline (L(c)) phase. Upon heating, the L(c) phase of dilauroyl phosphatidylserine converts directly to the L(alpha) phase at a temperature slightly higher than that of its original L(beta)/L(alpha) phase transition. Calorimetrically, this process is manifested by a less cooperative but considerably more energetic, higher-temperature phase transition, which replaces the weaker L(beta)/L(alpha) phase transition alluded to above. However, with the longer chain compounds, the L(c) phase first converts to the L(beta) phase at temperatures some 10-25 degrees C below that at which the L(beta) phase converts to the L(alpha) phase

  7. Assessment of amide I spectroscopic maps for a gas-phase peptide using IR-UV double-resonance spectroscopy and density functional theory calculations.

    PubMed

    Carr, J K; Zabuga, A V; Roy, S; Rizzo, T R; Skinner, J L

    2014-06-14

    The spectroscopy of amide I vibrations has become a powerful tool for exploring protein structure and dynamics. To help with spectral interpretation, it is often useful to perform molecular dynamics (MD) simulations. To connect spectroscopic experiments to simulations in an efficient manner, several researchers have proposed "maps," which relate observables in classical MD simulations to quantum spectroscopic variables. It can be difficult to discern whether errors in the theoretical results (compared to experiment) arise from inaccuracies in the MD trajectories or in the maps themselves. In this work, we evaluate spectroscopic maps independently from MD simulations by comparing experimental and theoretical spectra for a single conformation of the α-helical model peptide Ac-Phe-(Ala)5-Lys-H(+) in the gas phase. Conformation-specific experimental spectra are obtained for the unlabeled peptide and for several singly and doubly (13)C-labeled variants using infrared-ultraviolet double-resonance spectroscopy, and these spectra are found to be well-modeled by density functional theory (DFT) calculations at the B3LYP/6-31G** level. We then compare DFT results for the deuterated and (13)C(18)O-labeled peptide with those from spectroscopic maps developed and used previously by the Skinner group. We find that the maps are typically accurate to within a few cm(-1) for both frequencies and couplings, having larger errors only for the frequencies of terminal amides.

  8. Assessment of amide I spectroscopic maps for a gas-phase peptide using IR-UV double-resonance spectroscopy and density functional theory calculations

    SciTech Connect

    Carr, J. K.; Roy, S.; Skinner, J. L.; Zabuga, A. V.; Rizzo, T. R.

    2014-06-14

    The spectroscopy of amide I vibrations has become a powerful tool for exploring protein structure and dynamics. To help with spectral interpretation, it is often useful to perform molecular dynamics (MD) simulations. To connect spectroscopic experiments to simulations in an efficient manner, several researchers have proposed “maps,” which relate observables in classical MD simulations to quantum spectroscopic variables. It can be difficult to discern whether errors in the theoretical results (compared to experiment) arise from inaccuracies in the MD trajectories or in the maps themselves. In this work, we evaluate spectroscopic maps independently from MD simulations by comparing experimental and theoretical spectra for a single conformation of the α-helical model peptide Ac-Phe-(Ala){sub 5}-Lys-H{sup +} in the gas phase. Conformation-specific experimental spectra are obtained for the unlabeled peptide and for several singly and doubly {sup 13}C-labeled variants using infrared-ultraviolet double-resonance spectroscopy, and these spectra are found to be well-modeled by density functional theory (DFT) calculations at the B3LYP/6-31G** level. We then compare DFT results for the deuterated and {sup 13}C{sup 18}O-labeled peptide with those from spectroscopic maps developed and used previously by the Skinner group. We find that the maps are typically accurate to within a few cm{sup −1} for both frequencies and couplings, having larger errors only for the frequencies of terminal amides.

  9. Assessment of amide I spectroscopic maps for a gas-phase peptide using IR-UV double-resonance spectroscopy and density functional theory calculations

    PubMed Central

    Carr, J. K.; Zabuga, A. V.; Roy, S.; Rizzo, T. R.; Skinner, J. L.

    2014-01-01

    The spectroscopy of amide I vibrations has become a powerful tool for exploring protein structure and dynamics. To help with spectral interpretation, it is often useful to perform molecular dynamics (MD) simulations. To connect spectroscopic experiments to simulations in an efficient manner, several researchers have proposed “maps,” which relate observables in classical MD simulations to quantum spectroscopic variables. It can be difficult to discern whether errors in the theoretical results (compared to experiment) arise from inaccuracies in the MD trajectories or in the maps themselves. In this work, we evaluate spectroscopic maps independently from MD simulations by comparing experimental and theoretical spectra for a single conformation of the α-helical model peptide Ac-Phe-(Ala)5-Lys-H+ in the gas phase. Conformation-specific experimental spectra are obtained for the unlabeled peptide and for several singly and doubly 13C-labeled variants using infrared-ultraviolet double-resonance spectroscopy, and these spectra are found to be well-modeled by density functional theory (DFT) calculations at the B3LYP/6-31G** level. We then compare DFT results for the deuterated and 13C18O-labeled peptide with those from spectroscopic maps developed and used previously by the Skinner group. We find that the maps are typically accurate to within a few cm−1 for both frequencies and couplings, having larger errors only for the frequencies of terminal amides. PMID:24929378

  10. Assessment of amide I spectroscopic maps for a gas-phase peptide using IR-UV double-resonance spectroscopy and density functional theory calculations

    NASA Astrophysics Data System (ADS)

    Carr, J. K.; Zabuga, A. V.; Roy, S.; Rizzo, T. R.; Skinner, J. L.

    2014-06-01

    The spectroscopy of amide I vibrations has become a powerful tool for exploring protein structure and dynamics. To help with spectral interpretation, it is often useful to perform molecular dynamics (MD) simulations. To connect spectroscopic experiments to simulations in an efficient manner, several researchers have proposed "maps," which relate observables in classical MD simulations to quantum spectroscopic variables. It can be difficult to discern whether errors in the theoretical results (compared to experiment) arise from inaccuracies in the MD trajectories or in the maps themselves. In this work, we evaluate spectroscopic maps independently from MD simulations by comparing experimental and theoretical spectra for a single conformation of the α-helical model peptide Ac-Phe-(Ala)5-Lys-H+ in the gas phase. Conformation-specific experimental spectra are obtained for the unlabeled peptide and for several singly and doubly 13C-labeled variants using infrared-ultraviolet double-resonance spectroscopy, and these spectra are found to be well-modeled by density functional theory (DFT) calculations at the B3LYP/6-31G** level. We then compare DFT results for the deuterated and 13C18O-labeled peptide with those from spectroscopic maps developed and used previously by the Skinner group. We find that the maps are typically accurate to within a few cm-1 for both frequencies and couplings, having larger errors only for the frequencies of terminal amides.

  11. Dynamic phase microscopy, a new method to detect viable and killed spores and to estimate the heterogeneity of spore populations

    NASA Astrophysics Data System (ADS)

    Tychinsky, V. P.; Mulyukin, A. L.; Lisovskii, V. V.; Nikolaev, Yu. A.; Kretushev, A. V.; Vyshenskaya, T. V.; Suzina, N. E.; Duda, V. I.; El-Registan, G. I.

    One of the challenging tasks in monitoring studies is to estimate heterogeneity of microbial populations by the physiological state and potential viability of individual cells especially with regard of their ability to withstand various environmental assaults Previously we described some approaches based on electron microscopy methods to discriminate vegetative dormant and dead cells in both aged microbial cultures and environmental samples including permafrost In this communication we propose to extend the arsenal of microscopy methods for monitoring studies by a new non-invasive and informative method - dynamic phase microscopy DPM The substantial advantage of DPM is that it gives quantitative digitized data of un-destroyed living microscopic objects exemplified in our work by Bacillus licheniformis spores Using DPM made it possible to record interference images of objects spores and to produce picture of their phase thickness PT that is the optical path difference in nm Thus it was demonstrated the remarkable difference in the PT of spores at different physiological states dormant germinating and heat-killed spores had PT values of 80 nm 40-50 nm and 20 nm respectively The other found criterion to distinguish between spores was the PT fluctuations In contrast to dormant and killed spores the PT of germinating spores oscillated with amplitude of up to 7 nm with typical frequencies of 1 3 and 3 4 Hz A combination of the recorded PT values and PT fluctuations gave a key to detect viable and dead cells Under the conditions that did not

  12. Coherence-controlled holographic microscopy principle embodiment into Q-PHASE microscope: story of a successful technology transfer

    NASA Astrophysics Data System (ADS)

    Lostak, M.; Chmelik, R.

    2016-03-01

    Curiously, the coherence-controlled holographic microscopy (CCHM) was brought into the world owing to the endeavor of Chmelik's team at Brno University of Technology (BUT) to avoid scanning in confocal microscopy. As coherence gating seemed to be the way, the Leith & Upatnieks proposal of incoherent holography had been considered attractive. Their method made interference system free from strict dependence on both spatial and temporal coherence. Off axis holographic system proposed on such basis has been proved capable of coherence based depth discrimination in single wide-field shot in reflected-light arrangement. Consequently, extremely low-coherence holographic imaging had been found highly contributive also to the image quality depriving it from coherence artefacts and improving its transversal resolution. This is why CCHM promised high precision of quantitative phase imaging (QPI) in transmitted light set up that was realized for cell biology. However the cost of necessarily complicated optical design and need of very precise mechanics forced the team of prof Chmelik at BUT to search for a company capable of mastering the instrument. It was TESCAN ORSAY the highly successful scanning electron microscopes producer that finally took charge of the commercial design. Long-term collaboration of the company with BUT made possible both the CCHM technology successful transfer up to Q-PHASE microscope production as well as the company Light microscopy division reinforcement. This contribution merges views of CCHM technology author and the TESCAN development team.

  13. Phase-resolved imaging of edge-mode spin waves using scanning transmission x-ray microscopy

    NASA Astrophysics Data System (ADS)

    Cheng, C.; Cao, W.; Bailey, W. E.

    2017-02-01

    We have imaged the excitation of small-amplitude spin-wave eigenmodes, localized within ∼100 nm of the vertices of nanoscale Ni81Fe19 ellipses, using time-resolved scanning transmission x-ray microscopy (STXM) at 2 GHz and resolution of 70 nm. Taking advantage of the buried-layer sensitivity of STXM, we find that the magnetization precession at the two vertices changes from predominantly in-phase to out-of-phase in samples with and without a conductive layer deposited over the ellipses. As a plausible interpretation for the reversal in phase, we propose that unexpectedly strong Oersted fields are generated in the discontinuous overlayer, although effects of edge roughness cannot be fully excluded. The results demonstrate the capabilities of STXM to image small-amplitude, GHz magnetization dynamics with the potential to map rf magnetic fields on the nanoscale.

  14. Two-dimensional phase separation of block copolymer and homopolymer blend studied by scanning near-field optical microscopy

    NASA Astrophysics Data System (ADS)

    Aoki, H.; Kunai, Y.; Ito, S.; Yamada, H.; Matsushige, K.

    2002-03-01

    The phase separation of dye-labeled poly(isobutyl methacrylate) (PiBMA) and poly(octadecyl methacrylate) (PODMA) in two-dimensional monolayers was investigated by scanning near-field optical microscopy (SNOM). The energy transfer (ET) efficiency between dyes introduced to the polymers was mapped with SNOM for a PiBMA/PODMA homopolymer blend monolayer. The ET fluorescence images revealed that the phase boundary had a width of a few hundred nanometers, which was considerably larger than that expected in a three-dimensional bulk state. The micro-phase separation of a diblock copolymer PiBMA- block-PODMA was also discussed. Ribbon-like lamellar structures with a width of ca. 300 nm were observed, and the block polymer chain took a highly elongated conformation in two dimensions.

  15. Superconducting scanning tunneling microscopy tips in a magnetic field: Geometry-controlled order of the phase transition

    SciTech Connect

    Eltschka, Matthias Jäck, Berthold; Assig, Maximilian; Etzkorn, Markus; Ast, Christian R.; Kondrashov, Oleg V.; Skvortsov, Mikhail A.; Kern, Klaus

    2015-09-21

    The properties of geometrically confined superconductors significantly differ from their bulk counterparts. Here, we demonstrate the geometrical impact for superconducting scanning tunneling microscopy (STM) tips, where the confinement ranges from the atomic to the mesoscopic scale. To this end, we compare the experimentally determined magnetic field dependence for several vanadium tips to microscopic calculations based on the Usadel equation. For our theoretical model of a superconducting cone, we find a direct correlation between the geometry and the order of the superconducting phase transition. Increasing the opening angle of the cone changes the phase transition from first to second order. Comparing our experimental findings to the theory reveals first and second order quantum phase transitions in the vanadium STM tips. In addition, the theory also explains experimentally observed broadening effects by the specific tip geometry.

  16. Enhanced quantitative phase imaging in self-interference digital holographic microscopy using an electrically focus tunable lens

    PubMed Central

    Schubert, Robin; Vollmer, Angelika; Ketelhut, Steffi; Kemper, Björn

    2014-01-01

    Self-interference digital holographic microscopy (DHM) has been found particular suitable for simplified quantitative phase imaging of living cells. However, a main drawback of the self-interference DHM principle are scattering patterns that are induced by the coherent nature of the laser light which affect the resolution for detection of optical path length changes. We present a simple and efficient technique for the reduction of coherent disturbances in quantitative phase images. Therefore, amplitude and phase of the sample illumination are modulated by an electrically focus tunable lens. The proposed method is in particular convenient with the self-interference DHM concept. Results from the characterization of the method show that a reduction of coherence induced disturbances up to 70 percent can be achieved. Finally, the performance for enhanced quantitative imaging of living cells is demonstrated. PMID:25574433

  17. Enhanced quantitative phase imaging in self-interference digital holographic microscopy using an electrically focus tunable lens.

    PubMed

    Schubert, Robin; Vollmer, Angelika; Ketelhut, Steffi; Kemper, Björn

    2014-12-01

    Self-interference digital holographic microscopy (DHM) has been found particular suitable for simplified quantitative phase imaging of living cells. However, a main drawback of the self-interference DHM principle are scattering patterns that are induced by the coherent nature of the laser light which affect the resolution for detection of optical path length changes. We present a simple and efficient technique for the reduction of coherent disturbances in quantitative phase images. Therefore, amplitude and phase of the sample illumination are modulated by an electrically focus tunable lens. The proposed method is in particular convenient with the self-interference DHM concept. Results from the characterization of the method show that a reduction of coherence induced disturbances up to 70 percent can be achieved. Finally, the performance for enhanced quantitative imaging of living cells is demonstrated.

  18. Quartz tuning fork-based frequency modulation atomic force spectroscopy and microscopy with all digital phase-locked loop

    NASA Astrophysics Data System (ADS)

    An, Sangmin; Hong, Mun-heon; Kim, Jongwoo; Kwon, Soyoung; Lee, Kunyoung; Lee, Manhee; Jhe, Wonho

    2012-11-01

    We present a platform for the quartz tuning fork (QTF)-based, frequency modulation atomic force microscopy (FM-AFM) system for quantitative study of the mechanical or topographical properties of nanoscale materials, such as the nano-sized water bridge formed between the quartz tip (˜100 nm curvature) and the mica substrate. A thermally stable, all digital phase-locked loop is used to detect the small frequency shift of the QTF signal resulting from the nanomaterial-mediated interactions. The proposed and demonstrated novel FM-AFM technique provides high experimental sensitivity in the measurement of the viscoelastic forces associated with the confined nano-water meniscus, short response time, and insensitivity to amplitude noise, which are essential for precision dynamic force spectroscopy and microscopy.

  19. Quartz tuning fork-based frequency modulation atomic force spectroscopy and microscopy with all digital phase-locked loop.

    PubMed

    An, Sangmin; Hong, Mun-heon; Kim, Jongwoo; Kwon, Soyoung; Lee, Kunyoung; Lee, Manhee; Jhe, Wonho

    2012-11-01

    We present a platform for the quartz tuning fork (QTF)-based, frequency modulation atomic force microscopy (FM-AFM) system for quantitative study of the mechanical or topographical properties of nanoscale materials, such as the nano-sized water bridge formed between the quartz tip (~100 nm curvature) and the mica substrate. A thermally stable, all digital phase-locked loop is used to detect the small frequency shift of the QTF signal resulting from the nanomaterial-mediated interactions. The proposed and demonstrated novel FM-AFM technique provides high experimental sensitivity in the measurement of the viscoelastic forces associated with the confined nano-water meniscus, short response time, and insensitivity to amplitude noise, which are essential for precision dynamic force spectroscopy and microscopy.

  20. Super resolution microscopy of lipid bilayer phases and single molecule kinetic studies on merocyanine 540 bound lipid vesicles

    NASA Astrophysics Data System (ADS)

    Kuo, Chin-Kuei

    Recently, observing biological process and structural details in live cell became feasible after the introduction of super-resolution microscopy. Super-resolution microscopy by single molecule localization is the method that has commonly been used for such purpose. There are mainly three approaches to it: stochastic optical reconstruction microscopy (STORM), photoactivated localization microscopy (PALM), and point accumulation in nanoscale topology (PAINT). STORM and PALM rely on external laser control and use of photoactivable fluorescent protein or photoswitchable dyes and are technically challenging. The PAINT method relies on the control of thermal reaction rates to enable the switching between bright and dark states. Therefore, many conventional fluorescent probes can be applied in PAINT method and the images denote different information composed of interactions between the probe and its immediate environment by variations of probe parameters. The existence of lipid rafts has been under debates for decades due to the lack of a tool to directly visualize them in live cells. In the thesis, we combine PAINT with a phase sensitive dye, Merocyanine 540, to enable nanoscale observation of phase separation on supported lipid bilayers of mixed liquid/gel phases. The imaging results are presented in the chapter 3. Given that this is the first example of visualization of nanoscale phase separation of lipid bilayers using an optical microscope, we further looked into the kinetics of MC540 monomer dimer equilibrium in lipid bilayers using single molecule intensity time trajectory analysis and polarization dependent imaging. Our finding confirms that perpendicular monomeric MC540 (to the membrance surface) is the emitting speices in our system and it stays fluorescent for roughly 3 ms before it switches off to dark states. This part of analysis is presented in the chapter 4. All the materials, procedures to carry out experiments and data analysis, methods involved in our

  1. Calorimetric and spectroscopic evidence of chain-melting in smectic E and smectic A phases of 4-alkyl-4'-isothiocyanatobiphenyl (nTCB).

    PubMed

    Yamamura, Yasuhisa; Adachi, Takuya; Miyazawa, Takahito; Horiuchi, Katsuya; Sumita, Masato; Massalska-Arodź, Maria; Urban, Stanisław; Saito, Kazuya

    2012-08-02

    To confirm the molten state of the alkyl chain in soft crystalline phase, smectic E (SmE) phase, thermodynamic and spectroscopic analyses were performed on 4-n-alkyl-4'-isothiocyanatobiphenyl (nTCB, n: the number of carbon atoms in the alkyl group). DSC results of 11TCB and 12TCB, having extra smectic A phase besides smectic E phase, show that their chain-length dependence of entropies of transition (Δ(trs)S) from the ordered crystalline (OC) phase to the SmE phase matches the trend found for nTCB (n = 4-10), while no chain-length dependence is observed in Δ(trs)S at the SmE-to-SmA and SmA-to-isotropic liquid (IL) phase transitions in 11TCB and 12TCB. Temperature dependences of FT-IR spectra of six compounds (n = 2, 3, 5, 8, 10, and 12) were recorded. The CH stretching modes of the chain exhibited more pronounced change at the transition from the OC to the SmE phase than at the transition from the SmE phase to the IL or SmA phase. These results indicate that the alkyl chain is molten in the SmE phase as in IL. The disordering process of nTCB molecules from the OC to IL via anisotropic mesophases is discussed in terms of entropy.

  2. Dissecting eukaryotic cells by coherent phase microscopy: quantitative analysis of quiescent and activated T lymphocytes

    PubMed Central

    Kretushev, Alexander V.; Vyshenskaya, Tatiana V.; Shtil, Alexander A.

    2012-01-01

    Abstract. We present a concept for quantitative characterization of a functional state of an individual eukaryotic cell based on interference imaging. The informative parameters of the phase images of quiescent and mitogen-activated T lymphocytes included the phase thickness, phase volume, the area, and the size of organelles. These parameters were obtained without a special hypothesis about cell structure. Combinations of these parameters generated a “phase portrait” of the cell. A simplified spherical multilayer optic model of a T lymphocyte was used to calculate the refractivity profile, to identify structural elements of the image with the organelles, and to interpret the parameters of the phase portrait. The values of phase image parameters underwent characteristic changes in the course of mitogenic stimulation of T cells; thereby, the functional state of individual cells can be described using these parameters. Because the values of the components of the phase portrait are measured in absolute units, it is possible to compare the parameters of images obtained with different interference microscopes. Thus, the analysis of phase portraits provides a new and perspective approach for quantitative, real-time analysis of subcellular structure and physiologic state of an individual cell. PMID:22894503

  3. Dissecting eukaryotic cells by coherent phase microscopy: quantitative analysis of quiescent and activated T lymphocytes

    NASA Astrophysics Data System (ADS)

    Tychinsky, Vladimir P.; Kretushev, Alexander V.; Vyshenskaya, Tatiana V.; Shtil, Alexander A.

    2012-07-01

    We present a concept for quantitative characterization of a functional state of an individual eukaryotic cell based on interference imaging. The informative parameters of the phase images of quiescent and mitogen-activated T lymphocytes included the phase thickness, phase volume, the area, and the size of organelles. These parameters were obtained without a special hypothesis about cell structure. Combinations of these parameters generated a ``phase portrait'' of the cell. A simplified spherical multilayer optic model of a T lymphocyte was used to calculate the refractivity profile, to identify structural elements of the image with the organelles, and to interpret the parameters of the phase portrait. The values of phase image parameters underwent characteristic changes in the course of mitogenic stimulation of T cells; thereby, the functional state of individual cells can be described using these parameters. Because the values of the components of the phase portrait are measured in absolute units, it is possible to compare the parameters of images obtained with different interference microscopes. Thus, the analysis of phase portraits provides a new and perspective approach for quantitative, real-time analysis of subcellular structure and physiologic state of an individual cell.

  4. Physical compensation of phase curvature in digital holographic microscopy by use of programmable liquid lens.

    PubMed

    Doblas, Ana; Hincapie-Zuluaga, Diego; Saavedra, Genaro; Martínez-Corral, Manuel; Garcia-Sucerquia, Jorge

    2015-06-01

    Quantitative phase measurements obtained with digital holographic microscopes are strongly dependent on the optical arrangement of the imaging system. The nontelecentric operation provides phase measurements affected by a parabolic phase factor and requires numerical postprocessing, which does not always remove all the perturbation. Accurate phase measurements are achieved by using the imaging system in telecentric mode. Unfortunately, this condition is not accomplished when a commercial microscope is used as the imaging system. In this paper, we present an approach for obtaining accurate phase measurements in nontelecentric imaging systems without the need for numerical postprocessing. The method uses an electrically tunable liquid lens to illuminate the sample so that the perturbing parabolic wavefront is cancelled out. Experimental holograms of a Fresnel lens and a section of the thorax of a Drosophila melanogaster fly are captured to verify the proposed method.

  5. The HCP To BCC Phase Transformation in Ti Characterized by Nanosecond Electron Microscopy

    SciTech Connect

    Campbell, G; LaGrange, T; King, W; Colvin, J; Ziegler, A; Browning, N; Kleinschmidt, H; Bostanjoglo, O

    2005-06-21

    The general class of martensitic phase transformations occurs by a rapid lattice-distortive mechanism, where kinetics and morphology of the transformation are dominated by the strain energy. Since transformation is diffusionless, phase fronts propagate through a crystal with great speed that can approach the speed of sound. We have observed a particular example of this class of phase transformation, the hexagonal close packed (HCP) to body centered cubic (BCC) transformation in titanium that is driven by a rapid increase in temperature. We have used a novel nanosecond electron microscope (the dynamic transmission electron microscope, DTEM) to acquire diffraction and imaging information on the transformation, which is driven in-situ by nanosecond laser irradiation. Using nanosecond exposure times that are possible in the DTEM, data can be collected about the transient events in these fast transformations. We have identified the phase transformation with diffraction patterns and correlated the time of the phase transformation with calculated conditions in the sample.

  6. Characterization of bacterial spore germination using phase-contrast and fluorescence microscopy, Raman spectroscopy and optical tweezers.

    PubMed

    Kong, Lingbo; Zhang, Pengfei; Wang, Guiwen; Yu, Jing; Setlow, Peter; Li, Yong-qing

    2011-05-01

    This protocol describes a method combining phase-contrast and fluorescence microscopy, Raman spectroscopy and optical tweezers to characterize the germination of single bacterial spores. The characterization consists of the following steps: (i) loading heat-activated dormant spores into a temperature-controlled microscope sample holder containing a germinant solution plus a nucleic acid stain; (ii) capturing a single spore with optical tweezers; (iii) simultaneously measuring phase-contrast images, Raman spectra and fluorescence images of the optically captured spore at 2- to 10-s intervals; and (iv) analyzing the acquired data for the loss of spore refractility, changes in spore-specific molecules (in particular, dipicolinic acid) and uptake of the nucleic acid stain. This information leads to precise correlations between various germination events, and takes 1-2 h to complete. The method can also be adapted to use multi-trap Raman spectroscopy or phase-contrast microscopy of spores adhered on a cover slip to simultaneously obtain germination parameters for multiple individual spores.

  7. Direct observation of mesoscopic phase separation in KxFeySe2 by scanning microwave microscopy

    NASA Astrophysics Data System (ADS)

    Maeda, Atsutaka; Takahashi, Hideyuki; Imai, Yoshinori

    2015-03-01

    KxFeySe2 is isostructural to 122-FeAs compounds. However, its electronic structure is unique among Fe-based superconductors in the sense that hole Fermi pocket is absent at the center of the Brillouin zone. Therefore, it is important to study this compounds in terms of the mechanism of superconductivity since some pairing (for example, s +/- -wave) needs the interaction between hole and electron Fermi pockets. However, the phase separation in this material makes studies using conventional macroscopic measurement techniques very difficult. Scanning near-field microwave microscope (SMM), which can measure local electric property of inhomogeneous conducting samples, should be a powerful tool. Recently we developed the combined instrument of STM and SMM with high sensitivity, and investigated the local electric property of KxFeySe2 (x = 0.8, y = 1.6 ~2, Tc = 31 K) using this scanning tunneling/microwave microscope. The characteristic pattern of mesoscopic phase separation of the metallic and the semiconducting phase was observed. From the comparison with previously reported SEM/EDS result we identified the metallic phase and the semiconducting phase as the minor Fe-rich phase and the major K2Fe4Se5 phase, respectively.

  8. Single grating phase contrast imaging for x-ray microscopy and microtomography

    NASA Astrophysics Data System (ADS)

    Bruyndonckx, P.; Sasov, A.; Pauwels, B.

    2014-09-01

    The grating based approach to phase contrast imaging is rather inefficient in the use of the available x-ray flux due to the presence of two absorption gratings and it requires longer scan times compared to conventional CT because multiple images are needed at each projection angle. To avoid these drawbacks, a proof-of-principle experiment was developed to obtain absorption, phase contrast (DPC) and dark field images (DCI) in a single exposure using only a non-absorbing phase grating, a micro-focus source in cone-beam geometry and a highresolution x-ray detector.

  9. Ratioed scatter diagrams - An erotetic method for phase identification on complex surfaces using scanning Auger microscopy

    NASA Technical Reports Server (NTRS)

    Browning, R.

    1984-01-01

    By ratioing multiple Auger intensities and plotting a two-dimensional occupational scatter diagram while digitally scanning across an area, the number and elemental association of surface phases can be determined. This can prove a useful tool in scanning Auger microscopic analysis of complex materials. The technique is illustrated by results from an anomalous region on the reaction zone of a SiC/Ti-6Al-4V metal matrix composite material. The anomalous region is shown to be a single phase associated with sulphur and phosphorus impurities. Imaging of a selected phase from the ratioed scatter diagram is possible and may be a useful technique for presenting multiple scanning Auger images.

  10. Quantitative Carré differential interference contrast microscopy to assess phase and amplitude.

    PubMed

    Duncan, Donald D; Fischer, David G; Dayton, Amanda; Prahl, Scott A

    2011-06-01

    We present a method of using an unmodified differential interference contrast microscope to acquire quantitative information on scatter and absorption of thin tissue samples. A simple calibration process is discussed that uses a standard optical wedge. Subsequently, we present a phase-stepping procedure for acquiring phase gradient information exclusive of absorption effects. The procedure results in two-dimensional maps of the local angular (polar and azimuthal) ray deviation. We demonstrate the calibration process, discuss details of the phase-stepping algorithm, and present representative results for a porcine skin sample.

  11. Spectroscopic and chromatographic characterisation of a pentafluorophenylpropyl silica phase end-capped in supercritical carbon dioxide as a reaction solvent.

    PubMed

    Ashu-Arrah, Benjamin A; Glennon, Jeremy D; Albert, Klaus

    2013-07-12

    This research uses solid-state nuclear magnetic resonance (NMR) spectroscopy to characterise the nature and amount of different surface species, and chromatography to evaluate phase properties of a pentafluorophenylpropyl (PFPP) bonded silica phase prepared and end-capped using supercritical carbon dioxide (sc-CO2) as a reaction solvent. Under sc-CO2 reaction conditions (at temperature of 100 °C and pressure of 414 bar), a PFPP silica phase was prepared using 3-[(pentafluorophenyl)propyldimethylchlorosilane] within 1h. The bonded PFPP phase was subsequently end-capped with bis-N,O-trimethylsilylacetamide (BSA), hexamethyldisilazane (HMDS) and trimethylchlorosilane (TMCS) within 1h under the same sc-CO2 reaction conditions (100 °C/4141 bar). Elemental microanalysis, thermogravimetric analysis (TGA), and scanning electron microscopy (SEM) were used to provide support data to solid-state NMR and chromatographic evaluation. Results revealed a surface coverage of 2.2 μmol/m(2) for the non-end-capped PFPP silica phase while the PFPP phase end-capped with BSA gave a higher surface coverage (3.9 μmol/m(2)) compared to HMDS (2.9 μmol/m(2)) and TMCS (2.8 μmol/m(2)). (29)Si CP/MAS NMR analysis of the PFPP end-capped with BSA shows a significant decrease in the amount of Q(3) (free silanols) and Q(4) (siloxane groups) species, coupled with the absence of the most reactive Q(2) (geminal silanols) in addition to increased amount of a single resonance peak centred at +13 ppm (MH) corresponding to -Si-O-*Si-CH3 bond. (13)C CP/MAS NMR shows the resonance corresponding to the propyl linkage (CH3CH2CH2-) and methyl groups (Si(CH3)n) confirming successful silanisation and endcapping reactions in sc-CO2. Chromatographic evaluation of the BSA end-capped PFPP phase with Neue text mixture revealed improved chromatographic separation as evidenced in the enhanced retention of hydrophobic markers and decreased retention for basic solutes. Moreover, chromatography revealed a change in

  12. Electron microscopy analyses and electrical properties of the layered Bi{sub 2}WO{sub 6} phase

    SciTech Connect

    Taoufyq, A.; Ait Ahsaine, H.; Patout, L.; Benlhachemi, A.; Ezahri, M.; and others

    2013-07-15

    The bismuth tungstate Bi{sub 2}WO{sub 6} was synthesized using a classical coprecipitation method followed by a calcination process at different temperatures. The samples were characterized by X-ray diffraction, simultaneous thermogravimetry and differential thermal analysis (TGA/DTA), scanning and transmission electron microscopy (SEM, TEM) analyses. The Rietveld analysis and electron diffraction clearly confirmed the Pca2{sub 1} non centrosymmetric space group previously proposed for this phase. The layers Bi{sub 2}O{sub 2}{sup 2+} and WO{sub 4}{sup 2−} have been directly evidenced from the HRTEM images. The electrical properties of Bi{sub 2}WO{sub 6} compacted pellets systems were determined from electrical impedance spectrometry (EIS) and direct current (DC) analyses, under air and argon, between 350 and 700 °C. The direct current analyses showed that the conduction observed from EIS analyses was mainly ionic in this temperature range, with a small electronic contribution. Electrical change above the transition temperature of 660 °C is observed under air and argon atmospheres. The strong conductivity increase observed under argon is interpreted in terms of formation of additional oxygen vacancies coupled with electron conduction. - Graphical abstract: High resolution transmission electron microscopy: inverse fast Fourier transform giving the layered structure of the Bi{sub 2}WO{sub 6} phase, with a representation of the cell dimensions (b and c vectors). The Bi{sub 2}O{sub 2}{sup 2+} and WO{sub 4}{sup 2−} sandwiches are visible in the IFFT image. - Highlights: • Using transmission electron microscopy, we visualize the layered structure of Bi{sub 2}WO{sub 6}. • Electrical analyses under argon gas show some increase in conductivity. • The phase transition at 660 °C is evidenced from electrical modification.

  13. Direct observation by laser scanning confocal microscopy of microstructure and phase migration of PVC gels in an applied electric field.

    PubMed

    Xia, Hong; Ueki, Takamitsu; Hirai, Toshihiro

    2011-02-01

    The fluorescent probe lucigenin was incorporated in poly(vinyl chloride) (PVC) gels, and laser scanning confocal microscopy (LSCM) was used to clarify the internal structures of the gels. From the two-dimensional and three-dimensional information by LSCM, we first observed the internal structure of the PVC gel at a wet status, where the PVC gels comprised a polymer-rich phase and a polymer-poor phase uniformly with a three-dimensional network structure. After an electric field was applied, an effect of the electric field resulted in the change of internal structure in the gels. The polymer-poor phase moved from the cathode to the anode and the polymer-rich phase formed linelike arrangement between electrodes due to the attraction force. On the other hand, the freeze-dried PVC gels with/without in-situ dc voltage casting were particularly fabricated to confirm above results by the field emission scanning electron microscopy (FE-SEM). It was found that many craters remained on the surface of the gel near the anode due to sublimation in freeze-drying. This phenomenon did not appear on the surface near the cathode. The results of in-situ dc voltage casting also suggested that a substantial amount of polymer-poor phase was moved and fixed at the anode. Thus, results of both LSCM and in-situ dc voltage casting corresponded to the effect of electric field on PVC gels and provided a convincing evidence for the interpretation of the deformation mechanism of PVC gel actuators by an applied electric field.

  14. Integrated quantitative phase and birefringence microscopy for imaging malaria-infected red blood cells.

    PubMed

    Li, Chengshuai; Chen, Shichao; Klemba, Michael; Zhu, Yizheng

    2016-09-01

    A dual-modality birefringence/phase imaging system is presented. The system features a crystal retarder that provides polarization mixing and generates two interferometric carrier waves in a single signal spectrum. The retardation and orientation of sample birefringence can then be measured simultaneously based on spectral multiplexing interferometry. Further, with the addition of a Nomarski prism, the same setup can be used for quantitative differential interference contrast (DIC) imaging. Sample phase can then be obtained with two-dimensional integration. In addition, birefringence-induced phase error can be corrected using the birefringence data. This dual-modality approach is analyzed theoretically with Jones calculus and validated experimentally with malaria-infected red blood cells. The system generates not only corrected DIC and phase images, but a birefringence map that highlights the distribution of hemozoin crystals.

  15. Integrated quantitative phase and birefringence microscopy for imaging malaria-infected red blood cells

    NASA Astrophysics Data System (ADS)

    Li, Chengshuai; Chen, Shichao; Klemba, Michael; Zhu, Yizheng

    2016-09-01

    A dual-modality birefringence/phase imaging system is presented. The system features a crystal retarder that provides polarization mixing and generates two interferometric carrier waves in a single signal spectrum. The retardation and orientation of sample birefringence can then be measured simultaneously based on spectral multiplexing interferometry. Further, with the addition of a Nomarski prism, the same setup can be used for quantitative differential interference contrast (DIC) imaging. Sample phase can then be obtained with two-dimensional integration. In addition, birefringence-induced phase error can be corrected using the birefringence data. This dual-modality approach is analyzed theoretically with Jones calculus and validated experimentally with malaria-infected red blood cells. The system generates not only corrected DIC and phase images, but a birefringence map that highlights the distribution of hemozoin crystals.

  16. Interface morphology studies of liquid phase epitaxy grown HgCdTe films by atomic force microscopy

    NASA Astrophysics Data System (ADS)

    Azoulay, M.; George, M. A.; Burger, A.; Collins, W. E.; Silberman, E.

    1994-04-01

    In this paper we report an investigation of the morphology of the interfaces of liquid phase epitaxy (LPE) grown HgCdTe thin films on CdTe and CdZnTe substrates by atomic force microscopy (AFM) on freshly cleaved (110) crystallographic planes. An empirical observation which may be linked to lattice mismatch was indicated by an angle between the cleavage steps of the substrate to those of the film. The precipitates with size ranging from 5 nm to 20 nm were found to be most apparent near the interface.

  17. Chemotaxis of cancer cells in three-dimensional environment monitored label-free by quantitative phase digital holographic microscopy

    NASA Astrophysics Data System (ADS)

    Kemper, Björn; Schnekenburger, Jürgen; Ketelhut, Steffi

    2017-02-01

    We investigated the capabilities of digital holographic microscopy (DHM) for label-free quantification of the response of living single cells to chemical stimuli in 3D assays. Fibro sarcoma cells were observed in a collagen matrix inside 3D chemotaxis chambers with a Mach-Zehnder interferometer-based DHM setup. From the obtained series of quantitative phase images, the migration trajectories of single cells were retrieved by automated cell tracking and subsequently analyzed for maximum migration distance and motility. Our results demonstrate DHM as a highly reliable and efficient tool for label-free quantification of chemotaxis in 2D and 3D environments.

  18. Sensing dynamic cytoplasm refractive index changes of adherent cells with quantitative phase microscopy using incorporated microspheres as optical probes

    NASA Astrophysics Data System (ADS)

    Przibilla, Sabine; Dartmann, Sebastian; Vollmer, Angelika; Ketelhut, Steffi; Greve, Burkhard; von Bally, Gert; Kemper, Björn

    2012-09-01

    The intracellular refractive index is an important parameter that describes the optical density of the cytoplasm and the concentration of the intracellular solutes. The refractive index of adherently grown cells is difficult to access. We present a method in which silica microspheres in living cells are used to determine the cytoplasm refractive index with quantitative phase microscopy. The reliability of our approach for refractive index retrieval is shown by data from a comparative study on osmotically stimulated adherent and suspended human pancreatic tumor cells. Results from adherent human fibro sarcoma cells demonstrate the capability of the method for sensing of dynamic refractive index changes and its usage with microfluidics.

  19. Differences in estimates of size distribution of beryllium powder materials using phase contrast microscopy, scanning electron microscopy, and liquid suspension counter techniques.

    PubMed

    Stefaniak, Aleksandr B; Hoover, Mark D; Dickerson, Robert M; Day, Gregory A; Breysse, Patrick N; Scripsick, Ronald C

    2007-02-28

    Accurate characterization of the physicochemical properties of aerosols generated for inhalation toxicology studies is essential for obtaining meaningful results. Great emphasis must also be placed on characterizing particle properties of materials as administered in inhalation studies. Thus, research is needed to identify a suite of techniques capable of characterizing the multiple particle properties (i.e., size, mass, surface area, number) of a material that may influence toxicity. The purpose of this study was to characterize the morphology and investigate the size distribution of a model toxicant, beryllium. Beryllium metal, oxides, and alloy particles were aerodynamically size-separated using an aerosol cyclone, imaged dry using scanning electron microscopy (SEM), then characterized using phase contrast microscopy (PCM), a liquid suspension particle counter (LPC), and computer-controlled SEM (CCSEM). Beryllium metal powder was compact with smaller sub-micrometer size particles attached to the surface of larger particles, whereas the beryllium oxides and alloy particles were clusters of primary particles. As expected, the geometric mean (GM) diameter of metal powder determined using PCM decreased with aerodynamic size, but when suspended in liquid for LPC or CCSEM analysis, the GM diameter decreased by a factor of two (p < 0.001). This observation suggested that the smaller submicrometer size particles attached to the surface of larger particles and/or particle agglomerates detach in liquid, thereby shifting the particle size distribution downward. The GM diameters of the oxide materials were similar regardless of sizing technique, but observed differences were generally significant (p < 0.001). For oxides, aerodynamic cluster size will dictate deposition in the lung, but primary particle size may influence biological activity. The GM diameter of alloy particles determined using PCM became smaller with decreasing aerodynamic size fraction; however, when

  20. Differences in estimates of size distribution of beryllium powder materials using phase contrast microscopy, scanning electron microscopy, and liquid suspension counter techniques

    PubMed Central

    Stefaniak, Aleksandr B; Hoover, Mark D; Dickerson, Robert M; Day, Gregory A; Breysse, Patrick N; Scripsick, Ronald C

    2007-01-01

    Accurate characterization of the physicochemical properties of aerosols generated for inhalation toxicology studies is essential for obtaining meaningful results. Great emphasis must also be placed on characterizing particle properties of materials as administered in inhalation studies. Thus, research is needed to identify a suite of techniques capable of characterizing the multiple particle properties (i.e., size, mass, surface area, number) of a material that may influence toxicity. The purpose of this study was to characterize the morphology and investigate the size distribution of a model toxicant, beryllium. Beryllium metal, oxides, and alloy particles were aerodynamically size-separated using an aerosol cyclone, imaged dry using scanning electron microscopy (SEM), then characterized using phase contrast microscopy (PCM), a liquid suspension particle counter (LPC), and computer-controlled SEM (CCSEM). Beryllium metal powder was compact with smaller sub-micrometer size particles attached to the surface of larger particles, whereas the beryllium oxides and alloy particles were clusters of primary particles. As expected, the geometric mean (GM) diameter of metal powder determined using PCM decreased with aerodynamic size, but when suspended in liquid for LPC or CCSEM analysis, the GM diameter decreased by a factor of two (p < 0.001). This observation suggested that the smaller submicrometer size particles attached to the surface of larger particles and/or particle agglomerates detach in liquid, thereby shifting the particle size distribution downward. The GM diameters of the oxide materials were similar regardless of sizing technique, but observed differences were generally significant (p < 0.001). For oxides, aerodynamic cluster size will dictate deposition in the lung, but primary particle size may influence biological activity. The GM diameter of alloy particles determined using PCM became smaller with decreasing aerodynamic size fraction; however, when

  1. The nanoscale phase distinguishing of PCL-PB-PCL blended in epoxy resin by tapping mode atomic force microscopy

    PubMed Central

    2012-01-01

    In this work, we investigated the bulk phase distinguishing of the poly(ε-caprolactone)-polybutadiene-poly(ε-caprolactone) (PCL-PB-PCL) triblock copolymer blended in epoxy resin by tapping mode atomic force microscopy (TM-AFM). We found that at a set-point amplitude ratio (rsp) less than or equal to 0.85, a clear phase contrast could be obtained using a probe with a force constant of 40 N/m. When rsp was decreased to 0.1 or less, the measured size of the PB-rich domain relatively shrank; however, the height images of the PB-rich domain would take reverse (translating from the original light to dark) at rsp = 0.85. Force-probe measurements were carried out on the phase-separated regions by TM-AFM. According to the phase shift angle vs. rsp curve, it could be concluded that the different force exerting on the epoxy matrix or on the PB-rich domain might result in the height and phase image reversion. Furthermore, the indentation depth vs. rsp plot showed that with large tapping force (lower rsp), the indentation depth for the PB-rich domain was nearly identical for the epoxy resin matrix. PMID:22360980

  2. Phase transitions in a LiMn2O4 nanowire battery observed by operando electron microscopy.

    PubMed

    Lee, Soyeon; Oshima, Yoshifumi; Hosono, Eiji; Zhou, Haoshen; Kim, Kyungsu; Chang, Hansen M; Kanno, Ryoji; Takayanagi, Kunio

    2015-01-27

    Fast charge-discharge process has been reported to give a high capacity loss. A nanobattery consisting of a single LiMn2O4 nanowire cathode, ionic liquid electrolyte and lithium titanium oxide anode was developed for in situ transmission electron microscopy. When it was fully charged or discharged within a range of 4 V in less than half an hour (corresponding average C rate: 2.5C), Li-rich and Li-poor phases were observed to be separated by a transition region, and coexisted during whole process. The phase transition region moved reversibly along the nanowire axis which corresponds to the [011] direction, allowing the volume fraction of both phases to change. In the electron diffraction patterns, the Li-rich phase was seen to have the (100) orientation with respect to the incident electron beam, while the Li-poor phase had the (111̅) orientation. The orientation was changed as the transition region moved. However, the nanowire did not fracture. This suggests that a LiMn2O4 nanowire has the advantage of preventing capacity fading at high charge rates.

  3. Photothermal nanoparticles as molecular specificity agents in interferometric phase microscopy (Conference Presentation)

    NASA Astrophysics Data System (ADS)

    Shaked, Natan T.

    2017-02-01

    I review our latest advances in wide-field interferometric imaging of biological cells with molecular specificity, obtained by time-modulated photothermal excitation of gold nanoparticles. Heat emitted from the nanoparticles affects the measured phase signal via both the nanoparticle surrounding refractive-index and thickness changes. These nanoparticles can be bio-functionalized to bind certain biological cell components; thus, they can be used for biomedical imaging with molecular specificity, as new nanoscopy labels, and for photothermal therapy. Predicting the ideal nanoparticle parameters requires a model that computes the thermal and phase distributions around the particle, enabling more efficient phase imaging of plasmonic nanoparticles, and sparing trial and error experiments of using unsuitable nanoparticles. We thus developed a new model for predicting phase signatures from photothermal nanoparticles with arbitrary parameters. We also present a dual-modality technique based on wide-field photothermal interferometric phase imaging and simultaneous ablation to selectively deplete specific cell populations labelled by plasmonic nanoparticles. We experimentally demonstrated our ability to detect and specifically ablate in vitro cancer cells over-expressing epidermal growth factor receptors (EGFRs), labelled with plasmonic nanoparticles, in the presence of either EGFR under-expressing cancer cells or white blood cells. This demonstration established an initial model for depletion of circulating tumour cells in blood. The proposed system is able to image in wide field the label-free quantitative phase profile together with the photothermal phase profile of the sample, and provides the ability of both detection and ablation of chosen cells after their selective imaging.

  4. Combined use of X-ray fluorescence microscopy, phase contrast imaging for high resolution quantitative iron mapping in inflamed cells

    NASA Astrophysics Data System (ADS)

    Gramaccioni, C.; Procopio, A.; Farruggia, G.; Malucelli, E.; Iotti, S.; Notargiacomo, A.; Fratini, M.; Yang, Y.; Pacureanu, A.; Cloetens, P.; Bohic, S.; Massimi, L.; Cutone, A.; Valenti, P.; Rosa, L.; Berlutti, F.; Lagomarsino, S.

    2017-06-01

    X-ray fluorescence microscopy (XRFM) is a powerful technique to detect and localize elements in cells. To derive information useful for biology and medicine, it is essential not only to localize, but also to map quantitatively the element concentration. Here we applied quantitative XRFM to iron in phagocytic cells. Iron, a primary component of living cells, can become toxic when present in excess. In human fluids, free iron is maintained at 10-18 M concentration thanks to iron binding proteins as lactoferrin (Lf). The iron homeostasis, involving the physiological ratio of iron between tissues/secretions and blood, is strictly regulated by ferroportin, the sole protein able to export iron from cells to blood. Inflammatory processes induced by lipopolysaccharide (LPS) or bacterial pathoge inhibit ferroportin synthesis in epithelial and phagocytic cells thus hindering iron export, increasing intracellular iron and bacterial multiplication. In this respect, Lf is emerging as an important regulator of both iron and inflammatory homeostasis. Here we studied phagocytic cells inflamed by bacterial LPS and untreated or treated with milk derived bovine Lf. Quantitative mapping of iron concentration and mass fraction at high spatial resolution is obtained combining X-ray fluorescence microscopy, atomic force microscopy and synchrotron phase contrast imaging.

  5. Effects of phase and coupling between the vibrational modes on selective excitation in coherent anti-Stokes Raman scattering microscopy

    SciTech Connect

    Patel, Vishesha; Malinovsky, Vladimir S.; Malinovskaya, Svetlana

    2010-06-15

    Coherent anti-Stokes Raman scattering (CARS) microscopy has been a major tool of investigation of biological structures as it contains the vibrational signature of molecules. A quantum control method based on chirped pulse adiabatic passage was recently proposed for selective excitation of a predetermined vibrational mode in CARS microscopy [Malinovskaya and Malinovsky, Opt. Lett. 32, 707 (2007)]. The method utilizes the chirp sign variation at the peak pulse amplitude and gives a robust adiabatic excitation of the desired vibrational mode. Using this method, we investigate the impact of coupling between vibrational modes in molecules on controllability of excitation of the CARS signal. We analyze two models of two coupled two-level systems (TLSs) having slightly different transitional frequencies. The first model, featuring degenerate ground states of the TLSs, gives robust adiabatic excitation and maximum coherence in the resonant TLS for positive value of the chirp. In the second model, implying nondegenerate ground states in the TLSs, a population distribution is observed in both TLSs, resulting in a lack of selectivity of excitation and low coherence. It is shown that the relative phase and coupling between the TLSs play an important role in optimizing coherence in the desired vibrational mode and suppressing unwanted transitions in CARS microscopy.

  6. Optical detection and measurement of living cell morphometric features with single-shot quantitative phase microscopy

    NASA Astrophysics Data System (ADS)

    Bon, Pierre; Savatier, Julien; Merlin, Marine; Wattellier, Benoît; Monneret, Serge

    2012-07-01

    We present a quadriwave lateral shearing interferometer used as a wavefront sensor and mounted on a commercial non-modified transmission white-light microscope as a quantitative phase imaging technique. The setup is designed to simultaneously make measurements with both quantitative transmission phase and fluorescence modes: phase enables enhanced contrasted visualization of the cell structure including intracellular organelles, while fluorescence allows a complete and precise identification of each component. After the characterization of the phase measurement reliability and sensitivity on calibrated samples, we use these two imaging modes to measure the characteristic optical path difference between subcellular elements (mitochondria, actin fibers, and vesicles) and cell medium, and demonstrate that phase-only information should be sufficient to identify some organelles without any labeling, like lysosomes. Proof of principle results show that the technique could be used either as a qualitative tool for the control of cells before an experiment, or for quantitative studies on morphology, behavior, and dynamics of cells or cellular components.

  7. Disturbance-free digital holographic microscopy via a micro-phase-step approach

    NASA Astrophysics Data System (ADS)

    Chang, Chi-Ching; Shiu, Ming-Tzung; Wang, Je-Chung; Wu, Chia-Hao; Chew, Yang-Kun

    2015-05-01

    This work proposes a cost-effective, simple, micro-phase-step (MPS) method for suppressing the zero-order diffraction and conjugate-image interferences that are caused during digital holographic microscopic image reconstruction. The proposed MPS method replaces the conventional phase modulation approach; it uses a rotatable cover glass that enables smooth modification of the incidence angle and the optical path of the reference beam. This setup allows the phase step to be accurately estimated by shifting the reference wave phase more freely close to π/2, at which the background noise can be suppressed more effectively. In the proposed MPS method, the optimal conditions for suppressing conjugate-image interference are identified using a relatively moderate intensity distribution and suppression of noise in the numerically reconstructed object wave-field. In addition, the proposed method mitigates the effect of disturbances that are caused by environmental factors, such as minor vibrations and small changes in temperature and humidity. Importantly, only two holograms are required to satisfy the objective of image reconstruction. The results in this work reveal that even with intentional interference caused by minor vibrations, conjugate-image interference can still be suppressed by determining the phase deviation between the two original holograms.

  8. Spectroscopic and theoretical study of the "azo"-dye E124 in condensate phase: evidence of a dominant hydrazo form.

    PubMed

    Almeida, Mariana R; Stephani, Rodrigo; Dos Santos, Hélio F; de Oliveira, Luiz Fernando C

    2010-01-14

    Spectroscopic techniques, including Raman, IR, UV/vis, and NMR were used to characterize the samples of the azo dye Ponceau 4R (also known as E124, New Coccine; Cochineal Red; C.I. no. 16255; Food Red No. 102), which is 1,3-naphthalenedisulfonic acid, 7-hydroxy-8-[(4-sulfo-1-naphthalenyl) azo] trisodium salt in aqueous solution and solid state. In addition, first principle calculations were carried out for the azo (OH) and hydrazo (NH) tautomers in order to assist in the assignment of the experimental data. The two intense bands observed in the UV/vis spectrum, centered at 332 and 507 nm, can be compared to the calculated values at 296 and 474 nm for azo and 315 and 500 nm for hydrazo isomer, with the latter in closer agreement to the experiment. The Raman spectrum is quite sensitive to tautomeric equilibrium; in solid state and aqueous solution, three bands were observed around 1574, 1515, and 1364 cm(-1), assigned to mixed modes including deltaNH + betaCH + nuCC, deltaNH + nuC horizontal lineO + nuC horizontal lineN + betaCH and nuCC vibrations, respectively. These assignments are predicted only for the NH species centered at 1606, 1554, and 1375 cm(-1). The calculated Raman spectrum for the azo (OH) tautomer showed two strong bands at 1468 (nuN = N + deltaOH) and 1324 cm(-1) (nuCC + nuC-N), which were not obtained experimentally. The (13)C NMR spectrum showed a very characteristic peak at 192 ppm assigned to the carbon bound to oxygen in the naphthol ring; the predicted values were 165 ppm for OH and 187 for NH isomer, supporting once again the predominance of NH species in solution. Therefore, all of the experimental and theoretical results strongly suggest the food dye Ponceau 4R or E124 has a major contribution of the hydrazo structure instead of the azo form as the most abundant in condensate phase.

  9. Repeated crack healing in MAX-phase ceramics revealed by 4D in situ synchrotron X-ray tomographic microscopy

    PubMed Central

    Sloof, Willem G.; Pei, Ruizhi; McDonald, Samuel A.; Fife, Julie L.; Shen, Lu; Boatemaa, Linda; Farle, Ann-Sophie; Yan, Kun; Zhang, Xun; van der Zwaag, Sybrand; Lee, Peter D.; Withers, Philip J.

    2016-01-01

    MAX phase materials are emerging as attractive engineering materials in applications where the material is exposed to severe thermal and mechanical conditions in an oxidative environment. The Ti2AlC MAX phase possesses attractive thermomechanical properties even beyond a temperature of 1000 K. An attractive feature of this material is its capacity for the autonomous healing of cracks when operating at high temperatures. Coupling a specialized thermomechanical setup to a synchrotron X-ray tomographic microscopy endstation at the TOMCAT beamline, we captured the temporal evolution of local crack opening and healing during multiple cracking and autonomous repair cycles at a temperature of 1500 K. For the first time, the rate and position dependence of crack repair in pristine Ti2AlC material and in previously healed cracks has been quantified. Our results demonstrate that healed cracks can have sufficient mechanical integrity to make subsequent cracks form elsewhere upon reloading after healing. PMID:26972608

  10. Phase variance optical coherence microscopy for label-free imaging of the developing vasculature in zebrafish embryos

    NASA Astrophysics Data System (ADS)

    Chen, Yu; Trinh, Le A.; Fingler, Jeff; Fraser, Scott E.

    2016-12-01

    A phase variance optical coherence microscope (pvOCM) has been created to image blood flow in the microvasculature of zebrafish embryos, without the use of exogenous labels. The pvOCM imaging system has axial and lateral resolutions of 2.8 μm in tissue and imaging depth of more than 100 μm. Images of 2 to 5 days postfertilization zebrafish embryos identified the detailed anatomical structure based on OCM intensity contrast. Phase variance contrast offered visualization of blood flow in the arteries, veins, and capillaries. The pvOCM images of the vasculature were confirmed by direct comparisons with fluorescence microscopy images of transgenic embryos in which the vascular endothelium is labeled with green fluorescent protein. The ability of pvOCM to capture activities of regional blood flow permits it to reveal functional information that is of great utility for the study of vascular development.

  11. Repeated crack healing in MAX-phase ceramics revealed by 4D in situ synchrotron X-ray tomographic microscopy.

    PubMed

    Sloof, Willem G; Pei, Ruizhi; McDonald, Samuel A; Fife, Julie L; Shen, Lu; Boatemaa, Linda; Farle, Ann-Sophie; Yan, Kun; Zhang, Xun; van der Zwaag, Sybrand; Lee, Peter D; Withers, Philip J

    2016-03-14

    MAX phase materials are emerging as attractive engineering materials in applications where the material is exposed to severe thermal and mechanical conditions in an oxidative environment. The Ti2AlC MAX phase possesses attractive thermomechanical properties even beyond a temperature of 1000 K. An attractive feature of this material is its capacity for the autonomous healing of cracks when operating at high temperatures. Coupling a specialized thermomechanical setup to a synchrotron X-ray tomographic microscopy endstation at the TOMCAT beamline, we captured the temporal evolution of local crack opening and healing during multiple cracking and autonomous repair cycles at a temperature of 1500 K. For the first time, the rate and position dependence of crack repair in pristine Ti2AlC material and in previously healed cracks has been quantified. Our results demonstrate that healed cracks can have sufficient mechanical integrity to make subsequent cracks form elsewhere upon reloading after healing.

  12. Multiscale phase mapping of LiFePO4-based electrodes by transmission electron microscopy and electron forward scattering diffraction.

    PubMed

    Robert, Donatien; Douillard, Thierry; Boulineau, Adrien; Brunetti, Guillaume; Nowakowski, Pawel; Venet, Denis; Bayle-Guillemaud, Pascale; Cayron, Cyril

    2013-12-23

    LiFePO4 and FePO4 phase distributions of entire cross-sectioned electrodes with various Li content are investigated from nanoscale to mesoscale, by transmission electron microscopy and by the new electron forward scattering diffraction technique. The distributions of the fully delithiated (FePO4) or lithiated particles (LiFePO4) are mapped on large fields of view (>100 × 100 μm(2)). Heterogeneities in thin and thick electrodes are highlighted at different scales. At the nanoscale, the statistical analysis of 64 000 particles unambiguously shows that the small particles delithiate first. At the mesoscale, the phase maps reveal a core-shell mechanism at the scale of the agglomerates with a preferential pathway along the electrode porosities. At larger scale, lithiation occurs in thick electrodes "stratum by stratum" from the surface in contact with electrolyte toward the current collector.

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

    PubMed

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

    2015-06-01

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

  14. Field-Induced Reversible Phase Manipulation in Metal-Insulator Transition using Scanning Tunneling Microscopy

    NASA Astrophysics Data System (ADS)

    Park, Se Jun

    2005-03-01

    Reversible electronic switching between insulating and metallic phases is a novel idea that may allow new types of field effect devices feasible.^1 Here we demonstrate the reversible manipulation between metallic and insulating phases in two-dimensional In nanowire arrays on Si(111) surface near the metal-insulator transition temperature (Tc). The electronic switching of phases was induced by local electric field applied by the probe tip of a scanning tunneling microscope. The field-dependent hysteresis behavior was also observed in tip height measurements as a function of the sample bias, under the constraint of constant tunneling current. A model including the intrinsic bi-stability of the nanometer-scale domains of In nanowire arrays will be discussed. ^1C. Ahn, J. Triscone, J. Mannhart, Nature 6952, 1015 (2003)

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

    PubMed Central

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

    2011-01-01

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

  16. Color-coded LED microscopy for multi-contrast and quantitative phase-gradient imaging

    PubMed Central

    Lee, Donghak; Ryu, Suho; Kim, Uihan; Jung, Daeseong; Joo, Chulmin

    2015-01-01

    We present a multi-contrast microscope based on color-coded illumination and computation. A programmable three-color light-emitting diode (LED) array illuminates a specimen, in which each color corresponds to a different illumination angle. A single color image sensor records light transmitted through the specimen, and images at each color channel are then separated and utilized to obtain bright-field, dark-field, and differential phase contrast (DPC) images simultaneously. Quantitative phase imaging is also achieved based on DPC images acquired with two different LED illumination patterns. The multi-contrast and quantitative phase imaging capabilities of our method are demonstrated by presenting images of various transparent biological samples. PMID:26713205

  17. Optofluidic bioimaging platform for quantitative phase imaging of lab on a chip devices using digital holographic microscopy.

    PubMed

    Pandiyan, Vimal Prabhu; John, Renu

    2016-01-20

    We propose a versatile 3D phase-imaging microscope platform for real-time imaging of optomicrofluidic devices based on the principle of digital holographic microscopy (DHM). Lab-on-chip microfluidic devices fabricated on transparent polydimethylsiloxane (PDMS) and glass substrates have attained wide popularity in biological sensing applications. However, monitoring, visualization, and characterization of microfluidic devices, microfluidic flows, and the biochemical kinetics happening in these devices is difficult due to the lack of proper techniques for real-time imaging and analysis. The traditional bright-field microscopic techniques fail in imaging applications, as the microfluidic channels and the fluids carrying biological samples are transparent and not visible in bright light. Phase-based microscopy techniques that can image the phase of the microfluidic channel and changes in refractive indices due to the fluids and biological samples present in the channel are ideal for imaging the fluid flow dynamics in a microfluidic channel at high resolutions. This paper demonstrates three-dimensional imaging of a microfluidic device with nanometric depth precisions and high SNR. We demonstrate imaging of microelectrodes of nanometric thickness patterned on glass substrate and the microfluidic channel. Three-dimensional imaging of a transparent PDMS optomicrofluidic channel, fluid flow, and live yeast cell flow in this channel has been demonstrated using DHM. We also quantify the average velocity of fluid flow through the channel. In comparison to any conventional bright-field microscope, the 3D depth information in the images illustrated in this work carry much information about the biological system under observation. The results demonstrated in this paper prove the high potential of DHM in imaging optofluidic devices; detection of pathogens, cells, and bioanalytes on lab-on-chip devices; and in studying microfluidic dynamics in real time based on phase changes.

  18. Microscopy imaging and quantitative phase contrast mapping in turbid microfluidic channels by digital holography.

    PubMed

    Paturzo, Melania; Finizio, Andrea; Memmolo, Pasquale; Puglisi, Roberto; Balduzzi, Donatella; Galli, Andrea; Ferraro, Pietro

    2012-09-07

    We show that sharp imaging and quantitative phase-contrast microcopy is possible in microfluidics in flowing turbid media by digital holography. In fact, in flowing liquids with suspended colloidal particles, clear vision is hindered and cannot be recovered by any other microscopic imaging technique. On the contrary, using digital holography, clear imaging is possible thanks to the Doppler frequency shift experienced by the photons scattered by the flowing colloidal particles, which do not contribute to the interference process, i.e. the recorded hologram. The method is illustrated and imaging results are demonstrated for pure phase objects, i.e. biological cells in microfluidic channels.

  19. In vivo confocal microscopy in the acute phase of corneal inflammation.

    PubMed

    Kobayashi, Akira; Maeda, Ari; Sugiyama, Kazuhisa

    2003-01-01

    Two cases of noninfectious keratitis were studied using in vivo corneal confocal microscopy in addition to routine slit-lamp biomicroscopy. A 10-year-old boy suffered from keratitis in his left eye due to a bee sting and a 20-year-old man had keratitis following corneal blunt trauma. In both cases, we found a honeycomb pattern at the anterior and mid-stromal level of the middle cornea. This honeycomb pattern disappeared in 1 week with steroid treatment. This pattern might be caused by syncytial cell bodies of activated keratocytes, by focal corneal stroma edema, or by polymorphonuclear leukocyte infiltration along with the keratocytes or along preformed channels within the corneal stroma known as Bowman channels. Further analysis in a large number of patients may aid in further understanding in vivo human corneal inflammation.

  20. Value of dysmorphic red cells and G1 cells by phase contrast microscopy in the diagnosis of glomerular diseases.

    PubMed

    Sultana, T; Sultana, T; Rahman, M Q; Rahman, F; Islam, M S; Ahmed, A N

    2011-01-01

    The morphology of red cells by phase-contrast microscopy (PCM) is a useful diagnostic marker for glomerular haematuria. This study evaluated the value of urinary dysmorphic red cells and G1 cells (special type of dysmorphic red cell) count by PCM providing a simple, cost effective and low risk technique in the diagnosis of glomerular diseases. Urine samples of 120 patients with haematuria and proteinuria were examined and the percentage of dysmorphic red cells and G1 cells were calculated. Cases were divided into two groups; group I (>20% dysmorphic red cells- glomerular group) and group II (≤20% dysmorphic red cells as non glomerular group). Renal histopathology was used as the gold standard method for the diagnosis of glomerulonephritis. Results from PCM showed a sensitivity of 92.7%, specificity 100% by the detection of dysmorphic red cell while by the detection of G1 cells, a sensitivity of 97.6% and specificity 100% were observed. The percentage of G1 cells is superior to counting dysmorphic red cells. The high sensitivity of phase contrast microscopy confirms its usefulness for the detection of dysmorphic red cells and G1 cells that can guide clinicians in the identification of the site of haematuria using non invasive tests.

  1. Phased-array of microcoils allows MR microscopy of ex vivo human skin samples at 9.4 T.

    PubMed

    Göbel, K; Gruschke, O G; Leupold, J; Kern, J S; Has, C; Bruckner-Tuderman, L; Hennig, J; von Elverfeldt, D; Baxan, N; Korvink, J G

    2015-02-01

    The aim of this study was to demonstrate the feasibility of a custom-made phased-array microcoil within a 400 MHz animal system for the morphological characterization of human skin tissue in correlation with histopathology. A dedicated 7-channel microcoil-based MR detector arranged in a phased-array geometry was developed to combine the advantages of both a large field of view and a high signal-to-noise ratio. Standard gradient echo sequences were adapted for the characterization of skin morphology ex vivo. In this study, the feasibility of using this type of microdetector, combined with specially manufactured sample holders, to achieve high-resolution MR images of fresh and formalin-fixed, normal and hidradenitis suppurativa diseased skin was successfully demonstrated. The setup presented in this work allows reliable acquisitions of high-resolution images with in-plane resolution up to 25 × 25 μm², and 100 μm in the orthogonal direction, thereby allowing the differentiation of typical layers of the skin, sebaceous glands and hair follicle. This study demonstrates that MR microscopy on skin biopsies can be applied at low cost on a standard animal MR imaging system. The successful imaging of different skin structures ex vivo is a prerequisite for non-invasive, in vivo application of skin MR microscopy for accurate complementary disease diagnosis in dermatology. © 2014 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

  2. Deciphering the internal complexity of living cells with quantitative phase microscopy: a multiscale approach

    NASA Astrophysics Data System (ADS)

    Martinez-Torres, Cristina; Laperrousaz, Bastien; Berguiga, Lotfi; Boyer-Provera, Elise; Elezgaray, Juan; Nicolini, Franck E.; Maguer-Satta, Veronique; Arneodo, Alain; Argoul, Françoise

    2015-09-01

    The distribution of refractive indices (RIs) of a living cell contributes in a nonintuitive manner to its optical phase image and quite rarely can be inverted to recover its internal structure. The interpretation of the quantitative phase images of living cells remains a difficult task because (1) we still have very little knowledge on the impact of its internal macromolecular complexes on the local RI and (2) phase changes produced by light propagation through the sample are mixed with diffraction effects by the internal cell bodies. We propose to implement a two-dimensional wavelet-based contour chain detection method to distinguish internal boundaries based on their greatest optical path difference gradients. These contour chains correspond to the highest image phase contrast and follow the local RI inhomogeneities linked to the intracellular structural intricacy. Their statistics and spatial distribution are the morphological indicators suited for comparing cells of different origins and/or to follow their transformation in pathologic situations. We use this method to compare nonadherent blood cells from primary and laboratory culture origins and to assess the internal transformation of hematopoietic stem cells by the transduction of the BCR-ABL oncogene responsible for the chronic myelogenous leukemia.

  3. Laser phase microscopy and functional imaging of living human cancer cells during the cell cycle

    NASA Astrophysics Data System (ADS)

    Perevedentseva, Elena V.; Graschew, Georgi; Balanos, Evangelos; Dressler, Cathrin; Beuthan, Juergen; Schlag, Peter M.

    2000-05-01

    The purpose of the investigation was to elaborate a new method of functional imaging of living tumor cells. Human colon carcinoma cells HCT116 were investigated with a conventional light microscope, confocal laser scanning microscope and with a laser phase microscope (LPM). The LPM is a functional imaging technique providing information about cell morphology which is imposed by the physiological inhomogeneity of the refractive index. The phase of the light wave passing through an object contains quantitative information about the object thickness, the shape, and the spatial distribution of the refractive index varying with morphology and chemical composition inhomogeneity inside the object. The new method of investigation of the cells in different stages of the cell cycle is developed. Every phase image of the investigated cells has been compared with conventional light microscopic and confocal microscopic images of the same cell. the relation between the cell state, their morphological peculiarities and the phase characteristics of the measured cell is determined. Data thus acquired, quantitatively characterizing intra- and intercellular processes during the cell cycle, and the method of measurements can be used to investigate with high optic resolution the mechanisms of different physical, chemical and biomolecular interactions with the tumor cells.

  4. Spectroscopic Imaging of Strongly Correlated Electronic States

    NASA Astrophysics Data System (ADS)

    Yazdani, Ali; da Silva Neto, Eduardo H.; Aynajian, Pegor

    2016-03-01

    The study of correlated electronic systems from high-Tc cuprates to heavy-fermion systems continues to motivate the development of experimental tools to probe electronic phenomena in new ways and with increasing precision. In the past two decades, spectroscopic imaging with scanning tunneling microscopy has emerged as a powerful experimental technique. The combination of high energy and spatial resolutions provided by this technique reveals unprecedented detail of the electronic properties of strongly correlated metals and superconductors. This review examines specific experiments, theoretical concepts, and measurement methods that have established the application of these techniques to correlated materials. A wide range of applications, such as the study of collective responses to single atomic impurities, the characterization of quasiparticle-like excitations through their interference, and the identification of competing electronic phases using spectroscopic imaging, are discussed.

  5. High-throughput label-free screening of euglena gracilis with optofluidic time-stretch quantitative phase microscopy

    NASA Astrophysics Data System (ADS)

    Guo, Baoshan; Lei, Cheng; Ito, Takuro; Yaxiaer, Yalikun; Kobayashi, Hirofumi; Jiang, Yiyue; Tanaka, Yo; Ozeki, Yasuyuki; Goda, Keisuke

    2017-02-01

    The development of reliable, sustainable, and economical sources of alternative fuels is an important, but challenging goal for the world. As an alternative to liquid fossil fuels, microalgal biofuel is expected to play a key role in reducing the detrimental effects of global warming since microalgae absorb atmospheric CO2 via photosynthesis. Unfortunately, conventional analytical methods only provide population-averaged lipid contents and fail to characterize a diverse population of microalgal cells with single-cell resolution in a noninvasive and interference-free manner. Here we demonstrate high-throughput label-free single-cell screening of lipid-producing microalgal cells with optofluidic time-stretch quantitative phase microscopy. In particular, we use Euglena gracilis - an attractive microalgal species that produces wax esters (suitable for biodiesel and aviation fuel after refinement) within lipid droplets. Our optofluidic time-stretch quantitative phase microscope is based on an integration of a hydrodynamic-focusing microfluidic chip, an optical time-stretch phase-contrast microscope, and a digital image processor equipped with machine learning. As a result, it provides both the opacity and phase contents of every single cell at a high throughput of 10,000 cells/s. We characterize heterogeneous populations of E. gracilis cells under two different culture conditions to evaluate their lipid production efficiency. Our method holds promise as an effective analytical tool for microalgaebased biofuel production.

  6. Detecting dynamic responses of materials and devices under an alternating electric potential by phase-locked transmission electron microscopy.

    PubMed

    Soma, Kentaro; Konings, Stan; Aso, Ryotaro; Kamiuchi, Naoto; Kobayashi, Genki; Yoshida, Hideto; Takeda, Seiji

    2017-10-01

    An apparatus is developed for transmission electron microscopy (TEM) to acquire image and spectral data, such as TEM images, electron holograms, and electron energy loss spectra, synchronized with the measurement of the dynamic response of a specimen under an applied alternating current (AC) electric potential (voltage, denoted VAC). From a VAC of frequency f, a shutter pulse signal is generated to open and close a pre-specimen shutter in a base TEM apparatus. A pulse is generated per VAC cycle from the targeted phase Φ to Φ +∆Φ with phase width ∆Φ (∆Φ <2π). ∆Φ corresponds to the temporal pulse width τ (τ < 1/f) of an electron beam; i.e., ∆Φ =2πfτ. Because of the high sensitivity of the TEM camera used in this study, the images and spectra that are acquired at the same target phase are integrated by means of stroboscopic illumination to obtain the final phase-locked images and spectra with sufficiently small S/N ratio. Phase-locked (strobe) images and/or spectra are obtained for model specimens of polycrystalline aluminum and an all-solid-state lithium ion battery (LIB). In the phase-locked TEM conditions, f ranges from 1Hz to about 40kHz and ∆Φ from 2π/80 to π. VAC ranges from 2mV to 1V depending on observation conditions. The quality of phase-locked strobe images can be improved markedly using a phase-locked strobe electron beam. Under specific conditions, the spatial resolution in images is better than 0.12nm, even though the spatial resolution generally depends on VAC, f, the base TEM, and the conductivity of the specimen. For the model specimens, it is shown that electrochemical impedance spectroscopy and cyclic voltammetry can be performed in a TEM apparatus, and could potentially be synchronized with phase-locked (strobe) imaging and spectroscopy. Severe electron irradiation damage is detected during phase-locked (strobe) electron holography of the model LIB. Copyright © 2017 Elsevier B.V. All rights reserved.

  7. Mapping phase diagrams of supported lipid bilayers by atomic force microscopy.

    PubMed

    Borrell, Jordi H; Montero, M Teresa; Domènech, Òscar

    2017-01-01

    In this work, we present the method followed to construct a pseudophase diagram of two phospholipids: 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphoethanolamine and 1-palmitoyl-2-oleoyl-sn-glycero-3-phospho-(1'-rac-glycerol). Two different techniques, DSC and AFM, have been used based in the determination of the onset (Tonset ) and completion (Toffset ) temperatures of the gel-to-liquid crystalline phases (Lβ →Lα ), the first from the endotherms from liposomes and the second from the topographic images of supported lipid bilayers. The features of both phase diagrams are discussed emphasizing the influence of Ca(2+) presence and the substrate (mica) on the transition undergone by the phospholipid mixture. Microsc. Res. Tech. 80:4-10, 2017. © 2016 Wiley Periodicals, Inc.

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

    NASA Astrophysics Data System (ADS)

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

    2016-05-01

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

  9. Complex Observation in Electron Microscopy. II. Direct Visualization of Phases and Amplitudes of Exit Wave Functions

    NASA Astrophysics Data System (ADS)

    Danev, Radostin; Nagayama, Kuniaki

    2001-03-01

    We report microscopic observation of complex waves emitted from objects. Two images sequentially detected at the image plane with two transmission electron microscopic (TEM) methods, the conventional and the phase-contrasted, are numerically combined to reproduce complex wave functions, which are holding complete optical information. The long lasting issue in conventional TEM that the contrast and the resolution of images are difficult to be simultaneously optimized was settled with the complex wave functions. The aberration-corrected phase images of ferritin (at 400 kV) and TMV (at 100 kV) did show an extraordinary high contrast free from image distortion without sacrificing the resolution compared with that obtained by conventional TEM.

  10. High-speed line-scanning confocal holographic microscopy for quantitative phase imaging (Conference Presentation)

    NASA Astrophysics Data System (ADS)

    Liu, Changgeng; Knitter, Sebastian; Cong, Zhilong; Sencan, Ikbal; Cao, Hui; Choma, Michael A.

    2016-03-01

    We present a high speed, phase-sensitive, line-scanning reflectance confocal interference microscope. We achieved rapid confocal imaging using a fast line-scan camera and quantitative phase imaging using off-axis digital holography on a 1D, line-by-line basis. In our prototype system, a He-Ne laser (~1.2 mW) was used to demonstrate the principle of operation. Using a 20 kHz line scan rate (1024 pixels per line scan), we achieved a video-rate frame rate of 20 Hz for 1024x500 pixel en-face confocal images (20 MHz total pixel rate). By using an objective lens of a NA 0.65, we achieved an axial and lateral resolution of ~3.5 micrometers and ~0.8 micrometers, respectively. By z-stack imaging of a custom silicon target with a stepped structure, we confirmed that the axial sectioning of the interference microscope is similar to that of a traditional line-scan confocal microscope (our microscope with the reference arm blocked). The utility of phase-sensitive holographic detection in line-scan confocal was demonstrated in two ways. First, using a custom axial height phantom fabricated using chrome deposition, we demonstrated variations in phase corresponding to heights in the 100 nm range with a contrast-to-noise ratio of ~31 dB. Second, we demonstrate digital refocusing of an out-of-focus holographic image. The mechanism of confocality in our line-scan system is 1D physical pinholing. Our ongoing work aims to add an additional mechanism of confocality by using low spatial coherence sources to impose interferometric pinholing.

  11. Comparison of fiber counting by TV screen and eyepieces of phase contrast microscopy.

    PubMed

    Mao, I-Fang; Yeh, Hui-Roung; Chen, Mei-Lien

    2002-01-01

    This study designed a modified light path of a phase-contrast microscope to evaluate the feasibility of fiber counting on a television screen (TVS). A comparison also was made of the fiber counts, fiber density, and precision derived from repeatedly counting fibers through eyepieces (EPs) and on a TVS connected to a phase contrast microscope. Thirty asbestos fiber samples were counted 10 times repeatedly. Ten samples were counted by viewing the same field through an EP and on a TVS alternately, whereas the other 20 samples were counted separately by using an EP and a TVS. The A rules of National Institute for Occupational Safety and Health (NIOSH) 7400 method was quoted. No statistically significant difference of fiber density (p = 0.39) or fiber counts (p > 0.05) was observed between the TVS and EP methods, though TVS gave a slightly lower value than EP did. The bias of the two methods was 7.7 +/- 8.0% on an average. The relative standard deviations (RSDs) for both methods were the same, 0.40, which resembled the theoretical RSD, 0.44, of NIOSH 7400. Meanwhile, the theoretical and experimental RSDs were not significantly different for either method (p > 0.05). The modified light path of a phase contrast microscope provided a compatible view with less eye strain on a TVS than a conventional EP. Moreover, operator biases and variability might be greatly reduced by training several counters simultaneously on the TVS.

  12. Phase imaging microscopy for the diagnostics of plasma-cell interaction

    NASA Astrophysics Data System (ADS)

    Ohene, Yolanda; Marinov, Ilya; de Laulanié, Lucie; Dupuy, Corinne; Wattelier, Benoit; Starikovskaia, Svetlana

    2015-06-01

    Phase images of biological specimens were obtained by the method of Quadriwave Lateral Shearing Interferometry (QWLSI). The QWLSI technique produces, at high resolution, phase images of the cells having been exposed to a plasma treatment and enables the quantitative analysis of the changes in the surface area of the cells over time. Morphological changes in the HTori normal thyroid cells were demonstrated using this method. There was a comparison of the cell behaviour between control cells, cells treated by plasma of a nanosecond dielectric barrier discharge, including cells pre-treated by catalase, and cells treated with an equivalent amount of H2O2. The major changes in the cell membrane morphology were observed at only 5 min after the plasma treatment. The primary role of reactive oxygen species (ROS) in this degradation is suggested. Deformation and condensation of the cell nucleus were observed 2-3 h after the treatment and are supposedly related to apoptosis induction. The coupling of the phase QWLSI with immunofluorescence imaging would give a deeper insight into the mechanisms of plasma induced cell death.

  13. Selective spectroscopic imaging of hyperpolarized pyruvate and its metabolites using a single-echo variable phase advance method in balanced SSFP

    PubMed Central

    Varma, Gopal; Wang, Xiaoen; Vinogradov, Elena; Bhatt, Rupal S.; Sukhatme, Vikas; Seth, Pankaj; Lenkinski, Robert E.; Alsop, David C.; Grant, Aaron K.

    2015-01-01

    Purpose In balanced steady state free precession (bSSFP), the signal intensity has a well-known dependence on the off-resonance frequency, or, equivalently, the phase advance between successive radiofrequency (RF) pulses. The signal profile can be used to resolve the contributions from the spectrally separated metabolites. This work describes a method based on use of a variable RF phase advance to acquire spatial and spectral data in a time-efficient manner for hyperpolarized 13C MRI. Theory and Methods The technique relies on the frequency response from a bSSFP acquisition to acquire relatively rapid, high-resolution images that may be reconstructed to separate contributions from different metabolites. The ability to produce images from spectrally separated metabolites was demonstrated in-vitro, as well as in-vivo following administration of hyperpolarized 1-13C pyruvate in mice with xenograft tumors. Results In-vivo images of pyruvate, alanine, pyruvate hydrate and lactate were reconstructed from 4 images acquired in 2 seconds with an in-plane resolution of 1.25 × 1.25mm2 and 5mm slice thickness. Conclusions The phase advance method allowed acquisition of spectroscopically selective images with high spatial and temporal resolution. This method provides an alternative approach to hyperpolarized 13C spectroscopic MRI that can be combined with other techniques such as multi-echo or fluctuating equilibrium bSSFP. PMID:26507361

  14. Lab on chip optical imaging of biological sample by quantitative phase microscopy

    NASA Astrophysics Data System (ADS)

    Memmolo, P.; Miccio, L.; Merola, F.; Gennari, O.; Mugnano, M.; Netti, P. A.; Ferraro, P.

    2015-03-01

    Quantitative imaging and three dimensional (3D) morphometric analysis of flowing and not-adherent cells is an important aspect for diagnostic purposes at Lab on Chip scale. Diagnostics tools need to be quantitative, label-free and, as much as possible, accurate. In recent years digital holography (DH) has been improved to be considered as suitable diagnostic method in several research field. In this paper we demonstrate that DH can be used for retrieving 3D morphometric data for sorting and diagnosis aims. Several techniques exist for 3D morphological study as optical coherent tomography and confocal microscopy, but they are not the best choice in case of dynamic events as flowing samples. Recently, a DH approach, based on shape from silhouette algorithm (SFS), has been developed for 3D shape display and calculation of cells biovolume. Such approach, adopted in combination with holographic optical tweezers (HOT) was successfully applied to cells with convex shape. Unfortunately, it's limited to cells with convex surface as sperm cells or diatoms. Here, we demonstrate an improvement of such procedure. By decoupling thickness information from refractive index ones and combining this with SFS analysis, 3D shape of concave cells is obtained. Specifically, the topography contour map is computed and used to adjust the 3D shape retrieved by the SFS algorithm. We prove the new procedure for healthy red blood cells having a concave surface in their central region. Experimental results are compared with theoretical model.

  15. Quantitative phase imaging of biological cells and tissues using singleshot white light interference microscopy and phase subtraction method for extended range of measurement

    NASA Astrophysics Data System (ADS)

    Mehta, Dalip Singh; Sharma, Anuradha; Dubey, Vishesh; Singh, Veena; Ahmad, Azeem

    2016-03-01

    We present a single-shot white light interference microscopy for the quantitative phase imaging (QPI) of biological cells and tissues. A common path white light interference microscope is developed and colorful white light interferogram is recorded by three-chip color CCD camera. The recorded white light interferogram is decomposed into the red, green and blue color wavelength component interferograms and processed it to find out the RI for different color wavelengths. The decomposed interferograms are analyzed using local model fitting (LMF)" algorithm developed for reconstructing the phase map from single interferogram. LMF is slightly off-axis interferometric QPI method which is a single-shot method that employs only a single image, so it is fast and accurate. The present method is very useful for dynamic process where path-length changes at millisecond level. From the single interferogram a wavelength-dependent quantitative phase imaging of human red blood cells (RBCs) are reconstructed and refractive index is determined. The LMF algorithm is simple to implement and is efficient in computation. The results are compared with the conventional phase shifting interferometry and Hilbert transform techniques.

  16. Zernike phase contrast cryo-electron microscopy reveals 100 kDa component in a protein complex

    NASA Astrophysics Data System (ADS)

    Wu, Yi-Min; Wang, Chun-Hsiung; Chang, Jen-wei; Chen, Yi-yun; Miyazaki, Naoyuki; Murata, Kazuyoshi; Nagayama, Kuniaki; Chang, Wei-Hau

    2013-12-01

    Cryo-electron microscopy (cryo-EM) has become a powerful technique for obtaining near atomic structures for large protein assemblies or large virus particles, but the application to protein particles smaller than 200-300 kDa has been hampered by the feeble phase contrast obtained for such small samples and the limited number of electrons tolerated by them without incurring excessive radiation damage. By implementing a thin-film quarter-wave phase plate to a cryo-EM, Nagayama, one of the present authors, has recently restored the long-lost very low spatial frequencies, generating in-focus phase contrast superior to that of conventional defocusing phase contrast, and successfully applied the so-called Zernike phase-plate cryo-EM to target various biological samples in native state. Nevertheless, the sought-after goal of using enhanced phase contrast to reveal a native protein as small as 100 kDa waits to be realized. Here, we report a study in which 200 kV Zernike phase-plate cryo-EM with a plate cut-on periodicity of 36 nm was applied to visualize 100 kDa components of various protein complexes, including the small domains on the surface of an icosahedral particle of ˜38 nm derived from the dragon grouper nervous necrosis virus (DGNNV) and the labile sub-complex dissociated from yeast RNA polymerase III of 17 nm. In the former case, we observed a phase contrast reversal phenomenon at the centre of the icosahedral particle and traced its root cause to the near matching of the cut-on size and the particle size. In summary, our work has demonstrated that Zernike phase-plate implementation can indeed expand the size range of proteins that can be successfully investigated by cryo-EM, opening the door for countless proteins. Finally, we briefly discuss the possibility of using a transfer lens system to enlarge the cut-on periodicity without further miniaturizing the plate pinhole.

  17. Characterization of LiBC by phase-contrast scanning transmission electron microscopy.

    PubMed

    Krumeich, Frank; Wörle, Michael; Reibisch, Philipp; Nesper, Reinhard

    2014-08-01

    LiBC was used as a model compound for probing the applicability of phase-contrast (PC) imaging in an aberration-corrected scanning transmission electron microscope (STEM) to visualize lithium distributions. In the LiBC structure, boron and carbon are arranged to hetero graphite layers between which lithium is incorporated. The crystal structure is reflected in the PC-STEM images recorded perpendicular to the layers. The experimental images and their defocus dependence match with multi-slice simulations calculated utilizing the reciprocity principle. The observation that a part of the Li positions is not occupied is likely an effect of the intense electron beam triggering Li displacement.

  18. Segmentation and Tracking of Lymphocytes Based on Modified Active Contour Models in Phase Contrast Microscopy Images

    PubMed Central

    Liu, Zhiwen

    2015-01-01

    The paper proposes an improved active contour model for segmenting and tracking accurate boundaries of the single lymphocyte in phase-contrast microscopic images. Active contour models have been widely used in object segmentation and tracking. However, current external-force-inspired methods are weak at handling low-contrast edges and suffer from initialization sensitivity. In order to segment low-contrast boundaries, we combine the region information of the object, extracted by morphology gray-scale reconstruction, and the edge information, extracted by the Laplacian of Gaussian filter, to obtain an improved feature map to compute the external force field for the evolution of active contours. To alleviate initial location sensitivity, we set the initial contour close to the real boundaries by performing morphological image processing. The proposed method was tested on live lymphocyte images acquired through the phase-contrast microscope from the blood samples of mice, and comparative experimental results showed the advantages of the proposed method in terms of the accuracy and the speed. Tracking experiments showed that the proposed method can accurately segment and track lymphocyte boundaries in microscopic images over time even in the presence of low-contrast edges, which will provide a good prerequisite for the quantitative analysis of lymphocyte morphology and motility. PMID:26089973

  19. Phase-contrast imaging in aberration-corrected scanning transmission electron microscopy.

    PubMed

    Krumeich, F; Müller, E; Wepf, R A

    2013-06-01

    Although the presence of phase-contrast information in bright field images recorded with a scanning transmission electron microscope (STEM) has been known for a long time, its systematic exploitation for the structural characterization of materials began only with the availability of aberration-corrected microscopes that allow sufficiently large illumination angles. Today, phase-contrast STEM (PC-STEM) imaging represents an increasingly important alternative to the well-established HRTEM method. In both methods, the image contrast is coherently generated and thus depends not only on illumination and collection angles but on defocus and specimen thickness as well. By PC-STEM, a projection of the crystal potential is obtained in thin areas, with the scattering sites being represented either with dark or bright contrast at two different defocus values which are both close to Gaussian defocus. This imaging behavior can be further investigated by image simulations performed with standard HRTEM simulation software based on the principle of reciprocity. As examples for the application of this method, PC-STEM results obtained on metal nanoparticles and dodecagonal quasicrystals dd-(Ta,V)₁.₆Te are discussed.

  20. A novel framework for cellular tracking and mitosis detection in dense phase contrast microscopy images.

    PubMed

    Thirusittampalam, Ketheesan; Hossain, M Julius; Ghita, Ovidiu; Whelan, Paul F

    2013-05-01

    The aim of this paper is to detail the development of a novel tracking framework that is able to extract the cell motility indicators and to determine the cellular division (mitosis) events in large time-lapse phase-contrast image sequences. To address the challenges induced by nonstructured (random) motion, cellular agglomeration, and cellular mitosis, the process of automatic (unsupervised) cell tracking is carried out in a sequential manner, where the interframe cell association is achieved by assessing the variation in the local cellular structures in consecutive frames of the image sequence. In our study, a strong emphasis has been placed on the robust use of the topological information in the cellular tracking process and in the development of targeted pattern recognition techniques that were designed to redress the problems caused by segmentation errors, and to precisely identify mitosis using a backward (reversed) tracking strategy. The proposed algorithm has been evaluated on dense phase-contrast cellular data and the experimental results indicate that the proposed algorithm is able to accurately track epithelial and endothelial cells in time-lapse image sequences that are characterized by low contrast and high level of noise. Our algorithm achieved 86.10% overall tracking accuracy and 90.12% mitosis detection accuracy.

  1. Analytical electron microscopy of fine-grained phases in primitive interplanetary dust particles and carbonaceous chondrites

    NASA Technical Reports Server (NTRS)

    Mackinnon, Ian D. R.; Rietmeijer, Frans J. M.; Mckay, David S.

    1987-01-01

    In order to describe the total mineralogical diversity within primitive extraterrestrial materials, individual interplanetary dust particles (IDPs) collected from the stratosphere as part of the JSC Cosmic Dust Curatorial Program were analyzed using a variety of AEM techniques. Identification of over 250 individual grains within one chondritic porous (CP) IDP shows that most phases could be formed by low temperature processes and that heating of the IDP during atmospheric entry is minimal and less than 600 C. In a review of the mineralogy of IDPs, it was suggested that the occurrence of other silicates such as enstatite whiskers is consistent with the formation in an early turbulent period of the solar nebula. Experimental confirmation of fundamental chemical and physical processes in a stellar environment, such as vapor phase condensation, nucleation, and growth by annealing, is an important aspect of astrophysical models for the evolution of the Solar System. A detailed comparison of chondritic IDP and carbonaceous chondrite mineralogies shows significant differences between the types of silicate minerals as well as the predominant oxides.

  2. Analytical electron microscopy of fine-grained phases in primitive interplanetary dust particles and carbonaceous chondrites

    NASA Astrophysics Data System (ADS)

    MacKinnon, Ian D. R.; Rietmeijer, Frans J. M.; McKay, David S.

    1987-05-01

    In order to describe the total mineralogical diversity within primitive extraterrestrial materials, individual interplanetary dust particles (IDPs) collected from the stratosphere as part of the JSC Cosmic Dust Curatorial Program were analyzed using a variety of AEM techniques. Identification of over 250 individual grains within one chondritic porous (CP) IDP shows that most phases could be formed by low temperature processes and that heating of the IDP during atmospheric entry is minimal and less than 600 C. In a review of the mineralogy of IDPs, it was suggested that the occurrence of other silicates such as enstatite whiskers is consistent with the formation in an early turbulent period of the solar nebula. Experimental confirmation of fundamental chemical and physical processes in a stellar environment, such as vapor phase condensation, nucleation, and growth by annealing, is an important aspect of astrophysical models for the evolution of the Solar System. A detailed comparison of chondritic IDP and carbonaceous chondrite mineralogies shows significant differences between the types of silicate minerals as well as the predominant oxides.

  3. Interaction potentials of anisotropic nanocrystals from the trajectory sampling of particle motion using in situ liquid phase transmission electron microscopy

    DOE PAGES

    Chen, Qian; Cho, Hoduk; Manthiram, Karthish; ...

    2015-03-23

    We demonstrate a generalizable strategy to use the relative trajectories of pairs and groups of nanocrystals, and potentially other nanoscale objects, moving in solution which can now be obtained by in situ liquid phase transmission electron microscopy (TEM) to determine the interaction potentials between nanocrystals. Such nanoscale interactions are crucial for collective behaviors and applications of synthetic nanocrystals and natural biomolecules, but have been very challenging to measure in situ at nanometer or sub-nanometer resolution. Here we use liquid phase TEM to extract the mathematical form of interaction potential between nanocrystals from their sampled trajectories. We show the power ofmore » this approach to reveal unanticipated features of nanocrystal–nanocrystal interactions by examining the anisotropic interaction potential between charged rod-shaped Au nanocrystals (Au nanorods); these Au nanorods assemble, in a tip-to-tip fashion in the liquid phase, in contrast to the well-known side-by-side arrangements commonly observed for drying-mediated assembly. These observations can be explained by a long-range and highly anisotropic electrostatic repulsion that leads to the tip-selective attachment. As a result, Au nanorods stay unassembled at a lower ionic strength, as the electrostatic repulsion is even longer-ranged. Our study not only provides a mechanistic understanding of the process by which metallic nanocrystals assemble but also demonstrates a method that can potentially quantify and elucidate a broad range of nanoscale interactions relevant to nanotechnology and biophysics.« less

  4. Liquid-solid phase transition of Ge-Sb-Te alloy observed by in-situ transmission electron microscopy.

    PubMed

    Berlin, Katja; Trampert, Achim

    2016-11-05

    Melting and crystallization dynamics of the multi-component Ge-Sb-Te alloy have been investigated by in-situ transmission electron microscopy (TEM). Starting point of the phase transition study is an ordered hexagonal Ge1Sb2Te4 thin film on Si(111) where the crystal structure and the chemical composition are verified by scanning TEM and electron energy-loss spectroscopy, respectively. The in-situ observation of the liquid phase at 600°C including the liquid-solid and liquid-vacuum interfaces and their movements was made possible due to an encapsulation of the TEM sample. The solid-liquid interface during melting displays a broad and diffuse transition zone characterized by a vacancy induced disordered state. Although the velocities of interface movements are measured to be in the nanometer per second scale, both, for crystallization and solidification, the underlying dynamic processes are considerably different. Melting reveals linear dependence on time, whereas crystallization exhibits a non-linear time-dependency featuring a superimposed start-stop motion. Our results may provide valuable insight into the atomic mechanisms at interfaces during the liquid-solid phase transition of Ge-Sb-Te alloys.

  5. Interaction Potentials of Anisotropic Nanocrystals from the Trajectory Sampling of Particle Motion using in Situ Liquid Phase Transmission Electron Microscopy

    PubMed Central

    2015-01-01

    We demonstrate a generalizable strategy to use the relative trajectories of pairs and groups of nanocrystals, and potentially other nanoscale objects, moving in solution which can now be obtained by in situ liquid phase transmission electron microscopy (TEM) to determine the interaction potentials between nanocrystals. Such nanoscale interactions are crucial for collective behaviors and applications of synthetic nanocrystals and natural biomolecules, but have been very challenging to measure in situ at nanometer or sub-nanometer resolution. Here we use liquid phase TEM to extract the mathematical form of interaction potential between nanocrystals from their sampled trajectories. We show the power of this approach to reveal unanticipated features of nanocrystal–nanocrystal interactions by examining the anisotropic interaction potential between charged rod-shaped Au nanocrystals (Au nanorods); these Au nanorods assemble, in a tip-to-tip fashion in the liquid phase, in contrast to the well-known side-by-side arrangements commonly observed for drying-mediated assembly. These observations can be explained by a long-range and highly anisotropic electrostatic repulsion that leads to the tip-selective attachment. As a result, Au nanorods stay unassembled at a lower ionic strength, as the electrostatic repulsion is even longer-ranged. Our study not only provides a mechanistic understanding of the process by which metallic nanocrystals assemble but also demonstrates a method that can potentially quantify and elucidate a broad range of nanoscale interactions relevant to nanotechnology and biophysics. PMID:27162944

  6. Direct observation of martensitic phase-transformation dynamics in iron by 4D single-pulse electron microscopy.

    PubMed

    Park, Hyun Soon; Kwon, Oh-Hoon; Baskin, J Spencer; Barwick, Brett; Zewail, Ahmed H

    2009-11-01

    The in situ martensitic phase transformation of iron, a complex solid-state transition involving collective atomic displacement and interface movement, is studied in real time by means of four-dimensional (4D) electron microscopy. The iron nanofilm specimen is heated at a maximum rate of approximately 10(11) K/s by a single heating pulse, and the evolution of the phase transformation from body-centered cubic to face-centered cubic crystal structure is followed by means of single-pulse, selected-area diffraction and real-space imaging. Two distinct components are revealed in the evolution of the crystal structure. The first, on the nanosecond time scale, is a direct martensitic transformation, which proceeds in regions heated into the temperature range of stability of the fcc phase, 1185-1667 K. The second, on the microsecond time scale, represents an indirect process for the hottest central zone of laser heating, where the temperature is initially above 1667 K and cooling is the rate-determining step. The mechanism of the direct transformation involves two steps, that of (barrier-crossing) nucleation on the reported nanosecond time scale, followed by a rapid grain growth typically in approximately 100 ps for 10 nm crystallites.

  7. Automated Method for the Rapid and Precise Estimation of Adherent Cell Culture Characteristics from Phase Contrast Microscopy Images

    PubMed Central

    Jaccard, Nicolas; Griffin, Lewis D; Keser, Ana; Macown, Rhys J; Super, Alexandre; Veraitch, Farlan S; Szita, Nicolas

    2014-01-01

    The quantitative determination of key adherent cell culture characteristics such as confluency, morphology, and cell density is necessary for the evaluation of experimental outcomes and to provide a suitable basis for the establishment of robust cell culture protocols. Automated processing of images acquired using phase contrast microscopy (PCM), an imaging modality widely used for the visual inspection of adherent cell cultures, could enable the non-invasive determination of these characteristics. We present an image-processing approach that accurately detects cellular objects in PCM images through a combination of local contrast thresholding and post hoc correction of halo artifacts. The method was thoroughly validated using a variety of cell lines, microscope models and imaging conditions, demonstrating consistently high segmentation performance in all cases and very short processing times (<1 s per 1,208 × 960 pixels image). Based on the high segmentation performance, it was possible to precisely determine culture confluency, cell density, and the morphology of cellular objects, demonstrating the wide applicability of our algorithm for typical microscopy image processing pipelines. Furthermore, PCM image segmentation was used to facilitate the interpretation and analysis of fluorescence microscopy data, enabling the determination of temporal and spatial expression patterns of a fluorescent reporter. We created a software toolbox (PHANTAST) that bundles all the algorithms and provides an easy to use graphical user interface. Source-code for MATLAB and ImageJ is freely available under a permissive open-source license. Biotechnol. Bioeng. 2014;111: 504–517. © 2013 Wiley Periodicals, Inc. PMID:24037521

  8. Multiplexed off-axis interferometric phase microscopy for dynamic cell measurements

    NASA Astrophysics Data System (ADS)

    Shaked, Natan T.; Girshovitz, Pinhas; Frenklach, Irena

    2015-03-01

    We present a new approach of optically multiplexing several off-axis interferograms on the same digital camera, each of which encodes a different field of view of the sample. Since the fringes of these interferograms are in different directions, as obtained experimentally by the optical system, we are able to double or even triple the amount of information that can be acquired in a single camera exposure, with the same number of camera pixels, while sharing the camera dynamic range. We show that this method can partially solve the problem of limited off-axis interferometric field of view due to low-coherence illumination. Our experimental demonstrations include quantitative phase imaging of microscopic diatom shells, fast swimming sperm cells and microorganisms, and contracting cardiomyocytes.

  9. Stain-Free Quantification of Chromosomes in Live Cells Using Regularized Tomographic Phase Microscopy

    PubMed Central

    Sung, Yongjin; Choi, Wonshik; Lue, Niyom; Dasari, Ramachandra R.; Yaqoob, Zahid

    2012-01-01

    Refractive index imaging is a label-free technique that enables long-term monitoring of the internal structures and molecular composition in living cells with minimal perturbation. Existing tomographic methods for the refractive index imaging lack 3-D resolution and result in artifacts that prevent accurate refractive index quantification. To overcome these limitations without compromising the capability to observe a sample in its most native condition, we have developed a regularized tomographic phase microscope (RTPM) enabling accurate refractive index imaging of organelles inside intact cells. With the enhanced accuracy, we quantify the mass of chromosomes in intact living cells, and differentiate two human colon cancer lines, HT-29 and T84 cells, solely based on the non-aqueous (dry) mass of chromosomes. In addition, we demonstrate chromosomal imaging using a dual-wavelength RTPM, which shows its potential to determine the molecular composition of cellular organelles in live cells. PMID:23166689

  10. Aberration-corrected transmission electron microscopy of the intergranular phase in magnetic recording media.

    PubMed

    Hossein-Babaei, Faraz; Koh, Ai Leen; Srinivasan, Kumar; Bertero, Gerardo A; Sinclair, Robert

    2012-05-09

    In perpendicular hard disk memory media, nanometric magnetic Co-rich grains are separated by a ∼1 nm thick nonmagnetic and preferably amorphous intergranular phase (IP). Attempts at observing the IP structure at high resolution using TEM have been obstructed by the superposition of lattice fringes from the crystalline grains extending into the IP region in images. Here we present the first images of a magnetic recording medium produced using a spherical aberration-corrected TEM showing the true amorphous IP structure in contrast to the crystalline grains, allowing the accurate determination of the grain-IP interface and the grain and IP dimensions. It is shown that these aberration-corrected TEM images are functionally superior for analyzing certain features of the ultrahigh capacity data recording media.

  11. Nanoscale nuclear architecture for cancer diagnosis beyond pathology via spatial-domain low-coherence quantitative phase microscopy

    NASA Astrophysics Data System (ADS)

    Wang, Pin; Bista, Rajan K.; Khalbuss, Walid E.; Qiu, Wei; Uttam, Shikhar; Staton, Kevin; Zhang, Lin; Brentnall, Teresa A.; Brand, Randall E.; Liu, Yang

    2010-11-01

    Definitive diagnosis of malignancy is often challenging due to limited availability of human cell or tissue samples and morphological similarity with certain benign conditions. Our recently developed novel technology-spatial-domain low-coherence quantitative phase microscopy (SL-QPM)-overcomes the technical difficulties and enables us to obtain quantitative information about cell nuclear architectural characteristics with nanoscale sensitivity. We explore its ability to improve the identification of malignancy, especially in cytopathologically non-cancerous-appearing cells. We perform proof-of-concept experiments with an animal model of colorectal carcinogenesis-APCMin mouse model and human cytology specimens of colorectal cancer. We show the ability of in situ nanoscale nuclear architectural characteristics in identifying cancerous cells, especially in those labeled as ``indeterminate or normal'' by expert cytopathologists. Our approach is based on the quantitative analysis of the cell nucleus on the original cytology slides without additional processing, which can be readily applied in a conventional clinical setting. Our simple and practical optical microscopy technique may lead to the development of novel methods for early detection of cancer.

  12. Helium ion microscopy based wall thickness and surface roughness analysis of polymer foams obtained from high internal phase emulsion.

    PubMed

    Rodenburg, C; Viswanathan, P; Jepson, M A E; Liu, X; Battaglia, G

    2014-04-01

    Due to their wide range of applications, porous polymers obtained from high internal phase emulsions have been widely studied using scanning electron microscopy. However, due to their lack of electrical conductivity, quantitative information of wall thicknesses and surface roughness, which are of particular interest to tissue engineering, has not been obtained. Here, Helium Ion Microscopy is used to examine uncoated polymer foams and some very strong but unexpected contrast is observed, the origin of which is established here. Based on this analysis, a method for the measurement of wall thickness variations and wall roughness measurements has been developed, based on the modeling of Helium ion transmission. The results presented here indicate that within the walls of the void structure there exist small features with height variations of ~30 nm and wall thickness variations from ~100 nm to larger 340 nm in regions surrounding interconnecting windows within the structure. The suggested imaging method is applicable to other porous carbon based structures with wall thicknesses in the range of 40-340 nm.

  13. Reflectivity and topography of cells grown on glass-coverslips measured with phase-shifted laser feedback interference microscopy

    PubMed Central

    Atılgan, Erdinç; Ovryn, Ben

    2011-01-01

    In spite of the advantages associated with the molecular specificity of fluorescence imaging, there is still a significant need to augment these approaches with label-free imaging. Therefore, we have implemented a form of interference microscopy based upon phase-shifted, laser-feedback interferometry and developed an algorithm that can be used to separate the contribution of the elastically scattered light by sub-cellular structures from the reflection at the coverslip-buffer interface. The method offers an opportunity to probe protein aggregation, index of refraction variations and structure. We measure the topography and reflection from calibration spheres and from stress fibers and adhesions in both fixed and motile cells. Unlike the data acquired with reflection interference contrast microscopy, where the reflection from adhesions can appear dark, our approach demonstrates that these regions have high reflectivity. The data acquired from fixed and live cells show the presence of a dense actin layer located ≈ 100 nm above the coverslip interface. Finally, the measured dynamics of filopodia and the lamella in a live cell supports retrograde flow as the dominate mechanism responsible for filopodia retraction. PMID:21833378

  14. Reflectivity and topography of cells grown on glass-coverslips measured with phase-shifted laser feedback interference microscopy.

    PubMed

    Atılgan, Erdinç; Ovryn, Ben

    2011-08-01

    In spite of the advantages associated with the molecular specificity of fluorescence imaging, there is still a significant need to augment these approaches with label-free imaging. Therefore, we have implemented a form of interference microscopy based upon phase-shifted, laser-feedback interferometry and developed an algorithm that can be used to separate the contribution of the elastically scattered light by sub-cellular structures from the reflection at the coverslip-buffer interface. The method offers an opportunity to probe protein aggregation, index of refraction variations and structure. We measure the topography and reflection from calibration spheres and from stress fibers and adhesions in both fixed and motile cells. Unlike the data acquired with reflection interference contrast microscopy, where the reflection from adhesions can appear dark, our approach demonstrates that these regions have high reflectivity. The data acquired from fixed and live cells show the presence of a dense actin layer located ≈ 100 nm above the coverslip interface. Finally, the measured dynamics of filopodia and the lamella in a live cell supports retrograde flow as the dominate mechanism responsible for filopodia retraction.

  15. Enhancing two-color absorption, self-phase modulation, and Raman microscopy signatures in tissue with femtosecond laser pulse shaping

    NASA Astrophysics Data System (ADS)

    Fischer, Martin C.; Piletic, Ivan; Fu, Dan; Matthews, Thomas E.; Liu, Henry; Samineni, Prathyush; Li, Baolei; Warren, Warren S.

    2009-02-01

    Nonlinear microscopies (most commonly, two-photon fluorescence, second harmonic generation, and coherent anti-Stokes Raman scattering (CARS)) have had notable successes in imaging a variety of endogenous and exogenous targets in recent years. These methods generate light at a color different from any of the exciting laser pulses, which makes the signal relatively easy to detect. Our work has focused on developing microscopy techniques using a wider range of nonlinear signatures (two-photon absorption of nonfluorescent species, self phase modulation) which have some specific advantages - for example, in recent papers we have shown that we can differentiate between different types of melanin in pigmented lesions, image hemoglobin and its oxygenation, and measure neuronal activity. In general, these signatures do not generate light at a different color and we rely on the advantages of femtosecond laser pulse shaping methods to amplify the signals and make them visible (for example, using heterodyne detection of the induced signal with one of the co-propagating laser pulses). Here we extend this work to stimulated Raman and CARS geometries. In the simplest experiments, both colors arise from filtering a single fs laser pulse, then modulating afterwards; in other cases, we demonstrate that spectral reshaping can retain high frequency resolution in Raman and CARS geometries with femtosecond laser pulses.

  16. Real-time and non-invasive measurements of cell mechanical behaviour with optical coherence phase microscopy

    NASA Astrophysics Data System (ADS)

    Gillies, D.; Gamal, W.; Downes, A.; Reinwald, Y.; Yang, Y.; El Haj, A.; Bagnaninchi, P. O.

    2017-02-01

    There is an unmet need in tissue engineering for non-invasive, label-free monitoring of cell mechanical behaviour in their physiological environment. Here, we describe a novel optical coherence phase microscopy (OCPM) set-up which can map relative cell mechanical behaviour in monolayers and 3D systems non-invasively, and in real-time. 3T3 and MCF-7 cells were investigated, with MCF-7 demonstrating an increased response to hydrostatic stimulus indicating MCF-7 being softer than 3T3. Thus, OCPM shows the ability to provide qualitative data on cell mechanical behaviour. Quantitative measurements of 6% agarose beads have been taken with commercial Cell Scale Microsquisher system demonstrating that their mechanical properties are in the same order of magnitude of cells, indicating that this is an appropriate test sample for the novel method described.

  17. Insights into the nanoscale lateral and vertical phase separation in organic bulk heterojunctions via scanning probe microscopy

    NASA Astrophysics Data System (ADS)

    Chintala, R.; Tait, J. G.; Eyben, P.; Voroshazi, E.; Surana, S.; Fleischmann, C.; Conard, T.; Vandervorst, W.

    2016-02-01

    Solution processed polymer (donor) and fullerene (acceptor) bulk heterojunctions are widely used as the photo active layer in organic solar cells. Intimate mixing of these two materials is essential for efficient charge separation and transport. Identifying relative positions of acceptor and donor rich regions in the bulk heterojunction with nanometer scale precision is crucial in understanding intricate details of operation. In this work, a combination of Ar+2000 gas cluster ion beam and scanning probe microscopy is used to examine the lateral and vertical phase separation within regio-regular poly(3-hexylthiophene)(P3HT):phenyl-C60-butyric acid methyl ester (PCBM) bulk heterojunction. While the Ar+2000 gas cluster ion beam is used as a sputter tool to expose the underneath layers, scanning probe microscopy techniques are used to obtain two-dimensional (2D) electrical maps (with sub-2 nm lateral resolution). The electrical mapping is decoded to chemical composition, essentially producing lateral and vertical maps of phase separation. Thermal stress causes large PCBM-rich hillocks to form, and consequently affecting the balance of P3HT:PCBM heterojunctions, hence a negative impact on the efficiency of the solar cell. We further developed a method to analyze the efficiency of exciton dissociation based on the current maps and a loss of 20% in efficiency is observed for thermally degraded samples compared to fresh un-annealed samples.Solution processed polymer (donor) and fullerene (acceptor) bulk heterojunctions are widely used as the photo active layer in organic solar cells. Intimate mixing of these two materials is essential for efficient charge separation and transport. Identifying relative positions of acceptor and donor rich regions in the bulk heterojunction with nanometer scale precision is crucial in understanding intricate details of operation. In this work, a combination of Ar+2000 gas cluster ion beam and scanning probe microscopy is used to examine the

  18. Rapid, High-Throughput Tracking of Bacterial Motility in 3D via Phase-Contrast Holographic Video Microscopy

    PubMed Central

    Cheong, Fook Chiong; Wong, Chui Ching; Gao, YunFeng; Nai, Mui Hoon; Cui, Yidan; Park, Sungsu; Kenney, Linda J.; Lim, Chwee Teck

    2015-01-01

    Tracking fast-swimming bacteria in three dimensions can be extremely challenging with current optical techniques and a microscopic approach that can rapidly acquire volumetric information is required. Here, we introduce phase-contrast holographic video microscopy as a solution for the simultaneous tracking of multiple fast moving cells in three dimensions. This technique uses interference patterns formed between the scattered and the incident field to infer the three-dimensional (3D) position and size of bacteria. Using this optical approach, motility dynamics of multiple bacteria in three dimensions, such as speed and turn angles, can be obtained within minutes. We demonstrated the feasibility of this method by effectively tracking multiple bacteria species, including Escherichia coli, Agrobacterium tumefaciens, and Pseudomonas aeruginosa. In addition, we combined our fast 3D imaging technique with a microfluidic device to present an example of a drug/chemical assay to study effects on bacterial motility. PMID:25762336

  19. Solid-phase immune electron microscopy with human immunoglobulin M for serotyping of Norwalk-like viruses.

    PubMed Central

    Lewis, D C; Lightfoot, N F; Pether, J V

    1988-01-01

    A solid-phase immune electron microscopy method that uses protein A, goat anti-human immunoglobulin M (IgM), and human serum is described. Evaluation of the method with different immunoglobulin fractions showed that human IgM constituted the major virus capture antibody. The method appeared to distinguish between two Norwalk-like virus serotypes and demonstrated specific IgM responses to these serotypes in infected individuals. Further work is being carried out to define the relationship of these two serotypes to the previously described Norwalk agent (A. Z. Kapikian, R. G. Wyatt, R. Dolin, T. S. Thornhill, A. R. Kalica, and R. M. Chanock, J. Virol. 10:1075-1081, 1972), and four subsequent hospital outbreaks are being studied. PMID:2838506

  20. Digital reconstruction based on angular spectrum diffraction with the ridge of wavelet transform in holographic phase-contrast microscopy.

    PubMed

    Weng, Jiawen; Zhong, Jiangang; Hu, Cuiying

    2008-12-22

    A numerical reconstruction technique of digital holography based on angular spectrum diffraction by means of the ridge of Gabor wavelet transform (GWT) is presented. Appling the GWT, the object wave can be reconstructed by calculating the wavelet coefficients of the hologram at the ridge of the GWT automatically even if the spectrum of the virtual image is disturbed by the other spectrum. It provides a way to eliminate the effect of the zero-order and the twin-image terms without the spatial filtering. In particular, based on the angular spectrum theory, GWT is applied to the digital holographic phase-contrast microscopy on biological specimens. The theory, the results of a simulation and an experiment of an onion specimen are shown.

  1. Alterations of filopodia by near infrared photoimmunotherapy: evaluation with 3D low-coherent quantitative phase microscopy

    PubMed Central

    Nakamura, Yuko; Nagaya, Tadanobu; Sato, Kazuhide; Harada, Toshiko; Okuyama, Shuhei; Choyke, Peter L.; Yamauchi, Toyohiko; Kobayashi, Hisataka

    2016-01-01

    Filopodia are highly organized cellular membrane structures that facilitate intercellular communication. Near infrared photoimmunotherapy (NIR-PIT) is a newly developed cancer treatment that causes necrotic cell death. Three-dimensional low-coherent quantitative phase microscopy (3D LC-QPM) is based on a newly established low-coherent interference microscope designed to obtain serial topographic images of the cellular membrane. Herein, we report rapid involution of filopodia after NIR-PIT using 3D LC-QPM. For 3T3/HER2 cells, the number of filopodia decreased immediately after treatment with significant differences. Volume and relative height of 3T3/HER2 cells increased immediately after NIR light exposure, but significant differences were not observed. Thus, disappearance of filopodia, evaluated by 3D LC-QPM, is an early indicator of cell membrane damage after NIR-PIT. PMID:27446702

  2. The measurement of red blood cell volume change induced by Ca2+ based on full field quantitative phase microscopy

    NASA Astrophysics Data System (ADS)

    Lee, Seungrag; Lee, Ji Yong; Yang, Wenzhong; Kim, Dug Young

    2009-02-01

    We present the measurement of red blood cell (RBC) volume change induced by Ca2+ for a live cell imaging with full field quantitative phase microscopy (FFQPM). FFQPM is based on the Mach-Zehnder interferometer combined with an inverted microscopy system. We present the effective method to obtain a clear image and an accurate volume of the cells. An edge detection technique is used to accurately resolve the boundary between the cell line and the suspension medium. The measurement of the polystyrene bead diameter and volume has been demonstrated the validity of our proposed method. The measured phase profile can be easily converted into thickness profile. The measured polystyrene bead volume and the simulated result are about 14.74 μm3 and 14.14 μm3, respectively. The experimental results of our proposed method agree well with the simulated results within less than 4 %. We have also measured the volume variation of a single RBC on a millisecond time scale. Its mean volume is 54.02 μm3 and its standard deviation is 0.52 μm3. With the proposed system, the shape and volume changes of RBC induced by the increased intracellular Ca2+ are measured after adding ionophore A23187. A discocyte RBC is deformed to a spherocyte due to the increased intracellular Ca2+ in RBC. The volume of the spherocyte is 47.88 μm3 and its standard deviation is 0.19 μm3. We have demonstrated that the volume measurement technique is easy, accurate, and robust method with high volume sensitivity (<0.0000452 μm3) and this provides the ability to study a biological phenomenon in Hematology.

  3. A phase-contrast microscopy-based method for modeling the mechanical behavior of mesenchymal stem cells.

    PubMed

    Saeed, Mayssam; Sharabani-Yosef, Orna; Weihs, Daphne; Gefen, Amit

    2016-10-01

    We present three-dimensional (3D) finite element (FE) models of single, mesenchymal stem cells (MSCs), generated from images obtained by optical phase-contrast microscopy and used to quantify the structural responses of the studied cells to externally applied mechanical loads. Mechanical loading has been shown to affect cell morphology and structure, phenotype, motility and other biological functions. Cells experience mechanical loads naturally, yet under prolonged or sizable loading, damage and cell death may occur, which motivates research regarding the structural behavior of loaded cells. For example, near the weight-bearing boney prominences of the buttocks of immobile persons, tissues may become highly loaded, eventually leading to massive cell death that manifests as pressure ulcers. Cell-specific computational models have previously been developed by our group, allowing simulations of cell deformations under compressive or stretching loads. These models were obtained by reconstructing specific cell structures from series of 2D fluorescence, confocal image-slices, requiring cell-specific fluorescent-staining protocols and costly (confocal) microscopy equipment. Alternative modeling approaches represent cells simply as half-spheres or half-ellipsoids (i.e. idealized geometries), which neglects the curvature details of the cell surfaces associated with changes in concentrations of strains and stresses. Thus, we introduce here for the first time an optical image-based FE modeling, where loads are simulated on reconstructed 3D geometrical cell models from a single 2D, phase-contrast image. Our novel modeling method eliminates the need for confocal imaging and fluorescent staining preparations (both expensive), and makes cell-specific FE modeling affordable and accessible to the biomechanics community. We demonstrate the utility of this cost-effective modeling method by performing simulations of compression of MSCs embedded in a gel.

  4. Determination of partition coefficients of biomolecules in a microfluidic aqueous two phase system platform using fluorescence microscopy.

    PubMed

    Silva, D F C; Azevedo, A M; Fernandes, P; Chu, V; Conde, J P; Aires-Barros, M R

    2017-03-03

    Aqueous two phase systems (ATPS) offer great potential for selective separation of a wide range of biomolecules by exploring differences in molecular solubility in each of the two immiscible phases. However, ATPS use has been limited due to the difficulty in predicting the behavior of a given biomolecule in the partition environment together with the empirical and time-consuming techniques that are used for the determination of partition and extraction parameters. In this work, a fast and novel technique based on a microfluidic platform and using fluorescence microscopy was developed to determine the partition coefficients of biomolecules in different ATPS. This method consists of using a microfluidic device with a single microchannel and three inlets. In two of the inlets, solutions containing the ATPS forming components were loaded while the third inlet was fed with the FITC tagged biomolecule of interest prepared in milli-Q water. Using fluorescence microscopy, it was possible to follow the location of the FITC-tagged biomolecule and, by simply varying the pumping rates of the solutions, to quickly test a wide variety of ATPS compositions. The ATPS system is allowed 4min for stabilization and fluorescence micrographs are used to determine the partition coefficient.The partition coefficients obtained were shown to be consistent with results from macroscale ATPS partition. This process allows for faster screening of partition coefficients using only a few microliters of material for each ATPS composition and is amenable to automation. The partitioning behavior of several biomolecules with molecular weights (MW) ranging from 5.8 to 150kDa, and isoelectric points (pI) ranging from 4.7 to 6.4 was investigated, as well as the effect of the molecular weight of the polymer ATPS component.

  5. Iodine-water-alkanol and potassium iodide-water-alkanol systems: Phase diagrams and regularities of association according to IR and NMR spectroscopic data

    NASA Astrophysics Data System (ADS)

    Monakhova, Yu. B.; Varlamova, T. M.; Rubtsova, E. M.; Mushtakova, S. P.

    2015-04-01

    The variation of the iodine and potassium iodide solubilities in water-monoatomic alcohol (ethanol, propanol, isopropanol) solvents is considered from the standpoint of IR spectroscopic and chemometric data on association in water-alkanol binary mixtures. The iodine and potassium iodide solubilities in the mixed solvents vary nonlinearly with solvent composition because of the formation of 1 : 1 and 1 : 3 water-alcohol heteroassociates and alcohol homoassociates. Different kinds of phase diagram are observed for the iodine-water-alcohol systems: the I2-H2O-1-C3H7OH and the I2-H2O-2-C3H7OH diagrams have a phase separation region, while the I2-H2O-C2H5OH diagram does not. This fact is explained in terms of the interaction between the components of the systems. The variation of the potassium iodide solubility in the mixed solvent is discussed: a decrease in the KI solubility is symbatic to an increase in the relative concentration of 1 : 1 associates in the water-alcohol solution. The run of the iodine and potassium iodide solubility curves at low alcohol concentrations is explained on the basis of NMR spectroscopic data on association in aqueous solutions of the monoatomic alcohols.

  6. Gel-gel phase separation within milk sphingomyelin domains revealed at the nanoscale using atomic force microscopy.

    PubMed

    Guyomarc'h, Fanny; Chen, Maohui; Et-Thakafy, Oumaima; Zou, Shan; Lopez, Christelle

    2017-05-01

    The milk sphingomyelin (MSM) is involved in the formation of ordered lipid domains in the biological milk fat globule membrane (MFGM), where it accounts for about 30%wt of the polar lipids. Moreover, MSM exhibits a large variety in saturated acyl chain lengths (from C16:0 to C24:0-SM) compared to other natural sphingomyelins, which may impact the packing of MSM molecular species in the gel phase domains and the topography of the MFGM. To investigate this, supported lipid bilayers of synthetic sphingomyelins or of MSM-containing mixtures, including a MFGM polar lipid extract, were imaged at temperatures below the Tm of MSM (i.e. <34°C for which MSM is in the gel phase) in hydrated conditions using atomic force microscopy. In all compositions containing MSM, the MSM-rich gel phase domains exhibited lower and upper height levels H, interpreted as two distinct gel phases with ∆H~0.5-1.1nm. Two (lower and upper) gel phases were also found for pure C24:0-SM bilayers or for bilayers of a C16:0-SM/C24:0-SM equimolar mixture, while C16:0-SM bilayers were uniformly flat and less thick than C24:0-SM bilayers. The upper gel phase of MSM-containing bilayers was interpreted as mixed interdigitated C24:0-SM molecules, while the lower gel phase was attributed both to fully interdigitated C24:0-SM molecules and non-interdigitated C16:0-SM molecules. These results show that the composition of natural sphingomyelins, inducing a mismatch between the d18:1 sphingosine and the acyl chains, is important in both the internal organization and the topography of biological membranes, especially that of the MFGM. This organization could be involved in specific biological functions, e.g. the insertion of proteins. Copyright © 2017 Elsevier B.V. All rights reserved.

  7. Structured illumination multimodal 3D-resolved quantitative phase and fluorescence sub-diffraction microscopy

    PubMed Central

    Chowdhury, Shwetadwip; Eldridge, Will J.; Wax, Adam; Izatt, Joseph A.

    2017-01-01

    Sub-diffraction resolution imaging has played a pivotal role in biological research by visualizing key, but previously unresolvable, sub-cellular structures. Unfortunately, applications of far-field sub-diffraction resolution are currently divided between fluorescent and coherent-diffraction regimes, and a multimodal sub-diffraction technique that bridges this gap has not yet been demonstrated. Here we report that structured illumination (SI) allows multimodal sub-diffraction imaging of both coherent quantitative-phase (QP) and fluorescence. Due to SI’s conventionally fluorescent applications, we first demonstrate the principle of SI-enabled three-dimensional (3D) QP sub-diffraction imaging with calibration microspheres. Image analysis confirmed enhanced lateral and axial resolutions over diffraction-limited QP imaging, and established striking parallels between coherent SI and conventional optical diffraction tomography. We next introduce an optical system utilizing SI to achieve 3D sub-diffraction, multimodal QP/fluorescent visualization of A549 biological cells fluorescently tagged for F-actin. Our results suggest that SI has a unique utility in studying biological phenomena with significant molecular, biophysical, and biochemical components. PMID:28663887

  8. Phase and Index of Refraction Imaging by Hyperspectral Reflectance Confocal Microscopy.

    PubMed

    Selci, Stefano

    2016-12-16

    A hyperspectral reflectance confocal microscope (HSCM) was realized by CNR-ISC (Consiglio Nazionale delle Ricerche-Istituto dei Sistemi Complessi) a few years ago. The instrument and data have been already presented and discussed. The main activity of this HSCM has been within biology, and reflectance data have shown good matching between spectral signatures and the nature or evolution on many types of cells. Such a relationship has been demonstrated mainly with statistical tools like Principal Component Analysis (PCA), or similar concepts, which represent a very common approach for hyperspectral imaging. However, the point is that reflectance data contains much more useful information and, moreover, there is an obvious interest to go from reflectance, bound to the single experiment, to reflectivity, or other physical quantities, related to the sample alone. To accomplish this aim, we can follow well-established analyses and methods used in reflectance spectroscopy. Therefore, we show methods of calculations for index of refraction n, extinction coefficient k and local thicknesses of frequency starting from phase images by fast Kramers-Kronig (KK) algorithms and the Abeles matrix formalism. Details, limitations and problems of the presented calculations as well as alternative procedures are given for an example of HSCM images of red blood cells (RBC).

  9. Time-gated digital phase conjugation for two-photon excitation microscopy through multimode optical fibers

    NASA Astrophysics Data System (ADS)

    Morales-Delgado, Edgar E.; Farahi, Salma; Papadopoulos, Ioannis N.; Psaltis, Demetri; Moser, Christophe

    2015-12-01

    The large number of modes supported by multimode optical fibers potentially allows the transmission of larger amounts of information compared to single mode fibers. However, when pulsed light is transmitted through multimode fibers, the spatio-temporal profile of the incident beam is altered upon propagation, leading to a highly scrambled spatial profile and a broadened temporal duration due to modal and material dispersion. We present a digital phase conjugation method to counter-propagate through a multimode optical fiber only a group of modes of similar propagation constants which interfere constructively at a single location at the other side of the fiber, generating a spatially focused pulse. Since only modes with the same speed are excited, temporal broadening due to modal dispersion is suppressed, preserving the ultrashort duration of the propagating pulse. Using this technique, we experimentally demonstrate the transmission of focused pulses of 500 fs through a 30 cm length, 200 micrometer core step-index multimode fiber. Additionally, using a graded-index fiber, which allows the propagation of a larger number of modes of the same speed than a graded index fiber (hence a better focusing capability), we have been able to deliver and scan high-intensity focused femtosecond pulses. Moreover, based on the described principle, we demonstrate for the first time two-photon excitation imaging through a multimode optical fiber.

  10. Analysis of holographic photopolymers for integrated optical systems via quantitative phase microscopy

    NASA Astrophysics Data System (ADS)

    Glugla, David J.; Alim, Marvin D.; Chosy, Madeline B.; Sullivan, Amy C.; McLeod, Robert R.

    2017-05-01

    Optically-driven diffusion of high refractive index molecules within a transparent thermoset polymer matrix is a promising platform for hybrid optics that combines a wide range of optical structures from large scale holograms to micron-scale gradient index waveguides in a single integrated optical system. Design of such a system requires characterization of the optical response of the material at a wide range of spatial scales and intensities. While holographic analysis of the photopolymers is appropriate to probe the smaller spatial scales and lower intensity optical response, quantitative phase mapping of isolated structures is needed to probe the response to the higher intensities and lower spatial frequencies used in direct write lithography of waveguides. We apply the transport of intensity equation (TIE) to demonstrate quantitative refractive index measurements of 10 μm-scale localized gradient index structures written into diffusive photopolymer materials using both single- and two-photon polymerization. These quantitative measurements allow us to study the effect of different exposure conditions and material parameters such as writing beam power, exposure time, and wt% loading of the writing monomer on the overall profile of the refractive index structure. We use these measurements to probe the time scales over which diffusion is significant, and take advantage of the diffusion of monomer with a multiple-write scheme that achieves a peak refractive index contrast of 0.025.

  11. High-resolution scanning tunneling microscopy of fully hydrated ripple-phase bilayers.

    PubMed

    Woodward, J T; Zasadzinski, J A

    1997-02-01

    A modified freeze-fracture replication technique for use with the scanning tunneling microscope (STM) has provided a quantitative, high-resolution description of the waveform and amplitude of rippled bilayers in the P beta' phase of dimyristoylphosphatidylcholine (DMPC) in excess water. The ripples are uniaxial and asymmetrical, with a temperature-dependent amplitude of 2.4 nm near the chain melting temperature that decreases to zero at the chain crystallization temperature. The wavelength of 11 nm does not change with temperature. The observed ripple shape and the temperature-induced structural changes are not predicted by any current theory. Calibration and reproducibility of the STM/replica technique were tested with replicas of well-characterized bilayers of cadmium arachidate on mica that provide regular 5.5-nm steps. STM images were analyzed using a cross-correlation averaging program to eliminate the effects of noise and the finite size and shapes of the metal grains that make up the replica. The correlation averaging allowed us to develop a composite ripple profile averaged over hundreds of individual ripples measured on different samples with different STM tips. The STM/replica technique avoids many of the previous artifacts of biological STM imaging and can be used to examine a variety of periodic hydrated lipid and protein samples at a lateral resolution of about 1 nm and a vertical resolution of about 0.3 nm. This resolution is superior to conventional and tapping mode AFM to soft biological materials; the technique is substrate-free, and the conductive and chemically uniform replicas make image interpretation simple and direct.

  12. High-Resolution Scanning Tunneling Microscopy of Fully Hydrated Ripple-Phase Bilayers

    PubMed Central

    Woodward IV, J. T.; Zasadzinski, J. A.

    1997-01-01

    A modified freeze-fracture replication technique for use with the scanning tunneling microscope (STM) has provided a quantitative, high-resolution description of the waveform and amplitude of rippled bilayers in the Pβ, phase of dimyristoylphosphatidylcholine (DMPC) in excess water. The ripples are uniaxial and asymmetrical, with a temperature-dependent amplitude of 2.4 nm near the chain melting temperature that decreases to zero at the chain crystallization temperature. The wavelength of 11 nm does not change with temperature. The observed ripple shape and the temperature-induced structural changes are not predicted by any current theory. Calibration and reproducibility of the STM/replica technique were tested with replicas of well-characterized bilayers of cadmium arachidate on mica that provide regular 5.5-nm steps. STM images were analyzed using a cross-correlation averaging program to eliminate the effects of noise and the finite size and shapes of the metal grains that make up the replica. The correlation averaging allowed us to develop a composite ripple profile averaged over hundreds of individual ripples measured on different samples with different STM tips. The STM/replica technique avoids many of the previous artifacts of biological STM imaging and can be used to examine a variety of periodic hydrated lipid and protein samples at a lateral resolution of about 1 nm and a vertical resolution of about 0.3 nm. This resolution is superior to conventional and tapping mode AFM of soft biological materials; the technique is substrate-free, and the conductive and chemically uniform replicas make image interpretation simple and direct. ImagesFIGURE 1FIGURE 2FIGURE 3FIGURE 5 PMID:9017222

  13. One- and two-dimensional infrared spectroscopic studies of solution-phase homogeneous catalysis and spin-forbidden reactions

    SciTech Connect

    Sawyer, Karma Rae

    2008-12-01

    Understanding chemical reactions requires the knowledge of the elementary steps of breaking and making bonds, and often a variety of experimental techniques are needed to achieve this goal. The initial steps occur on the femto- through picosecond time-scales, requiring the use of ultrafast spectroscopic methods, while the rate-limiting steps often occur more slowly, requiring alternative techniques. Ultrafast one and two-dimensional infrared and step-scan FTIR spectroscopies are used to investigate the photochemical reactions of four organometallic complexes. The analysis leads to a detailed understanding of mechanisms that are general in nature and may be applicable to a variety of reactions.

  14. ViriChip: a solid phase assay for detection and identification of viruses by atomic force microscopy

    NASA Astrophysics Data System (ADS)

    Nettikadan, Saju R.; Johnson, James C.; Vengasandra, Srikanth G.; Muys, James; Henderson, Eric

    2004-03-01

    Bionanotechnology can be viewed as the integration of tools and concepts in nanotechnology with the attributes of biomolecules. We report here on an atomic force microscopy-immunosensor assay (AFMIA) that couples AFM with solid phase affinity capture of biological entities for the rapid detection and identification of group B coxsackievirus particles. Virus identification is based on type-specific immunocapture and the morphological properties of the captured viruses as obtained by the AFM. Representatives of the six group B coxsackieviruses have been specifically captured from 1 µl volumes of clarified cell lysates, body fluids and environmental samples. Concentration and kinetic profiles for capture indicate that detection is possible at 103 TCID50 µl-1 and the dynamic range of the assay spans three logs. The results demonstrate that the melding of a nanotechnological tool (AFM) with biotechnology (solid phase immunocapture of virus particles) can create a clinically relevant platform, useful for the detection and identification of enterovirus particles in a variety of samples.

  15. GPU-based rapid reconstruction of cellular 3D refractive index maps from tomographic phase microscopy (Conference Presentation)

    NASA Astrophysics Data System (ADS)

    Dardikman, Gili; Shaked, Natan T.

    2016-03-01

    We present highly parallel and efficient algorithms for real-time reconstruction of the quantitative three-dimensional (3-D) refractive-index maps of biological cells without labeling, as obtained from the interferometric projections acquired by tomographic phase microscopy (TPM). The new algorithms are implemented on the graphic processing unit (GPU) of the computer using CUDA programming environment. The reconstruction process includes two main parts. First, we used parallel complex wave-front reconstruction of the TPM-based interferometric projections acquired at various angles. The complex wave front reconstructions are done on the GPU in parallel, while minimizing the calculation time of the Fourier transforms and phase unwrapping needed. Next, we implemented on the GPU in parallel the 3-D refractive index map retrieval using the TPM filtered-back projection algorithm. The incorporation of algorithms that are inherently parallel with a programming environment such as Nvidia's CUDA makes it possible to obtain real-time processing rate, and enables high-throughput platform for label-free, 3-D cell visualization and diagnosis.

  16. A hybrid analog-digital phase-locked loop for frequency mode non-contact scanning probe microscopy.

    PubMed

    Mehta, M M; Chandrasekhar, V

    2014-01-01

    Non-contact scanning probe microscopy (SPM) has developed into a powerful technique to image many different properties of samples. The conventional method involves monitoring the amplitude, phase, or frequency of a cantilever oscillating at or near its resonant frequency as it is scanned across the surface of a sample. For high Q factor cantilevers, monitoring the resonant frequency is the preferred method in order to obtain reasonable scan times. This can be done by using a phase-locked-loop (PLL). PLLs can be obtained as commercial integrated circuits, but these do not have the frequency resolution required for SPM. To increase the resolution, all-digital PLLs requiring sophisticated digital signal processors or field programmable gate arrays have also been implemented. We describe here a hybrid analog/digital PLL where most of the components are implemented using discrete analog integrated circuits, but the frequency resolution is provided by a direct digital synthesis chip controlled by a simple peripheral interface controller (PIC) microcontroller. The PLL has excellent frequency resolution and noise, and can be controlled and read by a computer via a universal serial bus connection.

  17. Construction of the Magnetic Phase Diagram of FeMn/Ni/Cu(001) Using Photoemission Electron Microscopy

    SciTech Connect

    Wu, J.; Scholl, A.; Arenholz, E.; Hwang, C.; Qiu, Z. Q.

    2011-01-04

    Single crystalline FeMn/Ni bilayer was epitaxially grown on Cu(001) substrate and investigated by photoemission electron microscopy (PEEM). The FeMn and Ni films were grown into two cross wedges to facilitate an independent control of the FeMn (0-20 ML) and Ni (0-20 ML) film thicknesses. The Ni magnetic phases were determined by Ni domain images as a function of the Ni thickness (d{sub Ni}) and the FeMn thickness (d{sub FeMn}). The result shows that as the Ni thickness increases, the Ni film undergoes a paramagnetic-to-ferromagnetic state transition at a critical thickness of d{sub FM} and an in-plane to out-of-plane spin reorientation transition at a thicker thickness d{sub SRT}. The phase diagram shows that both d{sub FM} and d{sub SRT} increase as the FeMn film establishes its antiferromagnetic order.

  18. Optofluidic time-stretch quantitative phase microscopy for high-throughput label-free single-cell analysis (Conference Presentation)

    NASA Astrophysics Data System (ADS)

    Guo, Baoshan; Lei, Cheng; Ito, Takuro; Jiang, Yiyue; Ozeki, Yasuyuki; Goda, Keisuke

    2017-02-01

    The ability to sift through a large heterogeneous population of cells is of paramount importance in a diverse range of biomedical and green applications. Furthermore, the capability of identifying various features of cells in a label-free manner is useful for high-throughput screening. Here we present optofluidic time-stretch quantitative phase microscopy for high-throughput label-free single-cell screening. This method is based on an integration of a hydrodynamic-focusing microfluidic chip, an optical time-stretch microscope for high-speed imaging with a spatial resolution of 800 nm at a frame rate of 10 million frames per second, and a digital image processor for image-based characterization, classification, and statistical analysis of biological cells such as blood cells and microalgae. It provides both the opacity (amplitude) and thickness (phase) content of every cell at a high throughput of 10,000 cells per second. This method is expected to be effective for a diverse range of applications such as cancer detection and biofuel production.

  19. Quantitative Phase Microscopy of microstructures with extended measurement range and correction of chromatic aberrations by multiwavelength digital holography.

    PubMed

    Ferraro, P; Miccio, L; Grilli, S; Paturzo, M; De Nicola, S; Finizio, A; Osellame, R; Laporta, P

    2007-10-29

    Quantitative Phase Microscopy (QPM) by interferometric techniques can require a multiwavelength configuration to remove 2pi ambiguity and improve accuracy. However, severe chromatic aberration can affect the resulting phase-contrast map. By means of classical interference microscope configuration it is quite unpractical to correct such aberration. We propose and demonstrate that by Digital Holography (DH) in a microscope configuration it is possible to clear out the QPM map from the chromatic aberration in a simpler and more effective way with respect to other approaches. The proposed method takes benefit of the unique feature of DH to record in a plane out-of-focus and subsequently reconstruct numerically at the right focal image plane. In fact, the main effect of the chromatic aberration is to shift differently the correct focal image plane at each wavelength and this can be readily compensated by adjusting the corresponding reconstruction distance for each wavelength. A procedure is described in order to determine easily the relative focal shift among different imaging wavelengths by performing a scanning of the numerical reconstruction along the optical axis, to find out the focus and to remove at the same time the chromatic aberration.

  20. Determination of electrostatic force and its characteristics based on phase difference by amplitude modulation atomic force microscopy

    NASA Astrophysics Data System (ADS)

    Wang, Kesheng; Cheng, Jia; Yao, Shiji; Lu, Yijia; Ji, Linhong; Xu, Dengfeng

    2016-12-01

    Electrostatic force measurement at the micro/nano scale is of great significance in science and engineering. In this paper, a reasonable way of applying voltage is put forward by taking an electrostatic chuck in a real integrated circuit manufacturing process as a sample, applying voltage in the probe and the sample electrode, respectively, and comparing the measurement effect of the probe oscillation phase difference by amplitude modulation atomic force microscopy. Based on the phase difference obtained from the experiment, the quantitative dependence of the absolute magnitude of the electrostatic force on the tip-sample distance and applied voltage is established by means of theoretical analysis and numerical simulation. The results show that the varying characteristics of the electrostatic force with the distance and voltage at the micro/nano scale are similar to those at the macroscopic scale. Electrostatic force gradually decays with increasing distance. Electrostatic force is basically proportional to the square of applied voltage. Meanwhile, the applicable conditions of the above laws are discussed. In addition, a comparison of the results in this paper with the results of the energy dissipation method shows the two are consistent in general. The error decreases with increasing distance, and the effect of voltage on the error is small.