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Sample records for video-rate confocal scanning

  1. A video rate laser scanning confocal microscope

    Science.gov (United States)

    Ma, Hongzhou; Jiang, James; Ren, Hongwu; Cable, Alex E.

    2008-02-01

    A video-rate laser scanning microscope was developed as an imaging engine to integrate with other photonic building blocks to fulfill various microscopic imaging applications. The system is quipped with diode laser source, resonant scanner, galvo scanner, control electronic and computer loaded with data acquisition boards and imaging software. Based on an open frame design, the system can be combined with varies optics to perform the functions of fluorescence confocal microscopy, multi-photon microscopy and backscattering confocal microscopy. Mounted to the camera port, it allows a traditional microscope to obtain confocal images at video rate. In this paper, we will describe the design principle and demonstrate examples of applications.

  2. A Video Rate Confocal Laser Beam Scanning Light Microscope Using An Image Dissector

    Science.gov (United States)

    Goldstein, Seth R.; Hubin, Thomas; Rosenthal, Scott; Washburn, Clayton

    1989-12-01

    A video rate confocal reflected light microscope with no moving parts has been developed. Return light from an acousto-optically raster scanned laser beam is imaged from the microscope stage onto the photocathode of an Image Dissector Tube (IDT). Confocal operation is achieved by appropriately raster scanning with the IDT x and y deflection coils so as to continuously "sample" that portion of the photocathode that is being instantaneously illuminated by the return image of the scanning laser spot. Optimum IDT scan parameters and geometric distortion correction parameters are determined under computer control within seconds and are then continuously applied to insure system alignment. The system is operational and reflected light images from a variety of objects have been obtained. The operating principle can be extended to fluorescence and transmission microscopy.

  3. Video-rate in vivo fluorescence imaging with a line-scanned dual-axis confocal microscope

    Science.gov (United States)

    Chen, Ye; Wang, Danni; Khan, Altaz; Wang, Yu; Borwege, Sabine; Sanai, Nader; Liu, Jonathan T. C.

    2015-10-01

    Video-rate optical-sectioning microscopy of living organisms would allow for the investigation of dynamic biological processes and would also reduce motion artifacts, especially for in vivo imaging applications. Previous feasibility studies, with a slow stage-scanned line-scanned dual-axis confocal (LS-DAC) microscope, have demonstrated that LS-DAC microscopy is capable of imaging tissues with subcellular resolution and high contrast at moderate depths of up to several hundred microns. However, the sensitivity and performance of a video-rate LS-DAC imaging system, with low-numerical aperture optics, have yet to be demonstrated. Here, we report on the construction and validation of a video-rate LS-DAC system that possesses sufficient sensitivity to visualize fluorescent contrast agents that are topically applied or systemically delivered in animal and human tissues. We present images of murine oral mucosa that are topically stained with methylene blue, and images of protoporphyrin IX-expressing brain tumor from glioma patients that have been administered 5-aminolevulinic acid prior to surgery. In addition, we demonstrate in vivo fluorescence imaging of red blood cells trafficking within the capillaries of a mouse ear, at frame rates of up to 30 fps. These results can serve as a benchmark for miniature in vivo microscopy devices under development.

  4. Acousto-optically scanned video-rate image dissector tube confocal microscope suitable for use with multiple wavelengths

    Science.gov (United States)

    Hubin, Thomas; Goldstein, Seth R.; Smith, Thomas G., Jr.

    1994-04-01

    We will present the design of a greatly simplified acousto-optic (A/O) scanning system which allows a change of wavelength in less than a second (and in principal between 2 TV lines). A/O deflectors now available with a 9.3 mm circular entrance pupil (rather than the 2 mm X 12 mm pupils previously used) eliminate the need for costly anamorphic beam shaping optics. The resulting simplified optic system can be straightforwardly corrected for different excitation wavelengths.

  5. Video rate near-field scanning optical microscopy

    Science.gov (United States)

    Bukofsky, S. J.; Grober, R. D.

    1997-11-01

    The enhanced transmission efficiency of chemically etched near-field optical fiber probes makes it possible to greatly increase the scanning speed of near-field optical microscopes. This increase in system bandwidth allows sub-diffraction limit imaging of samples at video rates. We demonstrate image acquisition at 10 frames/s, rate-limited by mechanical resonances in our scanner. It is demonstrated that the optical signal to noise ratio is large enough for megahertz single pixel acquisition rates.

  6. Scanning probe microscopes go video rate and beyond

    Science.gov (United States)

    Rost, M. J.; Crama, L.; Schakel, P.; van Tol, E.; van Velzen-Williams, G. B. E. M.; Overgauw, C. F.; ter Horst, H.; Dekker, H.; Okhuijsen, B.; Seynen, M.; Vijftigschild, A.; Han, P.; Katan, A. J.; Schoots, K.; Schumm, R.; van Loo, W.; Oosterkamp, T. H.; Frenken, J. W. M.

    2005-05-01

    In this article we introduce a, video-rate, control system that can be used with any type of scanning probe microscope, and that allows frame rates up to 200images/s. These electronics are capable of measuring in a fast, completely analog mode as well as in the more conventional digital mode. The latter allows measurements at low speeds and options, such as, e.g., atom manipulation, current-voltage spectroscopy, or force-distance curves. For scanning tunneling microscope (STM) application we implemented a hybrid mode between the well-known constant-height and constant-current modes. This hybrid mode not only increases the maximum speed at which the surface can be imaged, but also improves the resolution at lower speeds. Acceptable image quality at high speeds could only be obtained by pushing the performance of each individual part of the electronics to its limit: we developed a preamplifier with a bandwidth of 600kHz, a feedback electronics with a bandwidth of 1MHz, a home-built bus structure for the fast data transfer, fast analog to digital converters, and low-noise drivers. Future improvements and extensions to the control electronics can be realized easily and quickly, because of its open architecture with its modular plug-in units. In the second part of this article we show our high-speed results. The ultrahigh vacuum application of these control electronics on our (UHV)-STM enabled imaging speeds up to 0.3mm/s, while still obtaining atomic step resolution. At high frame rates, the images suffered from noticeable distortions, which we have been able to analyze by virtue of the unique access to the error (dZ) signal. The distortions have all been associated with mechanical resonances in the scan head of the UHV-STM. In order to reduce such resonance effects, we have designed and built a scan head with high resonance frequencies (⩾64kHz), especially for the purpose of testing the fast electronics. Using this scanner we have reached video-rate imaging speeds

  7. Confocal scanning Mueller polarimeter

    Science.gov (United States)

    Lompado, Arthur

    2009-08-01

    We describe the design, construction, calibration and testing of a confocal scanning Mueller polarimeter. A polarization state generator and polarization state analyzer have been inserted into the optical path of a conventional confocal scanning imager to collect the reflectance Muller matrix of samples measuring up to 6.26 mm on a side. Four sources are available for sample interrogation using diode lasers centered at 532 nm, 635 nm, 670 nm, and 785 nm. The device captures all required imagery to calculate the Mueller matrix of each image pixel in approximately 90 s. These matrices are then reduced into polarization imagery such as the diattenuation, retardance and depolarization index. Oftentimes this polarization imagery is quite different and potentially more informative than a conventional intensity image. There are a number of fields that can benefit from alternative/enhanced imagery, most notably in the biomedical, discrimination, and target recognition communities. The sensor has been designed for biomedical applications aimed at improving the technique of noninvasive detection of melanoma lesions.

  8. Confocal scanning microscopy

    DEFF Research Database (Denmark)

    Bariani, Paolo

    This report is based on a metrological investigation on confocal microscopy technique carried out by Uffe Rolf Arlø Theilade and Paolo Bariani. The purpose of the experimental activity was twofold a metrological instrument characterization and application to assessment of rough PP injection moulded...... replicated topography. Confocal microscopy is seen to be a promising technique in metrology of microstructures. Some limitations with respect to surface metrology were found during the experiments. The experiments were carried out using a Zeiss LSM 5 Pascal microscope owned by the Danish Polymer Centre...

  9. Improved depth resolution in video-rate line-scanning multiphoton microscopy using temporal focusing

    Science.gov (United States)

    Tal, Eran; Oron, Dan; Silberberg, Yaron

    2005-07-01

    By introducing spatiotemporal pulse shaping techniques to multiphoton microscopy it is possible to obtain video-rate images with depth resolution similar to point-by-point scanning multiphoton microscopy while mechanically scanning in only one dimension. This is achieved by temporal focusing of the illumination pulse: The pulsed excitation field is compressed as it propagates through the sample, reaching its shortest duration (and highest peak intensity) at the focal plane before stretching again beyond it. This method is applied to produce, in a simple and scalable setup, video-rate two-photon excitation fluorescence images of Drosophila egg chambers with nearly 100,000 effective pixels and 1.5 μm depth resolution.

  10. QUANTITATIVE CONFOCAL LASER SCANNING MICROSCOPY

    Directory of Open Access Journals (Sweden)

    Merete Krog Raarup

    2011-05-01

    Full Text Available This paper discusses recent advances in confocal laser scanning microscopy (CLSM for imaging of 3D structure as well as quantitative characterization of biomolecular interactions and diffusion behaviour by means of one- and two-photon excitation. The use of CLSM for improved stereological length estimation in thick (up to 0.5 mm tissue is proposed. The techniques of FRET (Fluorescence Resonance Energy Transfer, FLIM (Fluorescence Lifetime Imaging Microscopy, FCS (Fluorescence Correlation Spectroscopy and FRAP (Fluorescence Recovery After Photobleaching are introduced and their applicability for quantitative imaging of biomolecular (co-localization and trafficking in live cells described. The advantage of two-photon versus one-photon excitation in relation to these techniques is discussed.

  11. Synchronous-digitization for Video Rate Polarization Modulated Beam Scanning Second Harmonic Generation Microscopy.

    Science.gov (United States)

    Sullivan, Shane Z; DeWalt, Emma L; Schmitt, Paul D; Muir, Ryan M; Simpson, Garth J

    2015-03-09

    Fast beam-scanning non-linear optical microscopy, coupled with fast (8 MHz) polarization modulation and analytical modeling have enabled simultaneous nonlinear optical Stokes ellipsometry (NOSE) and linear Stokes ellipsometry imaging at video rate (15 Hz). NOSE enables recovery of the complex-valued Jones tensor that describes the polarization-dependent observables, in contrast to polarimetry, in which the polarization stated of the exciting beam is recorded. Each data acquisition consists of 30 images (10 for each detector, with three detectors operating in parallel), each of which corresponds to polarization-dependent results. Processing of this image set by linear fitting contracts down each set of 10 images to a set of 5 parameters for each detector in second harmonic generation (SHG) and three parameters for the transmittance of the fundamental laser beam. Using these parameters, it is possible to recover the Jones tensor elements of the sample at video rate. Video rate imaging is enabled by performing synchronous digitization (SD), in which a PCIe digital oscilloscope card is synchronized to the laser (the laser is the master clock.) Fast polarization modulation was achieved by modulating an electro-optic modulator synchronously with the laser and digitizer, with a simple sine-wave at 1/10th the period of the laser, producing a repeating pattern of 10 polarization states. This approach was validated using Z-cut quartz, and NOSE microscopy was performed for micro-crystals of naproxen.

  12. Synchronous-digitization for video rate polarization modulated beam scanning second harmonic generation microscopy

    Science.gov (United States)

    Sullivan, Shane Z.; DeWalt, Emma L.; Schmitt, Paul D.; Muir, Ryan D.; Simpson, Garth J.

    2015-03-01

    Fast beam-scanning non-linear optical microscopy, coupled with fast (8 MHz) polarization modulation and analytical modeling have enabled simultaneous nonlinear optical Stokes ellipsometry (NOSE) and linear Stokes ellipsometry imaging at video rate (15 Hz). NOSE enables recovery of the complex-valued Jones tensor that describes the polarization-dependent observables, in contrast to polarimetry, in which the polarization stated of the exciting beam is recorded. Each data acquisition consists of 30 images (10 for each detector, with three detectors operating in parallel), each of which corresponds to polarization-dependent results. Processing of this image set by linear fitting contracts down each set of 10 images to a set of 5 parameters for each detector in second harmonic generation (SHG) and three parameters for the transmittance of the fundamental laser beam. Using these parameters, it is possible to recover the Jones tensor elements of the sample at video rate. Video rate imaging is enabled by performing synchronous digitization (SD), in which a PCIe digital oscilloscope card is synchronized to the laser (the laser is the master clock.) Fast polarization modulation was achieved by modulating an electro-optic modulator synchronously with the laser and digitizer, with a simple sine-wave at 1/10th the period of the laser, producing a repeating pattern of 10 polarization states. This approach was validated using Z-cut quartz, and NOSE microscopy was performed for micro-crystals of naproxen.

  13. GPU accelerated OCT processing at megahertz axial scan rate and high resolution video rate volumetric rendering

    Science.gov (United States)

    Jian, Yifan; Wong, Kevin; Sarunic, Marinko V.

    2013-03-01

    In this report, we describe how to highly optimize a CUDA based platform to perform real time optical coherence tomography data processing and 3D volumetric rendering using commercially-available cost-effective graphic processing units (GPUs). The maximum complete attainable axial scan processing rate (including memory transfer and rendering frame) was 2.2 megahertz for 16 bits pixel depth and 2048 pixels/A-scan, the maximum 3D volumetric rendering speed is 23 volumes/second (size:1024×256×200). To the best of our knowledge, this is the fastest processing rate reported to date with single-chip GPU and the first implementation of real time video rate volumetric OCT processing and rendering that is capable of matching the ultrahigh-speed OCT acquisition rates.

  14. A confocal laser scanning microscopic study on thermoresponsive ...

    Indian Academy of Sciences (India)

    CdTe QDs composites using a fluorescence confocal laser scanning microscope. These composites have potential applications both in material science and biology. Keywords. Confocal ... of binary colloidal alloys and other soft matter systems.

  15. The FAST module: An add-on unit for driving commercial scanning probe microscopes at video rate and beyond

    Science.gov (United States)

    Esch, Friedrich; Dri, Carlo; Spessot, Alessio; Africh, Cristina; Cautero, Giuseppe; Giuressi, Dario; Sergo, Rudi; Tommasini, Riccardo; Comelli, Giovanni

    2011-05-01

    We present the design and the performance of the FAST (Fast Acquisition of SPM Timeseries) module, an add-on instrument that can drive commercial scanning probe microscopes (SPM) at and beyond video rate image frequencies. In the design of this module, we adopted and integrated several technical solutions previously proposed by different groups in order to overcome the problems encountered when driving SPMs at high scanning frequencies. The fast probe motion control and signal acquisition are implemented in a way that is totally transparent to the existing control electronics, allowing the user to switch immediately and seamlessly to the fast scanning mode when imaging in the conventional slow mode. The unit provides a completely non-invasive, fast scanning upgrade to common SPM instruments that are not specifically designed for high speed scanning. To test its performance, we used this module to drive a commercial scanning tunneling microscope (STM) system in a quasi-constant height mode to frame rates of 100 Hz and above, demonstrating extremely stable and high resolution imaging capabilities. The module is extremely versatile and its application is not limited to STM setups but can, in principle, be generalized to any scanning probe instrument.

  16. Graphics processing unit accelerated optical coherence tomography processing at megahertz axial scan rate and high resolution video rate volumetric rendering.

    Science.gov (United States)

    Jian, Yifan; Wong, Kevin; Sarunic, Marinko V

    2013-02-01

    In this report, we describe how to highly optimize a computer unified device architecture based platform to perform real-time processing of optical coherence tomography interferometric data and three-dimensional (3-D) volumetric rendering using a commercially available, cost-effective, graphics processing unit (GPU). The maximum complete attainable axial scan processing rate, including memory transfer and displaying B-scan frame, was 2.24 MHz for 16 bits pixel depth and 2048 fast Fourier transform size; the maximum 3-D volumetric rendering rate, including B-scan, en face view display, and 3-D rendering, was ~23 volumes/second (volume size: 1024×256×200). To the best of our knowledge, this is the fastest processing rate reported to date with a single-chip GPU and the first implementation of real-time video-rate volumetric optical coherence tomography (OCT) processing and rendering that is capable of matching the acquisition rates of ultrahigh-speed OCT.

  17. Graphics processing unit accelerated optical coherence tomography processing at megahertz axial scan rate and high resolution video rate volumetric rendering

    Science.gov (United States)

    Jian, Yifan; Wong, Kevin; Sarunic, Marinko V.

    2013-02-01

    In this report, we describe how to highly optimize a computer unified device architecture based platform to perform real-time processing of optical coherence tomography interferometric data and three-dimensional (3-D) volumetric rendering using a commercially available, cost-effective, graphics processing unit (GPU). The maximum complete attainable axial scan processing rate, including memory transfer and displaying B-scan frame, was 2.24 MHz for 16 bits pixel depth and 2048 fast Fourier transform size; the maximum 3-D volumetric rendering rate, including B-scan, en face view display, and 3-D rendering, was ˜23 volumes/second (volume size: 1024×256×200). To the best of our knowledge, this is the fastest processing rate reported to date with a single-chip GPU and the first implementation of real-time video-rate volumetric optical coherence tomography (OCT) processing and rendering that is capable of matching the acquisition rates of ultrahigh-speed OCT.

  18. Pupil engineering for a confocal reflectance line-scanning microscope

    Science.gov (United States)

    Patel, Yogesh G.; Rajadhyaksha, Milind; DiMarzio, Charles A.

    2011-03-01

    Confocal reflectance microscopy may enable screening and diagnosis of skin cancers noninvasively and in real-time, as an adjunct to biopsy and pathology. Current confocal point-scanning systems are large, complex, and expensive. A confocal line-scanning microscope, utilizing a of linear array detector can be simpler, smaller, less expensive, and may accelerate the translation of confocal microscopy in clinical and surgical dermatology. A line scanner may be implemented with a divided-pupil, half used for transmission and half for detection, or with a full-pupil using a beamsplitter. The premise is that a confocal line-scanner with either a divided-pupil or a full-pupil will provide high resolution and optical sectioning that would be competitive to that of the standard confocal point-scanner. We have developed a confocal line-scanner that combines both divided-pupil and full-pupil configurations. This combined-pupil prototype is being evaluated to determine the advantages and limitations of each configuration for imaging skin, and comparison of performance to that of commercially available standard confocal point-scanning microscopes. With the combined configuration, experimental evaluation of line spread functions (LSFs), contrast, signal-to-noise ratio, and imaging performance is in progress under identical optical and skin conditions. Experimental comparisons between divided-pupil and full-pupil LSFs will be used to determine imaging performance. Both results will be compared to theoretical calculations using our previously reported Fourier analysis model and to the confocal point spread function (PSF). These results may lead to a simpler class of confocal reflectance scanning microscopes for clinical and surgical dermatology.

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

    Science.gov (United States)

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

    2017-03-01

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

  20. [Current application of confocal laser scanning microscope (CLSM) in stomatology].

    Science.gov (United States)

    Zhang, Yu-sen; Li, Ning-yi

    2007-04-01

    Confocal laser scanning microscopy is one kind of modern Hi-tech on the basis of confocal imaging which is characterized by depth discrimination capability. It has been widely used in the field of stomatology due to its great advantages of non-destructive and non-invasive optical sectioning and three-dimensional reconstruction of the vital objects, in situ and dynamic real-time observation of the tissues and cells can be performed at high resolution. This paper reviews the fundamentals of confocal imaging and the application of CLSM in the fields of dental material, caries, dentin bonding interface and other basic researches in stomatology in recent years.

  1. [Advances of in vivo confocal scanning laser microscopy].

    Science.gov (United States)

    Tian, Ke-bin; Zhou, Guo-yu

    2006-02-01

    In vivo confocal scanning laser microscopy is being widely established as a time-saving, non-invasive, investigative methods in the study of body surfaces. Skin can be observed in its native state in vivo without the fixing, sectioning and staining that is necessary for routine histology. It is a new technology that can provide detailed images of tissue architecture and cellular morphology of living tissue. This paper reviews the fundamentals of in vivo confocal imaging and its clinical applications.

  2. A New Multichannel Spectral Imaging Laser Scanning Confocal Microscope

    Directory of Open Access Journals (Sweden)

    Yunhai Zhang

    2013-01-01

    Full Text Available We have developed a new multichannel spectral imaging laser scanning confocal microscope for effective detection of multiple fluorescent labeling in the research of biological tissues. In this paper, the design and key technologies of the system are introduced. Representative results on confocal imaging, 3-dimensional sectioning imaging, and spectral imaging are demonstrated. The results indicated that the system is applicable to multiple fluorescent labeling in biological experiments.

  3. Optomechatronics Design and Control for Confocal Laser Scanning Microscopy

    NARCIS (Netherlands)

    Yoo, H.W.

    2015-01-01

    Confocal laser scanning microscopy (CLSM) is considered as one of the major advancements in microscopy in the last century and is widely accepted as a 3D fluorescence imaging tool for biological studies. For the emerging biological questions CLSM requires fast imaging to detect rapid biological

  4. Modular Scanning Confocal Microscope with Digital Image Processing.

    Science.gov (United States)

    Ye, Xianjun; McCluskey, Matthew D

    2016-01-01

    In conventional confocal microscopy, a physical pinhole is placed at the image plane prior to the detector to limit the observation volume. In this work, we present a modular design of a scanning confocal microscope which uses a CCD camera to replace the physical pinhole for materials science applications. Experimental scans were performed on a microscope resolution target, a semiconductor chip carrier, and a piece of etched silicon wafer. The data collected by the CCD were processed to yield images of the specimen. By selecting effective pixels in the recorded CCD images, a virtual pinhole is created. By analyzing the image moments of the imaging data, a lateral resolution enhancement is achieved by using a 20 × / NA = 0.4 microscope objective at 532 nm laser wavelength.

  5. Line-scanning confocal microendoscope for nuclear morphometry imaging

    Science.gov (United States)

    Tang, Yubo; Carns, Jennifer; Richards-Kortum, Rebecca R.

    2017-11-01

    Fiber-optic endomicroscopy is a minimally invasive method to image cellular morphology in vivo. Using a coherent fiber bundle as an image relay, it allows additional imaging optics to be placed at the distal end of the fiber outside the body. In this research, we use this approach to demonstrate a compact, low-cost line-scanning confocal fluorescence microendoscope that can be constructed for pathological conditions.

  6. Quantitative single-molecule imaging by confocal laser scanning microscopy.

    Science.gov (United States)

    Vukojevic, Vladana; Heidkamp, Marcus; Ming, Yu; Johansson, Björn; Terenius, Lars; Rigler, Rudolf

    2008-11-25

    A new approach to quantitative single-molecule imaging by confocal laser scanning microscopy (CLSM) is presented. It relies on fluorescence intensity distribution to analyze the molecular occurrence statistics captured by digital imaging and enables direct determination of the number of fluorescent molecules and their diffusion rates without resorting to temporal or spatial autocorrelation analyses. Digital images of fluorescent molecules were recorded by using fast scanning and avalanche photodiode detectors. In this way the signal-to-background ratio was significantly improved, enabling direct quantitative imaging by CLSM. The potential of the proposed approach is demonstrated by using standard solutions of fluorescent dyes, fluorescently labeled DNA molecules, quantum dots, and the Enhanced Green Fluorescent Protein in solution and in live cells. The method was verified by using fluorescence correlation spectroscopy. The relevance for biological applications, in particular, for live cell imaging, is discussed.

  7. Reflection across plant cell boundaries in confocal laser scanning microscopy.

    Science.gov (United States)

    Liu, D Y T; Kuhlmey, B T; Smith, P M C; Day, D A; Faulkner, C R; Overall, R L

    2008-08-01

    The fluorescence patterns of proteins tagged with the green fluorescent protein (GFP) and its derivatives are routinely used in conjunction with confocal laser scanning microscopy to identify their sub-cellular localization in plant cells. GFP-tagged proteins localized to plasmodesmata, the intercellular junctions of plants, are often identified by single or paired punctate labelling across the cell wall. The observation of paired puncta, or 'doublets', across cell boundaries in tissues that have been transformed through biolistic bombardment is unexpected if there is no intercellular movement of the GFP-tagged protein, since bombardment usually leads to the transformation of single, isolated cells. We expressed a putative plasmodesmal protein tagged with GFP by bombarding Allium porrum epidermal cells and assessed the nature of the doublets observed at the cell boundaries. Doublets were formed when fluorescent spots were abutting a cell boundary and were only observable at certain focal planes. Fluorescence emitted from the half of a doublet lying outside the transformed cells was polarized. Optical simulations performed using finite-difference time-domain computations showed a dramatic distortion of the confocal microscope's point spread function when imaging voxels close to the plant cell wall due to refractive index differences between the wall and the cytosol. Consequently, axially and radially out-of-focus light could be detected. A model of this phenomenon suggests how a doublet may form when imaging only a single real fluorescent body in the vicinity of a plant cell wall using confocal microscopy. We suggest, therefore, that the appearance of doublets across cell boundaries is insufficient evidence for plasmodesmal localization due to the effects of the cell wall on the reflection and scattering of light.

  8. Confocal laser scanning microscopy-guided surgery for neurofibroma.

    Science.gov (United States)

    Koller, S; Horn, M; Weger, W; Massone, C; Smolle, J; Gerger, A

    2009-12-01

    The neurofibromatoses comprise at least two separate genetic disorders with variable clinical features and an unpredictable course. The most common type, neurofibromatosis 1, is characterized by > or = 6 café-au-lait spots and the occurrence of neurofibromas, which may present as cutaneous, subcutaneous or plexiform lesions. Normally, excision of neurofibromas is only indicated in the presence of neurological symptoms, suspicion of malignancy or for exceptional cosmetic reasons. For a good functional and aesthetic result with the least danger of recurrence, the surgeon's goal is to excise as much tissue as necessary and as little tissue as possible. One of the main issues during the surgical procedure is to distinguish between neurofibroma and surrounding tissue. We report for the first time the use of confocal laser scanning microscopy to differentiate between neurofibroma and healthy skin.

  9. Integrated Confocal and Scanning Probe Microscopy for Biomedical Research

    Directory of Open Access Journals (Sweden)

    B.J. Haupt

    2006-01-01

    Full Text Available Atomic force microscopy (AFM continues to be developed, not only in design, but also in application. The new focus of using AFM is changing from pure material to biomedical studies. More frequently, it is being used in combination with other optical imaging methods, such as confocal laser scanning microscopy (CLSM and fluorescent imaging, to provide a more comprehensive understanding of biological systems. To date, AFM has been used increasingly as a precise micromanipulator, probing and altering the mechanobiological characteristics of living cells and tissues, in order to examine specific, receptor-ligand interactions, material properties, and cell behavior. In this review, we discuss the development of this new hybrid AFM, current research, and potential applications in diagnosis and the detection of disease.

  10. 3-D Reconstruction of Neurons from Multichannel Confocal Laser Scanning Image Series

    NARCIS (Netherlands)

    Wouterlood, F.G.

    2014-01-01

    A confocal laser scanning microscope (CLSM) collects information from a thin, focal plane and ignores out-of-focus information. Scanning of a specimen, with stepwise axial (Z-) movement of the stage in between each scan, produces Z-series of confocal images of a tissue volume, which then can be used

  11. Confocal laser scanning microscopy in study of bone calcification

    Science.gov (United States)

    Nishikawa, Tetsunari; Kokubu, Mayu; Kato, Hirohito; Imai, Koichi; Tanaka, Akio

    2012-12-01

    Bone regeneration in mandible and maxillae after extraction of teeth or tumor resection and the use of rough surface implants in bone induction must be investigated to elucidate the mechanism of calcification. The calcified tissues are subjected to chemical decalcification or physical grinding to observe their microscopic features with light microscopy and transmission electron microscopy where the microscopic tissue morphology is significantly altered. We investigated the usefulness of confocal laser scanning microscopy (CLSM) for this purpose. After staggering the time of administration of calcein and alizarin red to experimental rats and dogs, rat alveolar bone and dog femur grafted with coral as scaffold or dental implants were observed with CLSM. In rat alveolar bone, the calcification of newly-formed bone and net-like canaliculi was observed at the mesial bone from the roots progressed at the rate of 15 μm/day. In dog femur grafted with coral, newly-formed bones along the space of coral were observed in an orderly manner. In dog femur with dental implants, after 8 weeks, newly-formed bone proceeded along the rough surface of the implants. CLSM produced high-magnification images of newly-formed bone and thin sections were not needed.

  12. Confocal laser scanning microscopy in study of bone calcification

    Energy Technology Data Exchange (ETDEWEB)

    Nishikawa, Tetsunari, E-mail: tetsu-n@cc.osaka-dent.ac.jp [Department of Oral Pathology, Osaka Dental University, Osaka (Japan); Kokubu, Mayu; Kato, Hirohito [Department of Oral Pathology, Osaka Dental University, Osaka (Japan); Imai, Koichi [Department of Biomaterials, Osaka Dental University, Osaka (Japan); Tanaka, Akio [Department of Oral Pathology, Osaka Dental University, Osaka (Japan)

    2012-12-01

    Highlights: Black-Right-Pointing-Pointer High-magnification images with depth selection, and thin sections were observed using CLSM. Black-Right-Pointing-Pointer The direction and velocity of calcification of the bone was observed by administration of 2 fluorescent dyes. Black-Right-Pointing-Pointer In dog femora grafted with coral blocks, newly-formed bone was observed in the coral block space with a rough surface. Black-Right-Pointing-Pointer Twelve weeks after dental implant was grafted in dog femora, the space between screws was filled with newly-formed bones. - Abstract: Bone regeneration in mandible and maxillae after extraction of teeth or tumor resection and the use of rough surface implants in bone induction must be investigated to elucidate the mechanism of calcification. The calcified tissues are subjected to chemical decalcification or physical grinding to observe their microscopic features with light microscopy and transmission electron microscopy where the microscopic tissue morphology is significantly altered. We investigated the usefulness of confocal laser scanning microscopy (CLSM) for this purpose. After staggering the time of administration of calcein and alizarin red to experimental rats and dogs, rat alveolar bone and dog femur grafted with coral as scaffold or dental implants were observed with CLSM. In rat alveolar bone, the calcification of newly-formed bone and net-like canaliculi was observed at the mesial bone from the roots progressed at the rate of 15 {mu}m/day. In dog femur grafted with coral, newly-formed bones along the space of coral were observed in an orderly manner. In dog femur with dental implants, after 8 weeks, newly-formed bone proceeded along the rough surface of the implants. CLSM produced high-magnification images of newly-formed bone and thin sections were not needed.

  13. MEMS-BASED 3D CONFOCAL SCANNING MICROENDOSCOPE USING MEMS SCANNERS FOR BOTH LATERAL AND AXIAL SCAN.

    Science.gov (United States)

    Liu, Lin; Wang, Erkang; Zhang, Xiaoyang; Liang, Wenxuan; Li, Xingde; Xie, Huikai

    2014-08-15

    A fiber-optic 3D confocal scanning microendoscope employing MEMS scanners for both lateral and axial scan was designed and constructed. The MEMS 3D scan engine achieved a lateral scan range of over ± 26° with a 2D MEMS scanning micromirror and a depth scan of over 400 μm with a 1D MEMS tunable microlens. The lateral resolution and axial resolution of this system were experimentally measured as 1.0 μm and 7.0 μm, respectively. 2D and 3D confocal reflectance images of micro-patterns, micro-particles, onion skins and acute rat brain tissue were obtained by this MEMS-based 3D confocal scanning microendoscope.

  14. Improved axial point spread function in a two-frequency laser scanning confocal fluorescence microscope.

    Science.gov (United States)

    Wu, Jheng-Syong; Chung, Yung-Chin; Chien, Jun-Jei; Chou, Chien

    2018-01-01

    A two-frequency laser scanning confocal fluorescence microscope (TF-LSCFM) based on intensity modulated fluorescence signal detection was proposed. The specimen-induced spherical aberration and scattering effect were suppressed intrinsically, and high image contrast was presented due to heterodyne interference. An improved axial point spread function in a TF-LSCFM compared with a conventional laser scanning confocal fluorescence microscope was demonstrated and discussed. (2018) COPYRIGHT Society of Photo-Optical Instrumentation Engineers (SPIE).

  15. Reproducibility of fundus autofluorescence measurements obtained using a confocal scanning laser ophthalmoscope

    OpenAIRE

    Lois, N.; Halfyard, A.; Bunce, C.; Bird, A.; Fitzke, F.

    1999-01-01

    AIM—To evaluate the reproducibility of the background fundus autofluorescence measurements obtained using a confocal scanning laser ophthalmoscope.
METHODS—10 normal volunteers and 10 patients with retinal disease were included in the study. One eye per subject was chosen randomly. Five images of the same eye of each individual were obtained, after pupillary dilatation, by two investigators using a confocal scanning laser ophthalmoscope. Background fundus autofluorescence was measured at 7 de...

  16. The Enhancement of 3D Scans Depth Resolution Obtained by Confocal Scanning of Porous Materials

    Directory of Open Access Journals (Sweden)

    Martisek Dalibor

    2017-12-01

    Full Text Available The 3D reconstruction of simple structured materials using a confocal microscope is widely used in many different areas including civil engineering. Nonetheless, scans of porous materials such as concrete or cement paste are highly problematic. The well-known problem of these scans is low depth resolution in comparison to the horizontal and vertical resolution. The degradation of the image depth resolution is caused by systematic errors and especially by different random events. Our method is focused on the elimination of such random events, mainly the additive noise. We use an averaging method based on the Lindeberg-Lévy theorem that improves the final depth resolution to a level comparable with horizontal and vertical resolution. Moreover, using the least square method, we also precisely determine the limit value of a depth resolution. Therefore, we can continuously evaluate the difference between current resolution and the optimal one. This substantially simplifies the scanning process because the operator can easily determine the required number of scans.

  17. Development of an add-on kit for scanning confocal microscopy (Conference Presentation)

    Science.gov (United States)

    Guo, Kaikai; Zheng, Guoan

    2017-03-01

    Scanning confocal microscopy is a standard choice for many fluorescence imaging applications in basic biomedical research. It is able to produce optically sectioned images and provide acquisition versatility to address many samples and application demands. However, scanning a focused point across the specimen limits the speed of image acquisition. As a result, scanning confocal microscope only works well with stationary samples. Researchers have performed parallel confocal scanning using digital-micromirror-device (DMD), which was used to project a scanning multi-point pattern across the sample. The DMD based parallel confocal systems increase the imaging speed while maintaining the optical sectioning ability. In this paper, we report the development of an add-on kit for high-speed and low-cost confocal microscopy. By adapting this add-on kit to an existing regular microscope, one can convert it into a confocal microscope without significant hardware modifications. Compared with current DMD-based implementations, the reported approach is able to recover multiple layers along the z axis simultaneously. It may find applications in wafer inspection and 3D metrology of semiconductor circuit. The dissemination of the proposed add-on kit under $1000 budget could also lead to new types of experimental designs for biological research labs, e.g., cytology analysis in cell culture experiments, genetic studies on multicellular organisms, pharmaceutical drug profiling, RNA interference studies, investigation of microbial communities in environmental systems, and etc.

  18. Bright-field scanning confocal electron microscopy using a double aberration-corrected transmission electron microscope

    Energy Technology Data Exchange (ETDEWEB)

    Wang, Peng; Behan, Gavin; Kirkland, Angus I. [Department of Materials, University of Oxford, Parks Road, Oxford OX1 3PH (United Kingdom); Nellist, Peter D., E-mail: peter.nellist@materials.ox.ac.uk [Department of Materials, University of Oxford, Parks Road, Oxford OX1 3PH (United Kingdom); Cosgriff, Eireann C.; D' Alfonso, Adrian J.; Morgan, Andrew J.; Allen, Leslie J. [School of Physics, University of Melbourne, Parkville, Victoria 3010 (Australia); Hashimoto, Ayako [Advanced Nano-characterization Center, National Institute for Materials Science (NIMS), 3-13 Sakura, Tsukuba 305-0003 (Japan); Takeguchi, Masaki [Advanced Nano-characterization Center, National Institute for Materials Science (NIMS), 3-13 Sakura, Tsukuba 305-0003 (Japan); High Voltage Electron Microscopy Station, NIMS, 3-13 Sakura, Tsukuba 305-0003 (Japan); Mitsuishi, Kazutaka [Advanced Nano-characterization Center, National Institute for Materials Science (NIMS), 3-13 Sakura, Tsukuba 305-0003 (Japan); Quantum Dot Research Center, NIMS, 3-13 Sakura, Tsukuba 305-0003 (Japan); Shimojo, Masayuki [High Voltage Electron Microscopy Station, NIMS, 3-13 Sakura, Tsukuba 305-0003 (Japan); Advanced Science Research Laboratory, Saitama Institute of Technology, 1690 Fusaiji, Fukaya 369-0293 (Japan)

    2011-06-15

    Scanning confocal electron microscopy (SCEM) offers a mechanism for three-dimensional imaging of materials, which makes use of the reduced depth of field in an aberration-corrected transmission electron microscope. The simplest configuration of SCEM is the bright-field mode. In this paper we present experimental data and simulations showing the form of bright-field SCEM images. We show that the depth dependence of the three-dimensional image can be explained in terms of two-dimensional images formed in the detector plane. For a crystalline sample, this so-called probe image is shown to be similar to a conventional diffraction pattern. Experimental results and simulations show how the diffracted probes in this image are elongated in thicker crystals and the use of this elongation to estimate sample thickness is explored. -- Research Highlights: {yields} The confocal probe image in a scanning confocal electron microscopy image reveals information about the thickness and height of the crystalline layer. {yields} The form of the contrast in a three-dimensional bright-field scanning confocal electron microscopy image can be explained in terms of the confocal probe image. {yields} Despite the complicated form of the contrast in bright-field scanning confocal electron microscopy, we see that depth information is transferred on a 10 nm scale.

  19. Confocal laser scanning microscopy. Using new technology to answer old questions in forensic investigations.

    Science.gov (United States)

    Turillazzi, Emanuela; Karch, Steven B; Neri, Margherita; Pomara, Cristoforo; Riezzo, Irene; Fineschi, Vittorio

    2008-03-01

    Confocal laser scanning microscopy (CLSM) is a relatively new technique for microscopic imaging. It has found a wide field of application in the general sphere of biological sciences. It has completely changed the study of cells and tissues by allowing greater resolution, optical sectioning of the sample and three-dimensional sanoke reconstruction. Confocal microscopy represents a valid, precious and useful tool capable of providing data (images) of unrivalled clearness and definition. This review discusses the possible applications of confocal microscopy in specific fields of forensic investigation, with specific regard to ballistics, forensic histopathology and toxicological pathology.

  20. Design and Performance of a Multi-Point Scan Confocal Microendoscope

    Directory of Open Access Journals (Sweden)

    Matthew D. Risi

    2014-11-01

    Full Text Available Confocal fluorescence microendoscopy provides high-resolution cellular-level imaging via a minimally invasive procedure, but requires fast scanning to achieve real-time imaging in vivo. Ideal confocal imaging performance is obtained with a point scanning system, but the scan rates required for in vivo biomedical imaging can be difficult to achieve. By scanning a line of illumination in one direction in conjunction with a stationary confocal slit aperture, very high image acquisition speeds can be achieved, but at the cost of a reduction in image quality. Here, the design, implementation, and experimental verification of a custom multi-point aperture modification to a line-scanning multi-spectral confocal microendoscope is presented. This new design improves the axial resolution of a line-scan system while maintaining high imaging rates. In addition, compared to the line-scanning configuration, previously reported simulations predicted that the multi-point aperture geometry greatly reduces the effects of tissue scatter on image quality. Experimental results confirming this prediction are presented.

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

    Science.gov (United States)

    Yang, Yong-fa; Li, Qi

    2014-12-01

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

  2. Superresolution upgrade for confocal spinning disk systems using image scanning microscopy (Conference Presentation)

    Science.gov (United States)

    Isbaner, Sebastian; Hähnel, Dirk; Gregor, Ingo; Enderlein, Jörg

    2017-02-01

    Confocal Spinning Disk Systems are widely used for 3D cell imaging because they offer the advantage of optical sectioning at high framerates and are easy to use. However, as in confocal microscopy, the imaging resolution is diffraction limited, which can be theoretically improved by a factor of 2 using the principle of Image Scanning Microscopy (ISM) [1]. ISM with a Confocal Spinning Disk setup (CSDISM) has been shown to improve contrast as well as lateral resolution (FWHM) from 201 +/- 20 nm to 130 +/- 10 nm at 488 nm excitation. A minimum total acquisition time of one second per ISM image makes this method highly suitable for 3D live cell imaging [2]. Here, we present a multicolor implementation of CSDISM for the popular Micro-Manager Open Source Microscopy platform. Since changes in the optical path are not necessary, this will allow any researcher to easily upgrade their standard Confocal Spinning Disk system at remarkable low cost ( 5000 USD) with an ISM superresolution option. [1]. Müller, C.B. and Enderlein, J. Image Scanning Microscopy. Physical Review Letters 104, (2010). [2]. Schulz, O. et al. Resolution doubling in fluorescence microscopy with confocal spinning-disk image scanning microscopy. Proceedings of the National Academy of Sciences of the United States of America 110, 21000-5 (2013).

  3. DTAF: an efficient probe to study cyanobacterial-plant interaction using confocal laser scanning microscopy (CLSM)

    NARCIS (Netherlands)

    Ahmed, M.; Stal, L.J.; Hasnain, S.

    2011-01-01

    A variety of microscopic techniques have been utilized to study cyanobacterial associations with plant roots, but confocal laser scanning microscopy (CLSM) is the least used due to the unavailability of a suitable fluorescent dye. Commonly used lectins have problems with their binding ability with

  4. DTAF: an efficient probe to study cyanobacterial-plant interaction using confocal laser scanning microscopy (CLSM).

    NARCIS (Netherlands)

    Ahmed, M.; Stal, L.J.; Hasnain, S.

    2011-01-01

    A variety of microscopic techniques have been utilized to study cyanobacterial associations with plant roots, but confocal laser scanning microscopy (CLSM) is the least used due to the unavailability of a suitable fluorescent dye. Commonly used lectins have problems with their binding ability with

  5. Imaging inclusion complex formation in starch granules using confocal laser scanning microscopy

    NARCIS (Netherlands)

    Manca, Marianna; Woortman, Albert J. J.; Loos, Katja; Loi, Maria A.

    The tendency of amylose to form inclusion complexes with guest molecules has been an object of wide interest due to its fundamental role in food processing. Here we investigated the features of starch granules from several botanical sources using confocal laser scanning microscopy (CLSM) and

  6. Imaging inflammation in mouse colon using a rapid stage-scanning confocal fluorescence microscope.

    Science.gov (United States)

    Saldua, Meagan A; Olsovsky, Cory A; Callaway, Evelyn S; Chapkin, Robert S; Maitland, Kristen C

    2012-01-01

    Large area confocal microscopy may provide fast, high-resolution image acquisition for evaluation of tissue in pre-clinical studies with reduced tissue processing in comparison to histology. We present a rapid beam and stage-scanning confocal fluorescence microscope to image cellular and tissue features along the length of the entire excised mouse colon. The beam is scanned at 8,333 lines/sec by a polygon scanning mirror while the specimen is scanned in the orthogonal axis by a motorized translation stage with a maximum speed of 7 mm/sec. A single 1 × 60 mm(2) field of view image spanning the length of the mouse colon is acquired in 10 s. Z-projection images generated from axial image stacks allow high resolution imaging of the surface of non-flat specimens. In contrast to the uniform size, shape, and distribution of colon crypts in confocal images of normal colon, confocal images of chronic bowel inflammation exhibit heterogeneous tissue structure with localized severe crypt distortion.

  7. Dynamic experimentation on the confocal laser scanning microscope : application to soft-solid, composite food materials

    NARCIS (Netherlands)

    Plucknett, K.P.; Pomfret, S.J.; Normand, V.; Ferdinando, D.; Veerman, C.; Frith, W.J.; Norton, I.T.

    2001-01-01

    Confocal laser scanning microscopy (CLSM) is used to follow the dynamic structural evolution of several phase-separated mixed biopolymer gel composites. Two protein/polysaccharide mixed gel systems were examined: gelatin/maltodextrin and gelatin/agarose. These materials exhibit 'emulsion-like'

  8. Various confocal scan features of cysts and trophozoites in cases with Acanthamoeba keratitis.

    Science.gov (United States)

    Rezaei Kanavi, Mozhgan; Naghshgar, Nima; Javadi, Mohammad Ali; Sadat Hashemi, Marzieh

    2012-01-01

    To describe the various confocal scan features of cysts and trophozoites in patients with Acanthamoeba keratitis and to specify the associated findings. In a retrospective study of cases between June 2005 and June 2010, we reviewed all the recorded confocal scan images of patients given a high index in regards to clinical suspicion of Acanthamoeba keratitis, in order to specify the various morphometric and morphologic features of Acanthamoeba cysts and trophozoites and to characterize the associated findings in such cases. Confocal scan images of 170 eyes from 170 patients were reviewed. Bilayered, target-shaped, coffee-bean and rod-shaped appearances of the cysts were observed in 100%, 82.9%, 36.4%, and 17.5% of cases, respectively. Single file arrangement of the cysts was noticed in 22 cases. The mean size of the cysts was 18.9 µm (range 10-39.6). In all cases, trophozoites were observed as pear-shaped or irregularly wedge-shaped structures, some surrounded by a brilliant halo and some exhibiting fine pseudopodia-like extensions, with mean size of 30.2 µm (range 19.2-55.6). Keratoneuritis and the anterior stromal honeycomb pattern were seen in 28.2% and 5.9% of cases, respectively. To our knowledge, this is the largest case-series study on confocal scan features of Acanthamoeba cysts and trophozoites in cases with clinical diagnosis of Acanthamoeba keratitis specifying the morphologic and morphometric criteria of this infectious organism and the associated findings.

  9. Masked illumination scheme for a galvanometer scanning high-speed confocal fluorescence microscope.

    Science.gov (United States)

    Kim, Dong Uk; Moon, Sucbei; Song, Hoseong; Kwon, Hyuk-Sang; Kim, Dug Young

    2011-01-01

    High-speed beam scanning and data acquisition in a laser scanning confocal microscope system are normally implemented with a resonant galvanometer scanner and a frame grabber. However, the nonlinear scanning speed of a resonant galvanometer can generate nonuniform photobleaching in a fluorescence sample as well as image distortion near the edges of a galvanometer scanned fluorescence image. Besides, incompatibility of signal format between a frame grabber and a point detector can lead to digitization error during data acquisition. In this article, we introduce a masked illumination scheme which can effectively decrease drawbacks in fluorescence images taken by a laser scanning confocal microscope with a resonant galvanometer and a frame grabber. We have demonstrated that the difference of photobleaching between the center and the edge of a fluorescence image can be reduced from 26 to 5% in our confocal laser scanning microscope with a square illumination mask. Another advantage of our masked illumination scheme is that the zero level or the lowest input level of an analog signal in a frame grabber can be accurately set by the dark area of a mask in our masked illumination scheme. We have experimentally demonstrated the advantages of our masked illumination method in detail. Copyright © 2011 Wiley Periodicals, Inc.

  10. Confocal fluorescence microscope with dual-axis architecture and biaxial postobjective scanning.

    Science.gov (United States)

    Wang, Thomas D; Contag, Christopher H; Mandella, Michael J; Chan, Ning Y; Kino, Gordon S

    2004-01-01

    We present a novel confocal microscope that has dual-axis architecture and biaxial postobjective scanning for the collection of fluorescence images from biological specimens. This design uses two low-numerical-aperture lenses to achieve high axial resolution and long working distance, and the scanning mirror located distal to the lenses rotates along the orthogonal axes to produce arc-surface images over a large field of view (FOV). With fiber optic coupling, this microscope can potentially be scaled down to millimeter dimensions via microelectromechanical systems (MEMS) technology. We demonstrate a benchtop prototype with a spatial resolution < or =4.4 microm that collects fluorescence images with a high SNR and a good contrast ratio from specimens expressing GFP. Furthermore, the scanning mechanism produces only small differences in aberrations over the image FOV. These results demonstrate proof of concept of the dual-axis confocal architecture for in vivo molecular and cellular imaging.

  11. Conversion of biocytin labelled cells and structures for the confocal laser-scanning method.

    Science.gov (United States)

    Hilbig, H; Müller, A

    1997-05-23

    The method for converting biocytin preparations of brain sections fills a gap in the application of confocal laser-scanning microscopy. Both neuronal and non-neuronal structures are converted. The background remains free of staining. The protocol can be applied to old and already existing biocytin-(diaminobenzidine)-nickel preparations which are then made accessible to evaluation with the laser-scanning microscope by the substitution of nickel with silver-gold. Sodium thiosulphate is used to remove the unbound silver. The reflection image of the laser-scanning microscopy provides more information than the transmission image.

  12. Microfabricated optical fiber with microlens that produces large field-of-view video-rate optical beam scanning for microendoscopy applications

    Science.gov (United States)

    Seibel, Eric J.; Fauver, Mark; Crossman-Bosworth, Janet L.; Smithwick, Quinn Y. J.; Brown, Chris M.

    2003-07-01

    Our goal is to produce a micro-optical scanner at the tip of an ultrathin flexible endoscope with an overall diameter of 1 mm. Using a small diameter piezoelectric tube actuator, a cantilevered optical fiber can be driven in mechanical resonance to scan a beam of light in a space-filling, spiral scan pattern. By knowing and/or controlling the fiber position and acquiring backscattered intensity with a photodetector, an image is acquired. A microfabrication process of computer-controlled acid etching is used to reduce the mass along the fiber scanner shaft to allow for high scan amplitude and frequency. A microlens (50 degrees full angle), up to video scan rates (>10 KHz), while maintaining a scanner diameter of 1 mm. A comparison can be made to bi-axial mirror scanners being fabricated as a MEMS device (micro-electro-mechanical system). Based on the opto-mechanical performance of these microlensed fiber scanners, flexible catheter scopes are possible for new microendoscopies that combine imaging with laser diagnoses.

  13. Ribbon scanning confocal for high-speed high-resolution volume imaging of brain.

    Directory of Open Access Journals (Sweden)

    Alan M Watson

    Full Text Available Whole-brain imaging is becoming a fundamental means of experimental insight; however, achieving subcellular resolution imagery in a reasonable time window has not been possible. We describe the first application of multicolor ribbon scanning confocal methods to collect high-resolution volume images of chemically cleared brains. We demonstrate that ribbon scanning collects images over ten times faster than conventional high speed confocal systems but with equivalent spectral and spatial resolution. Further, using this technology, we reconstruct large volumes of mouse brain infected with encephalitic alphaviruses and demonstrate that regions of the brain with abundant viral replication were inaccessible to vascular perfusion. This reveals that the destruction or collapse of large regions of brain micro vasculature may contribute to the severe disease caused by Venezuelan equine encephalitis virus. Visualization of this fundamental impact of infection would not be possible without sampling at subcellular resolution within large brain volumes.

  14. Confocal scanning electroluminescence spectro-microscope for multidimensional light-emitting property analysis

    Science.gov (United States)

    Hong, S.; Onushkin, G.; Park, J. S.; Kim, B. K.; Lee, D.-Y.; Fomin, A.; Ko, K.; Kim, J. W.

    2007-02-01

    We report new type of micro-EL instrument and its applications for light emitting devices. Our new micro-EL, so-called confocal scanning electroluminescence sprctro-microscope (CSESM) has not only fast image acquisition time but also high image resolution. The newly developed CSESM is combined with confocal laser scanning photoluminescence micsoscope, i.e. micro-PL. Therefore, micro-EL distribution can be directly matched with micro-PL and mechanical chip structure of LED. It is fruitful for providing a fast and non-destructive method to analyze the homogeneity of LEDs in its completely proceeded form. Using this apparatus, we study local intensity and wavelength distribution of electroluminescence for InGaN/GaN blue LED chip. Our results represent that local fluctuations of electroluminescence intensity and wavelength position are closely connected with the fluctuation of local current density, i.e. current spreading features on LED chips.

  15. Evaluation of Yogurt Microstructure Using Confocal Laser Scanning Microscopy and Image Analysis

    DEFF Research Database (Denmark)

    Skytte, Jacob Lercke; Ghita, Ovidiu; Whelan, Paul F.

    2015-01-01

    The microstructure of protein networks in yogurts defines important physical properties of the yogurt and hereby partly its quality. Imaging this protein network using confocal scanning laser microscopy (CSLM) has shown good results, and CSLM has become a standard measuring technique for fermented...... to image texture description. Here, CSLM images from a yogurt fermentation study are investigated, where production factors including fat content, protein content, heat treatment, and incubation temperature are varied. The descriptors are evaluated through nearest neighbor classification, variance analysis...

  16. Confocal scanning laser evaluation of repeated Q-switched laser exposure and possible retinal NFL damage

    Science.gov (United States)

    Zwick, Harry; Gagliano, Donald A.; Zuclich, Joseph A.; Stuck, Bruce E.; Lund, David J.; Glickman, Randolph D.

    1995-05-01

    Repeated extended source Q-switched exposure centered on the macula has been shown to produce a Bullseye maculopathy. This paper provides a confocal scanning laser ophthalmoscopic evaluation with regard to the retinal nerve fiber layer (NFL) and deeper choroidal vascular network. Confocal imaging revealed that the punctate annular appearance of this lesion in the deeper retinal layers is associated with retinal nerve fiber bundle disruptions and small gaps in the retinal NFL. No choroidal dysfunction was noticed with Indocyanine green angiography. It is hypothesized that retinal NFL damage occurs either through disruption of retinal pigment epithelial cell layer support to the NFL or through direct exposure to high spatial peak powers within the extended source beam profile, causing direct microthermal injury to the NFL. The apparent sparring of the fovea reflects central retinal morphology rather than a lack of retinal damage to the fovea.

  17. Leica solution: CARS microscopy at video rates

    Science.gov (United States)

    Lurquin, V.

    2008-02-01

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

  18. Method and apparatus for a high-resolution three dimensional confocal scanning transmission electron microscope

    Science.gov (United States)

    de Jonge, Niels [Oak Ridge, TN

    2010-08-17

    A confocal scanning transmission electron microscope which includes an electron illumination device providing an incident electron beam propagating in a direction defining a propagation axis, and a precision specimen scanning stage positioned along the propagation axis and movable in at least one direction transverse to the propagation axis. The precision specimen scanning stage is configured for positioning a specimen relative to the incident electron beam. A projector lens receives a transmitted electron beam transmitted through at least part of the specimen and focuses this transmitted beam onto an image plane, where the transmitted beam results from the specimen being illuminated by the incident electron beam. A detection system is placed approximately in the image plane.

  19. The application of dermal papillary rings in dermatology by in vivo confocal laser scanning microscopy

    Science.gov (United States)

    Xiang, W. Z.; Xu, A. E.; Xu, J.; Bi, Z. G.; Shang, Y. B.; Ren, Q. S.

    2010-08-01

    Confocal laser scanning microscopy (CLSM) allows noninvasive visualization of human skin in vivo, without needing to fix or section the tissue. Melanocytes and pigmented keratinocytes at the level of the basal layer form bright dermal papillary rings which are readily amenable to identify in confocal images. Our purpose was to explore the role of dermal papillary rings in assessment of lesion location, the diagnosis, differential diagnosis of lesions and assessment of therapeutic efficacy by in vivo CLSM. Seventy-one patients were imaged with the VivaScope 1500 reflectance confocal microscope provided by Lucid, Inc. The results indicate that dermal papillary rings can assess the location of lesion; the application of dermal papillary rings can provide diagnostic support and differential diagnosis for vitiligo, nevus depigmentosus, tinea versicolor, halo nevus, common nevi, and assess the therapeutic efficacy of NBUVB phototherapy plus topical 0.1 percent tacrolimus ointment for vitiligo. In conclusion, our findings indicate that the dermal papillary rings play an important role in the assessment the location of lesion, diagnosis, differential diagnosis of lesions and assessment of therapeutic efficacy by in vivo CLSM. CLSM may be a promising tool for noninvasive examination in dermatology. However, larger studies are needed to expand the application of dermal papillary rings in dermatology.

  20. Confocal line scanning of a Bessel beam for fast 3D imaging.

    Science.gov (United States)

    Zhang, P; Phipps, M E; Goodwin, P M; Werner, J H

    2014-06-15

    We have developed a light-sheet illumination microscope that can perform fast 3D imaging of transparent biological samples with inexpensive visible lasers and a single galvo mirror (GM). The light-sheet is created by raster scanning a Bessel beam with a GM, with this same GM also being used to rescan the fluorescence across a chip of a camera to construct an image in real time. A slit is used to reject out-of-focus fluorescence such that the image formed in real time has minimal contribution from the sidelobes of the Bessel beam. Compared with two-photon Bessel beam excitation or other confocal line-scanning approaches, our method is of lower cost, is simpler, and does not require calibration and synchronization of multiple GMs. We demonstrated the optical sectioning and out-of-focus background rejection capabilities of this microscope by imaging fluorescently labeled actin filaments in fixed 3T3 cells.

  1. QUANTIFICATION OF BIOFILMS IN MULTI-SPECTRAL DIGITAL1 VOLUMES FROM CONFOCAL LASER-SCANNING MICROSCOPES

    Directory of Open Access Journals (Sweden)

    Karsten Rodenacker

    2011-05-01

    Full Text Available Populations of bacteria in sludge flocs and biofilm marked by fluorescence marked with fluorescent probes are digitised with a confocal laser scanning microscope. These data are used to analyse the microbial community structure, to obtain information on the localisation of specific bacterial groups and to examine gene expression. This information is urgently required for an in-depth understanding of the function and, more generally, the microbial ecology of biofilms. Methods derived from quantitative image analysis are applied to digitised data from confocal laser scanning microscopes to obtain quantitative descriptions of volumetric, topological (and topographical properties of different compartments of the components under research. In addition to free-moving flocs, also biofilms attached to a substratum in an experimental environment are analysed. Growth form as well as interaction of components are quantitatively described. Classical measurements of volume and intensity (shape, distribution and distance dependent interaction measurements using methods from mathematical morphology are performed. Mainly image (volume processing methods are outlined. Segmented volumes are globally and individually (in terms of 3Dconnected components measured and used for distance mapping transform as well as for estimation of geodesic distances from the substrate. All transformations are applied on the 3D data set. Resulting distance distributions are quantified and related to information on the identity and activity of the probe-identified bacteria.

  2. Confocal laser scanning microscopic investigation of ultrasonic, sonic, and rotary sealer placement techniques

    Science.gov (United States)

    Nikhil, Vineeta; Singh, Renuka

    2013-01-01

    Background: Sealers are used to attain an impervious seal between the core material and root canal walls. Aim: To compare the depth and percentage of sealer penetration with three different placement techniques using confocal laser scanning microscopy as the evaluative tool. Materials and Methods: Root canals of 30 single-rooted teeth were prepared to a size of F3 and AH plus sealer with Rhodamine B was applied with Ultlrasonic file (Gr-1), lentulospiral (Gr-2), and Endoactivator (Gr-3). Canals were obturated with gutta-percha. The roots were sectioned at the 3 and 6-mm levels from the apical foramen and were examined on a confocal microscope. Results: A statistical significant differences among Gr-1, Gr-2, and Gr-3 were found at the 3 and 6-mm level (P < 0.05; ANOVA-Tukey tests) for the depth and percentage of sealer penetration except for Gr-1 and Gr-2 at 3-mm level. Gr-1 showed maximum mean depth of penetration (810 μm) and maximum mean percentage of sealer penetration (64.5) while Gr-3 showed minimum mean depth of penetration (112.7 μm) and minimum mean percentage of sealer penetration (26.7). Conclusion: Depth and percentage of penetration of sealer is influenced by the type of placement technique and by the root canal level with penetration decreasing apically. PMID:23956528

  3. Scanning a microhabitat: plant-microbe interactions revealed by confocal laser microscopy.

    Science.gov (United States)

    Cardinale, Massimiliano

    2014-01-01

    No plant or cryptogam exists in nature without microorganisms associated with its tissues. Plants as microbial hosts are puzzles of different microhabitats, each of them colonized by specifically adapted microbiomes. The interactions with such microorganisms have drastic effects on the host fitness. Since the last 20 years, the combination of microscopic tools and molecular approaches contributed to new insights into microbe-host interactions. Particularly, confocal laser scanning microscopy (CLSM) facilitated the exploration of microbial habitats and allowed the observation of host-associated microorganisms in situ with an unprecedented accuracy. Here I present an overview of the progresses made in the study of the interactions between microorganisms and plants or plant-like organisms, focusing on the role of CLSM for the understanding of their significance. I critically discuss risks of misinterpretation when procedures of CLSM are not properly optimized. I also review approaches for quantitative and statistical analyses of CLSM images, the combination with other molecular and microscopic methods, and suggest the re-evaluation of natural autofluorescence. In this review, technical aspects were coupled with scientific outcomes, to facilitate the readers in identifying possible CLSM applications in their research or to expand their existing potential. The scope of this review is to highlight the importance of confocal microscopy in the study of plant-microbe interactions and also to be an inspiration for integrating microscopy with molecular techniques in future researches of microbial ecology.

  4. Scanning a microhabitat: plant-microbe interactions revealed by confocal laser microscopy

    Directory of Open Access Journals (Sweden)

    Massimiliano eCardinale

    2014-03-01

    Full Text Available No plant or cryptogam exists in nature without microorganisms associated with its tissues. Plants as microbial hosts are puzzles of different microhabitats, each of them colonized by specifically adapted microbiomes. The interactions with such microorganisms have drastic effects on the host fitness. Since the last 20 years, the combination of microscopic tools and molecular approaches contributed to new insights into microbe-host interactions. Particularly, confocal laser scanning microscopy (CLSM facilitated the exploration of microbial habitats and allowed the observation of host-associated microorganisms in situ with an unprecedented accuracy. Here I present an overview of the progresses made in the study of the interactions between microorganisms and plants or plant-like organisms, focusing on the role of CLSM for the understanding of their significance. I critically discuss risks of misinterpretation when procedures of CLSM are not properly optimized. I also review approaches for quantitative and statistical analyses of CLSM images, the combination with other molecular and microscopic methods, and suggest the re-evaluation of natural autofluorescence. In this review, technical aspects were coupled with scientific outcomes, to facilitate the readers in identifying possible CLSM applications in their research or to expand their existing potential. The scope of this review is to highlight the importance of confocal microscopy in the study of plant-microbe interactions and also to be an inspiration for integrating microscopy with molecular techniques in future researches of microbial ecology.

  5. Three-photon fluorescence imaging of melanin with a dual-wedge confocal scanning system

    Science.gov (United States)

    Mega, Yair; Kerimo, Joseph; Robinson, Joseph; Vakili, Ali; Johnson, Nicolette; DiMarzio, Charles

    2012-03-01

    Confocal microscopy can be used as a practical tool in non-invasive applications in medical diagnostics and evaluation. In particular, it is being used for the early detection of skin cancer to identify pathological cellular components and, potentially, replace conventional biopsies. The detection of melanin and its spatial location and distribution plays a crucial role in the detection and evaluation of skin cancer. Our previous work has shown that the visible emission from melanin is strong and can be easily observed with a near-infrared CW laser using low power. This is due to a unique step-wise, (SW) three-photon excitation of melanin. This paper shows that the same SW, 3-photon fluorescence can also be achieved with an inexpensive, continuous-wave laser using a dual-prism scanning system. This demonstrates that the technology could be integrated into a portable confocal microscope for clinical applications. The results presented here are in agreement with images obtained with the larger and more expensive femtosecond laser system used earlier.

  6. Imaging of whole tumor cut sections using a novel scanning beam confocal fluorescence MACROscope

    Science.gov (United States)

    Constantinou, Paul; Vukovic, Vojislav; Haugland, Hans K.; Nicklee, Trudey; Hedley, David W.; Wilson, Brian C.

    2001-07-01

    Hypoxia caused by inadequate structure and function of the tumor vasculature has been found to negatively determine the prognosis of cancer patients. Hence, understanding the biological basis of tumor hypoxia is of significant clinical interest. To study solid tumor microenvironments in sufficient detail, large areas (several mm in diameter) need to be imaged at micrometers resolutions. We have used a novel confocal scanning laser MACROscopeTM (CSLM) capable of acquiring images over fields of view up to 2 cm X 2 cm. To demonstrate its performance, frozen sections from a cervical carcinoma xenograft were triple labeled for tissue hypoxia, blood vessels and hypoxia-inducible transcription factor 1 alpha (HIF-1(alpha) ), imaged using the CSLM and compared to images obtained using a standard epifluorescence microscope imaging system. The results indicate that the CSLM is a useful instrument for imaging tissue-based fluorescence at resolutions comparable to standard low-power microscope objectives.

  7. Fluorescence confocal laser scanning microscopy for in vivo imaging of epidermal reactions to two experimental irritants

    DEFF Research Database (Denmark)

    Suihko, C.; Serup, J.

    2008-01-01

    Background: Fibre-optic fluorescence confocal laser scanning microscopy (CLSM) is a novel non-invasive technique for in vivo imaging of skin. The cellular structure of the epidermis can be studied. A fluorophore, e.g. fluorescein sodium, is introduced by an intradermal injection or applied...... dermatitis reactions caused by established model irritants, e.g. sodium lauryl sulphate (SLS) and pelargonic acid (PA). Methods: Twelve healthy individuals volunteered. The flexor aspect of the right and the left forearm was exposed to SLS in water and PA in isopropanol and occluded under Finn Chambers...... for 24 h. The reactions were rated clinically and, following epicutaneous and intra-dermal application of fluorescein sodium, studied by fluorescence CLSM, magnification x 1000. Results: Both irritants disturbed the epidermal intercellular borders, which became blurred, thickened and variably altered...

  8. Trypan blue as a fluorochrome for confocal laser scanning microscopy of arbuscular mycorrhizae in three mangroves.

    Science.gov (United States)

    Kumar, T; Majumdar, A; Das, P; Sarafis, V; Ghose, M

    2008-06-01

    Roots of three mangroves, Acanthus ilicifolius, Ceriops tagal and Excoecaria agallocha, collected from forests of the Sundarbans of India were stained with trypan blue to observe arbuscular mycorrhizal colonization. Spores of arbuscular mycorrhizal fungi isolated from rhizospheric soil, collected together with the root samples, also were stained for testing the suitability of the dye as a fluorochrome. Confocal laser scanning microscopy images were constructed. A. ilicifolius and E. agallocha exhibited "Arum" type colonization with highly branched arbuscules, whereas C. tagal showed "Paris" type association with clumped and collapsed arbuscules. We demonstrated that trypan blue is a suitable fluorochrome for staining arbuscular mycorrhizal fungal spores, fungal hyphae, arbuscules and vesicles, which presumably have a considerable amount of surface chitin. It appears that as the integration of chitin into the fungal cell wall changes, its accessibility to trypan blue dye also changes.

  9. Real-time line-scanning reflectance confocal endoscope to enhance sectioning and reduce speckle for intraoral imaging

    Science.gov (United States)

    Glazowski, Christopher; Abeytunge, Sanjeewa; Rajadhyaksha, Milind

    2012-02-01

    The line-scanning confocal microscope is simpler than a point-scanning confocal microscope and allows for a smaller and lower cost footprint, making it attractive for endoscopic clinical use. The optical configuration affects image fidelity. Here, we present a benchtop version of an endoscopic line-scanning confocal microscope for intraoral imaging, with a divided pupil and optimal detection configuration (magnification, pixel-to-resolution ratio) to enhance image fidelity. Improved sectioning performance and reduction of "speckle" noise are demonstrated. A topology for use of a deformable MEMs mirror-based optical axial focus control for imaging in depth is presented. Preliminary images of human oral mucosa in vivo demonstrate feasibility for clinical application.

  10. Large area mapping of excised breast tissue by fluorescence confocal strip scanning: a preliminary feasibility study

    Science.gov (United States)

    Larson, Bjorg A.; Abeytunge, Sanjee; Murray, Melissa; Rajadhyaksha, Milind

    2013-03-01

    Lumpectomy, in conjunction with radiation and chemotherapy drugs, together comprise breast-conserving treatment as an alternative to total mastectomy for patients with breast tumors. The tumor is removed in surgery and sent for pathology processing to assess the margins, a process that takes at minimum several hours, and generally days. If the margins are not clear of tumor, the patient must undergo a second surgery to remove residual tumor. This re-excision rate varies by institution, but can be as high as 60%. Currently, no intraoperative microscopic technique is used routinely to examine tumor margins in breast tissue. A new technique for rapidly scanning large areas of tissue has been developed, called confocal strip scanning, which provides high resolution and seamless mosaics over large areas of intact tissue, with nuclear and cellular resolution and optical sectioning of about 2 microns. Up to 3.5 x 3.5 cm2 of tissue is imaged in 13 minutes at current stage speeds. This technique is demonstrated in freshly excised breast tissue, using a mobile confocal microscope stationed in our pathology laboratory. Twenty-five lumpectomy and mastectomy cases were used as a testing ground for reflectance and fluorescence contrast modes, resolution requirements and tissue fixturing configurations. It was concluded that fluorescent imaging provides the needed contrast to distinguish ducts and lobules from surrounding stromal tissue. Therefore the system was configured with 488 nm illumination, with acridine orange fluorescent dye for nuclear contrast, with the aim of building an image library of malignant and benign breast pathologies.

  11. Optical coherence tomography, scanning laser polarimetry and confocal scanning laser ophthalmoscopy in retinal nerve fiber layer measurements of glaucoma patients.

    Science.gov (United States)

    Fanihagh, Farsad; Kremmer, Stephan; Anastassiou, Gerasimos; Schallenberg, Maurice

    2015-01-01

    To determine the correlations and strength of association between different imaging systems in analyzing the retinal nerve fiber layer (RNFL) of glaucoma patients: optical coherence tomography (OCT), scanning laser polarimetry (SLP) and confocal scanning laser ophthalmoscopy (CSLO). 114 eyes of patients with moderate open angle glaucoma underwent spectral domain OCT (Topcon SD-OCT 2000 and Zeiss Cirrus HD-OCT), SLP (GDx VCC and GDx Pro) and CSLO (Heidelberg Retina Tomograph, HRT 3). Correlation coefficients were calculated between the structural parameters yielded by these examinations. The quantitative relationship between the measured RNFL thickness globally and for the four regions (superior, inferior, nasal, temporal) were evaluated with different regression models for all used imaging systems. The strongest correlation of RNFL measurements was found between devices using the same technology like GDx VCC and GDx Pro as well as Topcon OCT and Cirrus OCT. In glaucoma patients, the strongest associations (R²) were found between RNFL measurements of the two optical coherence tomography devices Topcon OCT and Cirrus OCT (R² = 0.513) and between GDx VCC and GDx Pro (R² = 0.451). The results of the OCTs and GDX Pro also had a strong quantitative relationship (Topcon OCT R² = 0.339 and Cirrus OCT R² = 0.347). GDx VCC and the OCTs showed a mild to moderate association (Topcon OCT R² = 0.207 and Cirrus OCT R² = 0.258). The confocal scanning laser ophthalmoscopy (HRT 3) had the lowest association to all other devices (Topcon OCT R² = 0.254, Cirrus OCT R² = 0.158, GDx Pro R² = 0.086 and GDx VCC R² = 0.1). The measurements of the RNFL in glaucoma patients reveal a high correlation of OCT and GDx devices because OCTs can measure all major retinal layers and SLP can detect nerve fibers allowing a comparison between the results of this devices. However, CSLO by means of HRT topography can only measure height values of the retinal surface but it cannot distinguish

  12. Experimental setup for energy-filtered scanning confocal electron microscopy (EFSCEM) in a double aberration-corrected transmission electron microscope

    Energy Technology Data Exchange (ETDEWEB)

    Wang, P; Behan, G; Kirkland, A I; Nellist, P D, E-mail: peng.wang@materials.ox.ac.u [Department of Materials, University of Oxford, Parks Road, Oxford OX1 3PH (United Kingdom)

    2010-07-01

    Scanning confocal electron microscopy (SCEM) is a new imaging mode in electron microscopy. Spherical aberration corrected electron microscope instruments fitted with two aberration correctors can be used in this mode which provides improved depth resolution and selectivity compared to optical sectioning in a conventional scanning transmission geometry. In this article, we consider a confocal optical configuration for SCEM using inelastically scattered electrons. We lay out the necessary steps for achieving this new operational mode in a double aberration-corrected instrument with uncorrected chromatic aberration and present preliminary experimental results in such mode.

  13. An instrumental approach to combining confocal microspectroscopy and 3D scanning probe nanotomography.

    Science.gov (United States)

    Mochalov, Konstantin E; Chistyakov, Anton A; Solovyeva, Daria O; Mezin, Alexey V; Oleinikov, Vladimir A; Vaskan, Ivan S; Molinari, Michael; Agapov, Igor I; Nabiev, Igor; Efimov, Anton E

    2017-11-01

    In the past decade correlative microscopy, which combines the potentials of different types of high-resolution microscopies with a variety of optical microspectroscopy techniques, has been attracting increasing attention in material science and biological research. One of outstanding solutions in this area is the combination of scanning probe microscopy (SPM), which provides data on not only the topography, but also the spatial distribution of a wide range of physical properties (elasticity, conductivity, etc.), with ultramicrotomy, allowing 3D multiparametric examination of materials. The combination of SPM and ultramicrotomy (scanning probe nanotomography) is very appropriate for characterization of soft multicompound nanostructurized materials, such as polymer matrices and microstructures doped with different types of nanoparticles (magnetic nanoparticles, quantum dots, nanotubes, etc.), and biological materials. A serious problem of this technique is a lack of chemical and optical characterization tools, which may be solved by using optical microspectroscopy. Here, we report the development of an instrumental approach to combining confocal microspectroscopy and 3D scanning probe nanotomography in a single apparatus. This approach retains all the advantages of SPM and upright optical microspectroscopy and allows 3D multiparametric characterization using both techniques. As the first test of the system developed, we have performed correlative characterization of the morphology and the magnetic and fluorescent properties of fluorescent magnetic microspheres doped with a fluorescent dye and magnetic nanoparticles. The results of this study can be used to obtain 3D volume images of a specimen for most high-resolution near-field scanning probe microscopies: SNOM, TERS, AFM-IR, etc. This approach will result in development of unique techniques combining the advantages of SPM (nanoscale morphology and a wide range of physical parameters) and high-resolution optical

  14. Preliminary Study of In Vivo Formed Dental Plaque Using Confocal Microscopy and Scanning Electron Microscopy

    Directory of Open Access Journals (Sweden)

    KA. Al-Salihi

    2009-12-01

    Full Text Available Objective: Confocal laser scanning microscopy (CLSM is relatively a new light microscopical imaging technique with a wide range of applications in biological sciences. The primary value of CLSM for the biologist is its ability to provide optical sections from athree-dimensional specimen. The present study was designed to assess the thickness and content of in vivo accumulated dental plaque using CLSM and scanning electron microscopy (SEM.Materials and Methods: Acroflat lower arch splints (acrylic appliance were worn by five participants for three days without any disturbance. The formed plaques were assessed using CLSM combined with vital fluorescence technique and SEM.Results: In this study accumulated dental plaque revealed varied plaque microflora vitality and thickness according to participant’s oral hygiene. The thickness of plaque smears ranged from 40.32 to 140.72 μm and 65.00 to 128.88 μm for live (vital and dead accumulated microorganisms, respectively. Meanwhile, the thickness of plaque on the appliance ranged from 101 μm to 653 μm. CLSM revealed both dead and vital bacteria on the surface of the dental plaque. In addition, SEM revealed layers of various bacterial aggregations in all dental plaques.Conclusion: This study offers a potent non-invasive tool to evaluate and assess the dental plaque biofilm, which is a very important factor in the development of dental caries.

  15. Noninvasive in vivo detection and quantification of Demodex mites by confocal laser scanning microscopy.

    Science.gov (United States)

    Sattler, E C; Maier, T; Hoffmann, V S; Hegyi, J; Ruzicka, T; Berking, C

    2012-11-01

    In many Demodex-associated skin diseases Demodex mites are present in abundance and seem to be at least partially pathogenic. So far all diagnostic approaches such as scraping or standardized superficial skin biopsy are (semi-)invasive and may cause discomfort to the patient. To see whether confocal laser scanning microscopy (CLSM) - a noninvasive method for the visualization of superficial skin layers - is able to detect and quantify D. folliculorum in facial skin of patients with rosacea. Twenty-five patients (34-72 years of age) with facial rosacea and 25 age- and sex-matched normal controls were examined by CLSM. Mosaics of 8 × 8 mm and 5 × 5 mm were created by scanning horizontal layers of lesional skin and quantification of mites per follicle and per area as well as follicles per area was performed. In all patients D. folliculorum could be detected by CLSM and presented as roundish or lengthy cone-shaped structures. CLSM allowed the quantification of Demodex mites and revealed significant differences (P Demodex mites noninvasively in facial skin of patients with rosacea. © 2012 The Authors. BJD © 2012 British Association of Dermatologists.

  16. Plastic-to-Elastic Transition in Aggregated Emulsion Networks, Studied with Atomic Force Microscopy-Confocal Scanning Laser Microscopy Microrheology

    NARCIS (Netherlands)

    Filip, D.; Duits, Michael H.G.; Uricanu, V.I.; Mellema, J.

    2006-01-01

    In this paper, we demonstrate how the simultaneous application of atomic force microscopy (AFM) and confocal scanning laser microscopy (CSLM) can be used to characterize the (local) rheological properties of soft condensed matter at micrometer length scales. Measurement of AFM force curves as a

  17. Analysis of a marine phototrophic biofilm by confocal laser scanning microscopy using the new image quantification software PHLIP

    NARCIS (Netherlands)

    Müller, L.N.; de Brouwer, J.F.C.; Almeida, J.S.; Stal, L.J.; Xavier, J.B.

    2006-01-01

    Background Confocal laser scanning microscopy (CLSM) is the method of choice to study interfacial biofilms and acquires time-resolved three-dimensional data of the biofilm structure. CLSM can be used in a multi-channel modus where the different channels map individual biofilm components. This

  18. Microradiography and confocal laser scanning microscopy applied to enamel lesions formed in vivo with and without fluoride varnish treatment

    NARCIS (Netherlands)

    Ogaard, B; Duschner, H; Ruben, J; Arends, J

    The aim of the present investigation was to combine 2 techniques suitable for lesion characterization: quantitative microradiography (TMR) and confocal laser scanning microscopy (CLSM) on in vivo induced lesions with and without a fluoride varnish (Duraphat(R)) treatment. Orthodontic bands were

  19. Further observations on cerebellar climbing fibers. A study by means of light microscopy, confocal laser scanning microscopy and scanning and transmission electron microscopy.

    Science.gov (United States)

    Castejón, O J; Castejón, H V; Alvarado, M V

    2000-12-01

    The intracortical pathways of climbing fibers were traced in several vertebrate cerebella using light microscopy, confocal laser scanning microscopy, scanning and transmission electron microscopy. They were identified as fine fibers up to 1(micron thick, with a characteristic crossing-over bifurcation pattern. Climbing fiber collaterals were tridimensionally visualized forming thin climbing fiber glomeruli in the granular layer. Confocal laser scanning microscopy revealed three types of collateral processes at the interface between granular and Purkinje cell layers. Scanning electron microscopy showed climbing fiber retrograde collaterals in the molecular layer. Asymmetric synaptic contacts of climbing fibers with Purkinje dendritic spines and stellate neuron dendrites were characterized by transmission electron microscopy. Correlative microscopy allowed us to obtain the basic three-dimensional morphological features of climbing fibers in several vertebrates and to show with more accuracy a higher degree of lateral collateralization of these fibers within the cerebellar cortex. The correlative microscopy approach provides new views in the cerebellar cortex information processing.

  20. Three-dimensional imaging of plant cuticle architecture using confocal scanning laser microscopy.

    Science.gov (United States)

    Buda, Gregory J; Isaacson, Tal; Matas, Antonio J; Paolillo, Dominick J; Rose, Jocelyn K C

    2009-10-01

    Full appreciation of the roles of the plant cuticle in numerous aspects of physiology and development requires a comprehensive understanding of its biosynthesis and deposition; however, much is still not known about cuticle structure, trafficking and assembly. To date, assessment of cuticle organization has been dominated by 2D imaging, using histochemical stains in conjunction with light and fluorescence microscopy. This strategy, while providing valuable information, has limitations because it attempts to describe a complex 3D structure in 2D. An imaging technique that could accurately resolve 3D architecture would provide valuable additions to the growing body of information on cuticle molecular biology and biochemistry. We present a novel application of 3D confocal scanning laser microscopy for visualizing the architecture, deposition patterns and micro-structure of plant cuticles, using the fluorescent stain auramine O. We demonstrate the utility of this technique by contrasting the fruit cuticle of wild-type tomato (Solanum lycopersicum cv. M82) with those of cutin-deficient mutants. We also introduce 3D cuticle modeling based on reconstruction of serial optical sections, and describe its use in identification of several previously unreported features of the tomato fruit cuticle.

  1. Prognostic significance of vascularity in cutaneous melanoma: pilot study using in vivo confocal scanning laser microscopy.

    Science.gov (United States)

    Humphrey, Shannon; Walsh, Noreen M; Delaney, Laura; Propperova, Iva; Langley, Richard G B

    2006-01-01

    Tumor vascularity may be of strong prognostic significance in cutaneous melanoma. We are the first to use a novel, noninvasive, in vivo confocal scanning laser microscope (CSLM) to evaluate vascularity in cutaneous melanoma. Our purpose was to apply a CSLM to assess vascularity in melanoma and to evaluate the prognostic significance of these findings. Patients with a suspicious pigmented lesion were prospectively recruited to undergo CSLM prior to skin biopsy, and those diagnosed with melanoma were included in this study. A blinded observer graded tumor vascularity from still digital CSLM images. The CSLM vascularity grading was correlated to tumor thickness and ulceration as a proxy for clinical prognosis. Sixty-six patients and 67 lesions underwent imaging with CSLM. Eleven patients were diagnosed with melanoma, including six in situ and five invasive melanomas. Prominent vascularity was observed in all advanced melanomas. There was an overall increase in mean tumor thickness between the absent (x = 0.315 mm) to prominent (x = 1.51 mm) categories. In this pilot study, vascularity was readily detected in cutaneous melanomas using CSLM. Prominent vascularity was observed in patients with advanced cutaneous melanomas. Our preliminary results are encouraging and indicate potential for the use of CSLM to assess vascularity in cutaneous melanoma, with potential prognostic and therapeutic implications.

  2. High-speed adaptive optics line scan confocal retinal imaging for human eye.

    Science.gov (United States)

    Lu, Jing; Gu, Boyu; Wang, Xiaolin; Zhang, Yuhua

    2017-01-01

    Continuous and rapid eye movement causes significant intraframe distortion in adaptive optics high resolution retinal imaging. To minimize this artifact, we developed a high speed adaptive optics line scan confocal retinal imaging system. A high speed line camera was employed to acquire retinal image and custom adaptive optics was developed to compensate the wave aberration of the human eye's optics. The spatial resolution and signal to noise ratio were assessed in model eye and in living human eye. The improvement of imaging fidelity was estimated by reduction of intra-frame distortion of retinal images acquired in the living human eyes with frame rates at 30 frames/second (FPS), 100 FPS, and 200 FPS. The device produced retinal image with cellular level resolution at 200 FPS with a digitization of 512×512 pixels/frame in the living human eye. Cone photoreceptors in the central fovea and rod photoreceptors near the fovea were resolved in three human subjects in normal chorioretinal health. Compared with retinal images acquired at 30 FPS, the intra-frame distortion in images taken at 200 FPS was reduced by 50.9% to 79.7%. We demonstrated the feasibility of acquiring high resolution retinal images in the living human eye at a speed that minimizes retinal motion artifact. This device may facilitate research involving subjects with nystagmus or unsteady fixation due to central vision loss.

  3. Noninvasive imaging of the human rod photoreceptor mosaic using a confocal adaptive optics scanning ophthalmoscope

    Science.gov (United States)

    Dubra, Alfredo; Sulai, Yusufu; Norris, Jennifer L.; Cooper, Robert F.; Dubis, Adam M.; Williams, David R.; Carroll, Joseph

    2011-01-01

    The rod photoreceptors are implicated in a number of devastating retinal diseases. However, routine imaging of these cells has remained elusive, even with the advent of adaptive optics imaging. Here, we present the first in vivo images of the contiguous rod photoreceptor mosaic in nine healthy human subjects. The images were collected with three different confocal adaptive optics scanning ophthalmoscopes at two different institutions, using 680 and 775 nm superluminescent diodes for illumination. Estimates of photoreceptor density and rod:cone ratios in the 5°–15° retinal eccentricity range are consistent with histological findings, confirming our ability to resolve the rod mosaic by averaging multiple registered images, without the need for additional image processing. In one subject, we were able to identify the emergence of the first rods at approximately 190 μm from the foveal center, in agreement with previous histological studies. The rod and cone photoreceptor mosaics appear in focus at different retinal depths, with the rod mosaic best focus (i.e., brightest and sharpest) being at least 10 μm shallower than the cones at retinal eccentricities larger than 8°. This study represents an important step in bringing high-resolution imaging to bear on the study of rod disorders. PMID:21750765

  4. Studies of the microstructure of polymer-modified bitumen emulsions using confocal laser scanning microscopy.

    Science.gov (United States)

    Forbes, A; Haverkamp, R G; Robertson, T; Bryant, J; Bearsley, S

    2001-12-01

    Polymer-modified bitumen emulsions present a safer and more environmentally friendly binder for enhancing the properties of roads. Cationic bitumen emulsion binders containing polymer latex were investigated using confocal laser scanning microscopy. The latex was incorporated into the bitumen emulsion by using four different addition methods and all emulsions were processed with a conventional colloid mill. The emulsion binder films were studied after evaporation of the emulsion aqueous phase. We show how the microstructure and distribution of the polymer varies within the bitumen binder depending on latex addition method, and that the microstructure of the binder remains intact when exposed to elevated temperature. It was found that a distinctly fine dispersion of polymer results when the polymer is blended into the bitumen before the emulsifying process (a monophase emulsion). In contrast, bi-phase emulsion binders produced by either post-adding the latex to the bitumen emulsion, or by adding the latex into the emulsifier solution phase before processing, or by comilling the latex with the bitumen, water and emulsifier all resulted in a network formation of bitumen particles surrounded by a continuous polymer film. The use of emulsified binders appears to result in a more evenly distributed polymer network compared to the use of hot polymer-modified binders, and they therefore have greater potential for consistent binder cohesion strength, stone retention and therefore improved pavement performance.

  5. Confocal laser-scanning microscopy of capillaries in normal and psoriatic skin

    Science.gov (United States)

    Archid, Rami; Patzelt, Alexa; Lange-Asschenfeldt, Bernhard; Ahmad, Sufian S.; Ulrich, Martina; Stockfleth, Eggert; Philipp, Sandra; Sterry, Wolfram; Lademann, Juergen

    2012-10-01

    An important and most likely active role in the pathogenesis of psoriasis has been attributed to changes in cutaneous blood vessels. The purpose of this study was to use confocal laser-scanning microscopy (CLSM) to investigate dermal capillaries in psoriatic and normal skin. The structures of the capillary loops in 5 healthy participants were compared with those in affected skin of 13 psoriasis patients. The diameters of the capillaries and papillae were measured for each group with CLSM. All investigated psoriasis patients showed elongated, widened, and tortuous microvessels in the papillary dermis, whereas all healthy controls showed a single capillary loop in each dermal papilla. The capillaries of the papillary loop and the dermal papilla were significantly enlarged in the psoriatic skin lesions (diameters 24.39±2.34 and 146.46±28.52 μm, respectively) in comparison to healthy skin (diameters 9.53±1.8 and 69.48±17.16 μm, respectively) (P<0.001). CLSM appears to represent a promising noninvasive technique for evaluating dermal capillaries in patients with psoriasis. The diameter of the vessels could be seen as a well-quantifiable indicator for the state of psoriatic skin. CLSM could be useful for therapeutic monitoring to delay possible recurrences.

  6. Evaluation of Yogurt Microstructure Using Confocal Laser Scanning Microscopy and Image Analysis.

    Science.gov (United States)

    Skytte, Jacob L; Ghita, Ovidiu; Whelan, Paul F; Andersen, Ulf; Møller, Flemming; Dahl, Anders B; Larsen, Rasmus

    2015-06-01

    The microstructure of protein networks in yogurts defines important physical properties of the yogurt and hereby partly its quality. Imaging this protein network using confocal scanning laser microscopy (CSLM) has shown good results, and CSLM has become a standard measuring technique for fermented dairy products. When studying such networks, hundreds of images can be obtained, and here image analysis methods are essential for using the images in statistical analysis. Previously, methods including gray level co-occurrence matrix analysis and fractal analysis have been used with success. However, a range of other image texture characterization methods exists. These methods describe an image by a frequency distribution of predefined image features (denoted textons). Our contribution is an investigation of the choice of image analysis methods by performing a comparative study of 7 major approaches to image texture description. Here, CSLM images from a yogurt fermentation study are investigated, where production factors including fat content, protein content, heat treatment, and incubation temperature are varied. The descriptors are evaluated through nearest neighbor classification, variance analysis, and cluster analysis. Our investigation suggests that the texton-based descriptors provide a fuller description of the images compared to gray-level co-occurrence matrix descriptors and fractal analysis, while still being as applicable and in some cases as easy to tune. © 2015 Institute of Food Technologists®

  7. Comparison between optical-resolution photoacoustic microscopy and confocal laser scanning microscopy for turbid sample imaging.

    Science.gov (United States)

    U-Thainual, Paweena; Kim, Do-Hyun

    2015-12-01

    Optical-resolution photoacoustic microscopy (ORPAM) in theory provides lateral resolution equivalent to the optical diffraction limit. Scattering media, such as biological turbid media, attenuates the optical signal and also alters the diffraction-limited spot size of the focused beam. The ORPAM signal is generated only from a small voxel in scattering media with dimensions equivalent to the laser spot size after passing through scattering layers and is detected by an acoustic transducer, which is not affected by optical scattering. Thus, both ORPAM and confocal laser scanning microscopy (CLSM) reject scattered light. A multimodal optical microscopy platform that includes ORPAM and CLSM was constructed, and the lateral resolution of both modes was measured using patterned thin metal film with and without a scattering barrier. The effect of scattering media on the lateral resolution was studied using different scattering coefficients and was compared to computational results based on Monte Carlo simulations. It was found that degradation of lateral resolution due to optical scattering was not significant for either ORPAM or CLSM. The depth discrimination capability of ORPAM and CLSM was measured using microfiber embedded in a light scattering phantom material. ORPAM images demonstrated higher contrast compared to CLSM images partly due to reduced acoustic signal scattering.

  8. Subcellular localization of flavonol aglycone in hepatocytes visualized by confocal laser scanning fluorescence microscope.

    Science.gov (United States)

    Mukai, Rie; Shirai, Yasuhito; Saito, Naoaki; Yoshida, Ken-Ichi; Ashida, Hitoshi

    2009-04-01

    Flavonoids are widely distributed in the plant kingdom and show various biological activities. The bioavailability of flavonoids in biological samples has conventionally been quantified by high-performance liquid chromatography and mass spectrometry, but with these analytical techniques it is difficult to estimate the subcellular localization of flavonoids in intact cells. In this study, we attempted to examine the localization of flavonoids in cultured cells using a confocal laser scanning fluorescence microscope and mouse hepatoma Hepa-1c1c7 cells. Five flavonol aglycones showed autofluorescence in the cells under the conditions (Ex. 488 nm to Em. 515-535 nm), whereas three flavonol glycosides and eight compounds belonging to other flavonoid subclasses, i.e., flavones, flavanones, and catechins, did not. The autofluorescence of galangin and kaempferol appeared stronger in the nucleus than cytoplasm, suggesting that they are incorporated into the cells and accumulated in the nucleus. The proposed method provided evidence that flavonol aglycones are incorporated into, and accumulated in the nucleus of, hepatocytes.

  9. Homebuilt single-molecule scanning confocal fluorescence microscope studies of single DNA/protein interactions.

    Science.gov (United States)

    Zheng, Haocheng; Goldner, Lori S; Leuba, Sanford H

    2007-03-01

    Many technical improvements in fluorescence microscopy over the years have focused on decreasing background and increasing the signal to noise ratio (SNR). The scanning confocal fluorescence microscope (SCFM) represented a major improvement in these efforts. The SCFM acquires signal from a thin layer of a thick sample, rejecting light whose origin is not in the focal plane thereby dramatically decreasing the background signal. A second major innovation was the advent of high quantum-yield, low noise, single-photon counting detectors. The superior background rejection of SCFM combined with low-noise, high-yield detectors makes it possible to detect the fluorescence from single-dye molecules. By labeling a DNA molecule or a DNA/protein complex with a donor/acceptor dye pair, fluorescence resonance energy transfer (FRET) can be used to track conformational changes in the molecule/complex itself, on a single molecule/complex basis. In this methods paper, we describe the core concepts of SCFM in the context of a study that uses FRET to reveal conformational fluctuations in individual Holliday junction DNA molecules and nucleosomal particles. We also discuss data processing methods for SCFM.

  10. Starch/carrageenan/milk proteins interactions studied using multiple staining and Confocal Laser Scanning Microscopy.

    Science.gov (United States)

    Matignon, A; Moulin, G; Barey, P; Desprairies, M; Mauduit, S; Sieffermann, J M; Michon, C

    2014-01-01

    This study focused on the effects of the interactions between modified waxy maize starch, kappa carrageenan and skim milk on the microstructure of their mixed systems using Confocal Laser Scanning Microscopy (CLSM). A multiple staining of the components was set up with a view to improving starch covalent staining. In starch/carrageenan pasted mixtures, carrageenan was found to adsorb on and penetrate slightly into the starch granules, whereas no interactions were observed between starch and milk proteins. In ternary mixtures, interactions between starch granules and carrageenan were no longer observed, even when milk proteins were added after starch swelling in the carrageenan solution, thus showing preferential interactions between carrageenan/milk proteins in comparison to carrageenan/starch granules. Modifying the blending order of the components led to microstructure differences depending on several parameters such as starch/carrageenan interactions, carrageenan/milk proteins network structure, level of starch granules disruption and amylopectin contribution to the microstructure. Copyright © 2013 Elsevier Ltd. All rights reserved.

  11. Combining confocal laser scanning microscopy with serial section reconstruction in the study of adult neurogenesis.

    Directory of Open Access Journals (Sweden)

    Federico eLuzzati

    2011-05-01

    Full Text Available Current advances in imaging techniques have extended the possibility of visualizing small structures within large volumes of both fixed and live specimens without sectioning. These techniques have contributed valuable information to study neuronal plasticity in the adult brain. However, technical limits still hamper the use of these approaches to investigate neurogenic regions located far from the ventricular surface such as parenchymal neurogenic niches, or the scattered neuroblasts induced by brain lesions. Here, we present a method to combine confocal laser scanning microscopy (CLSM and serial section reconstruction in order to reconstruct large volumes of brain tissue at cellular resolution. In this method a series of thick sections are imaged with CLSM and the resulting stacks of images are registered and 3D reconstructed. This approach is based on existing freeware software and can be performed on ordinary laboratory personal computers (PC. By using this technique we have investigated the morphology and spatial organization of a group of doublecortin (DCX+ neuroblasts located in the lateral striatum of the late post-natal guinea pig. The 3D study unravelled a complex network of long and poorly ramified cell processes, often fascicled and mostly oriented along the internal capsule fibre bundles. These data support CLSM serial section reconstruction as a reliable alternative to the whole mount approaches to analyze cyto-architectural features of adult germinative niches.

  12. Use of biocytin as neuroanatomic tracer in harvested human pancreas: a confocal laser scanning microscopy analysis.

    Science.gov (United States)

    Spiga, Saturnino; Fattore, Liana; Puddu, Maria Cristina; Cappai, Antonello; Picciau, Susanna; Brotzu, Giovanni; Serra, Giuliana Paola; Petruzzo, Palmina

    2002-05-01

    To identify central neuroanatomic structure, biocytin labeling has recently been used. To date, there are no bibliographic references about the use of this molecule in investigations of the peripheral nervous system. In the present study, fresh, harvested human pancreas was used to evidence pancreatic innervations by biocytin. To investigate for the first time pancreatic innervation in harvested pancreas from human multiorgan cadaveric donors. Biocytin labeling was used as a neuroanatomic tracing method, and confocal laser scanning microscopy was used for analysis for description by means of high-resolution images. The application of biocytin-avidin staining in harvested human pancreas revealed numerous bundles of nervous fibers, intrapancreatic ganglia, few small solitary neurons, and a large number of positive supporting cells (glial-like cells). Biocytin appeared to pass through gap junctions between glial elements and neurons and among the neurons. In human pancreas, biocytin is rapidly transported in both anterograde and retrograde directions, with consequent visualization of fine details of pancreatic innervation morphology. Indeed, evidence of anterograde and retrograde transportation of biocytin has been demonstrated in the extensive labeling of pancreatic preganglionic and postganglionic fibers as well as a great number of chemical buds that wind through exocrine tissue or undetermined target cells. To our knowledge, this is the first report of the successful use of biocytin in neuronal retrograde and anterograde labeling in the human peripheral nervous system.

  13. Sarcoglycan immunoreactivity is lacking in infantile hypertrophic pyloric stenosis. A confocal laser scanning microscopic study.

    Science.gov (United States)

    Romeo, C; Santoro, G; Impellizzeri, P; Manganaro, A; Cutroneo, G; Trimarchi, E; Antonuccio, P; Anastasi, G; Zuccarello, B

    2007-01-01

    The Dystrophin-Glycoprotein Complex (DGC) is a large multisubunit complex that plays a crucial role in maintaining the structural integrity and physiology of muscle fibers. Dystrophin has been reported to be absent in the pyloric muscle of infantile hypertrophic pyloric stenosis (IHPS) patients. The present study was designed to investigate the other two patterns of DGC (dystroglycan and sarcoglycan complexes) in normal pyloric muscle and their possible modifications in IHPS patients. Ten pyloric muscle biopsies were obtained from babies operated for IHPS and five control pylorus biopsy taken at autopsy from cases without gastrointestinal disease. The DGC sub-complexes (beta-dystroglican and beta, delta- sarcoglycans) were localized immunohistochemically using specific monoclonal antibodies. The results were evaluated using a confocal laser scanning microscope. Positive immunolocalization of the two DGC sub complexes was demonstrated in the smooth muscle cells (SMCs) of the pyloric region of control patients. Similarly, a positive immune expression of beta-dystroglican was observed in the pyloric SMCs of IHPS patients. On the other hand a negative immunoreaction for sarcoglycans was recorded within the full thickness of the pyloric SMCs of these patients. The absence of sarcoglycans within the hypertrophied pyloric muscle may be a predisposing factor in the pathogenesis of IHPS since it could alter the normal physiology of SMCs through the modifications of structural integrity of sarcolemma and signaling between the extracellular and intracellular compartment.

  14. Enumeration of leukocyte infiltration in solid tumors by confocal laser scanning microscopy

    Directory of Open Access Journals (Sweden)

    Amirkhosravi A

    2006-07-01

    Full Text Available Abstract Background Leukocytes commonly infiltrate solid tumors, and have been implicated in the mechanism of spontaneous regression in some cancers. Conventional techniques for the quantitative estimation of leukocyte infiltrates in tumors rely on light microscopy of immunostained thin tissue sections, in which an arbitrary assessment (based on low, medium or high levels of infiltration of antigen density is made by the pathologist. These estimates are relatively subjective and often require the opinion of a second pathologist. In addition, since thin tissue sections are cut, no data regarding the three-dimensional distribution of antigen can be obtained. Results To overcome these problems, we have designed a method to enumerate leukocyte infiltration into tumors, using confocal laser scanning microscopy of fluorescently immunostained leukocytes in thick tissue sections. Using image analysis software, a threshold was applied to eliminate unstained tissue and residual noise. The total antigen volume in the scanned tissue was calculated and divided by the mean cell volume (calculated by "seeding" ten individual cells to obtain the cell count. Using this method, we compared the calculated leukocyte counts with those obtained manually by ten laboratory personnel. There was no significant difference (P > 0.05 between the cell counts obtained by either method. We then compared leukocyte infiltration into seven tumors and matched non-malignant tissue obtained from the periphery of the resected tissue. There was a significant increase in the infiltration of all leukocyte subsets into the tumors compared to minimal numbers in the non-malignant tissue. Conclusion From these results we conclude that this method may be of considerable use for the enumeration of cells in tissues. Furthermore, since it can be performed by laboratory technical staff, less time input is required by the pathologist in assessing the degree of leukocyte infiltration into tumors.

  15. Cost-effectiveness analysis of confocal scan laser ophthalmoscope (HRT II) versus GDX for diagnosing glaucoma.

    Science.gov (United States)

    Mokhtari-Payam, Mahdi; Moradi-Lakeh, Maziar; Yaghoubi, Mohsen; Moradijou, Mohammad

    2015-01-01

    The aim of this study was to assess the cost-effectiveness of confocal scan laser ophthalmoscopy (HRT II) and compare it with scanning laser polarimetry (GDx) for diagnosing glaucoma. A cost-effectiveness analysis was performed at two eye hospitals in Iran. The outcome was measured as the proportion of correctly diagnosed patients based on systematic review and Meta analysis. Costs were estimated at two hospitals that used the HRT II (Noor Hospital) and current diagnostic testing technology GDx (Farabi Hospital) from the perspective of the healthcare provider. The incremental cost-effectiveness ratio (ICER) was estimated on the base scenario. Annual average costs were estimated as 12.70 USD and 13.59 USD per HRT II and GDx test in 2012, respectively. It was assumed that 80% of the maximum feasible annual tests in a work shift would be performed using HRT II and GDx and that the glaucoma-positive (Gl+) proportion would be 56% in the referred eyes; the estimated diagnostic accuracies were 0.753 and 0.737 for GDx and HRT II, respectively. The incremental cost-effectiveness ratio (ICER) was estimated at USD44.18 per additional test accuracy. In a base sensitivity sampling analysis, we considered different proportions of Gl+ patients (30%-85%), one or two work shifts, and efficiency rate (60%-100%), and found that the ICER ranged from USD29.45to USD480.26, the lower and upper values in all scenarios. Based on ICER, HRT II as newer diagnostic technology is cost-effective according to the World Health Organization threshold of <1 Gross Domestic Product (GDP) per capita in Iran in 2012 (USD7228). Although GDx is more accurate and costly, the average cost-effectiveness ratio shows that HRT II provided diagnostic accuracy at a lower cost than GDx.

  16. [Use of the confocal laser scanning method for determining corneal topography and corneal tissue effects in refractive corneal surgery].

    Science.gov (United States)

    Koop, N; Brinkmann, R; Schirner, G

    1996-06-01

    Refraction of the cornea head been generally measured with ophthalmometers or computer disk keratometers. We therefore used a confocal laser scanning system for measurement of the corneal topography. Enucleated tonicized pig eyes were measured before and after laser thermokeratoplasty (LTK). The topographical data were used to determine refraction and refractive change; the data were stored digitally. The single images and their differences were displayed on a PC. Unlike conventional ophthalmometry, confocal laser scanning can demonstrate the topographical shape, showing the overall topography of the cornea and local corneal effects, e.g., coagulation, mechanical lesions or high-energy laser effects. Topographical laser scanning has proven to be a generally useful method of determining refraction and surface alterations in corneal refractive surgery.

  17. Thermal maturity of Tasmanites microfossils from confocal laser scanning fluorescence microscopy

    Science.gov (United States)

    Hackley, Paul C.; Kus, Jolanta

    2015-01-01

    We report here, for the first time, spectral properties of Tasmanites microfossils determined by confocal laser scanning fluorescence microscopy (CLSM, using Ar 458 nm excitation). The Tasmanites occur in a well-characterized natural maturation sequence (Ro 0.48–0.74%) of Devonian shale (n = 3 samples) from the Appalachian Basin. Spectral property λmax shows excellent agreement (r2 = 0.99) with extant spectra from interlaboratory studies which used conventional fluorescence microscopy techniques. This result suggests spectral measurements from CLSM can be used to infer thermal maturity of fluorescent organic materials in geologic samples. Spectra of regions with high fluorescence intensity at fold apices and flanks in individual Tasmanites are blue-shifted relative to less-deformed areas in the same body that have lower fluorescence intensity. This is interpreted to result from decreased quenching moiety concentration at these locations, and indicates caution is needed in the selection of measurement regions in conventional fluorescence microscopy, where it is common practice to select high intensity regions for improved signal intensity and better signal to noise ratios. This study also documents application of CLSM to microstructural characterization of Tasmanites microfossils. Finally, based on an extant empirical relation between conventional λmax values and bitumen reflectance, λmax values from CLSM of Tasmanites microfossils can be used to calculate a bitumen reflectance equivalent value. The results presented herein can be used as a basis to broaden the future application of CLSM in the geological sciences into hydrocarbon prospecting and basin analysis.

  18. Performance of confocal scanning laser tomograph Topographic Change Analysis (TCA) for assessing glaucomatous progression.

    Science.gov (United States)

    Bowd, Christopher; Balasubramanian, Madhusudhanan; Weinreb, Robert N; Vizzeri, Gianmarco; Alencar, Luciana M; O'Leary, Neil; Sample, Pamela A; Zangwill, Linda M

    2009-02-01

    To determine the sensitivity and specificity of confocal scanning laser ophthalmoscope's Topographic Change Analysis (TCA; Heidelberg Retina Tomograph [HRT]; Heidelberg Engineering, Heidelberg, Germany) parameters for discriminating between progressing glaucomatous and stable healthy eyes. The 0.90, 0.95, and 0.99 specificity cutoffs for various (n=70) TCA parameters were developed by using 1000 permuted topographic series derived from HRT images of 18 healthy eyes from Moorfields Eye Hospital, imaged at least four times. The cutoffs were then applied to topographic series from 36 eyes with known glaucomatous progression (by optic disc stereophotograph assessment and/or standard automated perimetry guided progression analysis, [GPA]) and 21 healthy eyes from the University of California, San Diego (UCSD) Diagnostic Innovations in Glaucoma Study (DIGS), all imaged at least four times, to determine TCA sensitivity and specificity. Cutoffs also were applied to 210 DIGS patients' eyes imaged at least four times with no evidence of progression (nonprogressed) by stereophotography or GPA. The TCA parameter providing the best sensitivity/specificity tradeoff using the 0.90, 0.95, and 0.99 cutoffs was the largest clustered superpixel area within the optic disc margin (CAREA(disc) mm(2)). Sensitivities/specificities for classifying progressing (by stereophotography and/or GPA) and healthy eyes were 0.778/0.809, 0.639/0.857, and 0.611/1.00, respectively. In nonprogressing eyes, specificities were 0.464, 0.570, and 0.647 (i.e., lower than in the healthy eyes). In addition, TCA parameter measurements of nonprogressing eyes were similar to those of progressing eyes. TCA parameters can discriminate between progressing and longitudinally observed healthy eyes. Low specificity in apparently nonprogressing patients' eyes suggests early progression detection using TCA.

  19. Effect of calcium on Staphylococcus aureus biofilm architecture: a confocal laser scanning microscopic study.

    Science.gov (United States)

    Shukla, Sudhir K; Rao, T Subba

    2013-03-01

    Bacterial adhesion is a threshold event in the formation of biofilms. Several studies on molecular and biochemical aspects have highlighted that the protein matrix of the biofilm is of interest in developing strategies to combat biofouling. The prevalent role of biofilm associated protein (Bap) of Staphylococcus aureus in early adhesion and the putative presence of Ca(2+) binding EF hand motif in Bap was the motivation for this study. Biofilm assays (S. aureus strains V329 and M556) were done in micro-titer plates and confocal laser scanning microscopy (CLSM) was used to study the biofilm architecture. The results showed that Ca(2+) did not influence planktonic growth of the cultures; however, it modulated the biofilm architecture of S. aureus V329 in a dose dependent manner. Strain M556 was found to be a weak biofilm former and showed no significant change in the presence of Ca(2+). When tested with increasing NaCl concentration, there was no reversal of the Bap-dependent Ca(2+) inhibition of S. aureus V329 biofilm. This indicates that the interaction of Bap and Ca(2+) is not mere electrostatic. CLSM images of V329 biofilm showed reduction in biofilm thickness as well as altered biofilm topography with varying Ca(2+) concentrations. The inhibition effect of Ca(2+) on strain V329 biofilm disappeared in the presence of chelating agent EDTA at a non-inhibiting concentration (0.15 mM). The paper elaborates the role of Ca(2+) in biofilm architecture of S. aureus. Copyright © 2012 Elsevier B.V. All rights reserved.

  20. Neutral Red as a Probe for Confocal Laser Scanning Microscopy Studies of Plant Roots

    Science.gov (United States)

    DUBROVSKY, JOSEPH G.; GUTTENBERGER, MARTIN; SARALEGUI, ANDRES; NAPSUCIALY-MENDIVIL, SELENE; VOIGT, BORIS; BALUŠKA, FRANTIŠEK; MENZEL, DIEDRIK

    2006-01-01

    • Background and Aims Neutral red (NR), a lipophilic phenazine dye, has been widely used in various biological systems as a vital stain for bright-field microscopy. In its unprotonated form it penetrates the plasma membrane and tonoplast of viable plant cells, then due to protonation it becomes trapped in acidic compartments. The possible applications of NR for confocal laser scanning microscopy (CLSM) studies were examined in various aspects of plant root biology. • Methods NR was used as a fluorochrome for living roots of Phaseolus vulgaris, Allium cepa, A. porrum and Arabidopsis thaliana (wild-type and transgenic GFP-carrying lines). The tissues were visualized using CLSM. The effect of NR on the integrity of the cytoskeleton and the growth rate of arabidopsis primary roots was analysed to judge potential toxic effects of the dye. • Key Results The main advantages of the use of NR are related to the fact that NR rapidly penetrates root tissues, has affinity to suberin and lignin, and accumulates in the vacuoles. It is shown that NR is a suitable probe for visualization of proto- and metaxylem elements, Casparian bands in the endodermis, and vacuoles in cells of living roots. The actin cytoskeleton and the microtubule system of the cells, as well as the dynamics of root growth, remain unchanged after short-term application of NR, indicating a relatively low toxicity of this chemical. It was also found that NR is a useful probe for the observation of the internal structures of root nodules and of fungal hyphae in vesicular–arbuscular mycorrhizas. • Conclusions Ease, low cost and absence of tissue processing make NR a useful probe for structural, developmental and vacuole-biogenetic studies of plant roots with CLSM. PMID:16520341

  1. CTC staining and counting of actively respiring bacteria in natural stone using confocal laser scanning microscopy.

    Science.gov (United States)

    Bartosch, S; Mansch, R; Knötzsch, K; Bock, E

    2003-01-01

    A method was established for staining and counting of actively respiring bacteria in natural stone by using the tetrazolium salt 5-cyano-2,3-ditolyltetrazolium chloride (CTC) in combination with confocal laser scanning microscopy (CLSM). Applying 5 mM CTC for 2 h to pure cultures of representative stone-inhabiting microorganisms showed that chemoorganotrophic bacteria and fungi-in contrast to lithoautotrophic nitrifying bacteria-were able to reduce CTC to CTF, the red fluorescing formazan crystals of CTC. Optimal staining conditions for microorganisms in stone material were found to be 15 mM CTC applied for 24 h. The cells could be visualized on transparent and nontransparent mineral materials by means of CLSM. A semi-automated method was used to count the cells within the pore system of the stone. The percentage of CTC-stained bacteria was dependent on temperature and humidity of the material. At 28 degrees C and high humidity (maximum water holding capacity) in the laboratory, about 58% of the total bacterial microflora was active. On natural stone exposed for 9 years at an urban exposure site in Germany, 52-56% of the bacterial microflora was active at the east, west, and north side of the specimen, while only 18% cells were active at the south side. This is consistent with microclimatic differences on the south side which was more exposed to sunshine thus causing UV and water stress as well as higher temperatures on a microscale level. In combination with CLSM, staining by CTC can be used as a fast method for monitoring the metabolic activity of chemoorganotrophic bacteria in monuments, buildings of historic interest or any art objects of natural stone. Due to the small size of samples required, the damage to these objects and buildings can be minimized.

  2. Correlative scanning electron and confocal microscopy imaging of labeled cells coated by indium-tin-oxide

    KAUST Repository

    Rodighiero, Simona

    2015-03-22

    Confocal microscopy imaging of cells allows to visualize the presence of specific antigens by using fluorescent tags or fluorescent proteins, with resolution of few hundreds of nanometers, providing their localization in a large field-of-view and the understanding of their cellular function. Conversely, in scanning electron microscopy (SEM), the surface morphology of cells is imaged down to nanometer scale using secondary electrons. Combining both imaging techniques have brought to the correlative light and electron microscopy, contributing to investigate the existing relationships between biological surface structures and functions. Furthermore, in SEM, backscattered electrons (BSE) can image local compositional differences, like those due to nanosized gold particles labeling cellular surface antigens. To perform SEM imaging of cells, they could be grown on conducting substrates, but obtaining images of limited quality. Alternatively, they could be rendered electrically conductive, coating them with a thin metal layer. However, when BSE are collected to detect gold-labeled surface antigens, heavy metals cannot be used as coating material, as they would mask the BSE signal produced by the markers. Cell surface could be then coated with a thin layer of chromium, but this results in a loss of conductivity due to the fast chromium oxidation, if the samples come in contact with air. In order to overcome these major limitations, a thin layer of indium-tin-oxide was deposited by ion-sputtering on gold-decorated HeLa cells and neurons. Indium-tin-oxide was able to provide stable electrical conductivity and preservation of the BSE signal coming from the gold-conjugated markers. © 2015 Wiley Periodicals, Inc.

  3. Correlative scanning electron and confocal microscopy imaging of labeled cells coated by indium-tin-oxide.

    Science.gov (United States)

    Rodighiero, Simona; Torre, Bruno; Sogne, Elisa; Ruffilli, Roberta; Cagnoli, Cinzia; Francolini, Maura; Di Fabrizio, Enzo; Falqui, Andrea

    2015-06-01

    Confocal microscopy imaging of cells allows to visualize the presence of specific antigens by using fluorescent tags or fluorescent proteins, with resolution of few hundreds of nanometers, providing their localization in a large field-of-view and the understanding of their cellular function. Conversely, in scanning electron microscopy (SEM), the surface morphology of cells is imaged down to nanometer scale using secondary electrons. Combining both imaging techniques have brought to the correlative light and electron microscopy, contributing to investigate the existing relationships between biological surface structures and functions. Furthermore, in SEM, backscattered electrons (BSE) can image local compositional differences, like those due to nanosized gold particles labeling cellular surface antigens. To perform SEM imaging of cells, they could be grown on conducting substrates, but obtaining images of limited quality. Alternatively, they could be rendered electrically conductive, coating them with a thin metal layer. However, when BSE are collected to detect gold-labeled surface antigens, heavy metals cannot be used as coating material, as they would mask the BSE signal produced by the markers. Cell surface could be then coated with a thin layer of chromium, but this results in a loss of conductivity due to the fast chromium oxidation, if the samples come in contact with air. In order to overcome these major limitations, a thin layer of indium-tin-oxide was deposited by ion-sputtering on gold-decorated HeLa cells and neurons. Indium-tin-oxide was able to provide stable electrical conductivity and preservation of the BSE signal coming from the gold-conjugated markers. © 2015 Wiley Periodicals, Inc.

  4. Cutting efficiency of apical preparation using ultrasonic tips with microprojections: confocal laser scanning microscopy study

    Directory of Open Access Journals (Sweden)

    Sang-Won Kwak

    2014-11-01

    Full Text Available Objectives The purpose of this study was to compare the cutting efficiency of a newly developed microprojection tip and a diamond-coated tip under two different engine powers. Materials and Methods The apical 3-mm of each root was resected, and root-end preparation was performed with upward and downward pressure using one of the ultrasonic tips, KIS-1D (Obtura Spartan or JT-5B (B&L Biotech Ltd.. The ultrasonic engine was set to power-1 or -4. Forty teeth were randomly divided into four groups: K1 (KIS-1D / Power-1, J1 (JT-5B / Power-1, K4 (KIS-1D / Power-4, and J4 (JT-5B / Power-4. The total time required for root-end preparation was recorded. All teeth were resected and the apical parts were evaluated for the number and length of cracks using a confocal scanning micrscope. The size of the root-end cavity and the width of the remaining dentin were recorded. The data were statistically analyzed using two-way analysis of variance and a Mann-Whitney test. Results There was no significant difference in the time required between the instrument groups, but the power-4 groups showed reduced preparation time for both instrument groups (p < 0.05. The K4 and J4 groups with a power-4 showed a significantly higher crack formation and a longer crack irrespective of the instruments. There was no significant difference in the remaining dentin thickness or any of the parameters after preparation. Conclusions Ultrasonic tips with microprojections would be an option to substitute for the conventional ultrasonic tips with a diamond coating with the same clinical efficiency.

  5. Imaging hyphal growth of Physisporinus vitreus in Norway spruce wood by means of confocal laser scanning microscopy (CLSM)

    OpenAIRE

    Schubert, Mark; Stührk, Chris; Fuhr, Matthias J.; Schwarze, Francis W.M.R.

    2017-01-01

    Light microscopy and electron microscopy are the most common methods for analyzing wood-decay fungi. However, the 3D visualization and quantification of the filamentous structure of fungi in wood is difficult to realize by means of these traditional techniques. In the present work, confocal laser scanning microscopy (CLSM) was further developed for the quantitative imaging of the 3D microscopic hyphal growth of Physisporinus vitreus, a versatile fungus for engineering value-added wood product...

  6. Confocal laser scanning microscopy of liesegang rings in odontogenic cysts: analysis of three-dimensional image reconstruction.

    Science.gov (United States)

    Scivetti, Michele; Lucchese, Alberta; Crincoli, Vito; Pilolli, Giovanni Pietro; Favia, Gianfranco

    2009-01-01

    Liesegang rings are concentric noncellular lamellar structures, occasionally found in inflammatory tissues. They have been confused with various parasites, algas, calcification, and psammoma bodies. The authors examined Liesegang rings from oral inflammatory cysts by both optical and confocal laser scanning microscopy, and perfomed a three-dimensional reconstruction. These investigations indicate that Liesegang rings are composed of multiple birefringent concentric rings, resulting from a progressive deposition of organic substances, with an unclear pathogenesis.

  7. Laser-scanning velocimetry: A confocal microscopy method for quantitative measurement of cardiovascular performance in zebrafish embryos and larvae

    Directory of Open Access Journals (Sweden)

    Linney Elwood

    2007-07-01

    Full Text Available Abstract Background The zebrafish Danio rerio is an important model system for drug discovery and to study cardiovascular development. Using a laser-scanning confocal microscope, we have developed a non-invasive method of measuring cardiac performance in zebrafish embryos and larvae that obtains cardiovascular parameters similar to those obtained using Doppler echocardiography in mammals. A laser scan line placed parallel to the path of blood in the dorsal aorta measures blood cell velocity, from which cardiac output and indices of vascular resistance and contractility are calculated. Results This technique, called laser-scanning velocimetry, was used to quantify the effects of pharmacological, developmental, and genetic modifiers of cardiac function. Laser-scanning velocimetry was applied to analyze the cardiovascular effects of morpholino knockdown of osmosensing scaffold for MEKK3 (OSM, which when mutated causes the human vascular disease cerebral cavernous malformations. OSM-deficient embryos had a constricted aortic arch and markedly increased peak cell velocity, a characteristic indicator of aortic stenosis. Conclusion These data validate laser-scanning velocimetry as a quantitative tool to measure cardiovascular performance for pharmacological and genetic analysis in zebrafish, which requires no specialized equipment other than a laser-scanning confocal microscope.

  8. A portable confocal hyperspectral microscope without any scan or tube lens and its application in fluorescence and Raman spectral imaging

    Science.gov (United States)

    Li, Jingwei; Cai, Fuhong; Dong, Yongjiang; Zhu, Zhenfeng; Sun, Xianhe; Zhang, Hequn; He, Sailing

    2017-06-01

    In this study, a portable confocal hyperspectral microscope is developed. In traditional confocal laser scanning microscopes, scan lens and tube lens are utilized to achieve a conjugate relationship between the galvanometer and the back focal plane of the objective, in order to achieve a better resolution. However, these lenses make it difficult to scale down the volume of the system. In our portable confocal hyperspectral microscope (PCHM), the objective is placed directly next to the galvomirror. Thus, scan lens and tube lens are not included in our system and the size of this system is greatly reduced. Furthermore, the resolution is also acceptable in many biomedical and food-safety applications. Through reducing the optical length of the system, the signal detection efficiency is enhanced. This is conducive to realizing both the fluorescence and Raman hyperspectral imaging. With a multimode fiber as a pinhole, an improved image contrast is also achieved. Fluorescent spectral images for HeLa cells/fingers and Raman spectral images of kumquat pericarp are present. The spectral resolution and spatial resolutions are about 0.4 nm and 2.19 μm, respectively. These results demonstrate that this portable hyperspectral microscope can be used in in-vivo fluorescence imaging and in situ Raman spectral imaging.

  9. Comparison of divided and full pupil configurations for line-scanning confocal microscopy in human skin and oral mucosa

    Science.gov (United States)

    Larson, Bjorg; Abeytunge, Sanjeewa; Glazowski, Chris; Rajadhyaksha, Milind

    2012-02-01

    Confocal point-scanning microscopy has been showing promise in the detection, diagnosing and mapping of skin lesions in clinical settings. The noninvasive technique allows provides optical sectioning and cellular resolution for in vivo diagnosis of melanoma and basal cell carcinoma and pre-operative and intra-operative mapping of margins. The imaging has also enabled more accurate "guided" biopsies while minimizing the otherwise large number of "blind" biopsies. Despite these translational advances, however, point-scanning technology remains relatively complex and expensive. Line-scanning technology may offer an alternative approach to accelerate translation to the clinic. Line-scanning, using fewer optical components, inexpensive linear-array detectors and custom electronics, may enable smaller, simpler and lower-cost confocal microscopes. A line is formed using a cylindrical lens and scanned through the back focal plane of the objective with a galvanometric scanner. A linear CCD is used for detection. Two pupil configurations were compared for performance in imaging human tissue. In the full-pupil configuration, illumination and detection is made through the full objective pupil. In the divided pupil approach, half the pupil is illuminated and the other half is used for detection. The divided pupil configuration loses spatial and axial resolution due to a diminished NA, but the sectioning capability and rejection of background is improved. Imaging in skin and oral mucosa illustrate the performance of the two configurations.

  10. The neuromuscular system of Pycnophyes kielensis (Kinorhyncha: Allomalorhagida investigated by confocal laser scanning microscopy

    Directory of Open Access Journals (Sweden)

    Andreas Altenburger

    2016-11-01

    Full Text Available Abstract Background Kinorhynchs are ecdysozoan animals with a phylogenetic position close to priapulids and loriciferans. To understand the nature of segmentation within Kinorhyncha and to infer a probable ancestry of segmentation within the last common ancestor of Ecdysozoa, the musculature and the nervous system of the allomalorhagid kinorhynch Pycnophyes kielensis were investigated by use of immunohistochemistry, confocal laser scanning microscopy, and 3D reconstruction software. Results The kinorhynch body plan comprises 11 trunk segments. Trunk musculature consists of paired ventral and dorsal longitudinal muscles in segments 1–10 as well as dorsoventral muscles in segments 1–11. Dorsal and ventral longitudinal muscles insert on apodemes of the cuticle inside the animal within each segment. Strands of longitudinal musculature extend over segment borders in segments 1–6. In segments 7–10, the trunk musculature is confined to the segments. Musculature of the digestive system comprises a strong pharyngeal bulb with attached mouth cone muscles as well as pharyngeal bulb protractors and retractors. The musculature of the digestive system shows no sign of segmentation. Judged by the size of the pharyngeal bulb protractors and retractors, the pharyngeal bulb, as well as the introvert, is moved passively by internal pressure caused by concerted action of the dorsoventral muscles. The nervous system comprises a neuropil ring anterior to the pharyngeal bulb. Associated with the neuropil ring are flask-shaped serotonergic somata extending anteriorly and posteriorly. A ventral nerve cord is connected to the neuropil ring and runs toward the anterior until an attachment point in segment 1, and from there toward the posterior with one ganglion in segment 6. Conclusions Segmentation within Kinorhyncha likely evolved from an unsegmented ancestor. This conclusion is supported by continuous trunk musculature in the anterior segments 1–6, continuous

  11. Reconstruction of confocal micro-X-ray fluorescence spectroscopy depth scans obtained with a laboratory setup.

    Science.gov (United States)

    Mantouvalou, Ioanna; Wolff, Timo; Seim, Christian; Stoytschew, Valentin; Malzer, Wolfgang; Kanngiesser, Birgit

    2014-10-07

    Depth profiling with confocal micro-X-ray fluorescence spectroscopy (confocal micro-XRF) is a nondestructive analytical method for obtaining elemental depth profiles in the micrometer region. Up until now, the quantitative reconstruction of thicknesses and elemental concentration of stratified samples has been only possible with monochromatic, thus, synchrotron radiation. In this work, we present a new calibration and reconstruction procedure, which renders quantification in the laboratory feasible. The proposed model uses the approximation of an effective spot size of the optic in the excitation channel and relies on the calibration of the transmission of this lens beforehand. Calibration issues are discussed and validation measurements on thick multielement reference material and a stratified system are presented.

  12. Effects of Naphthalene on DNA and RNA quantity in Amoeba proteus by using confocal laser scanning microscope

    Directory of Open Access Journals (Sweden)

    Khwanmuni, J.

    2006-01-01

    Full Text Available Effects of Naphthalene which is a carcinogen on changes of DNA and RNA quantities were studied with acridine orange stained cells under a confocal laser scanning microscope. It was found that DNA and RNA in amoebae nucleus and cytoplasm, reared in 0 (control, 3 and 8.85 mg/l (24h-LD50 at 0 and 12 h. showed a statistically significant difference (p<0.05. The more naphthalene concentrations and larger incubation periods had greater effects on DNA and RNA decreases in amoebae nucleus and cytoplasm.

  13. Applications of confocal laser scanning microscopy in research into organic semiconductor thin films

    DEFF Research Database (Denmark)

    Schiek, Manuela; Balzer, Frank

    2014-01-01

    At the center of opto-electronic devices are thin layers of organic semiconductors, which need to be sandwiched between planar electrodes. With the growing demand for opto-electronic devices now and in the future, new electrode materials are needed to meet the requirements of organic semiconductors....... Control of these interfaces directly impacts on the performance and here we show with basic growth studies of model compounds on dielectric and graphitic surfaces, the formation of distinctly textured films. Silver-nanowire meshes are presented as an alternative transparent electrode material. Confocal...

  14. Miniature in vivo MEMS-based line-scanned dual-axis confocal microscope for point-of-care pathology

    Science.gov (United States)

    Yin, C.; Glaser, A.K.; Leigh, S. Y.; Chen, Y.; Wei, L.; Pillai, P. C. S.; Rosenberg, M. C.; Abeytunge, S.; Peterson, G.; Glazowski, C.; Sanai, N.; Mandella, M. J.; Rajadhyaksha, M.; Liu, J. T. C.

    2016-01-01

    There is a need for miniature optical-sectioning microscopes to enable in vivo interrogation of tissues as a real-time and noninvasive alternative to gold-standard histopathology. Such devices could have a transformative impact for the early detection of cancer as well as for guiding tumor-resection procedures. Miniature confocal microscopes have been developed by various researchers and corporations to enable optical sectioning of highly scattering tissues, all of which have necessitated various trade-offs in size, speed, depth selectivity, field of view, resolution, image contrast, and sensitivity. In this study, a miniature line-scanned (LS) dual-axis confocal (DAC) microscope, with a 12-mm diameter distal tip, has been developed for clinical point-of-care pathology. The dual-axis architecture has demonstrated an advantage over the conventional single-axis confocal configuration for reducing background noise from out-of-focus and multiply scattered light. The use of line scanning enables fast frame rates (16 frames/sec is demonstrated here, but faster rates are possible), which mitigates motion artifacts of a hand-held device during clinical use. We have developed a method to actively align the illumination and collection beams in a DAC microscope through the use of a pair of rotatable alignment mirrors. Incorporation of a custom objective lens, with a small form factor for in vivo clinical use, enables our device to achieve an optical-sectioning thickness and lateral resolution of 2.0 and 1.1 microns respectively. Validation measurements with reflective targets, as well as in vivo and ex vivo images of tissues, demonstrate the clinical potential of this high-speed optical-sectioning microscopy device. PMID:26977337

  15. Real-time mapping of the corneal sub-basal nerve plexus by in vivo laser scanning confocal microscopy

    Science.gov (United States)

    Guthoff, Rudolf F.; Zhivov, Andrey; Stachs, Oliver

    2010-02-01

    The aim of the study was to produce two-dimensional reconstruction maps of the living corneal sub-basal nerve plexus by in vivo laser scanning confocal microscopy in real time. CLSM source data (frame rate 30Hz, 384x384 pixel) were used to create large-scale maps of the scanned area by selecting the Automatic Real Time (ART) composite mode. The mapping algorithm is based on an affine transformation. Microscopy of the sub-basal nerve plexus was performed on normal and LASIK eyes as well as on rabbit eyes. Real-time mapping of the sub-basal nerve plexus was performed in large-scale up to a size of 3.2mm x 3.2mm. The developed method enables a real-time in vivo mapping of the sub-basal nerve plexus which is stringently necessary for statistically firmed conclusions about morphometric plexus alterations.

  16. Design of an affordable fluorescence confocal laser scanning microscope for medical diagnostics

    Science.gov (United States)

    Bechtel, Christin; Knobbe, Jens; Grüger, Heinrich; Lakner, Hubert

    2012-12-01

    Confocal fluorescence microscopes are a promising imaging tool in medical diagnostics due to their capability to selectively survey cross-sections of individual layers from `thick' samples. Non-invasive depth resolved investigation of neoplastic skin disorders is one example among other applications. However these microscopes are at present uncommon in medical practice. This is due to their main application area in research. The instruments dealt with here are generally complex, stationary units and are accordingly cost-intensive. It is for this reason, that we have designed a robust and portable MEMS based confocal fluorescence microscope with a field of view of 0.6mm x 0.6mm. This has been made possible by the integration of a 2D micro scanner mirror developed at Fraunhofer IPMS. A variable acquisition depth of cross-sectional images of the fluorescence specimen is enabled by an integrated z-shifter. With the use of commercially available optics an optical demonstrator set up has been realized. To characterize and to demonstrate the ability of this system test measurements were performed. The resolution of the microscope is better than 228 lp/mm determined by 1951 USAF resolution test target. Images of various biological samples are presented and optical sectioning capabilities are shown. A comparison of the measured with the predicted system performance will be given.

  17. Impression Cytology in Eyes with Clinical and Confocal Scan Features of Acanthamoeba Keratitis

    Directory of Open Access Journals (Sweden)

    Mozhgan Rezaei Kanavi

    2013-01-01

    Full Text Available Purpose: To report impression cytology findings in specimens obtained from eyes with clinical and confocal microscopic features of Acanthamoeba keratitis (AK. Methods: In this interventional case series, impression cytology was obtained from corneas of patients with clinical and confocal microscopic features indicative of AK. Specimens were stained with Periodic acid-Schiff/Papanicolaou (PAS/PAP and examined for the presence of PAS-reactive Acanthamoeba cysts and/or hyperchromatic pear-shaped trophozoites. All specimens were then decolorized and re-stained with calcofluor white (CFW for the presence of chemofluorescent cysts. Results: Fifty-six eyes of 50 patients with mean age of 25.5±9.3 (range, 17 to 78 years were evaluated. Forty-one (82% cases were female and 51 (91.1% eyes had history of contact lens wear. PAS-reactive Acanthamoeba cysts and/or hyperchromatic pear-shaped trophozoites were identified in 53 eyes (94.6%, 2 of which demonstrated only trophozoitelike structures. CFW staining was able to reveal the presence of chemofluorescent cysts in all 51 specimens (91.1% in which cysts had been demonstrated with PAS/PAP staining. Trophozoites were not detected with CFW due to background staining of the cellulose acetate strip used for impression cytology. Conclusion: Corneal impression cytology, stained with PAS/PAP or with CFW, successfully detects Acanthamoeba and can be employed for early noninvasive diagnosis of AK.

  18. Impression cytology in eyes with clinical and confocal scan features of acanthamoeba keratitis.

    Science.gov (United States)

    Rezaei Kanavi, Mozhgan; Hosseini, Bagher; Javadi, Fatemeh; Rakhshani, Nasser; Javadi, Mohammad-Ali

    2013-07-01

    To report impression cytology findings in specimens obtained from eyes with clinical and confocal microscopic features of Acanthamoeba keratitis (AK). In this interventional case series, impression cytology was obtained from corneas of patients with clinical and confocal microscopic features indicative of AK. Specimens were stained with Periodic acid-Schiff/Papanicolaou (PAS/PAP) and examined for the presence of PAS-reactive Acanthamoeba cysts and/or hyperchromatic pear-shaped trophozoites. All specimens were then decolorized and re-stained with calcofluor white (CFW) for the presence of chemofluorescent cysts. Fifty-six eyes of 50 patients with mean age of 25.5±9.3 (range, 17 to 78) years were evaluated. Forty-one (82%) cases were female and 51 (91.1%) eyes had history of contact lens wear. PAS-reactive Acanthamoeba cysts and/or hyperchromatic pear-shaped trophozoites were identified in 53 eyes (94.6%), 2 of which demonstrated only trophozoite- like structures. CFW staining was able to reveal the presence of chemofluorescent cysts in all 51 specimens (91.1%) in which cysts had been demonstrated with PAS/PAP staining. Trophozoites were not detected with CFW due to background staining of the cellulose acetate strip used for impression cytology. Corneal impression cytology, stained with PAS/PAP or with CFW, successfully detects Acanthamoeba and can be employed for early noninvasive diagnosis of AK.

  19. An essential role for dendritic cells in vernal keratoconjunctivitis: analysis by laser scanning confocal microscopy.

    Science.gov (United States)

    Liu, M; Gao, H; Wang, T; Wang, S; Li, S; Shi, W

    2014-03-01

    CD4+ T helper type 2 cells play a central role in the pathogenesis of vernal keratoconjunctivitis (VKC), and antigen-presenting cells are required for the cell activation. In this study, we aimed to survey the density, distribution, and morphology of dendritic cells (DCs) in patients with VKC by in vivo confocal microscopy. Thirty-five patients (mean, 12.4 ± 5.3 years) affected by VKC were included. All patients were treated with 0.1% fluorometholone eye drops and 0.5% cyclosporine A eye drops. The density and morphological and distributional characteristics of DCs in each right eye were evaluated by in vivo confocal microscopy before treatment and at 1, 3, and 6 months after treatment. Thirty-five age-matched normal subjects (mean, 16.5 ± 1.8 years) were studied as controls. There was significant difference in age between the VKC group and the control group (F = 18.17, P < 0.05). Compared with normal eyes, increased numbers of DCs were found in patients with VKC, with mean cell densities of 244.09 ± 59.76 cells/mm(2) at the bulbar conjunctiva, 574.53 ± 87.34 cells/mm(2) at the limbus, and 403.32 ± 106.59 cells/mm(2) at the peripheral cornea before treatment. These DCs exhibited a typical dendritic shape. At 3 months after treatment, the DC density at the conjunctiva decreased significantly (P < 0.05), approximating that in the controls. At 3 and 6 months, the DC densities at the limbus and peripheral cornea also decreased significantly (P < 0.05), but were still statistically higher than those in the controls. These DCs, with small dendritic processes or irregular shapes, were observed to gradually locate at the epithelial basal membrane and subbasal nerve plexus. In vivo confocal microscopy appears to be a valuable tool in evaluating the dynamic change of DCs at the conjunctiva and cornea. DCs play an essential role in VKC and therefore may constitute a target for therapeutic intervention for VKC. © 2013 John Wiley & Sons Ltd.

  20. Aerogel Track Morphology: Measurement, Three Dimensional Reconstruction and Particle Location using Confocal Laser Scanning Microscopy

    Science.gov (United States)

    Kearsley, A. T.; Ball, A. D.; Wozniakiewicz, P. A.; Graham, G. A.; Burchell, M. J.; Cole, M. J.; Horz, F.; See, T. H.

    2007-01-01

    The Stardust spacecraft returned the first undoubted samples of cometary dust, with many grains embedded in the silica aerogel collector . Although many tracks contain one or more large terminal particles of a wide range of mineral compositions , there is also abundant material along the track walls. To help interpret the full particle size, structure and mass, both experimental simulation of impact by shots and numerical modeling of the impact process have been attempted. However, all approaches require accurate and precise measurement of impact track size parameters such as length, width and volume of specific portions. To make such measurements is not easy, especially if extensive aerogel fracturing and discoloration has occurred. In this paper we describe the application and limitations of laser confocal imagery for determination of aerogel track parameters, and for the location of particle remains.

  1. Upgrade of a Scanning Confocal Microscope to a Single-Beam Path STED Microscope.

    Directory of Open Access Journals (Sweden)

    André Klauss

    Full Text Available By overcoming the diffraction limit in light microscopy, super-resolution techniques, such as stimulated emission depletion (STED microscopy, are experiencing an increasing impact on life sciences. High costs and technically demanding setups, however, may still hinder a wider distribution of this innovation in biomedical research laboratories. As far-field microscopy is the most widely employed microscopy modality in the life sciences, upgrading already existing systems seems to be an attractive option for achieving diffraction-unlimited fluorescence microscopy in a cost-effective manner. Here, we demonstrate the successful upgrade of a commercial time-resolved confocal fluorescence microscope to an easy-to-align STED microscope in the single-beam path layout, previously proposed as "easy-STED", achieving lateral resolution < λ/10 corresponding to a five-fold improvement over a confocal modality. For this purpose, both the excitation and depletion laser beams pass through a commercially available segmented phase plate that creates the STED-doughnut light distribution in the focal plane, while leaving the excitation beam unaltered when implemented into the joint beam path. Diffraction-unlimited imaging of 20 nm-sized fluorescent beads as reference were achieved with the wavelength combination of 635 nm excitation and 766 nm depletion. To evaluate the STED performance in biological systems, we compared the popular phalloidin-coupled fluorescent dyes Atto647N and Abberior STAR635 by labeling F-actin filaments in vitro as well as through immunofluorescence recordings of microtubules in a complex epithelial tissue. Here, we applied a recently proposed deconvolution approach and showed that images obtained from time-gated pulsed STED microscopy may benefit concerning the signal-to-background ratio, from the joint deconvolution of sub-images with different spatial information which were extracted from offline time gating.

  2. Molecular characterization and confocal laser scanning microscopic study of Pygidiopsis macrostomum (Trematoda: Heterophyidae) parasites of guppies Poecilia vivipara.

    Science.gov (United States)

    Borges, J N; Costa, V S; Mantovani, C; Barros, E; Santos, E G N; Mafra, C L; Santos, C P

    2017-02-01

    Pygidiopsis macrostomum and Ascocotyle (Phagicola) pindoramensis (Digenea: Heterophyidae) parasitize guppies as intermediate hosts and, respectively, fish-eating mammals or birds as definitive hosts. Heterophyids have zoonotic potential, and molecular studies associated with morphological and ecological aspects have helped to clarify their taxonomy and phylogeny. Poecilia vivipara naturally parasitized by metacercariae of both species (100% prevalence) exhibit no external signs of parasitism. In this work, four new sequences of P. macrostomum (18S rDNA, 28S rDNA and ITS2 rDNA) and one new sequence of A. (P.) pindoramensis (mtDNA cox-1) are presented. Phylogeny reconstructions linked P. macrostomum to other heterophyids, but the separation of the Heterophyidae and Opisthorchiidae remains unclear. Additionally, we used indirect immunocytochemistry and the phalloidin-fluorescence techniques allied with confocal laser scanning microscopy to describe muscular and neuronal structures of P. macrostomum. A complex arrangement of muscular fibres is associated with the tegument, suckers, gut and reproductive system. Radial fibres around the ventral sucker are thick, branched and extend to the body wall. High-resolution confocal imaging revealed a typical digenean muscular arrangement and important heterophyid morphological traits. These data will support future control measures to reduce the parasitism in guppies reared in fish farming systems, especially for aquarium and experimental purposes. © 2016 John Wiley & Sons Ltd.

  3. A rapid method for combined laser scanning confocal microscopic and electron microscopic visualization of biocytin or neurobiotin-labeled neurons.

    Science.gov (United States)

    Sun, X J; Tolbert, L P; Hildebrand, J G; Meinertzhagen, I A

    1998-02-01

    Intracellular recording and dye filling are widely used to correlate the morphology of a neuron with its physiology. With laser scanning confocal microscopy, the complex shapes of labeled neurons in three dimensions can be reconstructed rapidly, but this requires fluorescent dyes. These dyes are neither permanent nor electron dense and therefore do not allow investigation by electron microscopy. Here we report a technique that quickly and easily converts a fluorescent label into a more stable and electron-dense stain. With this technique, a neuron is filled with Neurobiotin or biocytin, reacted with fluorophore-conjugated avidin, and imaged by confocal microscopy. To permit long-term storage or EM study, the fluorescent label is then converted to a stable electron-dense material by a single-step conversion using a commercially available ABC kit. We find that the method, which apparently relies on recognition of avidin's excess biotin binding sites by the biotin-peroxidase conjugate, is both faster and less labor intensive than photo-oxidation procedures in common use. The technique is readily adaptable to immunocytochemistry with biotinylated probes, as we demonstrate using anti-serotonin as an example.

  4. Evaluation of the presence of Enterococcus Faecalis in root cementum: A confocal laser scanning microscope analysis.

    Science.gov (United States)

    Halkai, Rahul; Hegde, Mithra N; Halkai, Kiran

    2014-03-01

    The aim of this study is to address the cause of persistent infection of root cementum by Enterococcus faecalis. A sample of 60 human single-rooted teeth were divided into three groups. Group I (control group) had no access opening and one-third of the apical root cementum was sealed using varnish. Group II had no preparation of teeth samples. In group III, apical root cementum was exposed to organic acid and roughened using diamond point to mimic apical resorption. After access opening in groups II and III, all teeth samples were sterilized using gamma irradiation (25 kGy). E. faecalis broth was placed in the root canal and apical one-third of the tooth was immersed in the broth for 8 weeks with alternate day refreshment followed by biomechanical preparation, obturation and coronal seal. Apical one-third of all teeth samples were again immersed in the broth for 8 weeks with alternate day refreshment to mimic secondary infection. The samples were observed under a confocal microscope after splitting the teeth into two halves. E. faecalis penetrated 160 μm deep into the root cementum in group III samples and only showed adhesion in group II samples. Penetration and survival of E. faecalis deep inside the cementum in extreme conditions could be the reason for persistent infection.

  5. Demons registration for in vivo and deformable laser scanning confocal endomicroscopy

    Science.gov (United States)

    Chiew, Wei Ming; Lin, Feng; Seah, Hock Soon

    2017-09-01

    A critical effect found in noninvasive in vivo endomicroscopic imaging modalities is image distortions due to sporadic movement exhibited by living organisms. In three-dimensional confocal imaging, this effect results in a dataset that is tilted across deeper slices. Apart from that, the sequential flow of the imaging-processing pipeline restricts real-time adjustments due to the unavailability of information obtainable only from subsequent stages. To solve these problems, we propose an approach to render Demons-registered datasets as they are being captured, focusing on the coupling between registration and visualization. To improve the acquisition process, we also propose a real-time visual analytics tool, which complements the imaging pipeline and the Demons registration pipeline with useful visual indicators to provide real-time feedback for immediate adjustments. We highlight the problem of deformation within the visualization pipeline for object-ordered and image-ordered rendering. Visualizations of critical information including registration forces and partial renderings of the captured data are also presented in the analytics system. We demonstrate the advantages of the algorithmic design through experimental results with both synthetically deformed datasets and actual in vivo, time-lapse tissue datasets expressing natural deformations. Remarkably, this algorithm design is for embedded implementation in intelligent biomedical imaging instrumentation with customizable circuitry.

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

    Energy Technology Data Exchange (ETDEWEB)

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

    2005-08-07

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

  7. Characterization by Confocal Laser Scanning Microscopy of the Phase Composition at Interfaces in Thick Films of Polymer Blends

    Directory of Open Access Journals (Sweden)

    Sandro Lattante

    2014-01-01

    Full Text Available Confocal Laser Scanning Microscopy (CLSM has been used as a fast, user-friendly, and noninvasive tool for characterizing the phase composition differences at the substrate and air interfaces in thick films of polymer blends. A clearly different phase composition at the blend/glass interface and at the blend/air interface has been detected. We show that PCBM preferentially accumulates at the glass/blend interface, while P3HT preferentially accumulates at the blend/air interface, by comparing the integrated signal intensity of the luminescence coming from both interfaces. Our results demonstrate that CLSM can be used conveniently for the fast identification of a preferential phase segregation at interfaces in polymer blends. This is useful in the research field on devices (like sensors or planar waveguides that are based on very thick layers (thickness higher than 1 μm.

  8. Morphological confocal laser scanning microscope evaluation of four different "etch and rinse" adhesives in post endodontic restoration.

    Science.gov (United States)

    Marigo, Luca; Lajolo, Carlo; Castagnola, Raffaella; Angerame, Daniele; Somma, Francesco

    2012-01-01

    The aim of this study was to perform a morphometric evaluation of the resin-root canal dentine interface of four "etch and rinse" adhesive systems. Forty human teeth were restored through a fiber post insertion using: Group 1: XP Bond adhesive; Group 2: Prime & Bond NT; Group 3: Surgi Prime Bond adhesive; Group 4: Scotchbond Multi-Purpose Plus. Two cross-sections (apical and coronal) were obtained and analyzed in morphological confocal laser scanning microscope. The numbers of resin tags (RT) and the thickness of the hybrid layer (HL) were evaluated. Quantitative variables were tested by a Shapiro-Wilk test. Parametric variables by an Anova test. Group 4 produced the thickest HL and Group 1 showed the thinnest. Considering RT, Group 1 produced the highest density, while Group 3 the lowest density. All the adhesives produced good results in terms of HL thickness and RT density.

  9. Penetration of tamoxifen citrate loaded ethosomes and liposomes across human skin: a comparative study with confocal laser scanning microscopy.

    Science.gov (United States)

    Sarwa, Khomendra K; Suresh, Preeti K; Rudrapal, Mithun; Verma, Vinod K

    2014-01-01

    In the present study, ethosomal and liposomal formulations containing tamoxifen citrate were prepared and evaluated for their penetration properties in human cadaver skin using Franz diffusion cell and confocal laser scanning microscope (CLSM). The results clearly revealed that ethosomal vesicles showed a better drug permeation profile than that of liposomal vesicles. In addition, low fluorescence intensity in CLSM was recorded with liposomes as compared to ethosomes, indicating lower cumulative amount of drug permeation from liposomal vesicles. Furthermore, CLSM showed uniform fluorescence intensity across the entire depth of skin in ethosomal treatment, indicating high penetrability of ethosomal vesicles through human cadaver skin. In contrast, low penetrability of conventional liposomal vesicles was recorded as penetration was limited to the 7(th) section (i.e. upper epidermis layer) of skin as evident from visualization of intact liposomal vesicles in CLSM.

  10. Extracting the ridge set as a graph for actin filament length estimation from confocal laser scanning microscopic images

    Science.gov (United States)

    Birkholz, Harald

    2012-04-01

    The progress in image acquisition techniques provides life sciences with an abundance of data. Image analysis facilitates the assessment. The actin cytoskeleton plays a crucial role in understanding the behavior of osteoblastic cells on biomaterials. In the flat basal part of the cells, it can be visualized by confocal laser scanning microscopy. In the microscopic images, the stained cytoskeleton appears as a dense network of bright ridges which is so far only qualitatively assessed. For its quantification, there is a need for ridge detection techniques that provide a geometrical description of this graph feature. The state of the art methods do not cope with the systematical degradation by noise, unspecific luminance, and uneven dye uptake. This work presents the key part of a ridge-tracking technique, which makes more efficient use of context information, and evaluate it by its length measurement accuracy. Two random models illustrate the performance against ground truth. Representative microscopic images confirm the applicability.

  11. Multimodal backside imaging of a microcontroller using confocal laser scanning and optical-beam-induced current imaging

    Science.gov (United States)

    Finkeldey, Markus; Göring, Lena; Schellenberg, Falk; Brenner, Carsten; Gerhardt, Nils C.; Hofmann, Martin

    2017-02-01

    Microscopy imaging with a single technology is usually restricted to a single contrast mechanism. Multimodal imaging is a promising technique to improve the structural information that could be obtained about a device under test (DUT). Due to the different contrast mechanisms of laser scanning microscopy (LSM), confocal laser scanning microscopy (CLSM) and optical beam induced current microscopy (OBICM), a combination could improve the detection of structures in integrated circuits (ICs) and helps to reveal their layout. While OBIC imaging is sensitive to the changes between differently doped areas and to semiconductor-metal transitions, CLSM imaging is mostly sensitive to changes in absorption and reflection. In this work we present the implementation of OBIC imaging into a CLSM. We show first results using industry standard Atmel microcontrollers (MCUs) with a feature size of about 250nm as DUTs. Analyzing these types of microcontrollers helps to improve in the field of side-channel attacks to find hardware Trojans, possible spots for laser fault attacks and for reverse engineering. For the experimental results the DUT is placed on a custom circuit board that allows us to measure the current while imaging it in our in-house built stage scanning microscope using a near infrared (NIR) laser diode as light source. The DUT is thinned and polished, allowing backside imaging through the Si-substrate. We demonstrate the possibilities using this optical setup by evaluating OBIC, LSM and CLSM images above and below the threshold of the laser source.

  12. A sensitive and versatile laser scanning confocal optical microscope for single-molecule fluorescence at 77 K.

    Science.gov (United States)

    Hirschfeld, V; Hübner, C G

    2010-11-01

    We developed a cryostat suitable for a laser scanning confocal microscope which allows for a short working distance and thus the usage of an objective with a high numerical aperture ensuring high collection efficiency. The in situ preparation of a thin layer of amorphous water is realized in a part of the cryostat, a Dewar vessel, which is put onto a custom-made, liquid-nitrogen immersed spin-coater. First tests on the setup are performed on a perylenemonoimide/polymethyl methacrylate model system using a standard oil objective and a dry objective at ambient temperature as well as a dry objective at liquid nitrogen temperature (77 K). Fluorescence resonance energy transfer (FRET) measurements on doubly labeled, freeze-quenched polyproline chains show the applicability of the new method on biomolecules. The alternating laser excitation (ALEX) is modified to a line-scanning process (slow ALEX) to optimize the sorting of the labeled molecules. Photophysics and photochemistry at liquid nitrogen temperature are investigated.

  13. A new approach for the spatially resolved qualitative analysis of the protein distribution in hydrogel beads based on confocal laser scanning microscopy

    NARCIS (Netherlands)

    Heinemann, Matthias; Wagner, Thomas; Doumèche, Bastien; Ansorge-Schumacher, Marion; Büchs, Jochen

    2002-01-01

    To investigate the spatial distribution of white egg albumin (WEA) in alginate beads, a new method based on confocal laser scanning microscopy (CLSM) was developed. In contrast to the existing CLSM methods, misleading conclusions are prevented with the application of the new method which does not

  14.   In situ identification of streptococci and other bacteria in initial dental biofilm by confocal laser scanning microscopy and fluorescence in situ hybridization

    DEFF Research Database (Denmark)

    Dige, Irene; Kilian, Mogens; Nilsson, Holger

    2007-01-01

    Confocal laser scanning microscopy (CLSM) has been employed as a method for studying intact natural biofilm. When combined with fluorescence in situ hybridization (FISH) it is possible to analyze spatial relationships and changes of specific members of microbial populations over time. The aim...

  15. Direct In Situ Viability Assessment of Bacteria in Probiotic Dairy Products Using Viability Staining in Conjunction with Confocal Scanning Laser Microscopy

    Science.gov (United States)

    Auty, M. A. E.; Gardiner, G. E.; McBrearty, S. J.; O'Sullivan, E. O.; Mulvihill, D. M.; Collins, J. K.; Fitzgerald, G. F.; Stanton, C.; Ross, R. P.

    2001-01-01

    The viability of the human probiotic strains Lactobacillus paracasei NFBC 338 and Bifidobacterium sp. strain UCC 35612 in reconstituted skim milk was assessed by confocal scanning laser microscopy using the LIVE/DEAD BacLight viability stain. The technique was rapid (diluent. PMID:11133474

  16. Evaluation of Enterococcus faecalis adhesion, penetration, and method to prevent the penetration of Enterococcus faecalis into root cementum: Confocal laser scanning microscope and scanning electron microscope analysis.

    Science.gov (United States)

    Halkai, Rahul S; Hegde, Mithra N; Halkai, Kiran R

    2016-01-01

    To ascertain the role of Enterococcus faecalis in persistent infection and a possible method to prevent the penetration of E. faecalis into root cementum. One hundred and twenty human single-rooted extracted teeth divided into five groups. Group I (control): intact teeth, Group II: no apical treatment done, Group III divided into two subgroups. In Groups IIIa and IIIb, root apex treated with lactic acid of acidic and neutral pH, respectively. Group IV: apical root cementum exposed to lactic acid and roughened to mimic the apical resorption. Group V: apical treatment done same as Group IV and root-end filling done using mineral trioxide aggregate (MTA). Apical one-third of all samples immersed in E. faecalis broth for 8 weeks followed by bone morphogenetic protein and obturation and again immersed into broth for 8 weeks. Teeth split into two halves and observed under confocal laser scanning microscope and scanning electron microscope, organism identified by culture and polymerase chain reaction techniques. Adhesion and penetration was observed in Group IIIa and Group IV. Only adhesion in Group II and IIIB and no adhesion and penetration in Group I and V. Adhesion and penetration of E. faecalis into root cementum providing a long-term nidus for subsequent infection are the possible reason for persistent infection and root-end filling with MTA prevents the adhesion and penetration.

  17. Evaluation of Enterococcus faecalis adhesion, penetration, and method to prevent the penetration of Enterococcus faecalis into root cementum: Confocal laser scanning microscope and scanning electron microscope analysis

    Science.gov (United States)

    Halkai, Rahul S.; Hegde, Mithra N.; Halkai, Kiran R.

    2016-01-01

    Aim: To ascertain the role of Enterococcus faecalis in persistent infection and a possible method to prevent the penetration of E. faecalis into root cementum. Methodology: One hundred and twenty human single-rooted extracted teeth divided into five groups. Group I (control): intact teeth, Group II: no apical treatment done, Group III divided into two subgroups. In Groups IIIa and IIIb, root apex treated with lactic acid of acidic and neutral pH, respectively. Group IV: apical root cementum exposed to lactic acid and roughened to mimic the apical resorption. Group V: apical treatment done same as Group IV and root-end filling done using mineral trioxide aggregate (MTA). Apical one-third of all samples immersed in E. faecalis broth for 8 weeks followed by bone morphogenetic protein and obturation and again immersed into broth for 8 weeks. Teeth split into two halves and observed under confocal laser scanning microscope and scanning electron microscope, organism identified by culture and polymerase chain reaction techniques. Results: Adhesion and penetration was observed in Group IIIa and Group IV. Only adhesion in Group II and IIIB and no adhesion and penetration in Group I and V. Conclusion: Adhesion and penetration of E. faecalis into root cementum providing a long-term nidus for subsequent infection are the possible reason for persistent infection and root-end filling with MTA prevents the adhesion and penetration. PMID:27994316

  18. Measuring ultra-sonic in-plane vibrations with the scanning confocal heterodyne interferometer

    Science.gov (United States)

    Rembe, C.; Ur-Rehman, F.; Heimes, F.; Boedecker, S.; Dräbenstedt, A.

    2010-05-01

    The advanced progress in miniaturization technologies of mechanical systems and structures has led to a growing demand of measurement tools for three-dimensional vibrations at ultra-high frequencies. Particularly radio-frequency, micro-electro-mechanical (RF-MEM) technology is a planar technology and, thus, the resonating structures are much larger in lateral dimensions compared to the height. Consequently, most ultra-high-frequency devices have larger inplane vibration amplitudes than out-of-plane amplitudes. Recently, we have presented a heterodyne interferometer for vibration frequencies up to 1.2 GHz. In this paper we demonstrate a new method to extract broad-bandwidth spectra of in-plane vibrations with our new heterodyne interferometer. To accomplish this goal we have combined heterodyne interferometry, scanning vibrometry, edge-knife technique, amplitude demodulation, and digital-image processing. With our experimental setup we can realize in-plane vibration measurements up to 600 MHz. We will also show our first measurements of a broad-bandwidth, in-plane vibration around 200 MHz. Our in-plane and out-of-plane vibration measurements are phase-correlated and, therefore, our technique is suitable for broad-bandwidth, full-3D vibration measurements of ultrasonic microdevices.

  19. Porosity of natural stone and use of confocal laser scanning microscopy on calcitic marble aged in laboratory

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    Ana Mladenovič

    2008-06-01

    Full Text Available Porosity is one of the key characteristics of natural stone, which influences ondurability as well as functionality of stone as building material. Further, deterioration processes themselves are also characterized by change of porosity. Different direct and indirect techniques can be used for porosity determination. In the following paper overview of these methods, as well as their advantages and disadvantages, is given. Confocal laser scanning microscopy (CLSM is indirect (microscopic technique. Despite its numerous advantages, among which 3D visualizationof pore structure is of major importance, this technique is less known in the area of building materials. An example how CLSM can be applied for qualitative and quantitative evaluation of porosity of calcitic polygonal granoblastic marble is given in this paper. Studied marble has been, despite of its poor durability, often used as building material, especially in the case of claddings. It is shown that thermal hydric factors of deterioration can influence porosity significantly,especially formation of intergranular cracks.This kind of deterioration can be successfully evaluated with use of CLSM method, if samples are suitable prepared and if suitable image analysis tools are developed.

  20. In Vitro Assessment of Enamel Permeability in Primary Teeth with and without Early Childhood Caries Using Laser Scanning Confocal Microscope.

    Science.gov (United States)

    Niviethitha, Sabarinathan; Muthu, Murugan Satta; Kavitha, Swaminathan

    To compare the permeability of the enamel of primary teeth from individuals free of Early Childhood Caries (ECC) with that from individuals affected with ECC by assessment of dye penetration using Laser Scanning Confocal Microscope (LSCM). Experimental in vitro study. Exfoliated primary maxillary anterior teeth (n = 44) were collected and divided into two groups (n=22 per group): samples with ECC (Group 1) and without ECC (Group 2). The samples were immersed in Rhodamine B dye solution for 1 day, cut longitudinally into 3 sections, observed using LSCM. Dye penetration depths in the incisal, middle, cervical thirds and on labial, lingual surfaces were recorded. Data were analyzed by the Mann-Whitney test (α = 5%, p < .005). The overall mean penetration depth for group 1 (100.6 μm ± 58.48 μm) was significantly higher than that of group 2 (31.55 μm ± 23.40 μm, p < .000). Mean penetration depth in the incisal, middle, and cervical thirds and on the labial and lingual surfaces of group 1 also presented significantly higher scores than in group 2 (p < .005). There was significantly more dye penetration in the ECC group than in the non-ECC group. This could be related to a higher level of enamel permeability in teeth affected with ECC.

  1. Precision automation of cell type classification and sub-cellular fluorescence quantification from laser scanning confocal images

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    Hardy Craig Hall

    2016-02-01

    Full Text Available While novel whole-plant phenotyping technologies have been successfully implemented into functional genomics and breeding programs, the potential of automated phenotyping with cellular resolution is largely unexploited. Laser scanning confocal microscopy has the potential to close this gap by providing spatially highly resolved images containing anatomic as well as chemical information on a subcellular basis. However, in the absence of automated methods, the assessment of the spatial patterns and abundance of fluorescent markers with subcellular resolution is still largely qualitative and time-consuming. Recent advances in image acquisition and analysis, coupled with improvements in microprocessor performance, have brought such automated methods within reach, so that information from thousands of cells per image for hundreds of images may be derived in an experimentally convenient time-frame. Here, we present a MATLAB-based analytical pipeline to 1 segment radial plant organs into individual cells, 2 classify cells into cell type categories based upon random forest classification, 3 divide each cell into sub-regions, and 4 quantify fluorescence intensity to a subcellular degree of precision for a separate fluorescence channel. In this research advance, we demonstrate the precision of this analytical process for the relatively complex tissues of Arabidopsis hypocotyls at various stages of development. High speed and robustness make our approach suitable for phenotyping of large collections of stem-like material and other tissue types.

  2. The noninvasive retro-mode imaging of confocal scanning laser ophthalmoscopy in myopic maculopathy: a prospective observational study

    Science.gov (United States)

    Su, Y; Zhang, X; Wu, K; Ji, Y; Zuo, C; Li, M; Wen, F

    2014-01-01

    Purpose To investigate the morphological features of myopic maculopathy with a new and noninvasive retro-mode imaging (RMI) technique using a confocal scanning laser ophthalmoscope. Methods A total of 42 patients (69 eyes) with myopic maculopathy were included. RMI combined with fundus photography, fundus fluorescein angiography, and optical coherence tomography together were used to observe and evaluate the morphological features of disease. Results Four in 4 eyes (100%) with macular retinoschisis were found with a characteristic pattern by RMI (firework pattern centrally with surrounding fingerprint pattern). Twenty-four in 24 eyes (100%) with pigment proliferation were found by RMI as dark plain patches, and 23 in 24 eyes with hemorrhage (95.8%) were found by RMI as gray bump. Atrophy of different degrees (12 in 14 eyes, 85.7%) was found by RMI as an area of pseudo-3D choroidal vessels or a fuzzy shadow but both without a clear boundary. Choroidal neovascularization (12 in 16 eyes, 75%) was identified laboriously by RMI as a vague raised region. Lacquer cracks were difficult to figure out in RMI. Conclusions Retinoschisis, pigment proliferation, hemorrhage, and atrophy secondary to myopic maculopathy have characteristic morphologic features in RMI; however, choroidal neovascularization and lacquer crack are not easily distinguishable in RMI. PMID:24924440

  3. The noninvasive retro-mode imaging of confocal scanning laser ophthalmoscopy in myopic maculopathy: a prospective observational study.

    Science.gov (United States)

    Su, Y; Zhang, X; Wu, K; Ji, Y; Zuo, C; Li, M; Wen, F

    2014-08-01

    To investigate the morphological features of myopic maculopathy with a new and noninvasive retro-mode imaging (RMI) technique using a confocal scanning laser ophthalmoscope. A total of 42 patients (69 eyes) with myopic maculopathy were included. RMI combined with fundus photography, fundus fluorescein angiography, and optical coherence tomography together were used to observe and evaluate the morphological features of disease. Four in 4 eyes (100%) with macular retinoschisis were found with a characteristic pattern by RMI (firework pattern centrally with surrounding fingerprint pattern). Twenty-four in 24 eyes (100%) with pigment proliferation were found by RMI as dark plain patches, and 23 in 24 eyes with hemorrhage (95.8%) were found by RMI as gray bump. Atrophy of different degrees (12 in 14 eyes, 85.7%) was found by RMI as an area of pseudo-3D choroidal vessels or a fuzzy shadow but both without a clear boundary. Choroidal neovascularization (12 in 16 eyes, 75%) was identified laboriously by RMI as a vague raised region. Lacquer cracks were difficult to figure out in RMI. Retinoschisis, pigment proliferation, hemorrhage, and atrophy secondary to myopic maculopathy have characteristic morphologic features in RMI; however, choroidal neovascularization and lacquer crack are not easily distinguishable in RMI.

  4. Evaluation of penetration depth of 2% chlorhexidine digluconate into root dentinal tubules using confocal laser scanning microscope.

    Science.gov (United States)

    Vadhana, Sekar; Latha, Jothi; Velmurugan, Natanasabapathy

    2015-05-01

    This study evaluated the penetration depth of 2% chlorhexidine digluconate (CHX) into root dentinal tubules and the influence of passive ultrasonic irrigation (PUI) using a confocal laser scanning microscope (CLSM). Twenty freshly extracted anterior teeth were decoronated and instrumented using Mtwo rotary files up to size 40, 4% taper. The samples were randomly divided into two groups (n = 10), that is, conventional syringe irrigation (CSI) and PUI. CHX was mixed with Rhodamine B dye and was used as the final irrigant. The teeth were sectioned at coronal, middle and apical levels and viewed under CLSM to record the penetration depth of CHX. The data were statistically analyzed using Kruskal-Wallis and Mann-Whitney U tests. The mean penetration depths of 2% CHX in coronal, middle and apical thirds were 138 µm, 80 µm and 44 µm in CSI group, respectively, whereas the mean penetration depths were 209 µm, 138 µm and 72 µm respectively in PUI group. Statistically significant difference was present between CSI group and PUI group at all three levels (p Penetration depth of 2% CHX into root dentinal tubules is deeper in coronal third when compared to middle and apical third. PUI aided in deeper penetration of 2% CHX into dentinal tubules when compared to conventional syringe irrigation at all three levels.

  5. Comparison of bacterial leakage resistance of various root canal filling materials and methods: Confocal laser-scanning microscope study.

    Science.gov (United States)

    Hwang, Ji Hee; Chung, Jin; Na, Hee-Sam; Park, Eunjoo; Kwak, Sangwon; Kim, Hyeon-Cheol

    2015-01-01

    This study evaluated the bacterial leakage resistance and root canal lining efficacy of various root canal filling materials and methods by using confocal laser-scanning microscope (CLSM). Sixty extracted human premolars with mature apex and single root canal were randomly divided into 2 control groups and 4 experimental groups. Group CW was filled with continuous wave technique using gutta-percha and AH Plus sealer. Group GC was coated with AH-Plus sealer and then obturated with soften GuttaCore. Group GF was obturated using GuttaFlow and gutta-percha. Group EM was filled with EndoSeal MTA and gutta-percha using ultrasonic vibration. The AH-Plus, GuttaFlow, and EndoSeal were labeled with Hoechst 33342 to facilitate fluorescence. The obturated root tip was incubated with Carboxyfluorescein diacetate succinimidyl ester (CFSE)-stained E. faecalis for 14 days. CLSM was performed to evaluate the sealer distribution and bacterial leakage for the apical 1-, 2-, 3-mm specimens. Statistically significant differences were determined by 1-way ANOVA with Tukey's post-hoc test and Pearson's correlation analysis. Group EM showed the better sealer distribution score than the other groups (p  0.05). Under the conditions of this study, different root canal filling materials and methods showed different efficacy for canal distribution and bacterial leakage resistance. © Wiley Periodicals, Inc.

  6. Video-rate two-photon excited fluorescence lifetime imaging system with interleaved digitization.

    Science.gov (United States)

    Dow, Ximeng Y; Sullivan, Shane Z; Muir, Ryan D; Simpson, Garth J

    2015-07-15

    A fast (up to video rate) two-photon excited fluorescence lifetime imaging system based on interleaved digitization is demonstrated. The system is compatible with existing beam-scanning microscopes with minor electronics and software modification. Proof-of-concept demonstrations were performed using laser dyes and biological tissue.

  7. Reflectance confocal microscope for imaging oral tissues in vivo, potentially with line scanning as a low-cost approach for clinical use

    Science.gov (United States)

    Peterson, Gary; Abeytunge, Sanjeewa; Eastman, Zachary; Rajadhyaksha, Milind

    2012-02-01

    Reflectance confocal microscopy with a line scanning approach potentially offers a smaller, simpler and less expensive approach than traditional methods of point scanning for imaging in living tissues. With one moving mechanical element (galvanometric scanner), a linear array detector and off-the-shelf optics, we designed a compact (102x102x76mm) line scanning confocal reflectance microscope (LSCRM) for imaging human tissues in vivo in a clinical setting. Custom-designed electronics, based on field programmable gate array (FPGA) logic has been developed. With 405 nm illumination and a custom objective lens of numerical aperture 0.5, lateral resolution was measured to be 0.8 um (calculated 0.64 um). The calculated optical sectioning is 3.2 um. Preliminary imaging shows nuclear and cellular detail in human skin and oral epithelium in vivo. Blood flow is also visualized in the deeper connective tissue (lamina propria) in oral mucosa. Since a line is confocal only in one dimension (parallel) but not in the other, the detection is more sensitive to multiply scattered out of focus background noise than in the traditional point scanning configuration. Based on the results of our translational studies thus far, a simpler, smaller and lower-cost approach based on a LSCRM appears to be promising for clinical imaging.

  8. Confocal laser scanning microscopic analysis of ectopic sublingual gland-like tissue inside the hamster submandibular gland.

    Science.gov (United States)

    Moriguchi, Keiichi; Utsumi, Michiya; Ohno, Norikazu

    2013-12-01

    Based on its histochemical properties, the secretory portion of the hamster submandibular gland has been classified as seromucous cells. The presence of endogenous peroxidase (PO) reaction was shown in the nuclear envelope, cisternae of endoplasmic reticulum and Golgi apparatus. The 3,3'-diaminobenzidene, tetrahydrochloride (DAB) method revealed bipartite secretory granules containing a PO-positive dense core surrounded by a less dense halo in these cells. In the present investigation, serous and mucous-like cells were found in resin-embedded semi-thin sections of the DAB-reacted hamster submandibular gland. These sections were already on glass slides for routine light microscopic observations, therefore electron microscopic analysis could be unrealizable. We then used reflectance-mode confocal laser scanning microscopy to visualize additional sites of PO activity as detected in these sections. Using this approach, we found mucous cells with PO activity-negative secretory granules and seromucous cells with PO activity-positive spot-like secretory granules of the regular sublingual gland most frequently adjacent to the serous cells with typical electron-dense secretory granules. These cells clearly differ from the seromucous cells with bipartite secretory granules and the granular duct cells with typical electron-dense secretory granules of the hamster submandibular gland. Additionally, secretory endpieces of the ectopic sublingual gland-like tissue empty into the duct of the hamster submandibular gland lobule. Thus, our findings suggest that a mass of sublingual gland tissue extends into the hamster submandibular gland during its development, and PO may be synthesized and secreted into the same duct. Copyright © 2013 Wiley Periodicals, Inc.

  9. Penetrability of AH plus and MTA fillapex after endodontic treatment and retreatment: a confocal laser scanning microscopy study.

    Science.gov (United States)

    Kok, Daniela; Rosa, Ricardo Abreu da; Barreto, Mirela Sangoi; Busanello, Fernanda Hoffmann; Santini, Manuela Favarin; Pereira, Jefferson Ricardo; Só, Marcus Vinícius Reis

    2014-06-01

    The aim of the study was to assess the penetrability of two endodontic sealers (AH Plus and MTA Fillapex) into dentinal tubules, submitted to endodontic treatment and subsequently to endodontic retreatment. Thirty ex vivo incisors were prepared using ProTaper rotary system up to F3 instrument and divided in three groups according to the endodontic sealer used for root canal filling: AH Plus (AHP), MTA Fillapex (MTAF), and control group (CG) without using EDTA previously to the root canal filling. Rhodamine B dye (red) was incorporated to the sealers in order to provide the fluorescence which will enable confocal laser scanning microscopy (CLSM) assessment. All specimens were filled with gutta-percha cones using the lateral compaction technique. The specimens were submitted to endodontic retreatment using ProTaper Retreatment system, re-prepared up to F5 instruments and filled with gutta-percha cones and the same sealer used during endodontic retreatment. Fluorescein dye (green) was incorporated to the sealer in order to distinguish from the first filling. The roots were sectioned 2 mm from the apex and assessed by CLSM. No difference was found between the two experimental groups (P > 0.05). On the other hand, in the control group the sealers were not capable to penetrate into dentinal tubules after endodontic treatment (P > 0.05). In retreatment cases, none of the sealers were able to penetrate into dentin tubules. It can be concluded that sealer penetrability is high during endodontic treatment. However, MTA Fillapex and AH Plus do not penetrate into dentinal tubules after endodontic retreatment. © 2014 Wiley Periodicals, Inc.

  10. Effect of the menstrual cycle on the optic nerve head in diabetes: analysis by confocal scanning laser ophthalmoscopy.

    Science.gov (United States)

    Akar, Munire Erman; Yucel, Iclal; Erdem, Uzeyir; Taskin, Omur; Ozel, Alper; Akar, Yusuf

    2005-04-01

    The purpose of this study was to examine and compare menstrual-cycle-dependent topographic changes in the optic nerve head of normally menstruating women with different grades of type 2 diabetes mellitus. We studied the right eyes of 123 normally menstruating women (36 with severe nonproliferative diabetic retinopathy [NPDR], 42 with mild NPDR and 45 healthy subjects). All subjects underwent a complete ocular examination at baseline. At 4 hormonally distinct phases of the menstrual cycle (early follicular, late follicular, mid-luteal and late luteal), we analysed the topography of the optic nerve head, using a confocal scanning laser ophthalmoscope, and measured the serum levels of estradiol, progesterone and luteinizing hormone. We excluded from analysis the data for 8 patients with severe NPDR, 10 patients with mild NPDR and 15 control subjects who were lost to follow-up examinations during the menstrual cycle. The mean age and optic disc area did not differ significantly among the 3 groups. The duration of diabetes was significantly longer in the patients with severe NPDR than in those with mild NPDR (p cup-shape measure, linear cup/disc ratio, cup/disc area ratio and cup area in the late luteal phase compared with the other phases of the menstrual cycle (p menstrual cycle. Severe NPDR is associated with significant topographic changes in the rim and cup of the optic nerve head during the menstrual cycle. This must be considered in the evaluation of women with both diabetes and glaucoma. The normal fluctuations in serum sex hormone levels during the menstrual cycle of diabetic women seem to affect the optic nerve head more when the disease is advanced.

  11. Analysis of a marine phototrophic biofilm by confocal laser scanning microscopy using the new image quantification software PHLIP

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    Almeida Jonas S

    2006-01-01

    Full Text Available Abstract Background Confocal laser scanning microscopy (CLSM is the method of choice to study interfacial biofilms and acquires time-resolved three-dimensional data of the biofilm structure. CLSM can be used in a multi-channel modus where the different channels map individual biofilm components. This communication presents a novel image quantification tool, PHLIP, for the quantitative analysis of large amounts of multichannel CLSM data in an automated way. PHLIP can be freely downloaded from http://phlip.sourceforge.net. Results PHLIP is an open source public license Matlab toolbox that includes functions for CLSM imaging data handling and ten image analysis operations describing various aspects of biofilm morphology. The use of PHLIP is here demonstrated by a study of the development of a natural marine phototrophic biofilm. It is shown how the examination of the individual biofilm components using the multi-channel capability of PHLIP allowed the description of the dynamic spatial and temporal separation of diatoms, bacteria and organic and inorganic matter during the shift from a bacteria-dominated to a diatom-dominated phototrophic biofilm. Reflection images and weight measurements complementing the PHLIP analyses suggest that a large part of the biofilm mass consisted of inorganic mineral material. Conclusion The presented case study reveals new insight into the temporal development of a phototrophic biofilm where multi-channel imaging allowed to parallel monitor the dynamics of the individual biofilm components over time. This application of PHLIP presents the power of biofilm image analysis by multi-channel CLSM software and demonstrates the importance of PHLIP for the scientific community as a flexible and extendable image analysis platform for automated image processing.

  12. The noninvasive retro-mode imaging modality of confocal scanning laser ophthalmoscopy in polypoidal choroidal vasculopathy: a preliminary application.

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    Renpan Zeng

    Full Text Available PURPOSE: To evaluate the validity of the novel and noninvasive retro-mode imaging modality of confocal scanning laser ophthalmoscopy (cSLO for detecting the morphological features of polypoidal choroidal vasculopathy (PCV. DESIGN: Prospective, observational, consecutive case series. METHODS: Twenty-six patients (29 eyes with PCV were enrolled in this study. All patients underwent comprehensive ophthalmologic examinations and imaging studies, including retro-mode imaging, fundus autofluorescence (FAF, fundus photography, fundus fluorescein angiography (FFA, indocyanine green angiography (ICGA and spectral-domain optical coherence tomography (SD-OCT. We investigated the retro-mode images and compared the results with those of SD-OCT, FFA and ICGA. RESULTS: In the 29 PCV eyes, the retro-mode images clearly revealed polypoidal lesions in 27 (93.1% eyes as well as branching vascular networks in 16 (55.2% eyes. Others findings, including pigment epithelial detachment (PED in 20 (69.0% eyes, neuroretinal detachment (NRD in 3 (10.3% eyes, cystoid macular edema (CME in 3 (10.3% eyes, drusen in 4 (13.8% eyes and minute granular changes of the retinal pigment epithelium (RPE in 12 (41.3% eyes, were also clearly visualized. When we compared the results with those of SD-OCT, FFA and ICGA, there was no significant difference between ICGA and retro-mode imaging for finding polypoidal lesions and (or branching choroidal vascular networks (P>0.05. However, the rate of PED detection was significantly better with retro-mode imaging than with the ICGA (P0.05. The differences were not statistically significant between FFA and retro-mode imaging for detecting PED, NRD, CME (P>0.05. CONCLUSIONS: The novel and noninvasive retro-mode imaging by cSLO is able to clearly visualize the morphological features of PCV.

  13. The simplicity of males: dwarf males of four species of Osedax (Siboglinidae; Annelida) investigated by confocal laser scanning microscopy.

    Science.gov (United States)

    Worsaae, Katrine; Rouse, Greg W

    2010-02-01

    Dwarf males of the bone-eating worms Osedax (Siboglinidae, Annelida) have been proposed to develop from larvae that settle on females rather than on bone. The apparent arrest in somatic development and resemblance of the males to trochophore larvae has been posited as an example of paedomorphosis. Here, we present the first investigation of the entire muscle and nervous system in dwarf males of Osedax frankpressi, O. roseus, O. rubiplumus, and O. "spiral" analyzed by multistaining and confocal laser scanning microscopy. Sperm shape and spermiogenesis, the sperm duct and internal and external ciliary patterns were likewise visualized. The males of all four species possess morphological traits typical of newly settled siboglinid larvae: a prostomium, a peristomium with a prototroch, one elongate segment and a second shorter segment. Each segment has a ring of eight long-handled hooked chaetae. The longitudinal muscles are distributed as evenly spaced strands forming a grid with the thin outer circular muscles. Oblique protractor and retractor muscles are associated with each of the chaetal sacs. The nervous system comprises a cerebral ganglion, a prototroch nerve ring, paired dorsolateral longitudinal nerves, five ventral longitudinal nerves with paired, posterior ganglia and a terminal commissure, as well as a net of fine peripheral transverse plexuses surrounding the first segment. Internal ciliation occurs as paired ventrolateral bands along the first segment. The bands appear to lead the free mature sperm to a ciliated duct and seminal vesicle lying just behind the prototroch region. A duct then runs from the seminal vesicle into the dorsal part of the prostomium. The similarity of Osedax males to the larvae of Osedax and other siboglinid annelids as well as similarities shown here to the neuromuscular organization seen in other annelid larvae supports the hypothesis of paedomorphosis in males of Osedax.

  14. Effects of two desensitizing dentifrices on dentinal tubule occlusion with citric acid challenge: Confocal laser scanning microscopy study

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    Sneha Anil Rajguru

    2017-01-01

    Full Text Available Background: Dentin hypersensitivity results when patent tubules are exposed to pain-inducing external stimuli. Aim: This study aims to compare the effects of two desensitizing dentifrices containing NovaMin and arginine on dentinal tubule occlusion with and without citric acid challenge in vitro using confocal laser scanning microscopy (CLSM. Materials and Methods: Forty dentin discs were randomly divided into Groups I and II containing twenty specimens each, treated with NovaMin and arginine-containing dentifrices, respectively. Groups I and II were divided into subgroups A and B where IA and IIA underwent CLSM analysis to determine the percentage of tubule occlusion while IB and IIB underwent 0.3% citric acid challenge and CLSM analysis. A novel grading system was devised to categorize tubule occlusion. Results: In Group II, the percentage of occluded tubules was highest for IIA (72.25% ± 10.57% and least for IIB (42.55% ± 8.65% having statistical significance (P < 0.0005. In Group I, the difference between IA (49.9% ± 12.96% and IB (43.15% ± 12.43% was statistically insignificant (P = 0.249. On the comparison between IB and IIB statistically indifferent result was obtained (P = 0.901, whereas the difference between IA and IIA was statistically significant (P < 0.001. The results of grading system were for IA 50% of samples belonged to Grade 2, for IIA 60% - Grade 3, and for IB 70% and for IIB 90% - Grade 2. Conclusion: Dentinal tubule occlusion with arginine-containing dentifrice was significantly higher than NovaMin. However, it could not resist citric acid challenge as effectively as NovaMin. The effects of NovaMin were more sustainable as compared to arginine-containing dentifrice, thus proving to be a better desensitizing agent.

  15. Evaluation of baseline structural factors for predicting glaucomatous visual-field progression using optical coherence tomography, scanning laser polarimetry and confocal scanning laser ophthalmoscopy.

    Science.gov (United States)

    Sehi, M; Bhardwaj, N; Chung, Y S; Greenfield, D S

    2012-12-01

    The objective of this study is to assess whether baseline optic nerve head (ONH) topography and retinal nerve fiber layer thickness (RNFLT) are predictive of glaucomatous visual-field progression in glaucoma suspect (GS) and glaucomatous eyes, and to calculate the level of risk associated with each of these parameters. Participants with ≥28 months of follow-up were recruited from the longitudinal Advanced Imaging for Glaucoma Study. All eyes underwent standard automated perimetry (SAP), confocal scanning laser ophthalmoscopy (CSLO), time-domain optical coherence tomography (TDOCT), and scanning laser polarimetry using enhanced corneal compensation (SLPECC) every 6 months. Visual-field progression was assessed using pointwise linear-regression analysis of SAP sensitivity values (progressor) and defined as significant sensitivity loss of >1 dB/year at ≥2 adjacent test locations in the same hemifield at P<0.01. Cox proportional hazard ratios (HR) were calculated to determine the predictive ability of baseline ONH and RNFL parameters for SAP progression using univariate and multivariate models. Seventy-three eyes of 73 patients (43 GS and 30 glaucoma, mean age 63.2±9.5 years) were enrolled (mean follow-up 51.5±11.3 months). Four of 43 GS (9.3%) and 6 of 30 (20%) glaucomatous eyes demonstrated progression. Mean time to progression was 50.8±11.4 months. Using multivariate models, abnormal CSLO temporal-inferior Moorfields classification (HR=3.76, 95% confidence interval (CI): 1.02-6.80, P=0.04), SLPECC inferior RNFLT (per -1 μm, HR=1.38, 95% CI: 1.02-2.2, P=0.02), and TDOCT inferior RNFLT (per -1 μm, HR=1.11, 95% CI: 1.04-1.2, P=0.001) had significant HRs for SAP progression. Abnormal baseline ONH topography and reduced inferior RNFL are predictive of SAP progression in GS and glaucomatous eyes.

  16. Spectral optical coherence tomography in video-rate and 3D imaging of contact lens wear.

    Science.gov (United States)

    Kaluzny, Bartlomiej J; Fojt, Wojciech; Szkulmowska, Anna; Bajraszewski, Tomasz; Wojtkowski, Maciej; Kowalczyk, Andrzej

    2007-12-01

    To present the applicability of spectral optical coherence tomography (SOCT) for video-rate and three-dimensional imaging of a contact lens on the eye surface. The SOCT prototype instrument constructed at Nicolaus Copernicus University (Torun, Poland) is based on Fourier domain detection, which enables high sensitivity (96 dB) and increases the speed of imaging 60 times compared with conventional optical coherence tomography techniques. Consequently, video-rate imaging and three-dimensional reconstructions can be achieved, preserving the high quality of the image. The instrument operates under clinical conditions in the Ophthalmology Department (Collegium Medicum Nicolaus Copernicus University, Bydgoszcz, Poland). A total of three eyes fitted with different contact lenses were examined with the aid of the instrument. Before SOCT measurements, slit lamp examinations were performed. Data, which are representative for each imaging mode, are presented. The instrument provided high-resolution (4 microm axial x 10 microm transverse) tomograms with an acquisition time of 40 micros per A-scan. Video-rate imaging allowed the simultaneous quantitative evaluation of the movement of the contact lens and assessment of the fitting relationship between the lens and the ocular surface. Three-dimensional scanning protocols further improved lens visualization and fit evaluation. SOCT allows video-rate and three-dimensional cross-sectional imaging of the eye fitted with a contact lens. The analysis of both imaging modes suggests the future applicability of this technology to the contact lens field.

  17. Distribution of biomolecules in porous nitrocellulose membrane pads using confocal laser scanning microscopy and high-speed cameras.

    Science.gov (United States)

    Mujawar, Liyakat Hamid; Maan, Abid Aslam; Khan, Muhammad Kashif Iqbal; Norde, Willem; van Amerongen, Aart

    2013-04-02

    The main focus of our research was to study the distribution of inkjet printed biomolecules in porous nitrocellulose membrane pads of different brands. We produced microarrays of fluorophore-labeled IgG and bovine serum albumin (BSA) on FAST, Unisart, and Oncyte-Avid slides and compared the spot morphology of the inkjet printed biomolecules. The distribution of these biomolecules within the spot embedded in the nitrocellulose membrane was analyzed by confocal laser scanning microscopy in the "Z" stack mode. By applying a "concentric ring" format, the distribution profile of the fluorescence intensity in each horizontal slice was measured and represented in a graphical color-coded way. Furthermore, a one-step diagnostic antibody assay was performed with a primary antibody, double-labeled amplicons, and fluorophore-labeled streptavidin in order to study the functionality and distribution of the immune complex in the nitrocellulose membrane slides. Under the conditions applied, the spot morphology and distribution of the primary labeled biomolecules was nonhomogenous and doughnut-like on the FAST and Unisart nitrocellulose slides, whereas a better spot morphology with more homogeneously distributed biomolecules was observed on the Oncyte-Avid slide. Similar morphologies and distribution patterns were observed when the diagnostic one-step nucleic acid microarray immunoassay was performed on these nitrocellulose slides. We also investigated possible reasons for the differences in the observed spot morphology by monitoring the dynamic behavior of a liquid droplet on and in these nitrocellulose slides. Using high speed cameras, we analyzed the wettability and fluid flow dynamics of a droplet on the various nitrocellulose substrates. The spreading of the liquid droplet was comparable for the FAST and Unisart slides but different, i.e., slower, for the Oncyte-Avid slide. The results of the spreading of the droplet and the penetration behavior of the liquid in the

  18. 3D digital image processing for biofilm quantification from confocal laser scanning microscopy: Multidimensional statistical analysis of biofilm modeling

    Science.gov (United States)

    Zielinski, Jerzy S.

    The dramatic increase in number and volume of digital images produced in medical diagnostics, and the escalating demand for rapid access to these relevant medical data, along with the need for interpretation and retrieval has become of paramount importance to a modern healthcare system. Therefore, there is an ever growing need for processed, interpreted and saved images of various types. Due to the high cost and unreliability of human-dependent image analysis, it is necessary to develop an automated method for feature extraction, using sophisticated mathematical algorithms and reasoning. This work is focused on digital image signal processing of biological and biomedical data in one- two- and three-dimensional space. Methods and algorithms presented in this work were used to acquire data from genomic sequences, breast cancer, and biofilm images. One-dimensional analysis was applied to DNA sequences which were presented as a non-stationary sequence and modeled by a time-dependent autoregressive moving average (TD-ARMA) model. Two-dimensional analyses used 2D-ARMA model and applied it to detect breast cancer from x-ray mammograms or ultrasound images. Three-dimensional detection and classification techniques were applied to biofilm images acquired using confocal laser scanning microscopy. Modern medical images are geometrically arranged arrays of data. The broadening scope of imaging as a way to organize our observations of the biophysical world has led to a dramatic increase in our ability to apply new processing techniques and to combine multiple channels of data into sophisticated and complex mathematical models of physiological function and dysfunction. With explosion of the amount of data produced in a field of biomedicine, it is crucial to be able to construct accurate mathematical models of the data at hand. Two main purposes of signal modeling are: data size conservation and parameter extraction. Specifically, in biomedical imaging we have four key problems

  19. Dynamic behavior of binary component ion-exchange displacement chromatography of proteins visualized by confocal laser scanning microscopy.

    Science.gov (United States)

    Shi, Qing-Hong; Shi, Zhi-Cong; Sun, Yan

    2012-09-28

    Confocal laser scanning microscopy (CLSM) was introduced to visualize particle-scale binary component protein displacement behavior in Q Sepharose HP column. To this end, displacement chromatography of two intrinsic fluorescent proteins, enhanced green fluorescent protein (eGFP) and red fluorescent protein (RFP), were developed using sodium saccharin (NaSac) as a displacer. The results indicated that RFP as well as eGFP could be effectively displaced in the single-component experiments by 50 mmol/L NaSac at 120 and 140 mmol/L NaCl whereas a fully developed displacement train with eGFP and RFP was only observed at 120 mmol/L NaCl in binary component displacement. At 140 mmol/L NaCl, there was a serious overlapping of the zones of the two proteins, indicating the importance of induced-salt effect on the formation of an isotachic displacement train. CLSM provided particle-scale evidence that induced-salt effect occurred likewise in the interior of an adsorbent and was synchronous to the introduction of the displacer. CLSM results at 140 mmol/L NaCl also demonstrated that both the proteins had the same fading rate at 50 mmol/L NaSac in the initial stage, suggesting the same displacement ability of NaSac to both the proteins. In the final stage, the fading rate of RFP in the adsorbent became slow, particularly at lower displacer concentrations. In the binary component displacement, the two proteins exhibited distinct fading rates as compared to the single component displacement and the remarkable lagging of the fading rate was observed in protein displacements. It suggested that the co-adsorbed proteins had significant influence on the formation of an isotachic train and the displacement chromatography of the proteins. Therefore, this research provided particle-scale insight into the dynamic behavior and complexity in the displacement of proteins. Copyright © 2012 Elsevier B.V. All rights reserved.

  20. In vivo laser scanning confocal microscopy of the cornea in patients with silicone oil tamponade after vitreoretinal surgery.

    Science.gov (United States)

    Le, Qihua; Wang, Xin; Lv, Jiahua; Sun, Xinghuai; Xu, Jianjiang

    2012-08-01

    To evaluate the morphological changes in the cornea by in vivo laser scanning confocal microscopy (LSCM) in a large case series with silicone oil endotamponade after vitreoretinal surgery and to explore the value of LSCM in the early detection of silicone keratopathy (SK). Ninety-nine patients (99 eyes) with silicone oil endotamponade after vitreoretinal surgery were included in the current study. Slit-lamp examination and measurement of intraocular pressure (IOP) were performed first. Then the central corneas of the subjects' eyes were examined by in vivo LSCM. The analysis of images of each corneal layer was performed and the endothelial cellular density (ECD), endothelial cellular area (ECA), coefficient of variation of cell size (CoV), and percentage of hexagonal cells (PHC) were measured. Moreover, the total size of stromal deposits was measured, and the correlation between the size of deposits and the parameters of endothelial cells was analyzed. Clinically recognizable abnormalities involving the cornea were identified in only 12 eyes (12.1%) under slit-lamp biomicroscopy, whereas in vivo LSCM revealed morphological abnormalities in 40 eyes (40.4%). The manifestations of endothelial lesions varied from decreased cellular density, increased polymegathism and pleomorphism to hyperreflective silicone oil membrane or droplets adhering to the endothelium. Moreover, hyperreflective deposits with various shapes could be identified in both posterior and anterior stroma, along with the infiltration of Langerhans cells beneath the epithelium. The average ECD and PHC of eyes with corneal abnormalities were significantly lower than those of normal corneas, whereas the average ECA and CoV were significantly larger (all Ps < 0.001). The patients with corneal abnormalities were significantly older than those others (P = 0.003). The rate of pseudophakic and aphakic eyes having corneal abnormalities was significantly higher than that of phakic eyes (P = 0.045). Interestingly

  1. Serial Sectioning Of Cells In Three Dimensions With Confocal Scanning Laser Fluorescence Microscopy (Fl-CSLM): Microtomoscopy

    Science.gov (United States)

    Stelzer, Ernst H.; Stricker, Reiner; Pick, Reinhard; Storz, Clemens; Wijnaendts-Van-Resandt, Roelof W.

    1988-06-01

    The discrimination of out of focus contributions in fluorescence microscopy possible in a confocal setup will establish itself as a supplement to conventional fluorescence microscopy. The improvement of the contrast compared with conventional fluorescence microscopy depends mainly on the density of the fluorescing material and the thickness of the sample. The term thickness, that which microscopists refer to as the size of the specimen along the optical axis, will gain a new quality since a confocal fluorescence microscope may reveal totally different features when recording data in planes that are 0.3μm apart. Differences that have in the past been neglected suddenly become important. The following article will outline important features in the application of confocal fluorescence microscopy in the biological sciences, point out its limitatk'ns, and draw attention to expected developments.

  2. In-vivo diagnosis and non-inasive monitoring of Imiquimod 5% cream for non-melanoma skin cancer using confocal laser scanning microscopy

    Science.gov (United States)

    Dietterle, S.; Lademann, J.; Röwert-Huber, H.-J.; Stockfleth, E.; Antoniou, C.; Sterry, W.; Astner, S.

    2008-10-01

    Basal cell carcinoma (BCC) is the most common cutaneous malignancy with increasing incidence rates worldwide. A number of established treatments are available, including surgical excision. The emergence of new non-invasive treatment modalities has prompted the development of non-invasive optical devices for therapeutic monitoring and evaluating treatment efficacy. This study was aimed to evaluate the clinical applicability of a fluorescence confocal laser scanning microscope (CFLSM) for non-invasive therapeutic monitoring of basal cell carcinoma treated with Imiquimod (Aldara®) as topical immune-response modifier. Eight participants with a diagnosis of basal cell carcinoma (BCC) were enrolled in this investigation. Sequential evaluation during treatment with Imiquimod showed progressive normalization of the confocal histomorphologic parameters in correlation with normal skin. Confocal laser scanning microscopy was able to identify characteristic features of BCC and allowed the visualization of therapeutic effects over time. Thus our results indicate the clinical applicability of CFLSM imaging to evaluate treatment efficacy in vivo and non-invasively.

  3. The effect of compression on clinical diagnosis of glaucoma based on non-analyzed confocal scanning laser ophthalmoscopy images

    NARCIS (Netherlands)

    Abramoff, M.D.

    2006-01-01

    Knowledge of the effect of compression of ophthalmic images on diagnostic reading is essential for effective tele-ophthalmology applications. It was therefore with great anticipation that I read the article “The Effect of Compression on Clinical Diagnosis of Glaucoma Based on Non-analyzed Confocal

  4. In situ microspatial imaging using two-photon and confocal laser scanning microscopy of bacteria and extracellular polymeric secretions (EPS) within marine stromatolites.

    Science.gov (United States)

    Kawaguchi, Tomohiro; Decho, Alan W

    2002-03-01

    The combination of a hydrophilic embedding resin, Nanoplast, with fluorescent probes, and subsequent imaging using two-photon and confocal laser scanning microscopy (2P-LSM and CLSM) has allowed in imaging of the in situ microspatial arrangements of microbial cells and their extracellular polymeric secretion (EPS) within marine stromatolites. Optical sectioning by 2P-LSM and CLSM allowed imaging of endolithic cyanobacteria cells, Solentia sp., seen within carbonate sand grains. 2P-LSM allowed very clear imaging with a high resolution of bacteria using DAPI, which normally require UV excitation and reduced photo-bleaching of fluorescent probes.

  5. Time-Lapse Förster Resonance Energy Transfer Imaging by Confocal Laser Scanning Microscopy for Analyzing Dynamic Molecular Interactions in the Plasma Membrane of B Cells.

    Science.gov (United States)

    Sohn, Hae Won; Brzostowski, Joseph

    2018-01-01

    For decades, various Förster resonance energy transfer (FRET) techniques have been developed to measure the distance between interacting molecules. FRET imaging by the sensitized acceptor emission method has been widely applied to study the dynamical association between two molecules at a nanometer scale in live cells. Here, we provide a detailed protocol for FRET imaging by sensitized emission using a confocal laser scanning microscope to analyze the interaction of the B cell receptor (BCR) with the Lyn-enriched lipid microdomain on the plasma membrane of live cells upon antigen binding, one of the earliest signaling events in BCR-mediated B cell activation.

  6. Noninvasive redox and back-scattered light imaging of keratocyte cells in the cornea: two-photon excitation and scanning slit confocal microscopy

    Science.gov (United States)

    Masters, Barry R.

    1995-04-01

    The ability to image and monitor the metabolic activity of keratocytes is important for the investigation of wound healing and repair mechanisms in the cornea. After laser refractive surgery there is activation of the stromal keratocytes in the human cornea. Two-photon excitation laser scanning microscopy was used to monitor the NAD(P)H levels in keratocytes in the cornea. The autofluorescence was confirmed to be mostly of NAD(P)H origin by treatment with cyanide which caused an increase in the fluorescence by a factor of two. We used a real-time scanning slit confocal microscope to image the distribution of keratocytes in the full thickness of the cornea. This microscope has the ability to image the cellular processes as well as the nuclei of the stromal keratocytes. Noninvasive optical imaging may provide a useful tool to investigate keratocyte activation after laser surgery or wound healing.

  7. Sealing ability of three root-end filling materials prepared using an erbium: Yttrium aluminium garnet laser and endosonic tip evaluated by confocal laser scanning microscopy

    Science.gov (United States)

    Nanjappa, A Salin; Ponnappa, KC; Nanjamma, KK; Ponappa, MC; Girish, Sabari; Nitin, Anita

    2015-01-01

    Aims: (1) To compare the sealing ability of mineral trioxide aggregate (MTA), Biodentine, and Chitra-calcium phosphate cement (CPC) when used as root-end filling, evaluated under confocal laser scanning microscope using Rhodamine B dye. (2) To evaluate effect of ultrasonic retroprep tip and an erbium:yttrium aluminium garnet (Er:YAG) laser on the integrity of three different root-end filling materials. Materials and Methods: The root canals of 80 extracted teeth were instrumented and obturated with gutta-percha. The apical 3 mm of each tooth was resected and 3 mm root-end preparation was made using ultrasonic tip (n = 30) and Er:YAG laser (n = 30). MTA, Biodentine, and Chitra-CPC were used to restore 10 teeth each. The samples were coated with varnish and after drying, they were immersed in Rhodamine B dye for 24 h. The teeth were then rinsed, sectioned longitudinally, and observed under confocal laser scanning microscope. Statistical Analysis Used: Data were analyzed using one-way analysis of variance (ANOVA) and a post-hoc Tukey's test at P ultrasonics, the difference was found to be statistically significant (P ultrasonics. PMID:26180420

  8. Real-time demonstration of split skin graft inosculation and integra dermal matrix neovascularization using confocal laser scanning microscopy.

    Science.gov (United States)

    Greenwood, John; Amjadi, Mahyar; Dearman, Bronwyn; Mackie, Ian

    2009-08-20

    During the first 48 hours after placement, an autograft "drinks" nutrients and dissolved oxygen from fluid exuding from the underlying recipient bed ("plasmatic imbibition"). The theory of inosculation (that skin grafts subsequently obtain nourishment via blood vessel "anastomosis" between new vessels invading from the wound bed and existing graft vessels) was hotly debated from the late 19th to mid-20th century. This study aimed to noninvasively observe blood flow in split skin grafts and Integra dermal regeneration matrix to provide further proof of inosculation and to contrast the structure of vascularization in both materials, reflecting mechanism. Observations were made both clinically and using confocal microscopy on normal skin, split skin graft, and Integra. The VivaScope allows noninvasive, real-time, in vivo images of tissue to be obtained. Observations of blood flow and tissue architecture in autologous skin graft and Integra suggest that 2 very different processes are occurring in the establishment of circulation in each case. Inosculation provides rapid circulatory return to skin grafts whereas slower neovascularization creates an unusual initial Integra circulation. The advent of confocal laser microscopy like the VivaScope 1500, together with "virtual" journals such as ePlasty, enables us to provide exciting images and distribute them widely to a "reading" audience. The development of the early Integra vasculature by neovascularization results in a large-vessel, high-volume, rapid flow circulation contrasting markedly from the inosculatory process in skin grafts and the capillary circulation in normal skin and merits further (planned) investigation.

  9. SU-E-T-98: Towards Cell Nucleus Microdosimetry: Construction of a Confocal Laser-Scanning Fluorescence Microscope to Readout Fluorescence Nuclear Track Detectors (FNTDs).

    Science.gov (United States)

    McFadden, C; Bartz, J; Akselrod, M; Sawakuchi, G

    2012-06-01

    To construct a custom confocal laser scanning microscope (CLSM) capable of resolving individual proton tracks in the volume of an Al2 O3 :C,Mg fluorescent nuclear track detector (FNTD). The spatial resolution of the FNTD technique is at the sub-micrometer scale. Therefore the FNTD technique has the potential to perform radiation measurements at the cell nucleus scale. The crystal volume of an FNTD contains defects which become fluorescent F2+ centers after trapping delta electrons from ionizing radiation. These centers have an absorption band centered at 620 nm and an emission band in the near infrared. Events of energy deposition in the crystal are read-out using a CLSM with sub-micrometer spatial resolution. Excitation light from a 635 nm laser is focused in the crystal volume by an objective lens. Fluorescence is collected back through the same path, filtered through a dichroic mirror, and focused through a small pinhole onto an avalanche photodiode. Lateral scanning of the focal point is performed with a scanning mirror galvanometer, and axial scanning is performed using a stepper-motor stage. Control of electronics and image acquisition was performed using a custom built LabVIEW VI and further image processing was done using Java. The system was used to scan FNTDs exposed to a 6 MV x-ray beam and an unexposed FNTD. Fluorescence images above the unexposed background were obtained at scan depths ranging from 5 - 10 micrometer below the crystal surface using a 100 micrometer pinhole size. Further work needs to be done to increase the resolution and the signal to noise ratio of the images so that energy deposition events may be identified more easily. Natural Sciences and Engineering Research Council of Canada. © 2012 American Association of Physicists in Medicine.

  10. Data on characterization of nano- and micro-structures resulting from glycine betaine surfactant/kappa-carrageenan interactions by Laser Scanning Confocal Microscopy and Transmission Electron Microscopy.

    Science.gov (United States)

    Gaillard, Cédric; Wang, Yunhui; Covis, Rudy; Vives, Thomas; Benoit, Maud; Benvegnu, Thierry

    2016-12-01

    This article contains data on the Laser Scanning Confocal Microscopy (LSCM) and Transmission Electron Microscopy (TEM) images related to multi-scaled self-assemblies resulting from 'green' cationic glycine betaine surfactant/anionic kappa-carrageenan interactions. These data gave clear evidence of the evolution of the micron-, nano-sized structures obtained at two surfactant/polymer molar ratios (3.5 and 0.8) and after the dilution of the aqueous dispersions with factors of 5 and 10 times. This data article is related to the research article entitled, "Monitoring the architecture of anionic ĸ-carrageenan/cationic glycine betaine amide surfactant assemblies by dilution: A multiscale approach" (Gaillard et al., 2017) [1].

  11. A handheld MEMS-based line-scanned dual-axis confocal microscope for early cancer detection and surgical guidance (Conference Presentation)

    Science.gov (United States)

    Chen, Ye; Yin, Chengbo; Wei, Linpeng; Glaser, Adam K.; Abeytunge, Sanjee; Peterson, Gary; Mandella, Michael J.; Sanai, Nader; Rajadhyaksha, Milind; Liu, Jonathan T.

    2017-02-01

    Considerable efforts have been recently undertaken to develop miniature optical-sectioning microscopes for in vivo microendoscopy and point-of-care pathology. These devices enable in vivo interrogation of disease as a real-time and noninvasive alternative to gold-standard histopathology, and therefore could have a transformative impact for the early detection of cancer as well as for guiding tumor-resection procedures. Regardless of the specific modality, various trade-offs in size, speed, field of view, resolution, contrast, and sensitivity are necessary to optimize a device for a particular application. Here, a miniature MEMS-based line-scanned dual-axis confocal (LS-DAC) microscope, with a 12-mm diameter distal tip, has been developed for point-of-care pathology. The dual-axis architecture has demonstrated superior rejection of out-of-focus and multiply scattered photons compared to a conventional single-axis confocal configuration. The use of line scanning enables fast frame rates (≥15 frames/sec), which mitigates motion artifacts of a handheld device during clinical use. We have developed a method to actively align the illumination and collection beams in this miniature LS-DAC microscope through the use of a pair of rotatable alignment mirrors. Incorporation of a custom objective lens, with a small form factor for in vivo application, enables the device to achieve an axial and lateral resolution of 2.0 and 1.1 microns, respectively. Validation measurements with reflective targets, as well as in vivo and ex vivo images of tissues, demonstrate that this high-speed LS-DAC microscope can achieve high-contrast imaging of fluorescently labeled tissues with sufficient sensitivity for applications such as oral cancer detection and guiding brain-tumor resections.

  12. Analysis of relationship between apoptosis and change of Ca2+ in HepG-2 induced by CSA with laser scanning confocal technology

    Science.gov (United States)

    Ji, Yu-bin; Yu, Lei

    2008-12-01

    Laser scanning confoncal technology was used to study relationship between apoptosis and change of Ca2+ induced by CSA (Capparis spinosa L. total alkaloid, CSA) on human heptocarcinoma cell HepG-2. Killing effect of CSA on human heptocarcinoma cell HepG-2 was measured by MTT method, while morphological observation of HepG-2 cells was completed by fluorescence microscope. Apoptosis induced by CSA on HepG-2 cells was measured by flowcytometry. In addition, change of intracellular Ca2+ level of CSA on HepG-2 cells was observed by laser scanning confocal microscope. As a result, CSA had obvious cytotoxicity on HepG-2 in a dose-dependent manner, and its IC50 was 162.4μg/ml. CSA could induce characteristic apoptosic morphology of HepG-2 cells, and apoptosis percentage was significantly higher than control one. Migration of cells cycle from S phase to G2 phase had been blocked by CSA. Concentration of Ca2+ in HepG-2 had been increased by CSA, which was positive correlation with drug dosage. CSA had obvious effect of killing and inducing apoptosis on human heptocarcinoma cell HepG-2, and overload of Ca2+ might be invovled in these events.

  13. Investigation of the cutaneous penetration behavior of dexamethasone loaded to nano-sized lipid particles by EPR spectroscopy, and confocal Raman and laser scanning microscopy.

    Science.gov (United States)

    Lohan, Silke B; Saeidpour, Siavash; Solik, Agnieszka; Schanzer, Sabine; Richter, Heike; Dong, Pin; Darvin, Maxim E; Bodmeier, Roland; Patzelt, Alexa; Zoubari, Gaith; Unbehauen, Michael; Haag, Rainer; Lademann, Jürgen; Teutloff, Christian; Bittl, Robert; Meinke, Martina C

    2017-07-01

    An improvement of the penetration efficiency combined with the controlled release of actives in the skin can facilitate the medical treatment of skin diseases immensely. Dexamethasone (Dx), a synthetic glucocorticoid, is frequently used for the treatment of inflammatory skin diseases. To investigate the penetration of nano-sized lipid particles (NLP) loaded with Dx in comparison to a commercially available base cream, different techniques were applied. Electron paramagnetic resonance (EPR) spectroscopy was used to monitor the penetration of Dx, which was covalently labeled with the spin probe 3-(Carboxy)-2,2,5,5-tetramethyl-1-pyrrolidinyloxy (PCA). The penetration into hair follicles was studied using confocal laser scanning microscopy (CLSM) with curcumin-loaded NLP. The penetration of the vehicle was followed by confocal Raman microscopy (CRM). Penetration studies using excised porcine skin revealed a more than twofold higher penetration efficiency for DxPCA into the stratum corneum (SC) after 24h incubation compared to 4h incubation when loaded to the NLP, whereas when applied in the base cream, almost no further penetration was observed beyond 4h. The distribution of DxPCA within the SC was investigated by consecutive tape stripping. The release of DxPCA from the base cream after 24h in deeper SC layers and the viable epidermis was shown by EPR. For NLP, no release from the carrier was observed, although DxPCA was detectable in the skin after the complete SC was removed. This phenomenon can be explained by the penetration of the NLP into the hair follicles. However, penetration profiles measured by CRM indicate that NLP did not penetrate as deeply into the SC as the base cream formulation. In conclusion, NLP can improve the accumulation of Dx in the skin and provide a reservoir within the SC and in the follicular infundibula. Copyright © 2017 Elsevier B.V. All rights reserved.

  14. Video-rate optical coherence tomography imaging with smart pixels

    Science.gov (United States)

    Beer, Stephan; Waldis, Severin; Seitz, Peter

    2003-10-01

    A novel concept for video-rate parallel acquisition of optical coherence tomography imaging is presented based on in-pixel demodulation. The main restrictions for parallel detection such as data rate, power consumption, circuit size and poor sensitivity are overcome with a smart pixel architecture incorporating an offset compensation circuit, a synchronous sampling stage, programmable time averaging and random pixel accessing, allowing envelope and phase detection in large 1D and 2D arrays.

  15. Scanning laser video camera/ microscope

    Science.gov (United States)

    Wang, C. P.; Bow, R. T.

    1984-10-01

    A laser scanning system capable of scanning at standard video rate has been developed. The scanning mirrors, circuit design and system performance, as well as its applications to video cameras and ultra-violet microscopes, are discussed.

  16. [Specifity of optic disc evaluation in healthy subjects with large optic discs and physiologic cupping using confocal scanning laser ophthalmoscopy].

    Science.gov (United States)

    Plange, N; Hirsch, T; Bienert, M; Remky, A

    2014-02-01

    Imaging methods of the optic nerve head appear to have an increasing impact in glaucoma diagnosis. The aim of this study is to evaluate the specifity of the Heidelberg Retina Tomograph (software version 1.7 and 3.0) in subjects with physiological cupping and large optic discs. 27 eyes of 27 subjects (mean age 41.3 ± 15.8 years) with bilateral physiological cupping and large optic discs (vertical cup-to-disc-ratio > 0.3, optic disc area 2.48 ± 0.45 mm2, max. 3.54 mm2) were included in a clinical study. All subjects had an intraocular pressure cupping by funduscopy and no nerve fibre layer defects (Scanning Laser Ophthalmoscope, Rodenstock, Germany). Standard achromatic perimetry (Humphrey Field Analyzer, Humphrey-Zeiss, 24/2 SITA or full threshold), short-wavelength automated perimetry (Humphrey Field Analyzer, Humphrey-Zeiss), and frequency doubling technology (FDT, Humphrey-Zeiss) revealed no visual field defects. Optic disc imaging was performed in all subjects using the Heidelberg Retina Tomograph II (HRT). Optic disc images were transferred to the software-update of the HRT 3 (Version 3.0, Heidelberg Engineering). Specifity was calculated using the Moorfields regression analysis (MRA, software version 1.7 and 3.0) and the glaucoma probability score (GPS analysis) using all disc sectors and omitting the nasal and 3 nasal sectors. Specifity of the MRA (software version 1.7) was 66.6 % (most specific criteria), and 22.2 % (least specific criteria). Specifity of the MRA (software version 3.0) was 33.3 % (most specific criteria), and 14.8 % (least specific criteria), whereas specifity of the GPS analysis was 37.0 % (most specific criteria), and 11.1 % (least specific criteria). When the nasal sectors were omitted for analysis, specifity increased for the MRA analysis, but not for the GPS analysis. Specifity of the MRA was unsatisfactory using the software version 1.7 and 3.0 in subjects with large optic discs and physiological cupping

  17. Comparative pattern of growth and development of Echinostoma paraensei (Digenea: Echinostomatidae) in hamster and Wistar rat using light and confocal laser scanning microscopy.

    Science.gov (United States)

    Souza, Joyce G R; Garcia, Juberlan S; Gomes, Ana Paula N; Machado-Silva, José Roberto; Maldonado, Arnaldo

    2017-12-01

    Echinostoma paraensei (Digenea: Echinostomatidae) lives in the duodenum and bile duct of rodents and is reported as a useful model for studies on the biology of flatworms. Here, we compared the growth and development of pre and post ovigerous worms collected 3, 7, 14 and 21 days post infection from experimentally infected hamster (permissive host) and Wistar rat (less permissive hosts). Linear measurements and ratios were examined by light (morphology and morphometry) and confocal laser scanning microscopy. At day 3, either worm from hamsters or rats were small with poorly developed gonads. At seven day, worms increased in size and morphometric differences between hosts are statistically significant after this time. In addition, adult worms (14 and 21 days of age) harvested from hamster showed developed gonads and vitelline glands laterally distributed on the body, whereas worms from rat showed atrophied reproductive system characterized by underdeveloped vitelline glands and stunted ovary. The worm rate recovery in rat decreased from 29.3% (day 7) to 20.6% (day 14) and 8% (day 21), whilst it remained around 37% in hamster. In conclusion, this is the first appointment demonstrating that low permissiveness influences the reproductive system of echinostome since the immature stages of development. The phenotypic analysis evidenced that hamster provides a more favorable microenvironment for gonads development than rat, confirming golden hamster as a permissive host, whereas Wistar rat is less permissive host. Copyright © 2017 Elsevier Inc. All rights reserved.

  18. Application of in vivo laser scanning confocal microscopy for evaluation of ocular surface diseases: lessons learned from pterygium, meibomian gland disease, and chemical burns.

    Science.gov (United States)

    Wang, Yan; Le, Qihua; Zhao, Feng; Hong, Jiaxu; Xu, Jianjiang; Zheng, Tianyu; Sun, Xinghuai

    2011-10-01

    In vivo laser scanning confocal microscopy (LSCM) has been widely used to evaluate the alterations caused by ocular surface diseases at a cellular level in the living eye. In this review, we focus on its use in the diagnosis of pterygium, meibomian gland (MG) disease, and chemical burns. Histopathologic changes occurring in pterygium can be examined in situ using in vivo LSCM. Alterations at the junction of the pterygium and the cornea, which cannot be observed in excised tissue samples, can be observed. MGs play an important role in maintaining the health of the ocular surface. Meibomian gland dysfunction (MGD) is one of the most common ocular surface diseases. The use of in vivo LSCM helps in the diagnosis of MGD and provides a way to examine the microstructure of MG acinar units and measure their size. In vivo LSCM also provides a new perspective in understanding the contribution of the MG to the health of the ocular surface. Chemical burns are one of the most common ocular injuries, and in vivo LSCM can provide images of the goblet cells on the corneal surface. This is a hallmark of limbal stem cell deficiency. The application of in vivo LSCM to assessing chemical burns requires extension, allowing for evaluation of the limbus structure and ocular surface changes after reconstructive ocular surgery.

  19. Biofilm formation on the Provox ActiValve: Composition and ingrowth analyzed by Illumina paired-end RNA sequencing, fluorescence in situ hybridization, and confocal laser scanning microscopy.

    Science.gov (United States)

    Timmermans, Adriana J; Harmsen, Hermie J M; Bus-Spoor, Carien; Buijssen, Kevin J D A; van As-Brooks, Corina; de Goffau, Marcus C; Tonk, Rudi H; van den Brekel, Michiel W M; Hilgers, Frans J M; van der Laan, Bernard F A M

    2016-04-01

    The most frequent cause of voice prosthesis failure is microbial biofilm formation on the silicone valve, leading to destruction of the material and transprosthetic leakage. The Provox ActiValve valve is made of fluoroplastic, which should be insusceptible to destruction. The purpose of this study was to determine if fluoroplastic is insusceptible to destruction by Candida species. Thirty-three dysfunctional Provox ActiValves (collected 2011-2013). Biofilm analysis was performed with Illumina paired-end sequencing (IPES), assessment of biofilm-material interaction with fluorescence in situ hybridization (FISH), and confocal laser scanning microscopy (CLSM). IPES (n = 10) showed that Candida albicans and Candida tropicalis are dominant populations on fluoroplastic and silicone. Microbial diversity is significantly lower on fluoroplastic. Lactobacillus gasseri is the prevalent bacterial strain on most voice prostheses. FISH and CLSM (n = 23): in none of the cases was ingrowth of Candida species present in the fluoroplastic. Fluoroplastic material of Provox ActiValve seems insusceptible to destruction by Candida species, which could help improve durability of voice prostheses. © 2015 Wiley Periodicals, Inc. Head Neck 38: E432-E440, 2016. © 2015 Wiley Periodicals, Inc.

  20. Benford's Law based detection of latent fingerprint forgeries on the example of artificial sweat printed fingerprints captured by confocal laser scanning microscopes

    Science.gov (United States)

    Hildebrandt, Mario; Dittmann, Jana

    2015-03-01

    The possibility of forging latent fingerprints at crime scenes is known for a long time. Ever since it has been stated that an expert is capable of recognizing the presence of multiple identical latent prints as an indicator towards forgeries. With the possibility of printing fingerprint patterns to arbitrary surfaces using affordable ink- jet printers equipped with artificial sweat, it is rather simple to create a multitude of fingerprints with slight variations to avoid raising any suspicion. Such artificially printed fingerprints are often hard to detect during the analysis procedure. Moreover, the visibility of particular detection properties might be decreased depending on the utilized enhancement and acquisition technique. In previous work primarily such detection properties are used in combination with non-destructive high resolution sensory and pattern recognition techniques to detect fingerprint forgeries. In this paper we apply Benford's Law in the spatial domain to differentiate between real latent fingerprints and printed fingerprints. This technique has been successfully applied in media forensics to detect image manipulations. We use the differences between Benford's Law and the distribution of the most significant digit of the intensity and topography data from a confocal laser scanning microscope as features for a pattern recognition based detection of printed fingerprints. Our evaluation based on 3000 printed and 3000 latent print samples shows a very good detection performance of up to 98.85% using WEKA's Bagging classifier in a 10-fold stratified cross-validation.

  1. In situ identification of streptococci and other bacteria in initial dental biofilm by confocal laser scanning microscopy and fluorescence in situ hybridization.

    Science.gov (United States)

    Dige, Irene; Nilsson, Holger; Kilian, Mogens; Nyvad, Bente

    2007-12-01

    Confocal laser scanning microscopy (CLSM) has been employed as a method for studying intact natural biofilm. When combined with fluorescence in situ hybridization (FISH) it is possible to analyze spatial relationships and changes of specific members of microbial populations over time. The aim of this study was to perform a systematic description of the pattern of initial dental biofilm formation by applying 16S rRNA-targeted oligonucleotide probes to the identification of streptococci and other bacteria, and to evaluate the usefulness of the combination of CLSM and FISH for structural studies of bacterial populations in dental biofilm. Biofilms were collected on standardized glass slabs mounted in intra-oral appliances and worn by 10 individuals for 6, 12, 24 or 48 h. After intra-oral exposure the biofilms were labelled with probes against either streptococci (STR405) or all bacteria (EUB338) and analysed by CLSM. The current approach of using FISH techniques enabled differentiation of streptococci from other bacteria and determination of their spatio-temporal organization. The presence of chimney-like multilayered microcolonies with different microbial compositions demonstrated by this methodology provided information supplementary to our previous knowledge obtained by classical electron microscopic methods and increased our understanding of the structure of developing biofilms.

  2. The determination of firing distance applying a microscopic quantitative method and confocal laser scanning microscopy for detection of gunshot residue particles.

    Science.gov (United States)

    Neri, Margherita; Turillazzi, Emanuela; Riezzo, Irene; Fineschi, Vittorio

    2007-07-01

    In this study, we applied a microscopic quantitative method based on the use of sodium rhodizonate to verify the presence of residues and their distribution on the cutis of gunshot wounds. A total of 250 skin samples were selected from cases in which the manner of death (accidental, suicide, and homicide) and the shooting distance could be reliably determined. The samples were examined under a light microscope, in transmitted bright field illumination and phase contrast mode, and with confocal laser scanning microscopy. In all skin specimens the area of each histological section was directly measured by an image analysis system. Both the number and the size of powder particles were measured. The distribution of gunshot residues (GSR) in the epidermal and subepidermal layers was also analyzed. The evaluation of the microscopic entrance wounds demonstrated different findings related to the range of fire. The data derived from the evaluation of both macroscopic and microscopic features demonstrated that the amount and the spatial distribution of GSR deposits in the skin surrounding entrance wounds strictly correlate with shooting distance.

  3. New insights into the painting stratigraphy of L'Homme blesse by Gustave Courbet combining scanning macro-XRF and confocal micro-XRF

    Energy Technology Data Exchange (ETDEWEB)

    Reiche, Ina [Staatliche Museen zu Berlin-Preussischer Kulturbesitz, Rathgen-Forschungslabor, Berlin (Germany); Laboratoire d' Archeologie Moleculaire et Structurale, Sorbonne Universites, Univ. Paris 06, CNRS, UMR 8220, Paris (France); Eveno, Myriam; Pichon, Laurent; Laval, Eric; Mottin, Bruno [Centre de Recherche et de Restauration des Musees de France (C2RMF), Paris (France); Mueller, Katharina [Laboratoire d' Archeologie Moleculaire et Structurale, Sorbonne Universites, Univ. Paris 06, CNRS, UMR 8220, Paris (France); Calligaro, Thomas [Centre de Recherche et de Restauration des Musees de France (C2RMF), Paris (France); PSL Research University, Chimie ParisTech-CNRS, Institut de Recherche Chimie Paris, Paris (France); Mysak, Erin [Centre de Recherche et de Restauration des Musees de France (C2RMF), Paris (France); Yale University, Institute for the Preservation of Cultural Heritage, New Haven, CT (United States)

    2016-11-15

    The painting L'Homme blesse by Gustave Courbet kept at the Musee d'Orsay in Paris has been recently studied by X-ray radiography, SEM-EDX observation of paint cross sections and confocal micro-X-ray fluorescence analyses (CXRF) at locations where the cross section samples were taken. This study allowed the establishment of the paint palette used by Courbet for the three paint compositions. Eight or more paint layers could be evidenced. In the view of the complexity of this painting, further analyses using two-dimensional scanning macro-X-ray fluorescence imaging (MA-XRF) providing chemical images corresponding to the superimposition of all detectable paint layers were employed. This method is combined with CXRF for depth-resolved paint layer analysis. Large elemental maps of Hg, Cu, As, Fe, Zn, Cr, Ba, Pb and Ca were obtained by MA-XRF on the painting and are discussed in combination with depth profiles obtained by CXRF on strategic points where three painting compositions overlap. The order of three successive compositions of this painting were determined in this study. This work also highlights the benefits of using complementary imaging methods to obtain a complete three-dimensional vision of the chemistry and stratigraphy of paintings. (orig.)

  4. Morphological aspects of Schistosoma mansoni adult worms isolated from nourished and undernourished mice: a comparative analysis by confocal laser scanning microscopy

    Directory of Open Access Journals (Sweden)

    Neves Renata Heisler

    2001-01-01

    Full Text Available Malnutrition hampers the course of schistosomiasis mansoni infection just as normal growth of adult worms. A comparative morphometric study on adult specimens (male and female recovered from undernourished (fed with a low protein diet - regional basic diet and nourished (rodent commercial laboratory food, NUVILAB white mice was performed. Tomographic images and morphometric analysis of the oral and ventral suckers, reproductive system and tegument were obtained by means of confocal laser scanning microscopy. Undernourished male specimens presented smaller morphometric values (length and width of the reproductive system (first, third and last testicular lobes and thickness of the tegument than controls. Besides that, it was demonstrated that the dorsal surface of the male worms bears large tubercles unevenly distributed, but kept grouped and flat. At the subtegumental region, vacuolated areas were detected. It was concluded that the inadequate nutritional status of the vertebrate host has a negative influence mainly in the reproductive system and topographical somatic development of male adult Schistosoma mansoni, inducing some alterations on the structure of the parasite.

  5. A new improved protocol for in vitro intratubular dentinal bacterial contamination for antimicrobial endodontic tests: standardization and validation by confocal laser scanning microscopy

    Directory of Open Access Journals (Sweden)

    Flaviana Bombarda de ANDRADE

    2015-01-01

    Full Text Available Objectives To compare three methods of intratubular contamination that simulate endodontic infections using confocal laser scanning microscopy (CLSM. Material and Methods Two pre-existing models of dentinal contamination were used to induce intratubular infection (groups A and B. These methods were modified in an attempt to improve the model (group C. Among the modifications it may be included: specimen contamination for five days, ultrasonic bath with BHI broth after specimen sterilization, use of E. faecalis during the exponential growth phase, greater concentration of inoculum, and two cycles of centrifugation on alternate days with changes of culture media. All specimens were longitudinally sectioned and stained with of LIVE/DEAD® for 20 min. Specimens were assessed using CLSM, which provided images of the depth of viable bacterial proliferation inside the dentinal tubules. Additionally, three examiners used scores to classify the CLSM images according to the following parameters: homogeneity, density, and depth of the bacterial contamination inside the dentinal tubules. Kruskal-Wallis and Dunn’s tests were used to evaluate the live and dead cells rates, and the scores obtained. Results The contamination scores revealed higher contamination levels in group C when compared with groups A and B (p0.05. The volume of live cells in group C was higher than in groups A and B (p<0.05. Conclusion The new protocol for intratubular infection resulted in high and uniform patterns of bacterial contamination and higher cell viability in all specimens when compared with the current methods.

  6. Comparison of fungiform taste-bud distribution among age groups using confocal laser scanning microscopy in vivo in combination with gustatory function.

    Science.gov (United States)

    Saito, Takehisa; Ito, Tetsufumi; Ito, Yumi; Manabe, Yasuhiro; Sano, Kazuo

    2016-04-01

    The aim of this study was to compare the distribution of taste buds in fungiform papillae (FP) and gustatory function between young and elderly age groups. Confocal laser scanning microscopy was used because it allows many FP to be observed non-invasively in a short period of time. The age of participants (n = 211) varied from 20 to 83 yr. The tip and midlateral region of the tongue were observed. Taste buds in an average of 10 FP in each area were counted. A total of 2,350 FP at the tongue tip and 2,592 FP in the midlateral region could be observed. The average number of taste buds was similar among all age groups both at the tongue tip and in the midlateral region. The taste function, measured by electrogustometry, among participants 20-29 yr of age was significantly lower than that in the other age groups; however, there was no difference among any other age groups in taste function. These results indicate that the peripheral gustatory system is well maintained anatomically and functionally in elderly people. © 2016 Eur J Oral Sci.

  7. In-vivo real-time tandem scanning confocal microscopic examination of wound healing in the cornea following an alkali burn

    Science.gov (United States)

    Chew, Sek J.; Beuerman, Roger W.; Kaufman, Herbert

    1994-06-01

    Chemical burns of the cornea can cause irreversible scarring, leading to visual impairment. The tandem scanning confocal microscope (TSM) was used to evaluate stromal changes in vivo following exposure to alkali, the most devastating form of chemical ocular injury. The corneas of anesthetized rabbits were exposed to filter papers impregnated with NaOH. A 25x water immersion objective lens was used with the TSM, and images captured with a CCD camera. Normal keratocytes appeared as ovoid nuclei. Collagen lamellae were not visible. Alkali led to immediate opacification of the extracellular matrix and loss of keratocytes. The former was quantified by en-face serial optical sectioning and subsequent off-line densitometry of the captured image. Wound healing was monitored as spindle-shaped cells appeared at the wound edge. This was accompanied by the production of fibrillary extracellular matrix. After 3 days, branched cellular processes 100 to 200 micrometers long were extended. By 1 week, dense aggregates of ovoid fibroblasts and whorls of collagen fibers had formed. Despite the overlying scarring, the deeper stromal layers and endothelium were still visible with this technique. We suggest that the TSM would be a useful clinical instrument for the evaluation and treatment evaluation of patients with chemical injuries of the cornea.

  8. Reproductive system abnormalities in Schistosoma mansoni adult worms isolated from Nectomys squamipes (Muridae: Sigmodontinae: brightfield and confocal laser scanning microscopy analysis

    Directory of Open Access Journals (Sweden)

    Neves Renata Heisler

    2003-01-01

    Full Text Available Schistosoma mansoni adult worms with genital anomalies isolated from Nectomys squamipes (Muridae: Sigmodontinae were studied by confocal laser scanning microscopy under the reflected mode. One male without testicular lobes (testicular agenesia/anorchism and two females, one with an atrophied ovary and another with 17 uterine eggs, were identified. The absence of testicular lobes occurred in a worm presenting otherwise normal male adult characteristics: tegument, tubercles and a gynaecophoric canal with spines. In both female specimens the digestive tube showed a vacuolated appearance, and the specimen with supernumerary uterine eggs exhibited a developing miracidium and an egg with a formed shell. The area of the ventral sucker was similar in both specimens however the tegument thickness, ovary and vitelline glands of the specimen with the atrophied ovary were smaller than those of the one with supernumerary eggs. These reported anomalies in the reproductive system call attention to the need to improve our understanding of genetic regulation and the possible role of environmental influences upon trematode development.

  9. Evaluation of transdermal delivery of nanoemulsions in ex vivo porcine skin using two-photon microscopy and confocal laser-scanning microscopy

    Science.gov (United States)

    Choi, Sanghoon; Kim, Jin Woong; Lee, Yong Joong; Delmas, Thomas; Kim, Changhwan; Park, Soyeun; Lee, Ho

    2014-10-01

    This study experimentally evaluates the self-targeting ability of asiaticoside-loaded nanoemulsions compared with nontargeted nanoemulsions in ex vivo experiments with porcine skin samples. Homebuilt two-photon and confocal laser-scanning microscopes were employed to noninvasively examine the transdermal delivery of two distinct nanoemulsions. Prior to the application of nanoemulsions, we noninvasively observed the morphology of porcine skin using two-photon microscopy. We have successfully visualized the distributions of the targeted and nontargeted nanoemulsions absorbed into the porcine skin samples. Asiaticoside-loaded nanoemulsions showed an improved ex vivo transdermal delivery through the stratum corneum compared with nonloaded nanoemulsions. As a secondary measure, nanoemulsions-applied samples were sliced in the depth direction with a surgical knife in order to obtain the complete depth-direction distribution profile of Nile red fluorescence. XZ images demonstrated that asiaticoside-loaded nanoemulsion penetrated deeper into the skin compared with nontargeted nanoemulsions. The basal layer boundary is clearly visible in the case of the asiaticoside-loaded skin sample. These results reaffirm the feasibility of using self-targeting ligands to improve permeation through the skin barrier for cosmetics and topical drug applications.

  10. Novel application of confocal laser scanning microscopy and 3D volume rendering toward improving the resolution of the fossil record of charcoal.

    Directory of Open Access Journals (Sweden)

    Claire M Belcher

    Full Text Available Variations in the abundance of fossil charcoals between rocks and sediments are assumed to reflect changes in fire activity in Earth's past. These variations in fire activity are often considered to be in response to environmental, ecological or climatic changes. The role that fire plays in feedbacks to such changes is becoming increasingly important to understand and highlights the need to create robust estimates of variations in fossil charcoal abundance. The majority of charcoal based fire reconstructions quantify the abundance of charcoal particles and do not consider the changes in the morphology of the individual particles that may have occurred due to fragmentation as part of their transport history. We have developed a novel application of confocal laser scanning microscopy coupled to image processing that enables the 3-dimensional reconstruction of individual charcoal particles. This method is able to measure the volume of both microfossil and mesofossil charcoal particles and allows the abundance of charcoal in a sample to be expressed as total volume of charcoal. The method further measures particle surface area and shape allowing both relationships between different size and shape metrics to be analysed and full consideration of variations in particle size and size sorting between different samples to be studied. We believe application of this new imaging approach could allow significant improvement in our ability to estimate variations in past fire activity using fossil charcoals.

  11. A novel method to analyse in vivo the physiological state and cell viability of phototrophic microorganisms by confocal laser scanning microscopy using a dual laser.

    Science.gov (United States)

    Millach, Laia; Obiol, Aleix; Solé, Antonio; Esteve, Isabel

    2017-10-01

    Phototrophic microorganisms are very abundant in extreme environments, where are subjected to frequent and strong changes in environmental parameters. Nevertheless, little is known about the physiological effects of these changing environmental conditions on viability of these microorganisms, which are difficult to grow in solid media and have the tendency to form aggregates. For that reason, it is essential to develop methodologies that provide data in short time consuming, in vivo and with minimal manipulating the samples, in response to distinct stress conditions. In this paper, we present a novel method using Confocal Laser Scanning Microscopy and a Dual Laser (CLSM-DL) for determining the cell viability of phototrophic microorganisms without the need of either staining or additional use of image treating software. In order to differentiate viable and nonviable Scenedesmus sp. DE2009 cells, a sequential scan in two different channels was carried out from each same xyz optical section. On the one hand, photosynthetic pigments fluorescence signal (living cells) was recorded at the red channel (625- to 785-nm fluorescence emission) exciting the samples with a 561-nm laser diode, and an acousto-optic tunable filter (AOTF) of 20%. On the other hand, nonphotosynthetic autofluorescence signal (dead cells) was recorded at the green channel (500- to 585-nm fluorescence emission) using a 405-nm UV laser, an AOTF of 15%. Both types of fluorescence signatures were captured with a hybrid detector. The validation of the CLSM-DL method was performed with SYTOX green fluorochrome and electron microscopic techniques, and it was also applied for studying the response of distinct light intensities, salinity doses and exposure times on a consortium of Scenedesmus sp. DE2009. © 2017 The Authors Journal of Microscopy © 2017 Royal Microscopical Society.

  12. Imaging of intracellular behavior of polymeric nanoparticles in Staphylococcus epidermidis biofilms by slit-scanning confocal Raman microscopy and scanning electron microscopy with energy-dispersive X-ray spectroscopy.

    Science.gov (United States)

    Takahashi, Chisato; Ueno, Kusuo; Aoyama, Junichi; Adachi, Mariko; Yamamoto, Hiromitsu

    2017-07-01

    In drug delivery systems employing polymeric nanoparticles, accurate delivery of drugs to target sites such as bacterial cells, cell tissues, and organelles is essential. In particular, when designing drug delivery systems for the treatment of the biofilm infections, evaluation of the interaction between polymeric nanoparticles and biofilm or bacterial cells using a simple technique is of significant importance. Here we develop two types of novel techniques for the biological imaging of the intracellular behavior of two types of polymeric nanoparticles, biodegradable chitosan-modified poly (dl-lactide-co-glycolide) (PLGA) nanoparticles and chitosan-modified polyvinyl caprolactam - polyvinyl acetate -polyethylene glycol graft copolymer (Soluplus®, Sol) nanoparticles, within a Staphylococcus epidermidis biofilm. As the first technique, Raman imaging of unstained biological materials using slit-scanning confocal Raman microscopy (unstained Raman imaging) was performed, and as the second, field-emission scanning electron microscopy with energy-dispersive X-ray spectroscopy analysis of biological materials labeled with quantum dots (SEM-QD imaging) was demonstrated. These analyses revealed differing localization of the respective nanoparticles within the biofilm in accordance with the specific interactions of PLGA nanoparticles and Sol nanoparticles with the biofilm. These novel techniques open the door to biological imaging and analyses with high spatial resolution, which will help to understand the efficacy of drug delivery to target materials. Copyright © 2017 Elsevier B.V. All rights reserved.

  13. Can scanning near-field optical microscopy be compared with confocal laser scanning microscopy? A preliminary study on alpha-sarcoglycan and beta1D-integrin in human skeletal muscle.

    Science.gov (United States)

    Anastasi, G; Cutroneo, G; Pisani, A; Bruschetta, D; Milardi, D; Princi, P; Gucciardi, P G; Bramanti, P; Soscia, L; Favaloro, A

    2007-12-01

    The dystrophin-glycoprotein complex and the vinculin-talin-integrin system constitute, together a protein machinery, called costameres. The dystrophin-glycoprotein complex contains, among other proteins, also dystrophin and the sarcoglycans subcomplex, proteins playing a key role in the pathogenesis of many muscular dystrophies and linking the cytoplasmic myofibrillar contractile elements to the signal transducing molecules of the extracellular matrix, also providing structural support to the sarcolemma. The vinculin-talin-integrin system connects some components of the extracellular matrix with intermediate filaments of desmin, forming transverse bridges between Z and M lines. In our previous reports we always studied these systems by confocal laser scanning microscopy (CLSM). In this paper we report on the first applications of optical near-field fluorescence microscopy to the spatial localization of alpha-sarcoglycan and beta1D-integrin in human skeletal muscle fibres in order to better compare and test the images obtained with conventional CLSM and with scanning near-field optical microscopy (SNOM). In addition, the analysis of the surface morphology, and the comparison with the fluorescence map is put forward and analyzed for the first time on human muscle fibres. In aperture-SNOM the sample is excited through the nanometre-scale aperture produced at the apex of an optical fibre after tapering and subsequent metal coating. The acquisition of the topography map, simultaneously to the optical signal, by SNOM, permits to exactly overlap the fluorescence images obtained from the two consecutive scans needed for the double localization. Besides, the differences between the topography and the optical spatial patterns permit to assess the absence of artefacts in the fluorescence maps. Although the SNOM represented a good method of analysis, this technique remains a complementary method to the CLSM and it can be accepted in order to confirm the hypothesis advanced by

  14. Confocal microscopy

    Indian Academy of Sciences (India)

    molecular aggregates in artificial light harvesting sys- tem it is important to elucidate the exciton dynamics of individual micro-rods which can be achieved by using confocal microscopy and polarization resolved single molecule fluorescence spectroscopy.30 41 In the present work, we have studied exciton dynamics of two.

  15. Can Visual Field Progression be Predicted by Confocal Scanning Laser Ophthalmoscopic Imaging of the Optic Nerve Head in Glaucoma? (An American Ophthalmological Society Thesis).

    Science.gov (United States)

    Danias, John; Serle, Janet

    2015-01-01

    To determine whether confocal scanning laser ophthalmoscopic imaging (Heidelberg retinal tomography [HRT]) can predict visual field change in glaucoma. The study included 561 patients with glaucoma or ocular hypertension whose clinical course was followed at the Mount Sinai Faculty practice. Humphrey visual fields (HVFs) and HRT images were collected on one randomly selected eye per patient. Glaucoma progression was determined by the presence of two sequential statistically significant negative slopes in mean deviation (MD) or visual field index (VFI) at any point during the study period. Trend-based analysis on HRT parameters was used to determine progressive changes and whether these occurred before or after HVF change. Sensitivity and specificity of HRT to predict HVF change were calculated. HVF rate of change was correlated to the rate of change detected by HRT imaging. Approximately 17% of patients progressed by either MD or VFI criteria. MD and VFI correlated highly and identified overlapping sets of patients as progressing. HRT global parameters had poor sensitivity (∼42%) and moderate specificity (∼67%) to predict HVF progression. Regional stereometric parameters were more sensitive (69%-78%) but significantly less specific (24%-27%). Sensitivity of global stereometric parameters in detecting HVF change was not significantly affected by the level of visual field damage (P=.3, Fisher exact test). HVF rate of change did not correlate with rate of change of HRT parameters. Trend-based analysis of HRT parameters has poor sensitivity and specificity in predicting HVF change. This may be related specifically to HRT imaging or may reflect the fact that in some patients with glaucoma, functional changes precede structural alterations.

  16. Imaging breast cancer morphology using probe-based confocal laser endomicroscopy: towards a real-time intraoperative imaging tool for cavity scanning.

    Science.gov (United States)

    Chang, Tou Pin; Leff, Daniel R; Shousha, Sami; Hadjiminas, Dimitri J; Ramakrishnan, Rathi; Hughes, Michael R; Yang, Guang-Zhong; Darzi, Ara

    2015-09-01

    Current techniques for assessing the adequacy of tumour excision during breast conserving surgery do not provide real-time direct cytopathological assessment of the internal cavity walls within the breast. This study investigates the ability of probe-based confocal laser endomicroscopy (pCLE), an emerging imaging tool, to image the morphology of neoplastic and non-neoplastic breast tissues, and determines the ability of histopathologists and surgeons to differentiate these images. Freshly excised tumour samples and adjacent non-diseased sections from 50 consenting patients were stained with 0.01 % acriflavine hydrochloride and imaged using pCLE. All discernible pCLE features were cross-examined with conventional histopathology. Following pattern recognition training, 17 histopathologists and surgeons with no pCLE experience interpreted 50 pCLE images independently whilst blinded to histopathology results. Three-hundred and fifty pCLE image mosaics were analysed. Consistent with histopathology findings, the glandular structures, adipocytes and collagen fibres of normal breast were readily visible on pCLE images. These were distinguishable from the morphological architecture exhibited by invasive and non-invasive carcinoma. The mean accuracy of pCLE image interpretation for histopathologists and surgeons was 94 and 92 %, respectively. Overall, inter-observer agreement for histopathologists was 'almost perfect', κ = 0.82; and 'substantial' for surgeons, κ = 0.74. pCLE morphological features of neoplastic and non-neoplastic breast tissues are readily visualized and distinguishable with high accuracy by both histopathologists and surgeons. Further research is required to investigate a potential role for the use of pCLE intraoperatively for in situ detection of residual cancerous foci, thereby guiding operating decision-making based on real-time breast cavity scanning.

  17. Influence of various herbal irrigants as a final rinse on the adherence of Enterococcus faecalis by fluorescence confocal laser scanning microscope

    Science.gov (United States)

    Rosaline, Hannah; Kandaswamy, D; Gogulnath, D; Rubin, MI

    2013-01-01

    Aim: The aim of this study was to assess the antibacterial efficacy of three different herbal irrigants against Enterococcus faecalis. Materials and Methods: Single rooted teeth were extracted due to orthodontic and periodontal reasons. The teeth were then inoculated with E. faecalis. The teeth were randomly divided into three experimental groups and two control groups of six samples each. Group 1 specimens were treated with 5.2% sodium hypochlorite (NaOCL) for 30 min followed by 5 mmol/L Ethylenediaminetetraacetic acid (EDTA) for 5 min and saline as final irrigant. Group 2 specimens were treated with and 5.2% NaOCl for 30 min as final irrigant. Group 3 were treated with Morinda citrifolia (MC) for 30 min as final irrigant. Group 4 were treated with Azadiracta indica (AI) as final irrigant. Group 5 were treated with green tea (GT) for 30 min as final irrigant. The dentin specimens were carefully spread onto a microscope slide and stained with BacLight and examined in a confocal laser scanning microscope set to monitor fluorescein isothiocyanate and propidium iodide. A total of nine fields were examined for each treatment and the bacteria presented were counted. Statistical Analysis: Using the one-way ANOVA with multiple comparison, significantly less bacteria were found adhering to the samples treated with Neem followed by NaOCL, GT, MC, Saline. Results: AI treatment produced the maximum reduction in adherence of E. faecalis to dentin (9.30%) followed by NaOCl (12.50%), GT (27.30%), MC (44.20%) and saline (86.70%). Conclusion: Neem is effective in preventing adhesion of E. faecalis to dentin. PMID:23956540

  18. In situ observation of the growth of biofouling layer in osmotic membrane bioreactors by multiple fluorescence labeling and confocal laser scanning microscopy.

    Science.gov (United States)

    Yuan, Bo; Wang, Xinhua; Tang, Chuyang; Li, Xiufen; Yu, Guanghui

    2015-05-15

    Since the concept of the osmotic membrane bioreactor (OMBR) was introduced in 2008, it has attracted growing interests for its potential applications in wastewater treatment and reclamation; however, the fouling mechanisms of forward osmosis (FO) membrane especially the development of biofouling layer in the OMBR are not yet clear. Here, the fouled FO membranes were obtained from the OMBRs on days 3, 8 and 25 in sequence, and then the structure and growing rule of the biofouling layer formed on the FO membrane samples were in-situ characterized by multiple fluorescence labeling and confocal laser scanning microscopy (CLSM). CLSM images indicated that the variations in abundance and distribution of polysaccharides, proteins and microorganisms in the biofouling layer during the operation of OMBRs were significantly different. Before the 8th day, their biovolume dramatically increased. Subsequently, the biovolumes of β-d-glucopyranose polysaccharides and proteins continued increasing and leveled off after 8 days, respectively, while the biovolumes of α-d-glucopyranose polysaccharides and microorganisms decreased. Extracellular polymeric substances (EPS) played a significant role in the formation and growth of biofouling layer, while the microorganisms were seldom detected on the upper fouling layer after 3 days. Based on the results obtained in this study, the growth of biofouling layer on the FO membrane surface in the OMBR could be divided into three stages. Initially, EPS was firstly deposited on the FO membrane surface, and then microorganisms associated with EPS located in the initial depositing layer to form clusters. After that, the dramatic increase of the clusters of EPS and microorganisms resulted in the quick growth of biofouling layer during the flux decline of the OMBR. However, when the water flux became stable in the OMBR, some microorganisms and EPS would be detached from the FO membrane surface. Copyright © 2015 Elsevier Ltd. All rights reserved.

  19. Analysis by confocal laser scanning microscopy of the MDPB bactericidal effect on S. mutans biofilm CLSM analysis of MDPB bactericidal effect on biofilm

    Directory of Open Access Journals (Sweden)

    Fabíola Galbiatti de Carvalho

    2012-10-01

    Full Text Available Since bacteria remain in the dentin following caries removal, restorative materials with antibacterial properties are desirable to help maintaining the residual microorganisms inactive. The adhesive system Clearfil Protect Bond (PB contains the antibacterial monomer 12-methacryloyloxydodecylpyridinium bromide (MDPB in its primer, which has shown antimicrobial activity. However, its bactericidal effect against biofilm on the dentin has been little investigated. Objective: The aim of this study was to analyze by confocal laser scanning microscopy (CLSM and viable bacteria counting (CFU the MDPB bactericidal effect against S. mutans biofilm on the dentin surface. Material and methods: Bovine dentin surfaces were obtained and subjected to S. mutans biofilm formation in BHI broth supplemented with 1% (w/v sucrose for 18 h. Samples were divided into three groups, according to the primer application (n=3: Clearfil Protect Bond (PB, Clearfil SE Bond, which does not contain MDPB, (SE and saline (control group. After the biofilm formation, Live/ Dead stain was applied directly to the surface of each sample. Next, 10 µL of each primer were applied on the samples during 590 s for the real-time CLSM analysis. The experiment was conducted in triplicate. The primers and saline were also applied on the other dentin samples during 20, 90, 300 and 590 s (n=9 for each group and period evaluated and the CFU were assessed by colonies counting. Results: The results of the CLSM showed that with the Se application, although non-viable bacteria were detected at 20 s, there was no increase in their count during 590 s. In contrast, after the PB application there was a gradual increase of non-viable bacteria over 590 s. Conclusions: The quantitative analysis demonstrated a significant decrease of S. mutans CFU at 90 s PB exposure and only after 300 s of Se application. Protect Bond showed an earlier antibacterial effect than Se Bond.

  20. Organic pollutant clustered in the plant cuticular membranes: visualizing the distribution of phenanthrene in leaf cuticle using two-photon confocal scanning laser microscopy.

    Science.gov (United States)

    Li, Qingqing; Chen, Baoliang

    2014-05-06

    Plants play a key role in the transport and fate of organic pollutants. Cuticles on plant surfaces represent the first resistance for the uptake of airborne toxicants. In this study, a confocal scanning microscope enhanced with a two-photon laser was applied as a direct and noninvasive probe to explore the in situ uptake of a model pollutant, phenanthrene (PHE), into the cuticular membrane of a hypostomatic plant, Photinia serrulata. On the leaf cuticle surfaces, PHE forms clusters instead of being evenly distributed. The PHE distribution was quantified by the PHE fluorescence intensity. When PHE concentrations in water varying over 5 orders of magnitude were applied to the isolated cuticle, the accumulated PHE level by the cuticle was not vastly different, whether PHE was applied to the outer or inner side of the cuticle. Notably, PHE was found to diffuse via a channel-like pathway into the middle layer of the cuticle matrix, where it was identified to be composed of polymeric lipids. The strong affinity of PHE for polymeric lipids is a major contributor of the fugacity gradient driving the diffusive uptake of PHE in the cuticular membrane. Membrane lipids constitute important domains for hydrophobic interaction with pollutants, determining significant differentials of fugacities within the membrane microsystem. These, under unsteady conditions, contribute to enhance net transport and clustering along the z dimension. Moreover, the liquid-like state of polymeric lipids may promote mobility by enhancing the diffusion rate. The proposed "diffusive uptake and storage" function of polymeric lipids within the membrane characterizes the modality of accumulation of the hydrophobic contaminant at the interface between the plant and the environment. Assessing the capacity of fugacity of these constituents in detail will bring about knowledge of contaminant fate in superior plants with a higher level of accuracy.

  1. Application of scanning cytometry and confocal-microscopy-based image analysis for investigation the role of cytoskeletal elements during equine herpesvirus type 1 (EHV-1) infection of primary murine neurons.

    Science.gov (United States)

    Słońska, A; Cymerys, J; Godlewski, M M; Bańbura, M W

    2016-11-01

    Equine herpesvirus type 1 (EHV-1), a member of Alphaherpesvirinae, has a broad host range in vitro, allowing for study of the mechanisms of productive viral infection, including intracellular transport in various cell cultures. In the current study, quantitative methods (scanning cytometry and real-time PCR) and confocal-microscopy-based image analysis were used to investigate the contribution of microtubules and neurofilaments in the transport of virus in primary murine neurons separately infected with two EHV-1 strains. Confocal-microscopy analysis revealed that viral antigen co-localized with the β-tubulin fibres within the neurites of infected cells. Alterations in β-tubulin and neurofilaments were evaluated by confocal microscopy and scanning cytometry. Real-time PCR analysis demonstrated that inhibitor-induced (nocodazole, EHNA) disruption of microtubules and dynein significantly reduced EHV-1 replication in neurons. Our results suggest that microtubules together with the motor protein - dynein, are involved in EHV-1 replication process in neurons. Moreover, the data presented here and our earlier results support the hypothesis that microtubules and actin filaments play an important role in the EHV-1 transport in primary murine neurons, and that both cytoskeletal structures complement each-other. Copyright © 2016 Elsevier B.V. All rights reserved.

  2. Confocal Raman Microscopy; applications in tissue engineering

    NARCIS (Netherlands)

    van Apeldoorn, Aart A.

    2005-01-01

    This dissertation describes the use of confocal Raman microscopy and spectroscopy in the field of tissue engineering. Moreover, it describes the combination of two already existing technologies, namely scanning electron microscopy and confocal Raman spectroscopy in one apparatus for the enhancement

  3. Realization of a video-rate distributed aperture millimeter-wave imaging system using optical upconversion

    Science.gov (United States)

    Schuetz, Christopher; Martin, Richard; Dillon, Thomas; Yao, Peng; Mackrides, Daniel; Harrity, Charles; Zablocki, Alicia; Shreve, Kevin; Bonnett, James; Curt, Petersen; Prather, Dennis

    2013-05-01

    Passive imaging using millimeter waves (mmWs) has many advantages and applications in the defense and security markets. All terrestrial bodies emit mmW radiation and these wavelengths are able to penetrate smoke, fog/clouds/marine layers, and even clothing. One primary obstacle to imaging in this spectrum is that longer wavelengths require larger apertures to achieve the resolutions desired for many applications. Accordingly, lens-based focal plane systems and scanning systems tend to require large aperture optics, which increase the achievable size and weight of such systems to beyond what can be supported by many applications. To overcome this limitation, a distributed aperture detection scheme is used in which the effective aperture size can be increased without the associated volumetric increase in imager size. This distributed aperture system is realized through conversion of the received mmW energy into sidebands on an optical carrier. This conversion serves, in essence, to scale the mmW sparse aperture array signals onto a complementary optical array. The side bands are subsequently stripped from the optical carrier and recombined to provide a real time snapshot of the mmW signal. Using this technique, we have constructed a real-time, video-rate imager operating at 75 GHz. A distributed aperture consisting of 220 upconversion channels is used to realize 2.5k pixels with passive sensitivity. Details of the construction and operation of this imager as well as field testing results will be presented herein.

  4. Spatially resolved analyses of uranium species using a coupled system made up of confocal laser-scanning microscopy (CLSM) and laser induced fluorescence spectroscopy (LIFS); Ortsaufgeloeste Analyse von Uranspezies mittels einem Gekoppelten System aus Konfokaler Laser-Scanning Mikroskopie (CLSM) und Laser Induzierter Fluoreszenzspektroskopie (LIFS)

    Energy Technology Data Exchange (ETDEWEB)

    Brockmann, S. [Verein fuer Kernverfahrenstechnik und Analytik Rossendorf e.V. (VKTA), Dresden (Germany); Grossmann, K.; Arnold, T. [Helmholtz-Zentrum Dresden-Rossendorf e.V. (Germany). Inst. fuer Ressourcenoekologie

    2014-01-15

    The fluorescent properties of uranium when excited by UV light are used increasingly for spectroscope analyses of uranium species within watery samples. Here, alongside the fluorescent properties of the hexavalent oxidation phases, the tetra and pentavalent oxidation phases also play an increasingly important role. The detection of fluorescent emission spectrums on solid and biological samples using (time-resolved) laser induced fluorescence spectroscopy (TRLFS or LIFS respectively) has, however, the disadvantage that no statements regarding the spatial localisation of the uranium can be made. However, particularly in complex, biological samples, such statements on the localisation of the uranium enrichment in the sample are desired, in order to e.g. be able to distinguish between intra and extra-cellular uranium bonds. The fluorescent properties of uranium (VI) compounds and minerals can also be used to detect their localisation within complex samples. So the application of fluorescent microscopic methods represents one possibility to localise and visualise uranium precipitates and enrichments in biological samples, such as biofilms or cells. The confocal laser-scanning microscopy (CLSM) is especially well suited to this purpose. Coupling confocal laser-scanning microscopy (CLSM) with laser induced fluorescence spectroscopy (LIFS) makes it possible to localise and visualise fluorescent signals spatially and three-dimensionally, while at the same time being able to detect spatially resolved, fluorescent-spectroscopic data. This technology is characterised by relatively low detection limits from up to 1.10{sup -6} M for uranium (VI) compounds within the confocal volume. (orig.)

  5. Evaluation of Corneal Cross-Linking for Treatment of Fungal Keratitis: Using Confocal Laser Scanning Microscopy on an Ex Vivo Human Corneal Model.

    Science.gov (United States)

    Alshehri, Jawaher M; Caballero-Lima, David; Hillarby, M Chantal; Shawcross, Susan G; Brahma, Arun; Carley, Fiona; Read, Nick D; Radhakrishnan, Hema

    2016-11-01

    Some previous reports have established the use of photoactivated chromophore-induced corneal cross-linking (PACK-CXL) in treating fungal keratitis. The results of these case reports have often been conflicting. To systematically study the effect of PACK-CXL in the management of Fusarium keratitis, we have developed an ex vivo model of human corneal infection using eye-banked human corneas. Sixteen healthy ex vivo human corneas were divided into four study groups: (1) untreated control, (2) cross-linked, (3) infected with fungal spores, and (4) infected with fungal spores and then cross-linked. All infected corneas were inoculated with Fusarium oxysporum spores. The PACK-CXL procedure was performed 24 hours post inoculation for group 4. For PACK-CXL treatment, the corneas were debrided of epithelium; then 1% (wt/vol) isotonic riboflavin was applied dropwise at 5-minute intervals for 30 minutes and during the course of UV-A cross-linking for another 30 minutes. The corneas were imaged using a confocal microscope at 48 hours post inoculation, and the Fusarium hyphal volume and spore concentration were calculated. The infected and then cross-linked group had a significantly lower volume of Fusarium hyphae, compared to the infected (P = 0.001) group. In the infected and then cross-linked group there was significant inhibition of Fusarium sporulation compared with the infected (P = 0.007) group. A model of human corneal infection was successfully developed for investigation of the effects of PACK-CXL on fungal keratitis. A treatment regimen of combined UV-A/riboflavin-induced corneal cross-linking appears to be a valuable approach to inhibit the growth and sporulation of Fusarium and suppress the progression of fungal keratitis.

  6. Combined in vivo confocal Raman spectroscopy and confocal microscopy of human skin

    NARCIS (Netherlands)

    P.J. Caspers (Peter); G.W. Lucassen (Gerald); G.J. Puppels (Gerwin)

    2003-01-01

    textabstractIn vivo confocal Raman spectroscopy is a noninvasive optical method to obtain detailed information about the molecular composition of the skin with high spatial resolution. In vivo confocal scanning laser microscopy is an imaging modality that provides optical sections

  7. Fluorescence (Multiwave) Confocal Microscopy.

    Science.gov (United States)

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

    2016-10-01

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

  8. Microscopia confocal de barrido láser y su relación con la morfología de la bula de filtración Confocal laser scanning microscopy and its association to the morphology of the filtering bleb

    Directory of Open Access Journals (Sweden)

    María Teresa Ferrer Guerra

    2012-01-01

    Full Text Available Objetivo: describir los hallazgos de la microscopia confocal de barrido láser y su relación con la morfología de la bula de filtración. Métodos: estudio observacional descriptivo de corte transversal en 100 ojos. Se les realizó microscopia confocal de barrido láser con el HRT II y el módulo corneal de Rostock. Se aplicó la prueba de significación estadística de Mc. Nemar para una p=0,05. Resultados: predominó el grupo entre 61 y 80 años de edad (55,8 % y el color de la piel blanca (46,8 %. En el grupo de descenso de la presión intraocular en más del 30 % se ubicó la mayor cantidad de bulas de todos los tamaños. Las bulas de mediano tamaño fueron las que más disminuyeron las cifras de presión intraocular, con 24 ojos (55,8 %, p=0,00, seguidas por las de pequeño tamaño (p=0,14. Las bulas de filtración aplanadas fueron las más frecuentes en 55 % de los casos (p=0,00, y 67,4 % de estas se ubicaron en el grupo de descenso de presión intraocular de más de 30 % (p=0,01. Las bulas medianas presentaron la mayor cantidad de estroma en malla porosa (60 % y de microquistes epiteliales (56 % p=0,00. Conclusiones: la configuración aplanada y el tamaño mediano de la bula de filtración se relacionaron con la presencia de variables histológicas que infieren buen funcionamiento de la bula (microquistes epiteliales y estroma en malla porosa. También se relacionaron con un mayor descenso de la presión intraocular.Objective: to describe the findings in the confocal laser scanning microscopy and its relation with the morphology of the filtering bleb. Methods: cross-sectional, observational and descriptive study of 100 eyes that underwent confocal laser scanning microscopy with the Retinal Heidelberg Tomography and the Rostock corneal module. Mc Nemar’s statistical significance test was made to obtain p=0,05. Results: the 61-80 years old age group (55,8 % and the caucasians predominated. The group of eyes with over 30 % decrease of

  9. Collection of trace evidence of explosive residues from the skin in a death due to a disguised letter bomb. The synergy between confocal laser scanning microscope and inductively coupled plasma atomic emission spectrometer analyses.

    Science.gov (United States)

    Turillazzi, Emanuela; Monaci, Fabrizio; Neri, Margherita; Pomara, Cristoforo; Riezzo, Irene; Baroni, Davide; Fineschi, Vittorio

    2010-04-15

    In most deaths caused by explosive, the victim's body becomes a depot for fragments of explosive materials, so contributing to the collection of trace evidence which may provide clues about the specific type of device used with explosion. Improvised explosive devices are used which contain "homemade" explosives rather than high explosives because of the relative ease with which such components can be procured. Many methods such as chromatography-mass spectrometry, scanning electron microscopy, stereomicroscopy, capillary electrophoresis are available for use in the identification of explosive residues on objects and bomb fragments. Identification and reconstruction of the distribution of explosive residues on the decedent's body may give additional hints in assessing the position of the victim in relation to the device. Traditionally these residues are retrieved by swabbing the body and clothing during the early phase, at autopsy. Gas chromatography-mass spectrometry and other analytical methods may be used to analyze the material swabbed from the victim body. The histological examination of explosive residues on skin samples collected during the autopsy may reveal significant details. The information about type, quantity and particularly about anatomical distribution of explosive residues obtained utilizing confocal laser scanning microscope (CLSM) together with inductively coupled plasma atomic emission spectrometer (ICP-AES), may provide very significant evidence in the clarification and reconstruction of the explosive-related events. Copyright 2009 Elsevier Ireland Ltd. All rights reserved.

  10. Scanning electron, 3D-confocal and stereotactic microscopies morpho-structural analysis of antibiotic-loaded cement porosity in three different formulations.

    Science.gov (United States)

    Fenga, D; Vermiglio, G; Traina, F; Favaloro, A; Rosa, M A

    2017-01-01

    Chronic osteoarticular infections such as osteomyelitis or periprosthetic joint infection (PJI) have become a growing problem over the years. The “gold standard” in local antibiotic administration is still the antibiotic-loaded acrylic bone cement (ALABC) which is used in both prophylaxis, because it has been shown it can reduce the risk of infection and used in therapy during a “two-stage surgery” in PJI or in chronic osteomyelitis. We performed morphological analysis of three different formulations of antibiotic-loaded cement (ALABC) using techniques of light microscopy, scanning electron microscopy (SEM) and 3D immunofluorescence, in order to explain how the morphological aspects of cement could influence and modulate antibiotic elution.

  11. High-speed multispectral confocal biomedical imaging.

    Science.gov (United States)

    Carver, Gary E; Locknar, Sarah A; Morrison, William A; Ramanujan, V Krishnan; Farkas, Daniel L

    2014-03-01

    A new approach for generating high-speed multispectral confocal images has been developed. The central concept is that spectra can be acquired for each pixel in a confocal spatial scan by using a fast spectrometer based on optical fiber delay lines. This approach merges fast spectroscopy with standard spatial scanning to create datacubes in real time. The spectrometer is based on a serial array of reflecting spectral elements, delay lines between these elements, and a single element detector. The spatial, spectral, and temporal resolution of the instrument is described and illustrated by multispectral images of laser-induced autofluorescence in biological tissues.

  12. Diffractive elements performance in chromatic confocal microscopy

    Energy Technology Data Exchange (ETDEWEB)

    Garzon, J; Duque, D; Alean, A; Toledo, M [Grupo de Optica y EspectroscopIa, Centro de Ciencia Basica, Universidad Pontificia Bolivariana. Medellin (Colombia); Meneses, J [Laboratorio de Optica y Tratamiento de Senales, Instituto de Fisica, Universidad Industrial de Santander, Bucaramanga (Colombia); Gharbi, T, E-mail: jgarzonr10@une.net.co [Laboratoire d' Optique P. M. Duffieux, UMR-6603 CNR/Universite de Franche-Comte. 16 route de Gray, 25030 Besancon Cedex (France)

    2011-01-01

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

  13. UNC Pembroke Laser Scanning Confocal Microscopy Facility

    Science.gov (United States)

    2016-04-29

    decision, unless so designated by other documentation. 9. SPONSORING/MONITORING AGENCY NAME(S) AND ADDRESS (ES) U.S. Army Research Office P.O. Box 12211...compare to histology images and to maximize data obtained from small groups of blast and toxin-treated samples, and 3) a guillotine and freezer for

  14. Sealing ability of mineral trioxide aggregate, calcium phosphate and polymethylmethacrylate bone cements on root ends prepared using an Erbium: Yttriumaluminium garnet laser and ultrasonics evaluated by confocal laser scanning microscopy.

    Science.gov (United States)

    Girish, C Sabari; Ponnappa, Kc; Girish, Tn; Ponappa, Mc

    2013-07-01

    Surgical endodontic therapy comprises of exposure of the involved root apex, resection of the apical end of the root, preparation of a class I cavity, and insertion of a root end filling material. Mineral trioxide aggregate (MTA) is now the gold standard among all root end filling materials. MTA is however difficult to handle, expensive and has a very slow setting reaction. (1) To compare the sealing ability of MTA, polymethylmethacrylate (PMMA) bone cement and CHITRA Calcium phosphate cement (CPC) when used as root end filling material using Rhodamine B dye evaluated under a confocal laser scanning microscope. (2) To compare the seal of root ends prepared using an ultrasonic retroprep tip and an Er: YAG laser using three different root end filling materials. Statistical analysis was performed using a one-way ANOVA and a two-way ANOVA, independent samples t-test and Scheffe's post hoc test using SPSS Version 16 for Windows. All the three materials, namely MTA, PMMA BONE CEMENT and CHITRA CPC, showed microleakage. Comparison of microleakage showed maximum peak value of 0.86 mm for MTA, 0.24 mm for PMMA bone cement and 1.37 mm for CHITRA CPC. The amount of dye penetration was found to be lesser in root ends prepared using Er: YAG laser when compared with ultrasonics, but the difference was found to be not statistically significant. PMMA bone cement is a better material as root end filling material to prevent apical microleakage. MTA still continues to be a gold standard root end filling material showing minimum microleakage. Er: YAG laser is a better alternative to ultrasonics for root end preparations.

  15. Confocal microscopy imaging of the biofilm matrix

    DEFF Research Database (Denmark)

    Schlafer, Sebastian; Meyer, Rikke L

    2017-01-01

    The extracellular matrix is an integral part of microbial biofilms and an important field of research. Confocal laser scanning microscopy is a valuable tool for the study of biofilms, and in particular of the biofilm matrix, as it allows real-time visualization of fully hydrated, living specimens...

  16. Ultrasensitive and selective detection of mercury (II) in serum based on the gold film sensor using a laser scanning confocal imaging-surface plasmon resonance system in real time

    Science.gov (United States)

    Liu, Sha; Zhang, Hongyan; Liu, Weimin; Wang, Pengfei

    2015-10-01

    Hg2+ ions are one of the most toxic heavy metal ion pollutants, and are caustic and carcinogenic materials with high cellular toxicity. The Hg2+ ions can accumulate in the human body through the food chain and cause serious and permanent damage to the brain with both acute and chronic toxicity. According to the US Environment Protection Agency (EPA) guidelines, Hg2+ ions must be at concentrations below 1 ng/ml (10 nM) in drinking water. If the Hg2+ ions are higher than 2.5 ng/ml in serum, that will bring mercury poisoning. The traditional testing for Hg2+ ions includes atomic absorption, atomic fluorescence, and inductively coupled plasma mass spectrometry. These methods are usually coupled with gas chromatography, high-performance liquid chromatography, and capillary electrophoresis. However, these instrument-based techniques are rather complicated, time-consuming, costly, and unsuitable for online and portable use. An ultrasensitive and selective detection of mercury (II) in serum was investigated using a laser scanning confocal imaging-surface plasmon resonance system (LSCI-SPR). The detection limit was as low as 0.01 ng/ml for Hg2+ ions in fetal calf serum and that is lower than that was required Hg2+ ions must be at concentrations below 1 ng/ml by the US Environment Protection Agency (EPA) guidelines. This sensor was designed on a T-rich, single-stranded DNA (ssDNA)-modified gold film, which can be individually manipulated using specific T-Hg2+-T complex formation. The quenching intensity of the fluorescence images for rhodamine-labeled ssDNA fitted well with the changes in SPR. The changes varied with the Hg2+ ion concentration, which is unaffected by the presence of other metal ions. A good liner relation was got with the coefficients of 0.9116 in 30% fetal calf serums with the linear part over a range of 0.01 ng/ml to10 ng/ml.

  17. Optimal pupil design for confocal microscopy

    Science.gov (United States)

    Patel, Yogesh G.; Rajadhyaksha, Milind; DiMarzio, Charles A.

    2010-02-01

    Confocal reflectance microscopy may enable screening and diagnosis of skin cancers noninvasively and in real-time, as an adjunct to biopsy and pathology. Current instruments are large, complex, and expensive. A simpler, confocal line-scanning microscope may accelerate the translation of confocal microscopy in clinical and surgical dermatology. A confocal reflectance microscope may use a beamsplitter, transmitting and detecting through the pupil, or a divided pupil, or theta configuration, with half used for transmission and half for detection. The divided pupil may offer better sectioning and contrast. We present a Fourier optics model and compare the on-axis irradiance of a confocal point-scanning microscope in both pupil configurations, optimizing the profile of a Gaussian beam in a circular or semicircular aperture. We repeat both calculations with a cylindrical lens which focuses the source to a line. The variable parameter is the fillfactor, h, the ratio of the 1/e2 diameter of the Gaussian beam to the diameter of the full aperture. The optimal values of h, for point scanning are 0.90 (full) and 0.66 for the half-aperture. For line-scanning, the fill-factors are 1.02 (full) and 0.52 (half). Additional parameters to consider are the optimal location of the point-source beam in the divided-pupil configuration, the optimal line width for the line-source, and the width of the aperture in the divided-pupil configuration. Additional figures of merit are field-of-view and sectioning. Use of optimal designs is critical in comparing the experimental performance of the different configurations.

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

    Science.gov (United States)

    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.

  19. Confocal microscopy in the diagnosis of melanoma

    Directory of Open Access Journals (Sweden)

    Apostolović-Stojanović Milica

    2013-01-01

    Full Text Available Melanoma is the most deadly form of skin cancer of melanocytic origin. The tumor has a high malignant potential and early metastasis. Prognosis is directly linked to the stage of the disease. Diagnosing thin melanoma at an early stage offers patients their best chance for survival. The crucial innovation in the early recognition of melanoma was the development of in vivo examination of the skin in high-resolution, by confocal microscopy. Confocal microscopy and its modifications provides a “virtual biopsy“, owing to melanosomes and melanin, which are a source of endogenous contrast. Confocal scanning laser microscopy (CSLM provides visualization of microanatomical structures and cellular detail in real time (pigmented keratinocytes, melanocytes, melanosomes and melanophages in the epidermis, dermoepidermal junction and superficial dermis at a resolution equivalent to the resolution of conventional microscopes. [Projekat Ministarstva nauke Republike Srbije, br. 41002

  20. Confocal Raman microscopy

    CERN Document Server

    Dieing, Thomas; Hollricher, Olaf

    2018-01-01

    This second edition provides a cutting-edge overview of physical, technical and scientific aspects related to the widely used analytical method of confocal Raman microscopy. The book includes expanded background information and adds insights into how confocal Raman microscopy, especially 3D Raman imaging, can be integrated with other methods to produce a variety of correlative microscopy combinations. The benefits are then demonstrated and supported by numerous examples from the fields of materials science, 2D materials, the life sciences, pharmaceutical research and development, as well as the geosciences.

  1. Image inpainting for the differential confocal microscope

    Science.gov (United States)

    Qiu, Lirong; Wang, Lei; Liu, Dali; Hou, Maosheng; Zhao, Weiqian

    2015-02-01

    In the process of zero-crossing trigger measurement of differential confocal microscope, the sample surface features or tilt will cause the edges can't be triggered. Meanwhile, environment vibration can also cause false triggering. In order to restore the invalid information of sample, and realize high-precision surface topography measurement, Total Variation (TV) inpainting model is applied to restore the scanning images. Emulation analysis and experimental verification of this method are investigated. The image inpainting algorithm based on TV model solves the minimization of the energy equation by calculus of variations, and it can effectively restore the non-textured image with noises. Using this algorithm, the simulation confocal laser intensity curve and height curve of standard step sample are restored. After inpainting the intensity curve below the threshold is repaired, the maximum deviation from ideal situation is 0.0042, the corresponding edge contour of height curve is restored, the maximum deviation is 0.1920, which proves the algorithm is effective. Experiment of grating inpainting indicates that the TV algorithm can restore the lost information caused by failed triggering and eliminate the noise caused by false triggering in zero-crossing trigger measurement of differential confocal microscope. The restored image is consistent with the scanning result of OLYMPUS confocal microscope, which can satisfy the request of follow-up measurement analysis.

  2. Video-rate bioluminescence imaging of matrix metalloproteinase-2 secreted from a migrating cell.

    Directory of Open Access Journals (Sweden)

    Takahiro Suzuki

    Full Text Available BACKGROUND: Matrix metalloproteinase-2 (MMP-2 plays an important role in cancer progression and metastasis. MMP-2 is secreted as a pro-enzyme, which is activated by the membrane-bound proteins, and the polarized distribution of secretory and the membrane-associated MMP-2 has been investigated. However, the real-time visualizations of both MMP-2 secretion from the front edge of a migration cell and its distribution on the cell surface have not been reported. METHODOLOGY/PRINCIPAL FINDINGS: The method of video-rate bioluminescence imaging was applied to visualize exocytosis of MMP-2 from a living cell using Gaussia luciferase (GLase as a reporter. The luminescence signals of GLase were detected by a high speed electron-multiplying charge-coupled device camera (EM-CCD camera with a time resolution within 500 ms per image. The fusion protein of MMP-2 to GLase was expressed in a HeLa cell and exocytosis of MMP-2 was detected in a few seconds along the leading edge of a migrating HeLa cell. The membrane-associated MMP-2 was observed at the specific sites on the bottom side of the cells, suggesting that the sites of MMP-2 secretion are different from that of MMP-2 binding. CONCLUSIONS: We were the first to successfully demonstrate secretory dynamics of MMP-2 and the specific sites for polarized distribution of MMP-2 on the cell surface. The video-rate bioluminescence imaging using GLase is a useful method to investigate distribution and dynamics of secreted proteins on the whole surface of polarized cells in real time.

  3. Confocal Raman Microscopy

    CERN Document Server

    Dieing, Thomas; Toporski, Jan

    2011-01-01

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

  4. Digital differential confocal microscopy based on spatial shift transformation.

    Science.gov (United States)

    Liu, J; Wang, Y; Liu, C; Wilson, T; Wang, H; Tan, J

    2014-11-01

    Differential confocal microscopy is a particularly powerful surface profilometry technique in industrial metrology due to its high axial sensitivity and insensitivity to noise. However, the practical implementation of the technique requires the accurate positioning of point detectors in three-dimensions. We describe a simple alternative based on spatial transformation of a through-focus series of images obtained from a homemade beam scanning confocal microscope. This digital differential confocal microscopy approach is described and compared with the traditional Differential confocal microscopy approach. The ease of use of the digital differential confocal microscopy system is illustrated by performing measurements on a 3D standard specimen. © 2014 The Authors Journal of Microscopy © 2014 Royal Microscopical Society.

  5. Confocal laser endomicroscopy

    DEFF Research Database (Denmark)

    Karstensen, John Gásdal; Săftoiu, Adrian; Brynskov, Jørn

    2016-01-01

    BACKGROUND AND STUDY AIMS: Confocal laser endomicroscopy (CLE) has been shown to predict relapse in ulcerative colitis in remission, but little is currently known about its role in Crohn's disease. The aim of this study was to identify reproducible CLE features in patients with Crohn's disease.......01, respectively). Inter- and intraobserver reproducibility was almost perfect (κ > 0.80) or substantial (κ > 0.60) for the majority of CLE parameters. CONCLUSIONS: CLE can identify reproducible microscopic changes in the terminal ileum that are risk factors for relapse in patients with otherwise inactive Crohn...

  6. ConfocalCheck--a software tool for the automated monitoring of confocal microscope performance.

    Directory of Open Access Journals (Sweden)

    Keng Imm Hng

    Full Text Available Laser scanning confocal microscopy has become an invaluable tool in biomedical research but regular quality testing is vital to maintain the system's performance for diagnostic and research purposes. Although many methods have been devised over the years to characterise specific aspects of a confocal microscope like measuring the optical point spread function or the field illumination, only very few analysis tools are available. Our aim was to develop a comprehensive quality assurance framework ranging from image acquisition to automated analysis and documentation. We created standardised test data to assess the performance of the lasers, the objective lenses and other key components required for optimum confocal operation. The ConfocalCheck software presented here analyses the data fully automatically. It creates numerous visual outputs indicating potential issues requiring further investigation. By storing results in a web browser compatible file format the software greatly simplifies record keeping allowing the operator to quickly compare old and new data and to spot developing trends. We demonstrate that the systematic monitoring of confocal performance is essential in a core facility environment and how the quantitative measurements obtained can be used for the detailed characterisation of system components as well as for comparisons across multiple instruments.

  7. ConfocalCheck - A Software Tool for the Automated Monitoring of Confocal Microscope Performance

    Science.gov (United States)

    Hng, Keng Imm; Dormann, Dirk

    2013-01-01

    Laser scanning confocal microscopy has become an invaluable tool in biomedical research but regular quality testing is vital to maintain the system’s performance for diagnostic and research purposes. Although many methods have been devised over the years to characterise specific aspects of a confocal microscope like measuring the optical point spread function or the field illumination, only very few analysis tools are available. Our aim was to develop a comprehensive quality assurance framework ranging from image acquisition to automated analysis and documentation. We created standardised test data to assess the performance of the lasers, the objective lenses and other key components required for optimum confocal operation. The ConfocalCheck software presented here analyses the data fully automatically. It creates numerous visual outputs indicating potential issues requiring further investigation. By storing results in a web browser compatible file format the software greatly simplifies record keeping allowing the operator to quickly compare old and new data and to spot developing trends. We demonstrate that the systematic monitoring of confocal performance is essential in a core facility environment and how the quantitative measurements obtained can be used for the detailed characterisation of system components as well as for comparisons across multiple instruments. PMID:24224017

  8. Ultraminiature video-rate forward-view spectrally encoded endoscopy with straight axis configuration

    Science.gov (United States)

    Wang, Zhuo; Wu, Tzu-Yu; Hamm, Mark A.; Altshuler, Alexander; Mach, Anderson T.; Gilbody, Donald I.; Wu, Bin; Ganesan, Santosh N.; Chung, James P.; Ikuta, Mitsuhiro; Brauer, Jacob S.; Takeuchi, Seiji; Honda, Tokuyuki

    2017-02-01

    As one of the smallest endoscopes that have been demonstrated, the spectrally encoded endoscope (SEE) shows potential for the use in minimally invasive surgeries. While the original SEE is designed for side-view applications, the forwardview (FV) scope is more desired by physicians for many clinical applications because it provides a more natural navigation. Several FV SEEs have been designed in the past, which involve either multiple optical elements or one optical element with multiple optically active surfaces. Here we report a complete FV SEE which comprises a rotating illumination probe within a drive cable, a sheath and a window to cover the optics, a customized spectrometer, hardware controllers for both motor control and synchronization, and a software suite to capture, process and store images and videos. In this solution, the optical axis is straight and the dispersion element, i.e. the grating, is designed such that the slightly focused light after the focusing element will be dispersed by the grating, covering forward view angles with high diffraction efficiencies. As such, the illumination probe is fabricated with a diameter of only 275 μm. The twodimensional video-rate image acquisition is realized by rotating the illumination optics at 30 Hz. In one finished design, the scope diameter including the window assembly is 1.2 mm.

  9. Video Rating in Neurodegenerative Disease Clinical Trials: The Experience of PRION-1

    Directory of Open Access Journals (Sweden)

    Christopher Carswell

    2012-08-01

    Full Text Available Background/Aims: Large clinical trials including patients with uncommon diseases involve assessors in different geographical locations, resulting in considerable inter-rater variability in assessment scores. As video recordings of examinations, which can be individually rated, may eliminate such variability, we measured the agreement between a single video rater and multiple examining physicians in the context of PRION-1, a clinical trial of the antimalarial drug quinacrine in human prion diseases. Methods: We analysed a 43-component neurocognitive assessment battery, on 101 patients with Creutzfeldt-Jakob disease, focusing on the correlation and agreement between examining physicians and a single video rater. Results: In total, 335 videos of examinations of 101 patients who were video-recorded over the 4-year trial period were assessed. For neurocognitive examination, inter-observer concordance was generally excellent. Highly visual neurological examination domains (e.g. finger-nose-finger assessment of ataxia had good inter-rater correlation, whereas those dependent on non-visual clues (e.g. power or reflexes correlated poorly. Some non-visual neurological domains were surprisingly concordant, such as limb muscle tone. Conclusion: Cognitive assessments and selected neurological domains can be practically and accurately recorded in a clinical trial using video rating. Video recording of examinations is a valuable addition to any trial provided appropriate selection of assessment instruments is used and rigorous training of assessors is undertaken.

  10. Digital confocal microscopy through a multimode fiber.

    Science.gov (United States)

    Loterie, Damien; Farahi, Salma; Papadopoulos, Ioannis; Goy, Alexandre; Psaltis, Demetri; Moser, Christophe

    2015-09-07

    Acquiring high-contrast optical images deep inside biological tissues is still a challenging problem. Confocal microscopy is an important tool for biomedical imaging since it improves image quality by rejecting background signals. However, it suffers from low sensitivity in deep tissues due to light scattering. Recently, multimode fibers have provided a new paradigm for minimally invasive endoscopic imaging by controlling light propagation through them. Here we introduce a combined imaging technique where confocal images are acquired through a multimode fiber. We achieve this by digitally engineering the excitation wavefront and then applying a virtual digital pinhole on the collected signal. In this way, we are able to acquire images through the fiber with significantly increased contrast. With a fiber of numerical aperture 0.22, we achieve a lateral resolution of 1.5µm, and an axial resolution of 12.7µm. The point-scanning rate is currently limited by our spatial light modulator (20Hz).

  11. Real time three-dimensional space video rate sensors for millimeter waves imaging based very inexpensive plasma LED lamps

    Science.gov (United States)

    Levanon, Assaf; Yitzhaky, Yitzhak; Kopeika, Natan S.; Rozban, Daniel; Abramovich, Amir

    2014-10-01

    In recent years, much effort has been invested to develop inexpensive but sensitive Millimeter Wave (MMW) detectors that can be used in focal plane arrays (FPAs), in order to implement real time MMW imaging. Real time MMW imaging systems are required for many varied applications in many fields as homeland security, medicine, communications, military products and space technology. It is mainly because this radiation has high penetration and good navigability through dust storm, fog, heavy rain, dielectric materials, biological tissue, and diverse materials. Moreover, the atmospheric attenuation in this range of the spectrum is relatively low and the scattering is also low compared to NIR and VIS. The lack of inexpensive room temperature imaging systems makes it difficult to provide a suitable MMW system for many of the above applications. In last few years we advanced in research and development of sensors using very inexpensive (30-50 cents) Glow Discharge Detector (GDD) plasma indicator lamps as MMW detectors. This paper presents three kinds of GDD sensor based lamp Focal Plane Arrays (FPA). Those three kinds of cameras are different in the number of detectors, scanning operation, and detection method. The 1st and 2nd generations are 8 × 8 pixel array and an 18 × 2 mono-rail scanner array respectively, both of them for direct detection and limited to fixed imaging. The last designed sensor is a multiplexing frame rate of 16x16 GDD FPA. It permits real time video rate imaging of 30 frames/ sec and comprehensive 3D MMW imaging. The principle of detection in this sensor is a frequency modulated continuous wave (FMCW) system while each of the 16 GDD pixel lines is sampled simultaneously. Direct detection is also possible and can be done with a friendly user interface. This FPA sensor is built over 256 commercial GDD lamps with 3 mm diameter International Light, Inc., Peabody, MA model 527 Ne indicator lamps as pixel detectors. All three sensors are fully supported

  12. Fluorescence confocal microscopy for pathologists.

    Science.gov (United States)

    Ragazzi, Moira; Piana, Simonetta; Longo, Caterina; Castagnetti, Fabio; Foroni, Monica; Ferrari, Guglielmo; Gardini, Giorgio; Pellacani, Giovanni

    2014-03-01

    Confocal microscopy is a non-invasive method of optical imaging that may provide microscopic images of untreated tissue that correspond almost perfectly to hematoxylin- and eosin-stained slides. Nowadays, following two confocal imaging systems are available: (1) reflectance confocal microscopy, based on the natural differences in refractive indices of subcellular structures within the tissues; (2) fluorescence confocal microscopy, based on the use of fluorochromes, such as acridine orange, to increase the contrast epithelium-stroma. In clinical practice to date, confocal microscopy has been used with the goal of obviating the need for excision biopsies, thereby reducing the need for pathological examination. The aim of our study was to test fluorescence confocal microscopy on different types of surgical specimens, specifically breast, lymph node, thyroid, and colon. The confocal images were correlated to the corresponding histological sections in order to provide a morphologic parallel and to highlight current limitations and possible applications of this technology for surgical pathology practice. As a result, neoplastic tissues were easily distinguishable from normal structures and reactive processes such as fibrosis; the use of fluorescence enhanced contrast and image quality in confocal microscopy without compromising final histologic evaluation. Finally, the fluorescence confocal microscopy images of the adipose tissue were as accurate as those of conventional histology and were devoid of the frozen-section-related artefacts that can compromise intraoperative evaluation. Despite some limitations mainly related to black/white images, which require training in imaging interpretation, this study confirms that fluorescence confocal microscopy may represent an alternative to frozen sections in the assessment of margin status in selected settings or when the conservation of the specimen is crucial. This is the first study to employ fluorescent confocal microscopy on

  13. A line-scanning semi-confocal multi-photon fluorescence microscope with a simultaneous broadband spectral acquisition and its application to the study of the thylakoid membrane of a cyanobacterium Anabaena PCC7120.

    Science.gov (United States)

    Kumazaki, Shigeichi; Hasegawa, Makoto; Ghoneim, Mohammad; Shimizu, Yugo; Okamoto, Kenji; Nishiyama, Masayoshi; Oh-Oka, Hirozo; Terazima, Masahide

    2007-11-01

    We describe the construction and characterization of a laser-line-scanning microscope capable of detection of broad fluorescence spectra with a resolution of 1 nm. A near-infrared femtosecond pulse train at 800 nm was illuminated on a line (one lateral axis, denoted as X axis) in a specimen by a resonant scanning mirror oscillating at 7.9 kHz, and total multi-photon-induced fluorescence from the linear region was focused on the slit of an imaging polychromator. An electron-multiplying CCD camera was used to resolve fluorescence of different colours at different horizontal pixels and fluorescence of different spatial positions in a specimen at different vertical pixels. Scanning on the other two axes (Y and Z) was achieved by a closed-loop controlled sample scanning stage and a piezo-driven objective actuator. The full widths at half maximum of the point-spread function of the system were estimated to be 0.39-0.40, 0.33 and 0.56-0.59 mum for the X (lateral axis along the line-scan), Y (the other lateral axis) and Z axes (the axial direction), respectively, at fluorescence wavelengths between 644 and 690 nm. A biological application of this microscope was demonstrated in a study of the sub-cellular fluorescence spectra of thylakoid membranes in a cyanobacterium, Anabaena PCC7120. It was found that the fluorescence intensity ratio between chlorophyll molecules mainly of photosystem II and phycobilin molecules of phycobilisome (chlorophyll/phycobilin), in the thylakoid membranes, became lower as one probed deeper inside the cells. This was attributable not to position dependence of re-absorption or scattering effects, but to an intrinsic change in the local physiological state of the thylakoid membrane, with the help of a transmission spectral measurement of sub-cellular domains. The efficiency of the new line-scanning spectromicroscope was estimated in comparison with our own point-by-point scanning spectromicroscope. Under typical conditions of observing

  14. Confocal laser endomicroscopy in the “in vivo” histological diagnosis of the gastrointestinal tract

    OpenAIRE

    Palma, Giovanni D De

    2009-01-01

    Recent technological advances in miniaturization have allowed for a confocal scanning microscope to be integrated into a conventional flexible endoscope, or into trans-endoscopic probes, a technique now known as confocal endomicroscopy or confocal laser endomicroscopy. This newly-developed technology has enabled endoscopists to collect real-time in vivo histological images or “virtual biopsies” of the gastrointestinal mucosa during endoscopy, and has stimulated significant interest in the app...

  15. An evaluation of confocal versus conventional imaging of biological structures by fluorescence light microscopy.

    Science.gov (United States)

    White, J G; Amos, W B; Fordham, M

    1987-07-01

    Scanning confocal microscopes offer improved rejection of out-of-focus noise and greater resolution than conventional imaging. In such a microscope, the imaging and condenser lenses are identical and confocal. These two lenses are replaced by a single lens when epi-illumination is used, making confocal imaging particularly applicable to incident light microscopy. We describe the results we have obtained with a confocal system in which scanning is performed by moving the light beam, rather than the stage. This system is considerably faster than the scanned stage microscope and is easy to use. We have found that confocal imaging gives greatly enhanced images of biological structures viewed with epifluorescence. The improvements are such that it is possible to optically section thick specimens with little degradation in the image quality of interior sections.

  16. Confocal direct imaging Raman microscope: Design and applications in biology

    NARCIS (Netherlands)

    Sijtsema, N M; Wouters, S D; Grauw, C J De; Otto, C; Greve, J

    1998-01-01

    A confocal direct imaging Raman microscope (CDIRM) based on two synchronized scanning mirrors, a monochromator, and two charge-coupled device (CCD) cameras has been developed. With this system it is possible to make both Raman spectra of a small measurement volume and images of a larger sample area

  17. Fluorescence Blinking and Photobleaching of Single Terrylenediimide Molecules Studied with a Confocal Microscope

    NARCIS (Netherlands)

    Göhde, Jr.; Fischer, U.C.; Fuchs, H.; Tittel, J.; Basché, Th.; Bräuchle, Ch.; Herrmann, A.; Müllen, K.

    1998-01-01

    Single terrylenediimide molecules diluted in a 20-nm-thick polyvinylbutyral polymer film were localized and observed by scanning confocal fluorescence microscopy. A modular and compact confocal microscope and the high optical stability of the molecules allowed a repeated imaging and observation over

  18. In vivo Confocal Microscopy Report after Lasik with Sequential Accelerated Corneal Collagen Cross-Linking Treatment

    National Research Council Canada - National Science Library

    Mazzotta, Cosimo; Balestrazzi, Angelo; Traversi, Claudio; Caragiuli, Stefano; Caporossi, Aldo

    2014-01-01

    ...) treatment combined with sequential high-fluence accelerated corneal collagen cross-linking, denominated Lasik XTra, by means of HRT II laser scanning in vivo confocal microscopy after a 6-month follow-up...

  19. A GRISM-based probe for spectrally encoded confocal microscopy.

    Science.gov (United States)

    Pitris, C; Bouma, B; Shiskov, M; Tearney, G

    2003-01-27

    Spectrally encoded confocal microscopy (SECM) is a novel approach for obtaining high resolution, depth-sectioned images of microstructure within turbid samples. By encoding one spatial dimension in wavelength, imaging probes can be greatly simplified compared to standard scanning confocal microscopes, potentially enabling endoscopic implementation. The use of a diffraction grating for spectral encoding, however, skews the optical axis through the probe, thus complicating the design of narrow diameter instruments. In this Letter, we describe a novel use of a single-optical-axis element based on high index-of-refraction prisms and a transmission holographic grating for the design of narrow diameter SECM devices. Confocal images obtained with a 10.0 mm probe demonstrate a transverse resolution of 1.1 microm and a field of view of 650 microm.

  20. Video rate nine-band multispectral short-wave infrared sensor.

    Science.gov (United States)

    Kutteruf, Mary R; Yetzbacher, Michael K; DePrenger, Michael J; Novak, Kyle M; Miller, Corey A; Downes, Trijntje Valerie; Kanaev, Andrey V

    2014-05-01

    Short-wave infrared (SWIR) imaging sensors are increasingly being used in surveillance and reconnaissance systems due to the reduced scatter in haze and the spectral response of materials over this wavelength range. Typically SWIR images have been provided either as full motion video from framing panchromatic systems or as spectral data cubes from line-scanning hyperspectral or multispectral systems. Here, we describe and characterize a system that bridges this divide, providing nine-band spectral images at 30 Hz. The system integrates a custom array of filters onto a commercial SWIR InGaAs array. We measure the filter placement and spectral response. We demonstrate a simple simulation technique to facilitate optimization of band selection for future sensors.

  1. Fungal keratitis - improving diagnostics by confocal microscopy.

    Science.gov (United States)

    Nielsen, E; Heegaard, S; Prause, J U; Ivarsen, A; Mortensen, K L; Hjortdal, J

    2013-09-01

    Introducing a simple image grading system to support the interpretation of in vivo confocal microscopy (IVCM) images in filamentous fungal keratitis. Clinical and confocal studies took place at the Department of Ophthalmology, Aarhus University Hospital, Denmark. Histopathological analysis was performed at the Eye Pathology Institute, Department of Neuroscience and Pharmacology, University of Copenhagen, Denmark. A recent series of consecutive patients with filamentous fungal keratitis is presented to demonstrate the results from in-house IVCM. Based upon our experience with IVCM and previously published images, we composed a grading system for interpreting IVCM images of filamentous fungal keratitis. A recent case series of filamentous fungal keratitis from 2011 to 2012 was examined. There were 3 male and 3 female patients. Mean age was 44.5 years (range 12-69), 6 out of 17 (35%) cultures were positive and a total of 6/7 (86%) IVCM scans were positive. Three different categories of IVCM results for the grading of diagnostic certainty were formed. IVCM is a valuable tool for diagnosing filamentous fungal keratitis. In order to improve the reliability of IVCM, we suggest implementing a simple and clinically applicable grading system for aiding the interpretation of IVCM images of filamentous fungal keratitis.

  2. Fungal Keratitis - Improving Diagnostics by Confocal Microscopy

    Directory of Open Access Journals (Sweden)

    Esben Nielsen

    2013-12-01

    Full Text Available Purpose: Introducing a simple image grading system to support the interpretation of in vivo confocal microscopy (IVCM images in filamentous fungal keratitis. Setting: Clinical and confocal studies took place at the Department of Ophthalmology, Aarhus University Hospital, Denmark. Histopathological analysis was performed at the Eye Pathology Institute, Department of Neuroscience and Pharmacology, University of Copenhagen, Denmark. Methods: A recent series of consecutive patients with filamentous fungal keratitis is presented to demonstrate the results from in-house IVCM. Based upon our experience with IVCM and previously published images, we composed a grading system for interpreting IVCM images of filamentous fungal keratitis. Results: A recent case series of filamentous fungal keratitis from 2011 to 2012 was examined. There were 3 male and 3 female patients. Mean age was 44.5 years (range 12-69, 6 out of 17 (35% cultures were positive and a total of 6/7 (86% IVCM scans were positive. Three different categories of IVCM results for the grading of diagnostic certainty were formed. Conclusion: IVCM is a valuable tool for diagnosing filamentous fungal keratitis. In order to improve the reliability of IVCM, we suggest implementing a simple and clinically applicable grading system for aiding the interpretation of IVCM images of filamentous fungal keratitis.

  3. Real-Time Confocal Imaging Of The Living Eye

    Science.gov (United States)

    Jester, James V.; Cavanagh, H. Dwight; Essepian, John; Shields, William J.; Lemp, Michael A.

    1989-12-01

    In 1986, we adapted the Tandem Scanning Reflected Light Microscope of Petran and Hadraysky to permit non-invasive, confocal imaging of the living eye in real-time. We were first to obtain stable, confocal optical sections in vivo, from human and animal eyes. Using confocal imaging systems we have now studied living, normal volunteers, rabbits, cats and primates sequentially, non-invasively, and in real-time. The continued development of real-time confocal imaging systems will unlock the door to a new field of cell biology involving for the first time the study of dynamic cellular processes in living organ systems. Towards this end we have concentrated our initial studies on three areas (1) evaluation of confocal microscope systems for real-time image acquisition, (2) studies of the living normal cornea (epithelium, stroma, endothelium) in human and other species; and (3) sequential wound-healing responses in the cornea in single animals to lamellar-keratectomy injury (cellular migration, inflammation, scarring). We believe that this instrument represents an important, new paradigm for research in cell biology and pathology and that it will fundamentally alter all experimental and clinical approaches in future years.

  4. Digital holographic interferometry accelerated with GPU: application in mechanical micro-deformation measurement operating at video rate

    Science.gov (United States)

    Múnera Ortiz, N.; Trujillo, C. A.; García-Sucerquia, J.

    2013-11-01

    The quantification of the deformations presented by mechanical parts is a useful tool for several applications in engineering; regularly this quantification is performed a posteriori. In this work, a digital holographic interferometer for measuring micro-deformation at video rate is presented. The interferometer is developed with the use of the parallel paradigm of CUDA™ (Compute Unified Device Architecture). A commercial Graphics Processor Unit (GPU) is used to accelerate phase processing from the recorded holograms. The proposed method can process record holograms of 1024x1024 pixels in 48 milliseconds. At the best performance of the method, it processes 21 frames per second (FPS). This benchmark surpasses 133-times the best performance of the method on a regular CPU.

  5. Feasibility of Radon projection acquisition for compressive imaging in MMW region based new video rate 16×16 GDD FPA camera

    Science.gov (United States)

    Levanon, Assaf; Konstantinovsky, Michael; Kopeika, Natan S.; Yitzhaky, Yitzhak; Stern, A.; Turak, Svetlana; Abramovich, Amir

    2015-05-01

    In this article we present preliminary results for the combination of two interesting fields in the last few years: 1) Compressed imaging (CI), which is a joint sensing and compressing process, that attempts to exploit the large redundancy in typical images in order to capture fewer samples than usual. 2) Millimeter Waves (MMW) imaging. MMW based imaging systems are required for a large variety of applications in many growing fields such as medical treatments, homeland security, concealed weapon detection, and space technology. Moreover, the possibility to create a reliable imaging in low visibility conditions such as heavy cloud, smoke, fog and sandstorms in the MMW region, generate high interest from military groups in order to be ready for new combat. The lack of inexpensive room temperature imaging sensors makes it difficult to provide a suitable MMW system for many of the above applications. A system based on Glow Discharge Detector (GDD) Focal Plane Arrays (FPA) can be very efficient in real time imaging with significant results. The GDD is located in free space and it can detect MMW radiation almost isotropically. In this article, we present a new approach of reconstruction MMW imaging by rotation scanning of the target. The Collection process here, based on Radon projections allows implementation of the compressive sensing principles into the MMW region. Feasibility of concept was obtained as radon line imaging results. MMW imaging results with our resent sensor are also presented for the first time. The multiplexing frame rate of 16×16 GDD FPA permits real time video rate imaging of 30 frames per second and comprehensive 3D MMW imaging. It uses commercial GDD lamps with 3mm diameter, Ne indicator lamps as pixel detectors. Combination of these two fields should make significant improvement in MMW region imaging research, and new various of possibilities in compressing sensing technique.

  6. Analysis of reactive oxygen species in the guard cell of wheat stoma with confocal microscope.

    Science.gov (United States)

    Liu, Dongwu; Chen, Zhiwei; Shi, Peiguo; Wang, Xue; Cai, Weiwei

    2011-09-01

    Recently, the laser-scanning confocal microscope has become a routine technique and indispensable tool for cell biological studies. Previous studies indicated that reactive oxygen species (ROS) were generated in tobacco epidermal cells with confocal microscope. In the present studies, the probe 2',7'-dichlorof luorescein diacetate (H₂DCF-DA) was used to research the change of ROS in the guard cell of wheat stoma, and catalase (CAT) was used to demonstrate that ROS had been labeled. The laser-scanning mode of confocal microscope was XYT, and the time interval between two sections was 1.6351 s. Sixty optical sections were acquired with the laser-scanning confocal microscope, and CAT (60,000 U mg⁻¹) was added after four optical sections were scanned. Furthermore, the region of interest (ROI) was circled and the fluorescence intensity of ROS was quantified with Leica Confocal Software. The quantitative data were exported and the trend chart was made with software Excell. The results indicated that ROS were produced intracellularly in stomatal guard cells, and the quantified fluorescence intensity of ROS was declined with CAT added. It is a good method to research the instantaneous change of ROS in plant cells with confocal microscope and fluorescence probe H₂DCF-DA. Copyright © 2010 Wiley-Liss, Inc.

  7. A confocal laser scanning microscopic study on thermoresponsive ...

    Indian Academy of Sciences (India)

    Monodisperse poly(N -isopropylacrylamide) (PNIPAM) particles loaded with cadmium telluride (CdTe) quantum dots (QDs) of two different sizes (4.7 nm and 5.6 nm) were synthesized in aqueous medium by bonding the capping agent on the quantum dots to the amide groups of PNIPAM and incubating the samples at 45° ...

  8. Real-time intravascular photoacoustic-ultrasound imaging of lipid-laden plaque at speed of video-rate level

    Science.gov (United States)

    Hui, Jie; Cao, Yingchun; Zhang, Yi; Kole, Ayeeshik; Wang, Pu; Yu, Guangli; Eakins, Gregory; Sturek, Michael; Chen, Weibiao; Cheng, Ji-Xin

    2017-03-01

    Intravascular photoacoustic-ultrasound (IVPA-US) imaging is an emerging hybrid modality for the detection of lipidladen plaques by providing simultaneous morphological and lipid-specific chemical information of an artery wall. The clinical utility of IVPA-US technology requires real-time imaging and display at speed of video-rate level. Here, we demonstrate a compact and portable IVPA-US system capable of imaging at up to 25 frames per second in real-time display mode. This unprecedented imaging speed was achieved by concurrent innovations in excitation laser source, rotary joint assembly, 1 mm IVPA-US catheter, differentiated A-line strategy, and real-time image processing and display algorithms. By imaging pulsatile motion at different imaging speeds, 16 frames per second was deemed to be adequate to suppress motion artifacts from cardiac pulsation for in vivo applications. Our lateral resolution results further verified the number of A-lines used for a cross-sectional IVPA image reconstruction. The translational capability of this system for the detection of lipid-laden plaques was validated by ex vivo imaging of an atherosclerotic human coronary artery at 16 frames per second, which showed strong correlation to gold-standard histopathology.

  9. Parallel detection experiment of fluorescence confocal microscopy using DMD.

    Science.gov (United States)

    Wang, Qingqing; Zheng, Jihong; Wang, Kangni; Gui, Kun; Guo, Hanming; Zhuang, Songlin

    2016-05-01

    Parallel detection of fluorescence confocal microscopy (PDFCM) system based on Digital Micromirror Device (DMD) is reported in this paper in order to realize simultaneous multi-channel imaging and improve detection speed. DMD is added into PDFCM system, working to take replace of the single traditional pinhole in the confocal system, which divides the laser source into multiple excitation beams. The PDFCM imaging system based on DMD is experimentally set up. The multi-channel image of fluorescence signal of potato cells sample is detected by parallel lateral scanning in order to verify the feasibility of introducing the DMD into fluorescence confocal microscope. In addition, for the purpose of characterizing the microscope, the depth response curve is also acquired. The experimental result shows that in contrast to conventional microscopy, the DMD-based PDFCM system has higher axial resolution and faster detection speed, which may bring some potential benefits in the biology and medicine analysis. SCANNING 38:234-239, 2016. © 2015 Wiley Periodicals, Inc. © Wiley Periodicals, Inc.

  10. Corneal Cell Morphology in Keratoconus: A Confocal Microscopic Observation

    Science.gov (United States)

    Ghosh, Somnath; Mutalib, Haliza Abdul; Kaur, Sharanjeet; Ghoshal, Rituparna; Retnasabapathy, Shamala

    2017-01-01

    Purpose To evaluate corneal cell morphology in patients with keratoconus using an in vivo slit scanning confocal microscope. Methods A cross-sectional study was conducted to evaluate the corneal cell morphology of 47 keratoconus patients and 32 healthy eyes without any ocular disease. New keratoconus patients with different disease severities and without any other ocular co-morbidity were recruited from the ophthalmology department of a public hospital in Malaysia from June 2013 to May 2014. Corneal cell morphology was evaluated using an in vivo slit-scanning confocal microscope. Qualitative and quantitative data were analysed using a grading scale and the Nidek Advanced Visual Information System software, respectively. Results The corneal cell morphology of patients with keratoconus was significantly different from that of healthy eyes except in endothelial cell density (P = 0.072). In the keratoconus group, increased level of stromal haze, alterations such as the elongation of keratocyte nuclei and clustering of cells at the anterior stroma, and dark bands in the posterior stroma were observed with increased severity of the disease. The mean anterior and posterior stromal keratocyte densities and cell areas among the different stages of keratoconus were significantly different (P 0.05) among the three stages of keratoconus. Conclusion Confocal microscopy observation showed significant changes in corneal cell morphology in keratoconic cornea from normal healthy cornea. Analysis also showed significant changes in different severities of keratoconus. Understanding the corneal cell morphology changes in keratoconus may help in the long-term monitoring and management of keratoconus. PMID:28894403

  11. Confocal Raman Microspectroscopy of Oral Streptococci

    Science.gov (United States)

    Beier, Brooke D.

    Raman spectroscopy has been used in a variety of applications throughout the field of biomedical optics. It has the ability to acquire chemically-specific information in a non-invasive manner, without the need for exogenous markers. This makes it useful in the identification of bacterial species, as well as in the study of tissues and other cells. In this work, a species identification model has been created in order to discriminate between the oral bacterial species Streptococcus sanguinis and Streptococcus mutans. These are two of the most prevalent species within the human mouth and their relative concentrations can be an indicator of a patient's oral health and risk of tooth decay. They are predominantly found within plaque on the tooth's surface. To study a simplified model for dental plaque, we have examined S. sanguinis and S. mutans grown in biofilm forms. Raman spectroscopy has been implemented here through a confocal microscope. The optical system has been equipped with computationally controlled stages to allow for automated scanning, including autofocusing to probe a consistent depth within a sample. A spectrum has been acquired from each position within a scan and sent for spectral preprocessing before being submitted for species identification. This preprocessing includes an algorithm that has been developed to remove fluorescence features from known contaminants within the confocal volume, to include signal from a fluorescent substrate. Species classification has been accomplished using a principal component score-fed logistic regression model constructed from a variety of biofilm samples that have been transferred and allowed to dry, as might occur with the study of plaque samples. This binary classification model has been validated on other samples with identical preparations. The model has also been transferred to determine the species of hydrated biofilms studied in situ. Artificially mixed biofilms have been examined to test the spatial

  12. Detecção de alterações na camada de fibras nervosas da retina por meio do laser confocal polarizado em hipertensão ocular antes do surgimento de defeitos perimétricos Detection of retinal nerve fiber layer changes in ocular hypertension with scanning laser polarimetry before the appearance of perimetric defects

    Directory of Open Access Journals (Sweden)

    Roberto Lauande-Pimentel

    2004-10-01

    Full Text Available OBJETIVO: Avaliar a capacidade do laser confocal polarizado (LCP em detectar alterações na camada de fibras nervosas (CFN de hipertensos oculares antes do aparecimento de alteração campimétrica. Desenho- Retrospectivo, caso-controle. MÉTODOS: Pacientes hipertensos oculares divididos em dois grupos: a estáveis e b conversores (que progrediram com dano perimétrico glaucomatoso. Parâmetros de retardo obtidos por meio do programa NFA/GDx. RESULTADOS: Um total de 108 pacientes estáveis e 13 conversores foram estudados por período médio de seguimento acima de 35 meses nos dois grupos. Diversos parâmetros do LCP mostraram diferenças significativas na espessura da CFN entre os dois grupos no inicio do seguimento (média de 27,4 meses antes do aparecimento de lesão perimétrica. Os parâmetros The Number, Maximum Modulation e Superior Average permaneceram diferentes entre os grupos no início e no final do seguimento. O odds ratio para desenvolvimento de conversão perimétrica, dado um resultado de The Number alterado (>32, foi estimado em 7,9 para esta série. CONCLUSÕES: O LCP foi capaz de detectar alterações significativas na CFN no grupo de hipertensos oculares que desenvolveram posteriormente lesão perimétrica glaucomatosa. Neste estudo, o resultado inicial anormal de The Number foi o principal fator de risco para desenvolvimento de alteração perimétrica futura em pacientes hipertensos oculares.OBJECTIVE: To evaluate the ability of the Confocal Scanning Laser Polarimeter (SLP to detect glaucoma alterations before the appearance of perimetric defects. Design- retrospective, case-control. METHODS: Ocular hipertensive patients divided in to two groups: a stable and b conversors (that have conversed to perimetric defined glaucoma. Nerve Fiber Analyser/GDx parameters of retardation. RESULTS: A total of 108 stable and 13 conversors were evaluated for a mean period over 35 months in each group. At the initial examination, several

  13. Automated identification of epidermal keratinocytes in reflectance confocal microscopy

    Science.gov (United States)

    Gareau, Dan

    2011-03-01

    Keratinocytes in skin epidermis, which have bright cytoplasmic contrast and dark nuclear contrast in reflectance confocal microscopy (RCM), were modeled with a simple error function reflectance profile: erf( ). Forty-two example keratinocytes were identified as a training set which characterized the nuclear size a = 8.6+/-2.8 μm and reflectance gradient b = 3.6+/-2.1 μm at the nuclear/cytoplasmic boundary. These mean a and b parameters were used to create a rotationally symmetric erf( ) mask that approximated the mean keratinocyte image. A computer vision algorithm used an erf( ) mask to scan RCM images, identifying the coordinates of keratinocytes. Applying the mask to the confocal data identified the positions of keratinocytes in the epidermis. This simple model may be used to noninvasively evaluate keratinocyte populations as a quantitative morphometric diagnostic in skin cancer detection and evaluation of dermatological cosmetics.

  14. Full-field interferometric confocal microscopy using a VCSEL array.

    Science.gov (United States)

    Redding, Brandon; Bromberg, Yaron; Choma, Michael A; Cao, Hui

    2014-08-01

    We present an interferometric confocal microscope using an array of 1200 vertical cavity surface emitting lasers (VCSELs) coupled to a multimode fiber. Spatial coherence gating provides ~18,000 continuous virtual pinholes, allowing an entire en face plane to be imaged in a snapshot. This approach maintains the same optical sectioning as a scanning confocal microscope without moving parts, while the high power of the VCSEL array (∼5  mW per laser) enables high-speed image acquisition with integration times as short as 100 μs. Interferometric detection also recovers the phase of the image, enabling quantitative phase measurements and improving the contrast when imaging phase objects.

  15. Deep stroma investigation by confocal microscopy

    Science.gov (United States)

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

    2015-03-01

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

  16. Comparison of confocal biomicroscopy and noncontact specular microscopy for evaluation of the corneal endothelium.

    Science.gov (United States)

    Hara, Makiko; Morishige, Naoyuki; Chikama, Tai-Ichiro; Nishida, Teruo

    2003-08-01

    To compare the clinical efficacy of confocal biomicroscopy with that of noncontact specular microscopy for the evaluation of the corneal endothelium. The corneal endothelium was examined in 14 normal subjects (28 eyes) and in 6 patients (11 eyes) with Fuchs corneal endothelial dystrophy using a noncontact specular microscope (SP-2000P, Topcon, Japan) and a confocal biomicroscope (ConfoScan, Tomey, Japan). The images and the calculated densities of corneal endothelial cells obtained by the 2 techniques were compared. For normal subjects, the images of corneal endothelial cells obtained by the 2 techniques were almost identical, although the density of these cells determined by confocal biomicroscopy (2916 +/- 334 cells/mm2) was slightly higher than that determined by noncontact specular microscopy (2765 +/- 323 cells/mm2). In contrast, whereas clear images of corneal endothelial cells, allowing the determination of cell density, were obtained for all 11 eyes of the patient group by confocal biomicroscopy, clear images were obtained for only 4 of these 11 eyes (36.4%) by noncontact specular microscopy. Both noncontact specular microscopy and confocal biomicroscopy revealed the shapes and number of endothelial cells in the normal cornea. However, for corneas with Fuchs dystrophy, clear images were obtained only by confocal biomicroscopy. Confocal biomicroscopy is thus an effective tool for evaluation of the diseased corneal endothelium.

  17. Calcium oxalate crystal growth modification; investigations with confocal Raman microscopy

    Science.gov (United States)

    McMulkin, Calum J.; Massi, Massimiliano; Jones, Franca

    2017-06-01

    Confocal Raman Microscopy (CRM) in combination with a photophysical investigation has been employed to give insight into the interaction between calcium oxalate monohydrate (COM) and a series of tetrazole containing crystal growth modifier's (CGM's), in conjunction with characterisation of morphological changes using scanning electron and optical microscopy. The tetrazole CGM's were found to interact by surface adsorption with minimal morphological changes to the COM crystals however, significant interactions via chemisorption were observed; it was discovered that the chemisorption is sufficiently strong for aggregation of the tetrazole species to occur within the crystal during crystallisation.

  18. Spatial scan statistics using elliptic windows

    DEFF Research Database (Denmark)

    Christiansen, Lasse Engbo; Andersen, Jens Strodl; Wegener, Henrik Caspar

    2006-01-01

    The spatial scan statistic is widely used to search for clusters. This article shows that the usually applied elimination of secondary clusters as implemented in SatScan is sensitive to smooth changes in the shape of the clusters. We present an algorithm for generation of a set of confocal elliptic...

  19. Generalized vector wave theory for ultrahigh resolution confocal optical microscopy.

    Science.gov (United States)

    Yang, Ken; Xie, Xiangsheng; Zhou, Jianying

    2017-01-01

    Polarization modulation of a tightly focused beam in a confocal imaging scheme is considered for incident and collected light fields. Rigorous vector wave theory of a confocal optical microscopy is developed, which provides clear physical pictures without the requirement for fragmentary calculations. Multiple spatial modulations on polarization, phase, or amplitude of the illuminating and the detected beams can be mathematically described by a uniform expression. Linear and nonlinear excitation schemes are derived with tailored excitation and detection fields within this generalized theory, whose results show that the ultimate resolution achieved with the linear excitation can reach one-fifth of the excitation wavelength (or λ/5), while the nonlinear excitation scheme gives rise to a resolution better than λ/12 for two-photon fluorescence excitation and λ/20 for three-photon fluorescence excitation. Hence the resolution of optical microscopy with a near-infrared excitation can routinely reach sub-60 nm. In addition, simulations for confocal laser scanning microscopy are carried out with the linear excitation scheme and the fluorescent one, respectively.

  20. Confocal Endomicroscopy of Colorectal Polyps

    Directory of Open Access Journals (Sweden)

    Vivian M. Ussui

    2012-01-01

    Full Text Available Confocal laser endomicroscopy (CLE is one of several novel methods that provide real-time, high-resolution imaging at a micron scale via endoscopes. CLE has the potential to be a disruptive technology in that it can change the current algorithms that depend on biopsy to perform surveillance of high-risk conditions. Furthermore, it allows on-table decision making that has the potential to guide therapy in real time and reduce the need for repeated procedures. CLE and related technologies are often termed “virtual biopsy” as they simulate the images seen in traditional histology. However, the imaging of living tissue allows more than just pragmatic convenience; it also allows imaging of living tissue such as active capillary circulation, cellular death, and vascular and endothelial translocation, thus extending beyond what is capable in traditional biopsy. Immediate potential applications of CLE are to guide biopsy sampling in Barrett's esophagus and inflammatory bowel disease surveillance, evaluation of colorectal polyps, and intraductal imaging of the pancreas and bile duct. Data on these applications is rapidly emerging, and more is needed to clearly demonstrate the optimal applications of CLE. In this paper, we will focus on the role of CLE as applied to colorectal polyps detected during colonoscopy.

  1. An FFT-based Method for Attenuation Correction in Fluorescence Confocal Microscopy

    NARCIS (Netherlands)

    Roerdink, J.B.T.M.; Bakker, M.

    1993-01-01

    A problem in three-dimensional imaging by a confocal scanning laser microscope (CSLM) in the (epi)fluorescence mode is the darkening of the deeper layers due to absorption and scattering of both the excitation and the fluorescence light. In this paper we propose a new method to correct for these

  2. An FFT-based method for attenuation correction in fluorescence confocal microscopy

    NARCIS (Netherlands)

    J.B.T.M. Roerdink (Jos); M. Bakker (Miente)

    1993-01-01

    htmlabstractA problem in three-dimensional imaging by a confocal scanning laser microscope (CSLM) in the (epi)fluorescence mode is the darkening of the deeper layers due to absorption and scattering of both the excitation and the fluorescence light. In this paper we propose a new method to correct

  3. FFT-Based Methods for Nonlinear Image Restoration in Confocal Microscopy

    NARCIS (Netherlands)

    Roerdink, J.B.T.M.

    1994-01-01

    Recently we developed a new method for attenuation correction in 3D imaging by a confocal scanning laser microscope (CSLM) in the (epi)fluorescence mode. The fundamental element in our approach consisted of multiplying the measured fluorescent intensity by a correction factor involving a convolution

  4. Live Confocal Imaging of Developing Arabidopsis Flowers

    OpenAIRE

    Prunet, Nathanaël

    2017-01-01

    The study of plant growth and development has long relied on experimental techniques using dead, fixed tissues and lacking proper cellular resolution. Recent advances in confocal microscopy, combined with the development of numerous fluorophores, have overcome these issues and opened the possibility to study the expression of several genes simultaneously, with a good cellular resolution, in live samples. Live confocal imaging provides plant biologists with a powerful tool to study development...

  5. In vivo confocal microscopy for the detection of canine fungal keratitis and monitoring of therapeutic response.

    Science.gov (United States)

    Ledbetter, Eric C; Norman, Mary L; Starr, Jennifer K

    2016-05-01

    To describe in vivo corneal confocal microscopy of dogs during the clinical course of fungal keratitis and correlate findings with clinical evaluations and an ex vivo experimental canine fungal keratitis model. Seven dogs with naturally acquired fungal keratitis and ex vivo canine corneas experimentally infected with clinical fungal isolates. Dogs with naturally acquired fungal keratitis were examined by in vivo laser scanning confocal microscopy. Initial confocal microscopic examinations were performed to assist in establishing the diagnosis of fungal keratitis. Serial confocal microscopic examinations were performed to guide antifungal chemotherapy. Confocal microscopy images of canine corneal fungal isolates were obtained by examination of experimentally infected ex vivo canine corneas to corroborate in vivo findings. Fungi cultured and detected by PCR from canine corneal samples included Candida albicans, Fusarium incarnatum-equiseti, Malassezia pachydermatis, and a Rhodotorula sp. Linear, branching, interlocking, hyperreflective structures were detected by confocal microscopy in dogs with filamentous fungal keratitis and round to oval hyperreflective structures were detected in dogs with yeast fungal keratitis. Antifungal chemotherapy was associated with a progressive reduction in the distribution and density of corneal fungal elements, alterations to fungal morphology, decreased leukocyte numbers, restoration of epithelial layers, and an increased number of visible keratocyte nuclei. No dogs had a recurrence of fungal keratitis following medication discontinuation. Confocal microscopic fungal morphologies were similar between in vivo and ex vivo examinations. In vivo corneal confocal microscopy is a rapid method of diagnosing fungal keratitis in dogs and provides a noninvasive mechanism for monitoring therapeutic response. © 2015 American College of Veterinary Ophthalmologists.

  6. A confocal optical microscope for detection of single impurities in a bulk crystal at cryogenic temperatures.

    Science.gov (United States)

    Karlsson, Jenny; Rippe, Lars; Kröll, Stefan

    2016-03-01

    A compact sample-scanning confocal optical microscope for detection of single impurities below the surface of a bulk crystal at cryogenic temperatures is described. The sample, lens, and scanners are mounted inside a helium bath cryostat and have a footprint of only 19 × 19 mm. Wide field imaging and confocal imaging using a Blu-ray lens immersed in liquid helium are demonstrated with excitation at 370 nm. A spatial resolution of 300 nm and a detection efficiency of 1.6% were achieved.

  7. Nuclear Scans

    Science.gov (United States)

    Nuclear scans use radioactive substances to see structures and functions inside your body. They use a special ... images. Most scans take 20 to 45 minutes. Nuclear scans can help doctors diagnose many conditions, including ...

  8. A novel method for enhancing the lateral resolution and image SNR in confocal microscopy

    Science.gov (United States)

    Chen, Youhua; Zhu, Dazhao; Fang, Yue; Kuang, Cuifang; Liu, Xu

    2017-12-01

    There is always a tradeoff between the resolution and the signal-to-noise ratio (SNR) in confocal microscopy. In particular, the pinhole size is very important for maintaining a balance between them. In this paper, we propose a method for improving the lateral resolution and image SNR in confocal microscopy without making any changes to the hardware. By using the fluorescence emission difference (FED) approach, we divide the images acquired by different pinhole sizes into one image acquired by the central pinhole and several images acquired by ring-shaped pinholes. Then, they are added together with the deconvolution method. Simulation and experimental results for fluorescent particles and cells show that our method can achieve a far better resolution than a large pinhole and a higher SNR than a small pinhole. Moreover, our method can improve the performance of classic confocal laser scanning microscopy (CLSM) to a certain extent, especially CLSM with a continuously variable pinhole.

  9. Application of digital image quality criteria to optimize the confocal microscope setup

    Science.gov (United States)

    Kriete, Andres

    1994-04-01

    This paper discusses how digital image quality criteria help to optimize image quality, in particular for applications in laser scanning microscopy. Image quality considerations offer a uniform description of the available transfer characteristics, which are summed up and weighted properly to finally represent the system by a single number. In the spatial domain we can measure sharpness and contrast of the (digital) volumes by analyzing intensities and their local dependencies in a statistical fashion. This includes sum modulus difference, gray level variance, and lateral inhibition. Based on information theory, the criterion volume fidelity takes into account the knowledge of the spatial structure of a test object and compares the intensities with those present in the final digital image. Applications presented here include measurement of image quality improvement when going from non-confocal to confocal imaging, testing of new confocal system designs and the evaluation of digital post-processing methods. Limitations in the presence of noise are discussed.

  10. Confocal laser endomicroscopy in the "in vivo" histological diagnosis of the gastrointestinal tract.

    Science.gov (United States)

    De Palma, Giovanni D

    2009-12-14

    Recent technological advances in miniaturization have allowed for a confocal scanning microscope to be integrated into a conventional flexible endoscope, or into trans-endoscopic probes, a technique now known as confocal endomicroscopy or confocal laser endomicroscopy. This newly-developed technology has enabled endoscopists to collect real-time in vivo histological images or "virtual biopsies" of the gastrointestinal mucosa during endoscopy, and has stimulated significant interest in the application of this technique in clinical gastroenterology. This review aims to evaluate the current data on the technical aspects and the utility of this new technology in clinical gastroenterology and its potential impact in the future, particularly in the screening or surveillance of gastrointestinal neoplasia.

  11. Confocal light scattering and absorption spectroscopic microscopy

    Science.gov (United States)

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

    2008-04-01

    We have developed a novel optical method for observing submicron intracellular structures in living cells which is called confocal light absorption and scattering spectroscopic (CLASS) microscopy. It combines confocal microscopy, a well-established high-resolution microscopic technique, with light scattering spectroscopy (LSS). 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. In addition, it provides not only size information but also information about the biochemical and physical properties of the cell.

  12. High resolution, high speed, long working distance, large field of view confocal fluorescence microscope.

    Science.gov (United States)

    Pacheco, Shaun; Wang, Chengliang; Chawla, Monica K; Nguyen, Minhkhoi; Baggett, Brend K; Utzinger, Urs; Barnes, Carol A; Liang, Rongguang

    2017-10-17

    Confocal fluorescence microscopy is often used in brain imaging experiments, however conventional confocal microscopes are limited in their field of view, working distance, and speed for high resolution imaging. We report here the development of a novel high resolution, high speed, long working distance, and large field of view confocal fluorescence microscope (H2L2-CFM) with the capability of multi-region and multifocal imaging. To demonstrate the concept, a 0.5 numerical aperture (NA) confocal fluorescence microscope is prototyped with a 3 mm × 3 mm field of view and 12 mm working distance, an array of 9 beams is scanned over the field of view in 9 different regions to speed up the acquisition time by a factor of 9. We test this custom designed confocal fluorescence microscope for future use with brain clarification methods to image large volumes of the brain at subcellular resolution. This multi-region and multi-spot imaging method can be used in other imaging modalities, such as multiphoton microscopes, and the field of view can be extended well beyond 12 mm × 12 mm.

  13. Fluorescence confocal polarizing microscopy: Three-dimensional ...

    Indian Academy of Sciences (India)

    Three-dimensional imaging of the director. O D LAVRENTOVICH. Chemical Physics ... cholesteric LCs. Keywords. 3D imaging; confocal microscopy; liquid crystals; dislocations. PACS Nos 07.60. ... magnetic resonance, x-ray diffraction, optical phase retardation, etc., suffer from the same deficiency: they produce only an ...

  14. Fluorescence confocal polarizing microscopy: Three-dimensional ...

    Indian Academy of Sciences (India)

    A PM image bears only two-dimensional (2D) information, integrating the 3D pattern of optical birefringence over the path of light. Recently, we proposed a technique to image 3D director patterns by fluorescence confocal polarizing microscopy (FCPM). The technique employs the property of LC to orient the fluorescent dye ...

  15. Confocal unrolled areal measurements of cylindrical surfaces

    Science.gov (United States)

    Matilla, A.; Bermudez, C.; Mariné, J.; Martínez, D.; Cadevall, C.; Artigas, R.

    2017-06-01

    Confocal microscopes are widely used for areal measurements thanks to its good height resolution and the capability to measure high local slopes. For the measurement of large areas while keeping few nm of system noise, it is needed to use high numerical aperture objectives, move the sample in the XY plane and stitch several fields together to cover the required surface. On cylindrical surfaces a rotational stage is used to measure fields along the round surface and stitch them in order to obtain a complete 3D measurement. The required amount of fields depends on the microscope's magnification, as well as on the cylinder diameter. However, for small diameters, if the local shape reaches slopes not suitable for the objective under use, the active field of the camera has to be reduced, leading to an increase of the required number of fields to be measured and stitched. In this paper we show a new approach for areal measurements of cylindrical surfaces that uses a rotational stage in combination with a slit projection confocal arrangement and a highspeed camera. An unrolled confocal image of the cylinder surface is built by rotating the sample and calculating the confocal intensity in the centre of the slit using a gradient algorithm. A set of 360º confocal images can be obtained at different heights of the sample relative to the sensor and used to calculate an unrolled areal measure of the cylinder. This method has several advantages over the conventional one such as no stitching required, or reduced measurement time. In addition, the result shows less residual flatness error since the surface lies flat in the measurement direction in comparison to field measures where the highest slope regions will show field distortion and non-constant sampling. We have also studied the influence on the areal measurements of wobble and run-out introduced by the clamping mechanism and the rotational axis.

  16. Parallel line scanning ophthalmoscope for retinal imaging

    NARCIS (Netherlands)

    Vienola, K.V.; Damodaran, M.; Braaf, B.; Vermeer, K.A.; de Boer, J.F.

    2015-01-01

    A parallel line scanning ophthalmoscope (PLSO) is presented using a digital micromirror device (DMD) for parallel confocal line imaging of the retina. The posterior part of the eye is illuminated using up to seven parallel lines, which were projected at 100 Hz. The DMD offers a high degree of

  17. Comparison of Noncontact Specular and Confocal Microscopy for Evaluation of Corneal Endothelium.

    Science.gov (United States)

    Huang, Jianyan; Maram, Jyotsna; Tepelus, Tudor C; Sadda, Srinivas R; Chopra, Vikas; Lee, Olivia L

    2017-03-24

    To compare endothelial cell analysis obtained by noncontact specular and confocal microscopy, using the Konan NSP-9900 and Nidek ConfoScan4 systems, respectively. Three groups including 70 healthy eyes, 49 eyes with Fuchs endothelial corneal dystrophy (FECD), and 78 eyes with glaucoma were examined with both the Konan NSP-9900 specular microscope and the Nidek ConfocScan4 confocal microscope. Certified graders at the Doheny Image Reading Center compared corneal endothelial images from both instruments side by side to assess image quality. Endothelial cell density (ECD) measurements were calculated and compared using three different modalities: (1) each instrument's fully automated analysis; (2) each instrument's semiautomatic analysis with grader input; and (3) manual grading methods by certified grader. All normal eyes yielded gradable endothelial images, and most but not all glaucomatous eyes yielded images with high enough image quality to allow grading. In addition, in corneas with severe FECD, poor image quality precluded ECD grading by specular microscopy in 20 eyes (40.8%) but in only 4 (8.2%) confocal images from the same eyes. For the gradable images, the ECD values obtained using the manual grading method from either device were comparable with no statistically significant difference (P>0.05) between specular and confocal devices. Machine-generated ECD values were significantly different from manual results, measuring greater in all cases with specular microscopy. Machine-generated ECD values from confocal microscopy also differed significantly from manual determinations, but not in a consistent direction. Semiautomatic methods for both instruments obtained clinically acceptable ECD values. Automatic machine-generated ECD measurements differed significantly from manual assessments of corneal endothelium by both specular and confocal microscopy, suggesting that automated results should be used with caution. But ECD values derived manually were comparable

  18. Non-mydriatic confocal retinal imaging using a digital light projector.

    Science.gov (United States)

    Muller, Matthew S; Green, Jason J; Baskaran, Karthikeyan; Ingling, Allen W; Clendenon, Jeffrey L; Gast, Thomas J; Elsner, Ann E

    2015-02-07

    A digital light projector is implemented as an integrated illumination source and scanning element in a confocal non-mydriatic retinal camera, the Digital Light Ophthalmoscope (DLO). To simulate scanning, a series of illumination lines are rapidly projected on the retina. The backscattered light is imaged onto a 2-dimensional rolling shutter CMOS sensor. By temporally and spatially overlapping the illumination lines with the rolling shutter, confocal imaging is achieved. This approach enables a low cost, flexible, and robust design with a small footprint. The 3(rd) generation DLO technical design is presented, using a DLP LightCrafter 4500 and USB3.0 CMOS sensor. Specific improvements over previous work include the use of yellow illumination, filtered from the broad green LED spectrum, to obtain strong blood absorption and high contrast images while reducing pupil constriction and patient discomfort.

  19. A simple way to identify non-viable cells within living plant tissue using confocal microscopy

    Directory of Open Access Journals (Sweden)

    Truernit Elisabeth

    2008-06-01

    Full Text Available Abstract Background Plant cell death is a normal process during plant development. Mutant plants may exhibit misregulation of this process, which can lead to severe growth defects. Simple ways of visualising cell death in living plant tissues can aid the study of plant development and physiology. Results Spectral variants of the fluorescent SYTOX dyes were tested for their usefulness for the detection of non-viable cells within plant embryos and roots using confocal laser-scanning microscopy. The dyes were selective for non-viable cells and showed very little background staining in living cells. Simultaneous detection of SYTOX dye and fluorescent protein (e.g. GFP fluorescence was possible. Conclusion The fluorescent SYTOX dyes are useful for an easy and quick first assay of plant cell viability in living plant samples using fluorescence and confocal laser-scanning microscopy.

  20. Renal scan

    Science.gov (United States)

    ... this page: //medlineplus.gov/ency/article/003790.htm Renal scan To use the sharing features on this ... anaphylaxis . Alternative Names Renogram; Kidney scan Images Kidney anatomy Kidney - blood and urine flow References Chernecky CC, ...

  1. CONFOCAL MICROSCOPY STUDY OF BIOLOGICAL PECULIARITIES OF SCAFFOLD MADE FROM RECOMBINANT SPIDER SILK

    Directory of Open Access Journals (Sweden)

    O. L. Pustovalova

    2009-01-01

    Full Text Available We studied the viability and dynamic of cell distribution during long-term cultivation of broblasts 3T3 in spider silk spidroin 1-based scaffold. Laser scanning confocal microscopy is shown to have advantages for visualization of cells situated on the external and internal surfaces of scaffold. Fibroblasts maintain high proliferative ability and viability during long term cultivation. Spidroin 1-based scaffold are the perspective materials for bioengineering. 

  2. In vivo confocal microscopy in dermatology: from research to clinical application

    Science.gov (United States)

    Ulrich, Martina; Lange-Asschenfeldt, Susanne

    2013-06-01

    Confocal laser scanning microscopy (CLSM) represents an emerging technique for the noninvasive histomorphological analysis of skin in vivo and has shown its applicability for dermatological research as well as its value as an adjunct tool in the clinical management of skin cancer patients. Herein, we aim to give an overview on the current clinical indications for CLSM in dermatology and also highlight the diverse applications of CLSM in dermatological research.

  3. Description and Performance of a Highly Sensitive Confocal Raman Microspectrometer

    NARCIS (Netherlands)

    Puppels, G.J.; Puppels, G.J.; Colier, W.; Olminkhof, J.H.F.; Otto, Cornelis; de Mul, F.F.M.; Greve, Jan

    1991-01-01

    A confocal Raman microspectrometer was developed for the study of small biological objects such as single living cells and metaphase and polytene chromosomes. It employs a confocal detection scheme, well known from confocal fluorescence microscopes, in order to avoid signal contributions from the

  4. Fungal keratitis - improving diagnostics by confocal microscopy

    DEFF Research Database (Denmark)

    Nielsen, Esben; Heegaard, S; Prause, J U

    2013-01-01

    Purpose: Introducing a simple image grading system to support the interpretation of in vivo confocal microscopy (IVCM) images in filamentous fungal keratitis. Setting: Clinical and confocal studies took place at the Department of Ophthalmology, Aarhus University Hospital, Denmark. Histopathological...... analysis was performed at the Eye Pathology Institute, Department of Neuroscience and Pharmacology, University of Copenhagen, Denmark. Methods: A recent series of consecutive patients with filamentous fungal keratitis is presented to demonstrate the results from in-house IVCM. Based upon our experience...... with IVCM and previously published images, we composed a grading system for interpreting IVCM images of filamentous fungal keratitis. Results: A recent case series of filamentous fungal keratitis from 2011 to 2012 was examined. There were 3 male and 3 female patients. Mean age was 44.5 years (range 12...

  5. Reflectance Confocal Microscopy in Lentigo Maligna.

    Science.gov (United States)

    Gamo, R; Pampín, A; Floristán, U

    2016-12-01

    Lentigo maligna is the most common type of facial melanoma. Diagnosis is complicated, however, as it shares clinical and dermoscopic characteristics with other cutaneous lesions of the face. Reflectance confocal microscopy is an imaging technique that permits the visualization of characteristic features of lentigo maligna. These include a disrupted honeycomb pattern and pagetoid cells with a tendency to show folliculotropism. These cells typically have a dendritic morphology, although they may also appear as round cells measuring over 20μm with atypical nuclei. Poorly defined dermal papillae and atypical cells may be seen at the dermal-epidermal junction and can form bridges resembling mitochondrial structures. Other characteristic findings include junctional swelling with atypical cells located around the follicles, resembling caput medusae. Reflectance confocal microscopy is a very useful tool for diagnosing lentigo maligna. Copyright © 2016 AEDV. Publicado por Elsevier España, S.L.U. All rights reserved.

  6. Confocal Laser Endomicroscopy in Inflammatory Bowel Disease

    DEFF Research Database (Denmark)

    Rasmussen, Ditlev Nytoft; Karstensen, John Gásdal; Riis, Lene Buhl

    2015-01-01

    BACKGROUND AND AIMS: Confocal laser endomicroscopy is an endoscopic method that provides in vivo real-time imaging of the mucosa at a cellular level, elucidating mucosal changes that are undetectable by white light endoscopy. This paper systematically reviews current indications and perspectives...... of intestinal barrier function. There is great heterogeneity in the literature and no single approach has been validated and reproduced to the level of general acceptance....

  7. Clinical applications of corneal confocal microscopy

    Directory of Open Access Journals (Sweden)

    Mitra Tavakoli

    2008-06-01

    Full Text Available Mitra Tavakoli1, Parwez Hossain2, Rayaz A Malik11Division of Cardiovascular Medicine, University of Manchester and Manchester Royal Infirmary, Manchester, UK; 2University of Southampton, Southampton Eye Unit, Southampton General Hospital, Southampton, UKAbstract: Corneal confocal microscopy is a novel clinical technique for the study of corneal cellular structure. It provides images which are comparable to in-vitro histochemical techniques delineating corneal epithelium, Bowman’s layer, stroma, Descemet’s membrane and the corneal endothelium. Because, corneal confocal microscopy is a non invasive technique for in vivo imaging of the living cornea it has huge clinical potential to investigate numerous corneal diseases. Thus far it has been used in the detection and management of pathologic and infectious conditions, corneal dystrophies and ecstasies, monitoring contact lens induced corneal changes and for pre and post surgical evaluation (PRK, LASIK and LASEK, flap evaluations and Radial Keratotomy, and penetrating keratoplasty. Most recently it has been used as a surrogate for peripheral nerve damage in a variety of peripheral neuropathies and may have potential in acting as a surrogate marker for endothelial abnormalities.Keywords: corneal confocal microscopy, cornea, infective keratitis, corneal dystrophy, neuropathy

  8. Line-scan Raman microspectrometry for biological applications

    NARCIS (Netherlands)

    de Grauw, C.J.; de Grauw, C.J.; Otto, Cornelis; Greve, Jan

    1997-01-01

    A high-resolution confocal line-scan Raman microscope was developed for the study of biological samples such as cells and chromosomes. With the illumination of a line on the sample, all the spectral information of the line is recorded. The line-scan microscope was attained by the introduction of one

  9. Demodex mites in acne rosacea: reflectance confocal microscopic study.

    Science.gov (United States)

    Turgut Erdemir, Asli; Gurel, Mehmet Salih; Koku Aksu, Ayse Esra; Falay, Tugba; Inan Yuksel, Esma; Sarikaya, Ebru

    2017-05-01

    Demodex mite density is emphasised in the aetiopathogenesis of acne rosacea. Reflectance confocal microscopy (RCM) has been shown to be a good method for determining demodex mite density. The objective was to determine demodex mite density using RCM in acne rosacea patients and compare them with controls. In all, 30 papulopustular rosacea (PPR) and 30 erythematotelangiectatic rosacea (ETR) totally 60 acne rosacea patients and 40 controls, were enrolled in the study. The right cheek was selected for imaging and RCM was used for scanning. Ten images of 1000 × 1000 μm (total 10 mm2 ) area were scanned from adjacent areas. The numbers of follicles, infested follicles and mites were counted. The mean numbers of mites per follicle and infested follicles were calculated and compared in the patients and control groups. The mean number of mites was 44.30 ± 23.22 in PPR, 14.57 ± 15.86 in ETR and 3.55 ± 6.48 in the control group (P Demodex mite density was markedly increased in both ETR and PPR patients. It is believed that the presence of demodex mites plays an important role in rosacea aetiopathogenesis. Demodex mite treatment may reduce the severity of the disease and slow its progressive nature. © 2016 The Australasian College of Dermatologists.

  10. Optical coherence tomography and confocal microscopy investigations of dental prostheses

    Science.gov (United States)

    Negrutiu, Meda L.; Sinescu, Cosmin; Hughes, Michael; Bradu, Adrian; Rominu, Mihai; Todea, Carmen; Dobre, George; Podoleanu, Adrian

    2008-09-01

    Dental prostheses are very complex systems, heterogenous in structure, made up from various materials, with different physical properties. An essential question mark is on the physical, chemical and mechanical compatibility between these materials. They have to satisfy high stress requirements as well as esthetic challenges. The masticatory stress may induce fractures of the prostheses, which may be triggered by initial materials defects or by alterations of the technological process. The failures of dental prostheses lead to functional, esthetic and phonetic disturbances which finally render the prosthetic treatment inefficient. The purpose of this study is to evaluate the capability of en-face optical coherence tomography as a possible non-invasive high resolution method in supplying the necessary information on the material defects of dental prostheses and microleakage at prosthetic interfaces. C-scan and B-scan OCT images as well as confocal images are acquired from a large range of samples. Gaps between the dental interfaces and material defects are clearly exposed. We conclude that OCT can successfully be used as a noninvasive analysis method.

  11. Confocal laser endomicroscopy in ulcerative colitis

    DEFF Research Database (Denmark)

    Karstensen, John Gásdal; Săftoiu, Adrian; Brynskov, Jørn

    2016-01-01

    was to correlate colonic confocal laser endomicroscopy (CLE) in ulcerative colitis with histopathology and macroscopic appearance before and after intensification of medical treatment. METHODS: Twenty-two patients with ulcerative colitis in clinical relapse and 7 control subjects referred for colonoscopy were...... colitis compared with inactive ulcerative colitis...... is an emerging endoscopic technique that reproducibly identifies mucosal changes in ulcerative colitis. With the exception of crypt changes, endomicroscopic features appear to improve slowly with time after medical treatment. ( CLINICAL TRIAL REGISTRATION NUMBER: NCT01684514.)....

  12. Cooperative scans

    NARCIS (Netherlands)

    M. Zukowski (Marcin); P.A. Boncz (Peter); M.L. Kersten (Martin)

    2004-01-01

    textabstractData mining, information retrieval and other application areas exhibit a query load with multiple concurrent queries touching a large fraction of a relation. This leads to individual query plans based on a table scan or large index scan. The implementation of this access path in most

  13. High resolution 3D confocal microscope imaging of volcanic ash particles.

    Science.gov (United States)

    Wertheim, David; Gillmore, Gavin; Gill, Ian; Petford, Nick

    2017-07-15

    We present initial results from a novel high resolution confocal microscopy study of the 3D surface structure of volcanic ash particles from two recent explosive basaltic eruptions, Eyjafjallajökull (2010) and Grimsvötn (2011), in Iceland. The majority of particles imaged are less than 100μm in size and include PM10s, known to be harmful to humans if inhaled. Previous studies have mainly used 2D microscopy to examine volcanic particles. The aim of this study was to test the potential of 3D laser scanning confocal microscopy as a reliable analysis tool for these materials and if so to what degree high resolution surface and volume data could be obtained that would further aid in their classification. First results obtained using an Olympus LEXT scanning confocal microscope with a ×50 and ×100 objective lens are highly encouraging. They reveal a range of discrete particle types characterised by sharp or concave edges consistent with explosive formation and sudden rupture of magma. Initial surface area/volume ratios are given that may prove useful in subsequent modelling of damage to aircraft engines and human tissue where inhalation has occurred. Copyright © 2017 Elsevier B.V. All rights reserved.

  14. Reflectance Confocal Microscopy for Inflammatory Skin Diseases.

    Science.gov (United States)

    Agozzino, M; Gonzalez, S; Ardigò, M

    2016-10-01

    In vivo reflectance confocal microscopy (RCM) is a relatively novel non-invasive tool for microscopic evaluation of the skin used prevalently for diagnosis and management of skin tumour. Its axial resolution, its non-invasive and easy clinical application represents the goals for a large diffusion of this technique. During the last 15 years, RCM has been demonstrated to be able to increase the sensibility and sensitivity of dermoscopy in the diagnosis of skin tumours integrating in real time clinic, dermoscopic and microscopic information useful for the definition of malignancy. Despite to date, no large comparative studies on inflammatory skin diseases has been published in the literature, several papers already showed that RCM has a potential for the evaluation of the descriptive features of the most common inflammatory skin diseases as psoriasis, lupus erythematosus, contact dermatitis and others. The aim of the application of this technique in non-neoplastic skin diseases has been prevalently focused on the possibility of clinical diagnosis confirmation, as well as therapeutic management. Moreover, the use of RCM as driver for an optimised skin biopsy has been also followed in order to reduce the number of unsuccessful histopathological examination. In this review article we describe the confocal features of the major groups of inflammatory skin disorders focusing on psoriasiform dermatitis, interface dermatitis and spongiotic dermatitis. Publicado por Elsevier España, S.L.U.

  15. Refractive index measurement based on confocal method

    Science.gov (United States)

    An, Zhe; Xu, XiPing; Yang, JinHua; Qiao, Yang; Liu, Yang

    2017-10-01

    The development of transparent materials is closed to optoelectronic technology. It plays an increasingly important role in various fields. It is not only widely used in optical lens, optical element, optical fiber grating, optoelectronics, but also widely used in the building material, pharmaceutical industry with vessel, aircraft windshield and daily wear glasses.Regard of solving the problem of refractive index measurement in optical transparent materials. We proposed that using the polychromatic confocal method to measuring the refractive index of transparent materials. In this article, we describes the principle of polychromatic confocal method for measuring the refractive index of glass,and sketched the optical system and its optimization. Then we establish the measurement model of the refractive index, and set up the experimental system. In this way, the refractive index of the glass has been calibrated for refractive index experiment. Due to the error in the experimental process, we manipulated the experiment data to compensate the refractive index measurement formula. The experiment taking the quartz glass for instance. The measurement accuracy of the refractive index of the glass is +/-1.8×10-5. This method is more practical and accurate, especially suitable for non-contact measurement occasions, which environmental requirements is not high. Environmental requirements are not high, the ordinary glass production line up to the ambient temperature can be fully adapted. There is no need for the color of the measured object that you can measure the white and a variety of colored glass.

  16. Microscopia confocal en operados de queratoplastia perforante Confocal microscopy in patients operated from penetrating keratoplasty

    Directory of Open Access Journals (Sweden)

    Zulema Gómez Castillo

    2009-06-01

    Full Text Available La microscopia confocal es un examen exploratorio, práctico y poco invasivo que permite conocer las características microscópicas del tejido corneal después del trasplante, por lo que constituye una herramienta muy útil en el manejo de los pacientes operados de queratoplastia. El presente trabajo tiene como finalidad describir las características del tejido corneal en pacientes operados de este tipo de trasplante, mediante la microscopia confocal in vivo. MÉTODOS: Se realizó un estudio descriptivo, de corte transversal, en 40 ojos de 40 pacientes operados de queratoplastia perforante, en el Servicio de Córnea del Instituto Cubano de Oftalmología "Ramón Pando Ferrer", de marzo de 2006 a marzo de 2007. Se confeccionó una historia clínica oftalmológica y se les realizó a todos el examen de microscopia confocal en el injerto corneal con el microscopio confocal CONFOSCAN 4. RESULTADOS: La queratopatía bullosa pseudofáquica fue la afección más frecuente previa a la cirugía y estuvo presente en el 77,5 % de los pacientes. En el 72,5 % de los intervenidos se encontró una disminución del grosor corneal. El epitelio presentó alteraciones en el 62,5 % de los pacientes. Todos presentaron afectación de la forma y el tamaño celular endotelial. En el 82,5 % de los pacientes se observó ausencia de plexos nerviosos. CONCLUSIONES: La microscopia confocal como nueva ciencia en el campo de la oftalmología, favorece el seguimiento evolutivo de las queratoplastias perforantes y con esto no solo a prevenir la aparición de posibles complicaciones, sino además de garantizar el éxito de la cirugía y la función refractiva de la córnea.Confocal microscopy is a practical, exploratory and less invassive examination that allows finding out the microscopic characteristics of the corneal tissue after transplantation, so it is a very useful tool for the management of patients operated from keratoplasty. The present paper was aimed at describing

  17. Cellular resolution expression profiling using confocal detection of NBT/BCIP precipitate by reflection microscopy.

    Science.gov (United States)

    Jékely, Gáspár; Arendt, Detlev

    2007-06-01

    The determination of gene expression patterns in three dimensions with cellular resolution is an important goal in developmental biology. However the most sensitive, efficient, and widely used staining technique for whole-mount in situ hybridization (WMISH), nitroblue tetrazolium (NBT)/5-bromo-4-chloro-3-indolyl phosphate (BCIP) precipitation by alkaline phosphatase, could not yet be combined with the most precise, high-resolution detection technique, confocal laser-scanning microscopy (CLSM). Here we report the efficient visualization of the NBT/BCIP precipitate using confocal reflection microscopy for WMISH samples of Drosophila, zebrafish, and the marine annelid worm, Platynereis dumerilii. In our simple WMISH protocol for reflection CLSM, NBT/BCIP staining can be combined with fluorescent WMISH, immunostainings, or transgenic green fluorescent protein (GFP) marker lines, allowing double labeling of cell types or of embryological structures of interest. Whole-mount reflection CLSM will thus greatly facilitate large-scale cellular resolution expression profiling in vertebrate and invertebrate model organisms.

  18. Interfacial shape and contact-angle measurement of transparent samples with confocal interference microscopy.

    Science.gov (United States)

    Fischer, D G; Ovryn, B

    2000-04-01

    A model has been developed that predicts the effective optical path through a thick, refractive specimen on a reflective substrate, as measured with a scanning confocal interference microscope equipped with a high-numerical-aperture objective. Assuming that the effective pinhole of the confocal microscope has an infinitesimal diameter, only one ray in the illumination bundle (the magic ray) contributes to the differential optical path length (OPL). A pinhole with finite diameter, however, allows rays within a small angular cone centered on the magic ray to contribute to the OPL. The model was incorporated into an iterative algorithm that allows the measured phase to be corrected for refractive errors by use of an a priori estimate of the sample profile. The algorithm was validated with a reflected-light microscope equipped with a phase-shifting laser-feedback interferometer to measure the interface shape and the 68 degrees contact angle of a silicone-oil drop on a coated silicon wafer.

  19. MRI Scans

    Science.gov (United States)

    Magnetic resonance imaging (MRI) uses a large magnet and radio waves to look at organs and structures inside your body. Health care professionals use MRI scans to diagnose a variety of conditions, from ...

  20. Bone Scan

    Science.gov (United States)

    ... posts Join Mayo Clinic Connect Bone scan About Advertisement Mayo Clinic does not endorse companies or products. ... a Job Site Map About This Site Twitter Facebook Google YouTube Pinterest Mayo Clinic is a not- ...

  1. Confocal imaging of confined quiescent and flowing colloid-polymer mixtures.

    Science.gov (United States)

    Pandey, Rahul; Spannuth, Melissa; Conrad, Jacinta C

    2014-05-20

    The behavior of confined colloidal suspensions with attractive interparticle interactions is critical to the rational design of materials for directed assembly(1-3), drug delivery(4), improved hydrocarbon recovery(5-7), and flowable electrodes for energy storage(8). Suspensions containing fluorescent colloids and non-adsorbing polymers are appealing model systems, as the ratio of the polymer radius of gyration to the particle radius and concentration of polymer control the range and strength of the interparticle attraction, respectively. By tuning the polymer properties and the volume fraction of the colloids, colloid fluids, fluids of clusters, gels, crystals, and glasses can be obtained(9). Confocal microscopy, a variant of fluorescence microscopy, allows an optically transparent and fluorescent sample to be imaged with high spatial and temporal resolution in three dimensions. In this technique, a small pinhole or slit blocks the emitted fluorescent light from regions of the sample that are outside the focal volume of the microscope optical system. As a result, only a thin section of the sample in the focal plane is imaged. This technique is particularly well suited to probe the structure and dynamics in dense colloidal suspensions at the single-particle scale: the particles are large enough to be resolved using visible light and diffuse slowly enough to be captured at typical scan speeds of commercial confocal systems(10). Improvements in scan speeds and analysis algorithms have also enabled quantitative confocal imaging of flowing suspensions(11-16,37). In this paper, we demonstrate confocal microscopy experiments to probe the confined phase behavior and flow properties of colloid-polymer mixtures. We first prepare colloid-polymer mixtures that are density- and refractive-index matched. Next, we report a standard protocol for imaging quiescent dense colloid-polymer mixtures under varying confinement in thin wedge-shaped cells. Finally, we demonstrate a protocol

  2. Confocal microscopy-based goniometry of barnacle cyprid permanent adhesive.

    Science.gov (United States)

    Aldred, Nick; Gohad, Neeraj V; Petrone, Luigi; Orihuela, Beatriz; Liedberg, Bo; Ederth, Thomas; Mount, Andrew; Rittschof, Dan; Clare, Anthony S

    2013-06-01

    Biological adhesives are materials of particular interest in the fields of bio-inspired technology and antifouling research. The adhesive of adult barnacles has received much attention over the years; however, the permanent adhesive of the cyprid - the colonisation stage of barnacles - is a material about which very little is presently known. We applied confocal laser-scanning microscopy to the measurement of contact angles between the permanent adhesive of barnacle cyprid larvae and self-assembled monolayers of OH- and CH3-terminated thiols. Measurement of contact angles between actual bioadhesives and surfaces has never previously been achieved and the data may provide insight into the physicochemical properties and mechanism of action of these functional materials. The adhesive is a dual-phase system post-secretion, with the behaviour of the components governed separately by the surface chemistry. The findings imply that the cyprid permanent adhesion process is more complex than previously thought, necessitating broad re-evaluation of the system. Improved understanding will have significant implications for the production of barnacle-resistant coatings as well as development of bio-inspired glues for niche applications.

  3. Spatial resolution of confocal XRF technique using capillary optics.

    Science.gov (United States)

    Dehlinger, Maël; Fauquet, Carole; Lavandier, Sebastien; Aumporn, Orawan; Jandard, Franck; Arkadiev, Vladimir; Bjeoumikhov, Aniouar; Tonneau, Didier

    2013-06-07

    XRF (X-ray fluorescence) is a powerful technique for elemental analysis with a high sensitivity. The resolution is presently limited by the size of the primary excitation X-ray beam. A test-bed for confocal-type XRF has been developed to estimate the ultimate lateral resolution which could be reached in chemical mapping using this technique. A polycapillary lens is used to tightly focus the primary X-ray beam of a low power rhodium X-ray source, while the fluorescence signal is collected by a SDD detector through a cylindrical monocapillary. This system was used to characterize the geometry of the fluorescent zone. Capillary radii ranging from 50 μm down to 5 μm were used to investigate the fluorescence signal maximum level This study allows to estimate the ultimate resolution which could be reached in-lab or on a synchrotron beamline. A new tool combining local XRF and scanning probe microscopy is finally proposed.

  4. Confocal Light Absorption and Scattering Spectroscopic (CLASS) imaging: From cancer detection to sub-cellular function

    Science.gov (United States)

    Qiu, Le

    Light scattering spectroscopy (LSS), an optical technique that relates the spectroscopic properties of light elastically scattered by small particles to their size, refractive index and shape, has been recently successfully employed for sensing morphological and biochemical properties of epithelial tissues and cells in vivo. LSS does not require exogenous markers, is non-invasive, and, due to its multispectral nature, can sense biological structures well beyond the diffraction limit. All that makes LSS be a very good candidate to be used both in clinical medicine for in vivo detection of disease and in cell biology to monitor cell function on the organelle scale. Recently we developed two LSS-based imaging modalities: clinical Polarized LSS (PLSS) Endoscopic Technique for locating early pre-cancerous changes in GI tract and Confocal Light Absorption and Scattering Spectroscopic (CLASS) Microscopy for studying cells in vivo without exogenous markers. One important application of the clinical PLSS endoscopic instrument, a noncontact scanning imaging device compatible with the standard clinical endoscopes and capable of detecting dysplastic changes, is to serve as a guide for biopsy in Barrett's esophagus (BE). The instrument detects parallel and perpendicular components of the polarized light, backscattered from epithelial tissues, and determines characteristics of epithelial nuclei from the residual spectra. It also can find tissue oxygenation, hemoglobin content and other properties from the diffuse light component. By rapidly scanning esophagus the PLSS endoscopic instrument makes sure the entire BE portion is scanned and examined for the presence of dysplasia. CLASS microscopy, on the other hand, combines principles of light scattering spectroscopy (LSS) with confocal microscopy. Its main purpose is to image cells on organelle scale in vivo without the use of exogenous labels which may affect the cell function. The confocal geometry selects specific region and

  5. [Fast 2-dimension scanning and line-scanning of intracellular Ca2+ transients in cardiac myocytes].

    Science.gov (United States)

    Shen, Jian-xin; Wang, Hai-yan; Li, Chao-yan; Xiao, Jian-feng

    2008-11-01

    Fast 2-dimension scanning and line-scanning of confocal imaging were employed for measurement of cardiac Ca2+ transients, and the advantages and disadvantages about these two scannings were discussed. Single adult SD rat cardiac myocytes were made freshly and loaded with fluo4-AM. Intracellular Ca2+ was imaging by the LSMS10 META system. The Ca2+ transients were evoked by electrical field stimulation from an electronic stimulator which was triggered to work synchronically with the confocal imaging system. Fast 2-dimension scanning showed the global Ca2+ signal clearly, which would be more helpful especially in monitoring a cell of Ca2+ overload or in other pathological conditions. And the images could be packaged into a vivid animation, which showed the process of Ca2+ transients and cell contraction visually and virtually. Line-scanning showed the Ca2+ transients in good temporal and spacial resolutions along the long axis of the cell. And the dynamic shortening of the cell length could be used for indicating the contraction of the cell. Data from line-scanning would be helpful for drawing some more exact conclusions. In general, fast 2-dimension scanning and line-scanning could work reciprocally to show a more perfect picture of the intracellular Ca2+ transients in cardiac myocytes.

  6. Miniaturized fiber-coupled confocal fluorescence microscope with an electrowetting variable focus lens using no moving parts.

    Science.gov (United States)

    Ozbay, Baris N; Losacco, Justin T; Cormack, Robert; Weir, Richard; Bright, Victor M; Gopinath, Juliet T; Restrepo, Diego; Gibson, Emily A

    2015-06-01

    We report a miniature, lightweight fiber-coupled confocal fluorescence microscope that incorporates an electrowetting variable focus lens to provide axial scanning for full three-dimensional (3D) imaging. Lateral scanning is accomplished by coupling our device to a laser-scanning confocal microscope through a coherent imaging fiber-bundle. The optical components of the device are combined in a custom 3D-printed adapter with an assembled weight of <2  g that can be mounted onto the head of a mouse. Confocal sectioning provides an axial resolution of ∼12  μm and an axial scan range of ∼80  μm. The lateral field-of-view is 300 μm, and the lateral resolution is 1.8 μm. We determined these parameters by imaging fixed sections of mouse neuronal tissue labeled with green fluorescent protein (GFP) and fluorescent bead samples in agarose gel. To demonstrate viability for imaging intact tissue, we resolved multiple optical sections of ex vivo mouse olfactory nerve fibers expressing yellow fluorescent protein (YFP).

  7. 4D confocal microscopy for visualisation of bone remodelling

    NARCIS (Netherlands)

    Konijn, GA; Vardaxis, NJ; Boon, ME; Kok, LP; Rietveld, DC; SCHUT, JJ

    Until recently it was very time consuming and difficult to make three-dimensional (3D) images of newly formed bone. With the advent of confocal technologies and increased computer power 3D imaging is greatly facilitated. In this paper we demonstrate that enhanced confocal visualisation of newly

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

    Science.gov (United States)

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

    2017-07-01

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

  9. Scanning table

    CERN Multimedia

    1960-01-01

    Before the invention of wire chambers, particles tracks were analysed on scanning tables like this one. Today, the process is electronic and much faster. Bubble chamber film - currently available - (links can be found below) was used for this analysis of the particle tracks.

  10. Scan Statistics

    CERN Document Server

    Glaz, Joseph

    2009-01-01

    Suitable for graduate students and researchers in applied probability and statistics, as well as for scientists in biology, computer science, pharmaceutical science and medicine, this title brings together a collection of chapters illustrating the depth and diversity of theory, methods and applications in the area of scan statistics.

  11. Combination of Small Molecule Microarray and Confocal Microscopy Techniques for Live Cell Staining Fluorescent Dye Discovery

    Directory of Open Access Journals (Sweden)

    Attila Bokros

    2013-08-01

    Full Text Available Discovering new fluorochromes is significantly advanced by high-throughput screening (HTS methods. In the present study a combination of small molecule microarray (SMM prescreening and confocal laser scanning microscopy (CLSM was developed in order to discover novel cell staining fluorescent dyes. Compounds with high native fluorescence were selected from a 14,585-member library and further tested on living cells under the microscope. Eleven compartment-specific, cell-permeable (or plasma membrane-targeted fluorochromes were identified. Their cytotoxicity was tested and found that between 1–10 micromolar range, they were non-toxic even during long-term incubations.

  12. In vivo Confocal Microscopy Report after Lasik with Sequential Accelerated Corneal Collagen Cross-Linking Treatment

    Directory of Open Access Journals (Sweden)

    Cosimo Mazzotta

    2014-04-01

    Full Text Available We report the first pilot qualitative confocal microscopic analysis of a laser in situ keratomileusis (Lasik treatment combined with sequential high-fluence accelerated corneal collagen cross-linking, denominated Lasik XTra, by means of HRT II laser scanning in vivo confocal microscopy after a 6-month follow-up. After obtaining approval from the Siena University Hospital Institutional Review Board, a 33-year-old female patient underwent a Lasik XTra procedure in her left eye. Confocal analysis demonstrated induced slight corneal microstructural changes by the interaction between UV-A, riboflavin and corneal stromal collagen, beyond the interface to a depth of 160 µm, without adverse events at the interface and endothelial levels. This application may be considered a prophylactic biomechanical treatment, stiffening the intermediate corneal stroma to prevent corneal ectasia and stabilizing the clinical results of refractive surgery. According to our preliminary experiences, this combined approach may be useful in higher-risk Lasik patients for hyperopic treatments, high myopia and lower corneal thicknesses.

  13. Tri-modal confocal mosaics detect residual invasive squamous cell carcinoma in Mohs surgical excisions.

    Science.gov (United States)

    Gareau, Dan; Bar, Anna; Snaveley, Nicholas; Lee, Ken; Chen, Nathaniel; Swanson, Neil; Simpson, Eric; Jacques, Steve

    2012-06-01

    For rapid, intra-operative pathological margin assessment to guide staged cancer excisions, multimodal confocal mosaic scan image wide surgical margins (approximately 1 cm) with sub-cellular resolution and mimic the appearance of conventional hematoxylin and eosin histopathology (H&E). The goal of this work is to combine three confocal imaging modes: acridine orange fluorescence (AO) for labeling nuclei, eosin fluorescence (Eo) for labeling cytoplasm, and endogenous reflectance (R) for marking collagen and keratin. Absorption contrast is achieved by alternating the excitation wavelength: 488 nm (AO fluorescence) and 532 nm (Eo fluorescence). Superposition and false-coloring of these modes mimics H&E, enabling detection of cutaneous squamous cell carcinomas (SCC). The sum of mosaic Eo+R is false-colored pink to mimic the appearance of eosin, while the AO mosaic is false-colored purple to mimic the appearance of hematoxylin in H&E. In this study, mosaics of 10 Mohs surgical excisions containing invasive SCC, and five containing only normal tissue were subdivided for digital presentation equivalent to 4 × histology. Of the total 50 SCC and 25 normal sub-mosaics presented, two reviewers made two and three type-2 errors (false positives), respectively. Limitations to precisely mimic H&E included occasional elastin staining by AO. These results suggest that confocal mosaics may effectively guide staged SCC excisions in skin and other tissues.

  14. Progress report towards a digital mirror device based confocal microscopic system

    Science.gov (United States)

    Yi, Dingrong; Lin, Shunhua; Huang, Simian; Xie, Shaochuan

    2013-12-01

    It is widely believed that by using a digital mirror device (DMD) as the spatial light modulator (SLM) of a programmable array microcopy (PAM), it is possible to achieve a cost-effective alternative to expensive confocal imaging devices. During the past decade, the design of such a DMD based PAM instrument has been frequently reported to enhance resolution and contrast, convincing images with improved quality are rare to be seen. The concrete implementation of a DMD based PAM instrument needs to successfully resolve multiple issues such as the adverse effects caused by the tilt angle of the micro-mirrors from the base board, the registration between a micro mirror of the DMD and the image pixel of the photo-detector and so on. In this paper, we report the design of a middle body consisting of a DMD as an independent attachment to a conventional microscope to convert the latter into a confocal imaging system, in a similar way as a filter turret that is placed below the head and the objectives of a regular microscopy to convert it into a fluorescent microscopy. Images of real objects with improved contrast are provided to demonstrate the effectiveness of using a DMD as SLM to improve the contrast of a PAM instrument. Such a PAM instrument has many advantages compared to conventional laser-scanning confocal systems including lower costs and higher imaging speeds. In addition, it allows convenient dynamic adjustments between imaging quality and imaging speed.

  15. Non-invasive in vivo visualization of enamel defects by reflectance confocal microscopy (RCM).

    Science.gov (United States)

    Contaldo, Maria; Di Stasio, Dario; Santoro, Rossella; Laino, Luigi; Perillo, Letizia; Petruzzi, Massimo; Lauritano, Dorina; Serpico, Rosario; Lucchese, Alberta

    2015-05-01

    The enamel defects (EDs) may present with a variety of clinical manifestations with increasing severity from the sole appearance of pale discoloration to remarkable structural alterations. EDs are responsible for higher caries receptivity. In vivo reflectance confocal microscopy (RCM) allows to image in vivo at microscopic resolution of the dental surface, thus avoiding the tooth extraction and the sample preparation because of its ability to optically scan living tissues along their depth. Aim of this study is the in vivo assessment at microscopic resolution of dental surfaces affected by EDs without resorting to invasive methods such as teeth extractions, to define histological findings occurring in chromatic and/or structural EDs. For the purpose, 15 children, referring at the Dental Clinic of the Second University of Naples, affected by several degrees of EDs, were enrolled and underwent in vivo RCM imaging to microscopically define the ED confocal features using a commercially available hand-held reflectance confocal microscope with neither injuries nor discomfort. Totally, 29 teeth were imaged. Results demonstrated images good in quality and the capability to detect EDs such as unevenness, grooves, and lack of mineralization according to their clinical degree of disarray. The present in vivo microscopic study on EDs allowed to highlight structural changes in dental enamel at microscopic resolution in real-time and in a non-invasive way, with no need for extraction or processing the samples. Further experiments could define the responsiveness to remineralizing procedures as therapeutic treatments.

  16. Confocal Microscopy of Unfixed Breast Needle Core Biopsies: A Comparison to Fixed and Stained Sections

    Science.gov (United States)

    2009-01-01

    Background Needle core biopsy, often in conjunction with ultrasonic or stereotactic guided techniques, is frequently used to diagnose breast carcinoma in women. Confocal scanning laser microscopy (CSLM) is a technology that provides real-time digital images of tissues with cellular resolution. This paper reports the progress in developing techniques to rapidly screen needle core breast biopsy and surgical specimens at the point of care. CSLM requires minimal tissue processing and has the potential to reduce the time from excision to diagnosis. Following imaging, specimens can still be submitted for standard histopathological preparation. Methods Needle core breast specimens from 49 patients were imaged at the time of biopsy. These lesions had been characterized under the Breast Imaging Reporting And Data System (BI-RADS) as category 3, 4 or 5. The core biopsies were imaged with the CSLM before fixation. Samples were treated with 5% citric acid and glycerin USP to enhance nuclear visibility in the reflectance confocal images. Immediately following imaging, the specimens were fixed in buffered formalin and submitted for histological processing and pathological diagnosis. CSLM images were then compared to the standard histology. Results The pathologic diagnoses by standard histology were 7 invasive ductal carcinomas, 2 invasive lobular carcinomas, 3 ductal carcinomas in-situ (CIS), 21 fibrocystic changes/proliferative conditions, 9 fibroadenomas, and 5 other/benign; two were excluded due to imaging difficulties. Morphologic and cellular features of benign and cancerous lesions were identified in the confocal images and were comparable to standard histologic sections of the same tissue. Conclusion CSLM is a technique with the potential to screen needle core biopsy specimens in real-time. The confocal images contained sufficient information to identify stromal reactions such as fibrosis and cellular proliferations such as intra-ductal and infiltrating carcinoma, and

  17. Probe-based confocal laser endomicroscopy (pCLE) - a new imaging technique for in situ localization of spermatozoa.

    Science.gov (United States)

    Trottmann, Matthias; Stepp, Herbert; Sroka, Ronald; Heide, Michael; Liedl, Bernhard; Reese, Sven; Becker, Armin J; Stief, Christian G; Kölle, Sabine

    2015-05-01

    In azoospermic patients, spermatozoa are routinely obtained by testicular sperm extraction (TESE). However, success rates of this technique are moderate, because the site of excision of testicular tissue is determined arbitrarily. Therefore the aim of this study was to establish probe-based laser endomicroscopy (pCLE) a noval biomedical imaging technique, which provides the opportunity of non-invasive, real-time visualisation of tissue at histological resolution. Using pCLE we clearly visualized longitudinal and horizontal views of the tubuli seminiferi contorti and localized vital spermatozoa. Obtained images and real-time videos were subsequently compared with confocal laser scanning microscopy (CLSM) of spermatozoa and tissues, respectively. Comparative visualization of single native Confocal laser scanning microscopy (CLSM, left) and probe-based laser endomicroscopy (pCLE, right) using Pro Flex(TM) UltraMini O after staining with acriflavine. © 2014 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  18. Chromatic confocal microscopy using staircase diffractive surface.

    Science.gov (United States)

    Rayer, Mathieu; Mansfield, Daniel

    2014-08-10

    A chromatic confocal microscope (CCM) is a high-dynamic-range noncontact distance measurement sensor; it is based on a hyperchromatic lens. The vast majority of commercial CCMs use refractive-based chromatic dispersion to chromatically code the optical axis. This approach significantly limits the range of applications and performance of the CCM. In order to be a suitable alternative to a laser triangulation gauge and laser encoder, the performance of the CCM must be improved. In this paper, it is shown how hybrid aspheric diffractive (HAD) lenses can bring the CCM to its full potential by increasing the dynamic range by a factor of 2 and the resolution by a factor of 5 while passively athermizing and increasing the light throughput efficiency of the optical head [M. Rayer, U.S. patent 1122052.2 (2011)]. The only commercially suitable manufacturing process is single-point diamond turning. However, the optical power carried by the diffractive side of a hybrid aspheric diffractive lens is limited by the manufacturing process. A theoretical study of manufacturing losses has revealed that the HAD configuration with the highest diffraction efficiency is for a staircase diffractive surface (SDS). SDS lenses have the potential to reduce light losses associated with manufacturing limits by a factor of 5 without increasing surface roughness, allowing scalar diffraction-limited optical design with a diffractive element.

  19. Microelectrophoresis of Silica Rods Using Confocal Microscopy

    Science.gov (United States)

    2017-01-01

    The electrophoretic mobility and the zeta potential (ζ) of fluorescently labeled colloidal silica rods, with an aspect ratio of 3.8 and 6.1, were determined with microelectrophoresis measurements using confocal microscopy. In the case where the colloidal particles all move at the same speed parallel to the direction of the electric field, we record a xyz-stack over the whole depth of the capillary. This method is faster and more robust compared to taking xyt-series at different depths inside the capillary to obtain the parabolic flow profile, as was done in previous work from our group. In some cases, rodlike particles do not move all at the same speed in the electric field, but exhibit a velocity that depends on the angle between the long axis of the rod and the electric field. We measured the orientation-dependent velocity of individual silica rods during electrophoresis as a function of κa, where κ–1 is the double layer thickness and a is the radius of the rod associated with the diameter. Thus, we determined the anisotropic electrophoretic mobility of the silica rods with different sized double layers. The size of the double layer was tuned by suspending silica rods in different solvents at different electrolyte concentrations. We compared these results with theoretical predictions. We show that even at already relatively high κa when the Smoluchowski limiting law is assumed to be valid (κa > 10), an orientation dependent velocity was measured. Furthermore, we observed that at decreasing values of κa the anisotropy in the electrophoretic mobility of the rods increases. However, in low polar solvents with κa mobility of the rods decreased. We argue that this decrease is due to end effects, which was already predicted theoretically. When end effects are not taken into account, this will lead to strong underestimation of the experimentally determined zeta potential. PMID:28045541

  20. Effects of acids used in the microabrasion technique: Microhardness and confocal microscopy analysis.

    Science.gov (United States)

    Pini, Núbia-Inocencya-Pavesi; Lima, Débora-Alves-Nunes-Leite; Ambrosano, Gláucia-Maria-Bovi; da Silva, Wander-José; Aguiar, Flávio-Henrique-Baggio; Lovadino, José-Roberto

    2015-10-01

    This study evaluated the effects of the acids used in the microabrasion on enamel. Seventy enamel/dentine blocks (25 mm2) of bovine incisors were divided into 7 groups (n=10). Experimental groups were treated by active/passive application of 35% H3PO4 (E1/E2) or 6.6% HCl (E3/E4). Control groups were treated by microabrasion with H3PO4+pumice (C5), HCl+silica (C6), or no treatment (C7). The superficial (SMH) and cross-sectional (CSMH; depths of 10, 25, 50, and 75 µm) microhardness of enamel were analyzed. Morphology was evaluated by confocal laser-scanning microscopy (CLSM). Data were analyzed by analysis of variance (Proc Mixed), Tukey, and Dunnet tests (α=5%). Active application (E1 and E3) resulted in higher microhardness than passive application (E2 and E4), with no difference between acids. For most groups, the CSMH decreased as the depth increased. All experimental groups and negative controls (C5 and C6) showed significantly reduced CSMH values compared to the control. A significantly higher mean CSMH result was obtained with the active application of H3PO4 (E1) compared to HCl (E3). Passive application did not result in CSMH differences between acids. CLSM revealed the conditioning pattern for each group. Although the acids displayed an erosive action, use of microabrasive mixture led to less damage to the enamel layers. Enamel microabrasion, enamel microhardness, confocal laser scanning microscopy.

  1. Handheld reflectance confocal microscopy for the diagnosis of conjunctival tumors.

    Science.gov (United States)

    Cinotti, Elisa; Perrot, Jean-Luc; Labeille, Bruno; Campolmi, Nelly; Espinasse, Marine; Grivet, Damien; Thuret, Gilles; Gain, Philippe; Douchet, Catherine; Andrea, Caroline; Haouas, Maher; Cambazard, Frédéric

    2015-02-01

    To evaluate whether the handheld in vivo reflectance confocal microscopy that has been recently developed for the study of skin tumors is suitable for the diagnosis of conjunctival tumors. Prospective study, observational case series. We prospectively evaluated the reflectance confocal microscopy features of 53 conjunctival lesions clinically suspicious for tumors of 46 patients referred to the University Hospital of Saint-Etienne (France) by using the handheld device. Twenty-three lesions were excised (3 nevi, 10 melanomas, 5 squamous cell carcinoma, 2 lymphomas, and 3 pinguecula/pterygium) while the other 30, presenting no reflectance confocal microscopy malignant features, were under follow-up for at least 1 year. Clinical reflectance confocal microscopy and histologic diagnosis were compared. In vivo reflectance confocal microscopy diagnosis was in agreement with the histologic diagnosis in all cases and none of the lesions that were not excised show any clinical progression under follow-up. In vivo reflectance confocal microscopy with a handheld dermatology-dedicated microscope can play a role in the noninvasive diagnosis of conjunctival lesions. Further studies should be performed to better define the diagnostic ability of this technique. Copyright © 2015 Elsevier Inc. All rights reserved.

  2. Thyroid Scan and Uptake

    Medline Plus

    Full Text Available ... News Physician Resources Professions Site Index A-Z Thyroid Scan and Uptake Thyroid scan and uptake uses ... the Thyroid Scan and Uptake? What is a Thyroid Scan and Uptake? A thyroid scan is a ...

  3. Thyroid Scan and Uptake

    Science.gov (United States)

    ... News Physician Resources Professions Site Index A-Z Thyroid Scan and Uptake Thyroid scan and uptake uses ... the Thyroid Scan and Uptake? What is a Thyroid Scan and Uptake? A thyroid scan is a ...

  4. Methods for studying biofilm formation: flow cells and confocal laser scanning microscopy

    DEFF Research Database (Denmark)

    Tolker-Nielsen, Tim; Sternberg, Claus

    2014-01-01

    In this chapter methods for growing and analyzing biofilms under hydrodynamic conditions in flow cells are described. Use of flow cells allows for direct microscopic investigation of biofilm formation. The flow in these chambers is essentially laminar, which means that the biofilms can be grown...

  5. Plasmon resonance and the imaging of metal-impregnated neurons with the laser scanning confocal microscope

    OpenAIRE

    Thompson, Karen J; Harley, Cynthia M.; Barthel, Grant M; Sanders, Mark A.; Mesce, Karen A.

    2015-01-01

    eLife digest A fresh slice of brain tissue has a fairly uniform appearance, even when viewed under a microscope. To study the neurons and other cells in the brain, scientists must therefore first prepare tissue samples using methods that make it easier to see certain kinds of cells, or particular features of them. One method that has been available for over a century is to use metal particles to stain some of the cells. For example, when the Spanish anatomist Santiago Ram?n y Cajal investigat...

  6. Sarcoglycan and integrin localization in normal human skeletal muscle: a confocal laser scanning microscope study

    Directory of Open Access Journals (Sweden)

    G Anastasi

    2009-06-01

    Full Text Available Many studies have been performed on the sarcoglycan subcomplex and a7B and b1D integrins, but their distribution and localization patterns along the non-junctional sarcolemma are still not clear. We have carried out an indirect immunofluorescence study on surgical biopsies of normal human skeletal muscle, performing double localization reactions with antibodies to sarcoglycans, integrins and sarcomeric actin. Our results indicate that the tested proteins colocalize with each other. In a few cases, a-sarcoglycan does not colocalize with the other sarcoglycans and integrins. We also demonstrated, by employing antibodies to all the tested proteins, that these proteins can be localized to regions of the sarcolemma corresponding either to the I-band or Aband. Our results seem to confirm the hypothesis of a correlation between the region of the sarcolemma occupied by costameric proteins and the metabolic type (fast or slow of muscle fibers. On this basis, we suggest that slow fibers are characterized by localization of costameric proteins to Ibands, while fast fibers are characterized by localization of costameric proteins to A-bands. The results open a new line of research in understanding interactions between the components of the DGC and vinculin-talin-integrin complexes in the context of different fiber types. Moreover, the same results may be extended to skeletal muscle fibers affected by neuromuscular diseases to detect possible structural alterations.

  7. Purchase of a Laser Scanning Confocal Microscope at Xavier University of Louisiana

    Science.gov (United States)

    2016-05-04

    TERMS b. ABSTRACT 2. REPORT TYPE 17. LIMITATION OF ABSTRACT 15. NUMBER OF PAGES 5d. PROJECT NUMBER 5e. TASK NUMBER 5f. WORK UNIT NUMBER 5c...at the Kinesin-Microtubule Interface". Annual Meeting of the American Society for Cell Biology , San Diego CA, December 2015. (c) Presentations...stained pathology specimens to further identify pathologies associated with unknown biopsied tissue. This gave them a hands-on experience with

  8. Confocal laser scanning microscopy evaluation of an acellular dermis tissue transplant (Epiflex®.

    Directory of Open Access Journals (Sweden)

    Eric Dominic Roessner

    Full Text Available The structure of a biological scaffold is a major determinant of its biological characteristics and its interaction with cells. An acellular dermis tissue transplant must undergo a series of processing steps, to remove cells and genetic material and provide the sterility required for surgical use. During manufacturing and sterilization the structure and composition of tissue transplants may change. The composition of the human cell-free dermis transplant Epiflex® was investigated with specific attention paid to its structure, matrix composition, cellular content and biomechanics. We demonstrated that after processing, the structure of Epiflex remains almost unchanged with an intact collagen network and extracellular matrix (ECM protein composition providing natural cell interactions. Although the ready to use transplant does contain some cellular and DNA debris, the processing procedure results in a total destruction of cells and active DNA which is a requirement for an immunologically inert and biologically safe substrate. Its biomechanical parameters do not change significantly during the processing.

  9. Sarcoglycan and integrin localization in normal human skeletal muscle: a confocal laser scanning microscope study.

    Science.gov (United States)

    Anastasi, G; Cutroneo, G; Rizzo, G; Arco, A; Santoro, G; Bramanti, P; Vitetta, A G; Pisani, A; Trimarchi, F; Favaloro, A

    2004-01-01

    Many studies have been performed on the sarcoglycan sub-complex and a7B and b1D integrins, but their distribution and localization patterns along the non-junctional sarcolemma are still not clear. We have carried out an indirect immunofluorescence study on surgical biopsies of normal human skeletal muscle, performing double localization reactions with antibodies to sarcoglycans, integrins and sarcomeric actin. Our results indicate that the tested proteins colocalize with each other. In a few cases, a-sarcoglycan does not colocalize with the other sarcoglycans and integrins. We also demonstrated, by employing antibodies to all the tested proteins, that these proteins can be localized to regions of the sarcolemma corresponding either to the I-band or A-band. Our results seem to confirm the hypothesis of a correlation between the region of the sarcolemma occupied by costameric proteins and the metabolic type (fast or slow) of muscle fibers. On this basis, we suggest that slow fibers are characterized by localization of costameric proteins to I-bands, while fast fibers are characterized by localization of costameric proteins to A-bands. The results open a new line of research in understanding interactions between the components of the DGC and vinculin-talin-integrin complexes in the context of different fiber types. Moreover, the same results may be extended to skeletal muscle fibers affected by neuromuscular diseases to detect possible structural alterations.

  10. Sarcoglycans in human skeletal muscle and human cardiac muscle: a confocal laser scanning microscope study.

    Science.gov (United States)

    Anastasi, G; Cutroneo, G; Trimarchi, F; Rizzo, G; Bramanti, P; Bruschetta, D; Fugazzotto, D; Cinelli, M P; Soscia, A; Santoro, G; Favaloro, A

    2003-01-01

    Sarcoglycans are a subcomplex of transmembrane proteins which are part of the dystrophin-glycoprotein complex. They are expressed in the skeletal, cardiac and smooth muscle. Although numerous studies have been conducted on the sarcoglycan subcomplex in skeletal and cardiac muscle, the manner of the distribution and localization of these proteins along the nonjunctional sarcolemma is not clear. We therefore carried out an indirect immunofluorescence study on surgical biopsies of normal human skeletal muscle and of healthy human atrial myocardium biopsies of patients affected by valvulopathy. Our results indicate that, in skeletal muscle, sarcoglycans have a costameric distribution and all colocalize with each other. Only in a few cases did the alpha-sarcoglycan not colocalize with other sarcoglycans. In addition, these glycoproteins can be localized in different fibers either in the regions of the sarcolemma over band I or band A. In cardiac muscle, our results show a costameric distribution of all proteins examined and, unlike in skeletal muscle, they show a constant colocalization of all sarcoglycans with each other, along with a consistent localization of these proteins in the region of the sarcolemma over band I. In our opinion, this situation seems to confirm the hypothesis of a correlation between the region of the sarcolemma occupied by costameric proteins and the metabolic type, fast or slow, of the muscular fibers. These data, besides opening a new line of research in understanding interactions between the sarcoglycans and other transmembrane proteins, could also be extended to skeletal and cardiac muscles affected by neuromuscular and cardiovascular pathologies to understand possible structural alterations. Copyright 2003 S. Karger AG, Basel

  11. Hyperchromatic laser scanning cytometry

    Science.gov (United States)

    Tárnok, Attila; Mittag, Anja

    2007-02-01

    In the emerging fields of high-content and high-throughput single cell analysis for Systems Biology and Cytomics multi- and polychromatic analysis of biological specimens has become increasingly important. Combining different technologies and staining methods polychromatic analysis (i.e. using 8 or more fluorescent colors at a time) can be pushed forward to measure anything stainable in a cell, an approach termed hyperchromatic cytometry. For cytometric cell analysis microscope based Slide Based Cytometry (SBC) technologies are ideal as, unlike flow cytometry, they are non-consumptive, i.e. the analyzed sample is fixed on the slide. Based on the feature of relocation identical cells can be subsequently reanalyzed. In this manner data on the single cell level after manipulation steps can be collected. In this overview various components for hyperchromatic cytometry are demonstrated for a SBC instrument, the Laser Scanning Cytometer (Compucyte Corp., Cambridge, MA): 1) polychromatic cytometry, 2) iterative restaining (using the same fluorochrome for restaining and subsequent reanalysis), 3) differential photobleaching (differentiating fluorochromes by their different photostability), 4) photoactivation (activating fluorescent nanoparticles or photocaged dyes), and 5) photodestruction (destruction of FRET dyes). With the intelligent combination of several of these techniques hyperchromatic cytometry allows to quantify and analyze virtually all components of relevance on the identical cell. The combination of high-throughput and high-content SBC analysis with high-resolution confocal imaging allows clear verification of phenotypically distinct subpopulations of cells with structural information. The information gained per specimen is only limited by the number of available antibodies and by sterical hindrance.

  12. Microstructural evaluation by confocal and electron microscopy in thrombi developed in central venous catheters.

    Science.gov (United States)

    Lucas, Thabata Coaglio; Silva, Eliata Ester da; Souza, Danilo Olzon Dionysio; Santos, Amanda Rodrigues Dos; Lara, Maristela Oliveira

    2017-08-28

    Evaluating thrombi microstructure developed in central venous catheters using confocal and electron microscopy. An experimental, descriptive study carrying out a microstructural evaluation of venous thrombi developed in central venous catheters using Scanning Electron Microscopy and Confocal Laser Scanning Microscopy. A total of 78 venous catheters were collected over a period of three months. Different fibrin structures were distinguished: fibrin plates, fibrin network, and fibrin fibers. It was observed that the thrombus had thick fibrin plates adhered to the catheter wall openings in both a catheter with three days of permanence as well as in a catheter with 20 days of insertion in the patient. However, a greater amount of erythrocytes and fibrin fibers were found in the central region of the thrombus. This study contributes to improving health care and can have a positive impact on clinical practice, as easy adherence of platelets and fibrins to the catheter wall demonstrated in this study makes it possible to adopt thrombus prevention strategies such as therapy discontinuation for an extended period, blood reflux by a catheter, slow infusion rate and hypercoagulo pathyclinical conditions. Avaliar a microestrutura por microscopia confocal e eletrônica em trombos desenvolvidos em cateteres venosos centrais. Pesquisa experimental, descritiva, em que foi feita uma avaliação microestrutural de trombos venosos desenvolvidos em cateteres venosos centrais por Microscopia Eletrônica de Varredura e Microscopia Confocal de Varredura a Laser. Foram coletados 78 cateteres venosos centrais num período de três meses. Distinguiram-se diferentes estruturas de fibrina: a placa de fibrina, a rede de fibrina e as fibras de fibrina. Observou-se que tanto em um cateter com três dias de permanência quanto em um cateter com 20 dias inserido no paciente o trombo apresentou placas de fibrina espessas aderidas às paredes dos orifícios dos cateteres. Na região central do

  13. Combining confocal microscopy and optical coherence tomography for imaging in developmental biology

    Science.gov (United States)

    Bradu, A.; Ma, Lisha; Bloor, J.; Podoleanu, A.

    2008-04-01

    In-vivo Optical Coherence Tomography (OCT) imaging of the fruit fly Drosophila melanogaster larval heart allows non invasive visualizations and assesment of its cardiac functions. To image Drosophila melanogaster heart, we have developed a dedicated imaging instrument able to provide simultaneous Optical Coherence Tomography (OCT) and Laser Confocal Scanning Microscopy (LCSM) or Laser Scanning Fluorescence Microscopy (LSFM) images and can be used to produce B-scan OCT images. With this dual imaging system, the image of heart can be easily located in the specimen and the change of the heart shape in a cardiac cycle monitored. This technique therefore provides an excellent tool for large scale screen of candidate genes responsible for the contractility of the Drosophila heart. As this technique can also image the dynamic process of the heartbeat in a non-invasive fashion, it provides a new avenue to study the physiology of the heart function. En-face and B-scan OCT images of the Drosophila melanogaster heart showing its chambers have been obtained with our imaging instruments. Our results are consistent with detailed anatomical studies from the literature.

  14. Freeze-thaw immobilization of liposomes in chromatographic gel beads: evaluation by confocal microscopy and effects of freezing rate.

    Science.gov (United States)

    Lundqvist, A; Ocklind, G; Haneskog, L; Lundahl, P

    1998-01-01

    Biological membranes immobilized in chromatographic gel beads constitute a multifunctional affinity matrix. Membrane protein-solute interactions and drug partitioning into the lipid bilayers can conveniently be studied. By the use of confocal laser-scanning microscopy (CLSM) the distribution of immobilized model membranes in the beads has been visualized for the first time. Freeze-thaw-immobilized liposomes in Superdex 200 gel beads were situated in a thick shell surrounding a liposome-free core. The amount of phospholipids immobilized by freeze-thawing was dependent on the temperature in the cooling bath and the type of test tube used. A bath temperature of -25 degrees C gave higher immobilization yield than freezing at -75 or -8 degrees C did. Freeze-thawing in the presence of liposomes did not affect the gel bead shape or the refractive index homogeneity of the agarose network of the beads, as shown by confocal microscopy.

  15. Handheld tunable focus confocal microscope utilizing a double-clad fiber coupler for in vivo imaging of oral epithelium

    Science.gov (United States)

    Olsovsky, Cory; Hinsdale, Taylor; Cuenca, Rodrigo; Cheng, Yi-Shing Lisa; Wright, John M.; Rees, Terry D.; Jo, Javier A.; Maitland, Kristen C.

    2017-05-01

    A reflectance confocal endomicroscope with double-clad fiber coupler and electrically tunable focus lens is applied to imaging of the oral mucosa. The instrument is designed to be lightweight and robust for clinical use. The tunable lens allows axial scanning through >250 μm in the epithelium when the probe tip is placed in contact with tissue. Images are acquired at 6.6 frames per second with a field of view diameter up to 850 μm. In vivo imaging of a wide range of normal sites in the oral cavity demonstrates the accessibility of the handheld probe. In vivo imaging of clinical lesions diagnosed as inflammation and dysplasia illustrates the ability of reflectance confocal endomicroscopy to image cellular changes associated with pathology.

  16. Second-harmonic patterned polarization-analyzed reflection confocal microscope

    Science.gov (United States)

    Okoro, Chukwuemeka; Toussaint, Kimani C.

    2017-08-01

    We introduce the second-harmonic patterned polarization-analyzed reflection confocal (SPPARC) microscope-a multimodal imaging platform that integrates Mueller matrix polarimetry with reflection confocal and second-harmonic generation (SHG) microscopy. SPPARC microscopy provides label-free three-dimensional (3-D), SHG-patterned confocal images that lend themselves to spatially dependent, linear polarimetric analysis for extraction of rich polarization information based on the Mueller calculus. To demonstrate its capabilities, we use SPPARC microscopy to analyze both porcine tendon and ligament samples and find differences in both circular degree-of-polarization and depolarization parameters. Moreover, using the collagen-generated SHG signal as an endogenous counterstain, we show that the technique can be used to provide 3-D polarimetric information of the surrounding extrafibrillar matrix plus cells or EFMC region. The unique characteristics of SPPARC microscopy holds strong potential for it to more accurately and quantitatively describe microstructural changes in collagen-rich samples in three spatial dimensions.

  17. Thyroid Scan and Uptake

    Medline Plus

    Full Text Available ... limitations of the Thyroid Scan and Uptake? What is a Thyroid Scan and Uptake? A thyroid scan is ... top of page What are some common uses of the procedure? The thyroid scan is used to ...

  18. Thyroid Scan and Uptake

    Medline Plus

    Full Text Available ... of the Thyroid Scan and Uptake? What is a Thyroid Scan and Uptake? A thyroid scan is ... of page What are some common uses of the procedure? The thyroid scan is used to determine ...

  19. Lumbar spine CT scan

    Science.gov (United States)

    CAT scan - lumbar spine; Computed axial tomography scan - lumbar spine; Computed tomography scan - lumbar spine; CT - lower ... The lumbar CT scan is good for evaluating large herniated disks, ... smaller ones. This test can be combined with a myelogram to get ...

  20. Arm CT scan

    Science.gov (United States)

    CAT scan - arm; Computed axial tomography scan - arm; Computed tomography scan - arm; CT scan - arm ... stopping.) A computer creates separate images of the arm area, called slices. These images can be stored, ...

  1. Thoracic spine CT scan

    Science.gov (United States)

    CAT scan - thoracic spine; Computed axial tomography scan - thoracic spine; Computed tomography scan - thoracic spine; CT scan - ... Philadelphia, PA: Elsevier Mosby; 2013:chap 44. US Food and Drug Administration. Computed tomography (CT). Updated August ...

  2. EUS-Guided Needle-Based Confocal Laser Endomicroscopy

    DEFF Research Database (Denmark)

    Bhutani, Manoop S; Koduru, Pramoda; Joshi, Virendra

    2015-01-01

    the gut, providing further diagnostic and staging information. Confocal laser endomicroscopy (CLE) is a novel endoscopic method that enables imaging at a subcellular level of resolution during endoscopy, allowing up to 1000-fold magnification of tissue and providing an optical biopsy. A new procedure...... that has been developed in the past few years is needle-based confocal laser endomicroscopy (nCLE), which involves a mini-CLE probe that can be passed through a 1 9-gauge needle during EUS-FNA. This enables the real-time visualization of tissue at a microscopic level, with the potential to further improve...

  3. Enhanced compton backscattering by confocal multipath laser cavity

    Energy Technology Data Exchange (ETDEWEB)

    Amano, Sho; Miyamoto, Shuji; Mochizuki, Takayasu [Himeji Institute of Technology, Laboratory of Advanced Science Technology for Industry, Kamigori, Hyogo (Japan)

    2001-10-01

    The design considerations of a confocal multipath laser cavity to enhance Compton backscattering are presented. Laser pulses are superposed at a confocal point of the cavity and enhance laser peak filed there. Ray trace simulation results predicted that the 29 - 14-fold enhanced laser filed could be achieved with the mode locked laser pulses whose repetition rate and duration time were 89.25 MHz and 10 ps, respectively. As a result, Compton backscattered X-rays generated by interaction of this intense laser field with a relativistic electron beam, will be enhanced efficiently by a factor of more than 10 at least. (author)

  4. Microscopia confocal in vivo na cistinose: relato de caso

    Directory of Open Access Journals (Sweden)

    Victor Gustavo

    2004-01-01

    Full Text Available A cistinose é doença autossômica recessiva rara caracterizada pelo acúmulo do aminoácido cistina livre dentro dos lisossomos e geralmente é fatal na primeira década de vida na ausência de transplante renal. O presente estudo tem por objetivo relatar os achados da microscopia confocal in vivo em paciente adulto com cistinose infantil. O exame de microscopia confocal in vivo revelou que há diferenças quanto à intensidade de acometimento, tamanho e forma dos depósitos nas diversas camadas corneanas.

  5. Confocal Raman microscopy for identification of bacterial species in biofilms

    Science.gov (United States)

    Beier, Brooke D.; Quivey, Robert G.; Berger, Andrew J.

    2011-03-01

    Implemented through a confocal microscope, Raman spectroscopy has been used to distinguish between biofilm samples of two common oral bacteria species, Streptococcus sanguinis and mutans, which are associated with healthy and cariogenic plaque, respectively. Biofilms of these species are studied as a model of dental plaque. A prediction model has been calibrated and validated using pure biofilms. This model has been used to identify the species of transferred and dehydrated samples (much like a plaque scraping) as well as hydrated biofilms in situ. Preliminary results of confocal Raman mapping of species in an intact two-species biofilm will be shown.

  6. Internal features of graphite in cast irons. Confocal microscopy: useful tool for graphite growth imaging.

    Science.gov (United States)

    Llorca-Isern, N; Tartera, J; Espanol, M; Marsal, M; Bertran, G; Castel, S

    2002-01-01

    Spherulitic crystallisation is a mode of growth of crystals from the melt. Considerable attention has been given to spheroidal graphite formation, providing detailed information about the internal microstructure of the spherulites in spheroidal (SG irons) and compacted graphite irons (CG irons) (Stefanescu, D., 1990. Cast Irons. ASM Handbook, 10th ed., vol. 1). Nevertheless, the mechanisms responsible for this mode of crystallisation are not fully understood. This study deals with the inoculation mechanisms, with particular emphasis on the study of the inclusions for the heterogeneous nucleation of graphite. It is shown that the graphite nuclei are sulfide products of the nodularizing treatment. It has been observed that when rare-earth treatment is applied, the central nucleus consists of a core and an envelope from which the graphite grows. Confocal Scanning Laser Microscopy (CSLM), in reflection mode, was used to study the internal features of the spheroidal graphite growth. Confocal reflection imaging, which has a capacity for optical sectioning of the sample, provides high-resolution images of surface and subsurface regions of interest contained within a semi-transparent sample. Furthermore, three-dimensional reconstruction of these optical sections can provide insight into the mechanism of graphite growth mechanism interpretation. With CSLM the radial growth of graphite was seen. Other techniques, such as TEM, SEM-EDS, WDS, AES and SAM were also used to corroborate the results.

  7. In vivo near-infrared dual-axis confocal microendoscopy in the human lower gastrointestinal tract.

    Science.gov (United States)

    Piyawattanametha, Wibool; Ra, Hyejun; Qiu, Zhen; Friedland, Shai; Liu, Jonathan T C; Loewke, Kevin; Kino, Gordon S; Solgaard, Olav; Wang, Thomas D; Mandella, Michael J; Contag, Christopher H

    2012-02-01

    Near-infrared confocal microendoscopy is a promising technique for deep in vivo imaging of tissues and can generate high-resolution cross-sectional images at the micron-scale. We demonstrate the use of a dual-axis confocal (DAC) near-infrared fluorescence microendoscope with a 5.5-mm outer diameter for obtaining clinical images of human colorectal mucosa. High-speed two-dimensional en face scanning was achieved through a microelectromechanical systems (MEMS) scanner while a micromotor was used for adjusting the axial focus. In vivo images of human patients are collected at 5 frames/sec with a field of view of 362×212 μm(2) and a maximum imaging depth of 140 μm. During routine endoscopy, indocyanine green (ICG) was topically applied a nonspecific optical contrasting agent to regions of the human colon. The DAC microendoscope was then used to obtain microanatomic images of the mucosa by detecting near-infrared fluorescence from ICG. These results suggest that DAC microendoscopy may have utility for visualizing the anatomical and, perhaps, functional changes associated with colorectal pathology for the early detection of colorectal cancer.

  8. Comparison of Three Different Sealer Placement Techniques: An In vitro Confocal Laser Microscopic Study

    Science.gov (United States)

    Dash, Avoy Kumar; Farista, Shanin; Dash, Abhilasha; Bendre, Ajinkya; Farista, Sana

    2017-01-01

    Introduction: Three-dimensional obturation of the root canal system is the final objective of root canal therapy. Greater penetration of sealer in root dentine lesser will be the voids at the dentine–sealer interface. Hence, analysis of the dentin/sealer interface allows the determination of a filling technique which could obturate the root canals with least gaps and voids. Therefore, the aim of this study is to compare the depth and percentage of sealer penetration into root dentin using three different root canal sealer placement techniques under confocal laser scanning microscope. Materials and Methods: Thirty single-rooted teeth were selected and prepared. Adseal sealer (Meta Biomed, South Korea) was mixed with Rhodamine B dye and applied using lentulo spiral (Dentsply Maillefer, USA) as Group 1, bidirectional spiral (EZ-Fill– EDS, USA) as Group 2, and ultrasonic endodontic tip (Sonofile– Dentsply Tulsa, USA) as Group 3. Canals were then obturated with gutta-percha. The roots were sectioned at the 3 and 6-mm levels from the apical foramen and examined under confocal laser microscope. Results: Maximum mean depth and percentage of sealer penetration were observed for Group 1 and minimum for Group 3. Furthermore, statistical significant differences among Group 1 and Group 3 were found at 6-mm level and among Group 2 and Group 3 were found at 3-mm level (P ultrasonics. PMID:28839420

  9. Atomic force microscopy analysis and confocal Raman microimaging of coated pellets.

    Science.gov (United States)

    Ringqvist, Ann; Taylor, Lynne S; Ekelund, Katarina; Ragnarsson, Gert; Engström, Sven; Axelsson, Anders

    2003-11-28

    Polymer-coated pellets with different coating thicknesses have been studied regarding coating morphology and drug release properties with atomic force microscopy (AFM) and confocal Raman microscopy. The results were compared with those from scanning electron microscopy (SEM) and drug release profiles, which have been measured previously for these systems and found to vary depending on coating thickness. Results from AFM studies indicated that these pellets differ in the amount of crystalline material on the surface of the coating. The amount was found to be highest on the pellet with the thinnest coating. Confocal Raman microscopy studies confirmed that the active component (remoxipride hydrochloride monohydrate) is present at or close to the surface and that the amount is higher for the thinnest coating. AFM studies in aqueous media showed that the crystalline material on the surface was almost instantaneously dissolved and released into the liquid. AFM has proven to be a powerful tool in the study of the surface of dry formulations and in the study of the controlled release mechanism of a pharmaceutical in a liquid cell. The method can be combined with Raman, giving the added possibility to identify the chemical composition in selected small areas of the coating surface.

  10. Fully automatic evaluation of the corneal endothelium from in vivo confocal microscopy.

    Science.gov (United States)

    Selig, Bettina; Vermeer, Koenraad A; Rieger, Bernd; Hillenaar, Toine; Luengo Hendriks, Cris L

    2015-04-26

    Manual and semi-automatic analyses of images, acquired in vivo by confocal microscopy, are often used to determine the quality of corneal endothelium in the human eye. These procedures are highly time consuming. Here, we present two fully automatic methods to analyze and quantify corneal endothelium imaged by in vivo white light slit-scanning confocal microscopy. In the first approach, endothelial cell density is estimated with the help of spatial frequency analysis. We evaluate published methods, and propose a new, parameter-free method. In the second approach, based on the stochastic watershed, cells are automatically segmented and the result is used to estimate cell density, polymegathism (cell size variability) and pleomorphism (cell shape variation). We show how to determine optimal values for the three parameters of this algorithm, and compare its results to a semi-automatic delineation by a trained observer. The frequency analysis method proposed here is more precise than any published method. The segmentation method outperforms the fully automatic method in the NAVIS software (Nidek Technologies Srl, Padova, Italy), which significantly overestimates the number of cells for cell densities below approximately 1200 mm(-2), as well as previously published methods. The methods presented here provide a significant improvement over the state of the art, and make in vivo, automated assessment of corneal endothelium more accessible. The segmentation method proposed paves the way to many possible new morphometric parameters, which can quickly and precisely be determined from the segmented image.

  11. Probe-based confocal laser endomicroscopy in head and neck malignancies: early preclinical experience

    Science.gov (United States)

    Englhard, Anna; Girschick, Susanne; Mack, Brigitte; Volgger, Veronika; Gires, Oliver; Conderman, Christian; Stepp, Herbert; Betz, Christian Stephan

    2013-06-01

    Background: Malignancies of the upper aerodigestive tract (UADT) are conventionally diagnosed by white light endoscopy, biopsy and histopathology. Probe-based Confocal Laser Endomicroscopy (pCLE) is a novel non-invasive technique which offers in vivo surface and sub-surface imaging of tissue. It produces pictures of cellular architecture comparable to histology without the need for biopsy. It has already been successfully used in different clinical subspecialties to help in the diagnosis and treatment planning of inflammatory and neoplastic diseases. PCLE needs to be used in combination with specific or non-specific contrast agents. In this study we evaluated the potential use of pCLE in combination with non-specific and specific contrast agents to distinguish between healthy mucosa and invasive carcinoma. Methods: Tissue samples from healthy mucosa and squamous cell carcinoma of the head and neck were taken during surgery. After topical application of three different contrast agents, samples were examined using different pCLE-probes and a Confocal Laser Scanning Microscope (CLSM). Images were then compared to the corresponding histological slides and cryosections. Results: Initial results show that pCLE in combination with fluorophores allows visualization of cellular and structural components. Imaging of different layers was possible using three distinct pCLEprobes. Conclusion: pCLE is a promising non-invasive technique that may be a useful adjunct in the evaluation, diagnosis and treatment planning of head and neck malignancies.

  12. Detection and imaging of lipids ofScenedesmus obliquusbased on confocal Raman microspectroscopy.

    Science.gov (United States)

    Shao, Yongni; Fang, Hui; Zhou, Hong; Wang, Qi; Zhu, Yiming; He, Yong

    2017-01-01

    In this study, confocal Raman microspectroscopy was used to detect lipids in microalgae rapidly and non-destructively. Microalgae cells were cultured under nitrogen deficiency. The accumulation of lipids in Scenedesmus obliquus was observed by Nile red staining, and the total amount of lipids accumulated in the cells was measured by gravimetric method. The signals from different microalgae cells were collected by confocal Raman microspectroscopy to establish a prediction model of intracellular lipid content, and surface scanning signals for drawing pseudo color images of lipids distribution. The images can show the location of pyrenoid and lipid accumulation in cells. Analyze Raman spectrum data and build PCA-LDA model using four different bands (full bands, pigments, lipids, and mixed features). Models of full bands or pigment characteristic bands were capable of identifying S. obliquus cells under different nitrogen stress culture time. The prediction accuracy of model of lipid characteristic bands is relatively low. The correlation between the fatty acid content measured by the gravimetric method and the integral Raman intensity of the oil characteristic peak (1445 cm -1 ) measured by Raman spectroscopy was analyzed. There was significant correlation ( R 2  = 0.83), which means that Raman spectroscopy is applicable to semi-quantitative detection of microalgal lipid content.

  13. Experiments on terahertz 3D scanning microscopic imaging

    Science.gov (United States)

    Zhou, Yi; Li, Qi

    2016-10-01

    Compared with the visible light and infrared, terahertz (THz) radiation can penetrate nonpolar and nonmetallic materials. There are many studies on the THz coaxial transmission confocal microscopy currently. But few researches on the THz dual-axis reflective confocal microscopy were reported. In this paper, we utilized a dual-axis reflective confocal scanning microscope working at 2.52 THz. In contrast with the THz coaxial transmission confocal microscope, the microscope adopted in this paper can attain higher axial resolution at the expense of reduced lateral resolution, revealing more satisfying 3D imaging capability. Objects such as Chinese characters "Zhong-Hua" written in paper with a pencil and a combined sheet metal which has three layers were scanned. The experimental results indicate that the system can extract two Chinese characters "Zhong," "Hua" or three layers of the combined sheet metal. It can be predicted that the microscope can be applied to biology, medicine and other fields in the future due to its favorable 3D imaging capability.

  14. Adaptive optics in digital micromirror based confocal microscopy

    NARCIS (Netherlands)

    Pozzi, P.; Wilding, D.; Soloviev, O.A.; Vdovine, G.V.; Verhaegen, M.H.G.; Bifano, Thomas G.; Kubby, Joel; Gigan, Sylvain

    2016-01-01

    This proceeding reports early results in the development of a new technique for adaptive optics in confocal microscopy. The term adaptive optics refers to the branch of optics in which an active element in the optical system is used to correct inhomogeneities in the media through which light

  15. Axial Confocal Tomography of Capillary-Contained Colloidal Structures.

    Science.gov (United States)

    Liber, Shir R; Indech, Ganit; van der Wee, Ernest B; Butenko, Alexander V; Kodger, Thomas E; Lu, Peter J; Schofield, Andrew B; Weitz, David A; van Blaaderen, Alfons; Sloutskin, Eli

    2017-10-27

    Confocal microscopy is widely used for three-dimensional (3D) sample reconstructions. Arguably, the most significant challenge in such reconstructions is posed by the resolution along the optical axis being significantly lower than in the lateral directions. In addition, the imaging rate is lower along the optical axis in most confocal architectures, prohibiting reliable 3D reconstruction of dynamic samples. Here, we demonstrate a very simple, cheap, and generic method of multiangle microscopy, allowing high-resolution high-rate confocal slice collection to be carried out with capillary-contained colloidal samples in a wide range of slice orientations. This method, realizable with any common confocal architecture and recently implemented with macroscopic specimens enclosed in rotatable cylindrical capillaries, allows 3D reconstructions of colloidal structures to be verified by direct experiments and provides a solid testing ground for complex reconstruction algorithms. In this paper, we focus on the implementation of this method for dense nonrotatable colloidal samples, contained in complex-shaped capillaries. Additionally, we discuss strategies to minimize potential pitfalls of this method, such as the artificial appearance of chain-like particle structures.

  16. Confocal microscopy patterns in nonmelanoma skin cancer and clinical applications.

    Science.gov (United States)

    González, S; Sánchez, V; González-Rodríguez, A; Parrado, C; Ullrich, M

    2014-06-01

    Reflectance confocal microscopy is currently the most promising noninvasive diagnostic tool for studying cutaneous structures between the stratum corneum and the superficial reticular dermis. This tool gives real-time images parallel to the skin surface; the microscopic resolution is similar to that of conventional histology. Numerous studies have identified the main confocal features of various inflammatory skin diseases and tumors, demonstrating the good correlation of these features with certain dermatoscopic patterns and histologic findings. Confocal patterns and diagnostic algorithms have been shown to have high sensitivity and specificity in melanoma and nonmelanoma skin cancer. Possible present and future applications of this noninvasive technology are wide ranging and reach beyond its use in noninvasive diagnosis. This tool can also be used, for example, to evaluate dynamic skin processes that occur after UV exposure or to assess tumor response to noninvasive treatments such as photodynamic therapy. We explain the characteristic confocal features found in the main nonmelanoma skin tumors and discuss possible applications for this novel diagnostic technique in routine dermatology practice. Copyright © 2012 Elsevier España, S.L. and AEDV. All rights reserved.

  17. Magnetically Triggered Release From Giant Unilamellar Vesicles: Visualization By Means Of Confocal Microscopy

    KAUST Repository

    Nappini, Silvia

    2011-04-07

    Magnetically triggered release from magnetic giant unilamellar vesicles (GUVs) loaded with Alexa fluorescent dye was studied by means of confocal laser scanning microscopy (CLSM) under a low-frequency alternating magnetic field (LF-AMF). Core/shell cobalt ferrite nanoparticles coated with rhodamine B isothiocyanate (MP@SiO 2(RITC)) were prepared and adsorbed on the GUV membrane. The MP@SiO 2(RITC) location and distribution on giant lipid vesicles were determined by 3D-CLSM projections, and their effect on the release properties and GUV permeability under a LF-AMF was investigated by CLSM time-resolved experiments. We show that the mechanism of release of the fluorescent dye during the LF-AMF exposure is induced by magnetic nanoparticle energy and mechanical vibration, which promote the perturbation of the GUV membrane without its collapse. © 2011 American Chemical Society.

  18. Local order in a supercooled colloidal fluid observed by confocal microscopy

    CERN Document Server

    Gasser, U; Weitz, D A

    2003-01-01

    The local order in a supercooled monodisperse colloidal fluid is studied by direct imaging of the particles with a laser scanning confocal microscope. The local structure is analysed with a bond order parameter method, which allows one to discern simple structures that are relevant in this system. As expected for samples that crystallize eventually, a large fraction of the particles are found to sit in surroundings with dominant face-centred cubic or hexagonally close-packed character. Evidence for local structures that contain fragments of icosahedra is found, and, moreover, the icosahedral character increases with volume fraction phi, which indicates that it might play an important role at volume fractions near the glass transition.

  19. Electric field and energy of a point electric charge between confocal hyperbolaidal electrodes

    Energy Technology Data Exchange (ETDEWEB)

    Ley-Koo, E. [Universidad Nacional Autonoma de Mexico, Mexico, D. F. (Mexico)

    2001-06-01

    The electric potential and intensity field, as well as the energy of a point electric charge between confocal hyperboloidal electrodes is evaluated as a superposition of prolate spheroidal harmonics using the Green-function technique. This study is motivated by the need to model the electric field between the tip and the sample in a scanning tunnelling microscope, and it can also be applied to a conductor-insulator-conductor junction. [Spanish] Los campos de potencial y de intensidad electrica, asi como la energia de una carga electrica puntual entre electrodos hiperboloidales confocales se evaluan como superposiciones de armonicos esferoidales prolatos usando la tecnica de la funcion de Green. Este estudio ha sido motivado por la necesidad de modelar el campo electrico entre la punta y la muestra de un microscopio de tunelamiento y barrido, y se puede aplicar tambien a una union de conductor-aislante-conductor.

  20. Fiber optic confocal microscope: In vivo precancer detection

    Science.gov (United States)

    Carlson, Kristen Dawn

    Cancer is a significant public health problem worldwide. Many cancers originate as precancerous lesions in the epithelium which, when removed in sufficient time, can prevent progression to cancer. However, current detection techniques are typically time-consuming and expensive, limiting their acceptance and accessibility. Optical techniques, such as confocal microscopy, have significant potential to provide clinicians with real-time, high-resolution images of cells and tissue without tissue removal. These images of cell morphology and tissue architecture can be used to characterize tissue and determine the presence or extent of precancer and cancer. This dissertation explores the instrumentation and application of fiber optic reflectance confocal microscopy for in vivo precancer detection. The first part of the dissertation presents in vivo imaging of suspicious lesions in the human uterine cervix and oral mucosa using a fiber bundle based confocal microscope with a complex glass miniature objective lens. Images are analyzed quantitatively and qualitatively to determine the potential of this technology in vivo. An analysis of nuclear density from images of 30 cervical epithelium sites shows differentiation between normal and precancerous sites. Similarly, images from 20 oral mucosa sites demonstrate changes in nuclear density and tissue architecture indicative of progression of precancer and cancer. In addition to this multi-fiber confocal microscope used with a glass objective lens for the clinical studies, imaging of tissue samples has been accomplished with the same confocal system using an injection molded plastic miniature objective lens demonstrating comparable optical quality for a significantly less expensive optical component. Finally, a benchtop prototype of a single fiber confocal microscope using a gimbaled two-axis MEMS scanner has been designed and constructed. Imaging of a resolution target and cellular samples demonstrates sufficient resolution and

  1. Confocal reflectance quantitative phase microscope system for cellular membranes dynamics study (Conference Presentation)

    Science.gov (United States)

    Singh, Vijay Raj; Yaqoob, Zahid; So, Peter T. C.

    2017-02-01

    Quantitative phase microscopy (QPM) techniques developed so far primarily belongs to high speed transmitted light based systems that has enough sensitivity to resolve membrane fluctuations and dynamics, but has no depth resolution. Therefore, most biomechanics studies using QPM today is confined to simple cells, such as RBCs, without internal organelles. An important instrument that will greatly extend the biomedical applications of QPM is to develop next generation microscope with 3D capability and sufficient temporal resolution to study biomechanics of complex eukaryotic cells including the mechanics of their internal compartments. For eukaryotic cells, the depth sectioning capability is critical and should be sufficient to distinguish nucleic membrane fluctuations from plasma membrane fluctuations. Further, this microscope must provide high temporal resolution since typical eukaryotes membranes are substantially stiffer than RBCs. A confocal reflectance quantitative phase microscope is presented based on multi-pinhole scanning, with the capabilities of higher temporal resolution and sensitivity for nucleic and plasma membranes of eukaryotic cells. System hardware is developed based on an array of confocal pinhole generated by using the `ON' state of subset of micro-mirrors of digital micro-mirror device (DMD, from Texas Instruments) and high-speed raster scanning provides 14ms imaging speed in wide-field mode. A common path interferometer is integrated at the imaging arm for detection of specimens' quantitative phase information. Theoretical investigation of quantitative phase reconstructed from system is investigated and application of system is presented for dimensional fluctuations measurements of both cellular plasma and nucleic membranes of embryonic stem cells.

  2. New Applications of Scanning Tunneling Microscopy

    Science.gov (United States)

    Smith, Douglas Philip Edward

    This dissertation describes the application of the scanning tunneling microscope (STM) technique to four new fields of study: thin organic films, phonon spectroscopy of bulk surfaces, the vibrational spectroscopy of molecules, and the tribological forces which occur between STM tip and sample. Images with atomic resolution were obtained with speeds approaching video rates. Two types of microscopes were used: one operated at room temperature in air, another at 4.2K in liquid helium. At room temperature, the STM was able to image molecules of cadmium arachidate deposited onto graphite by the Langmuir-Blodgett technique. The packing of molecules in the lipid bilayer was found to be partially ordered, with density of 1 molecule per 19.4 square angstroms. At liquid-helium temperature, inelastic electron processes were detected, and it was possible to determine within an area of a few square angstroms where the vibrational excitations occurred. On a bare graphite substrate, phonons of the sample and tip caused step increases in the tunneling conductivity at the phonon energies. Molecules of sorbic acid could be resolved when deposited onto graphite, and these molecules caused spatially localized peaks in conductivity at the energies of the bond vibrations. Although the STM is usually considered a non-contact instrument, under certain circumstances the tip and sample exerted strong forces on each other. With a tungsten tip and a graphite sample, friction and mechanical deformations on the atomic scale were observed.

  3. Solar Confocal interferometers for Sub-Picometer-Resolution Spectral Filters

    Science.gov (United States)

    Gary, G. Allen; Pietraszewski, Chris; West, Edward A.; Dines. Terence C.

    2007-01-01

    The confocal Fabry-Perot interferometer allows sub-picometer spectral resolution of Fraunhofer line profiles. Such high spectral resolution is needed to keep pace with the higher spatial resolution of the new set of large-aperture solar telescopes. The line-of-sight spatial resolution derived for line profile inversions would then track the improvements of the transverse spatial scale provided by the larger apertures. In particular, profile inversion allows improved velocity and magnetic field gradients to be determined independent of multiple line analysis using different energy levels and ions. The confocal interferometer's unique properties allow a simultaneous increase in both etendue and spectral power. The higher throughput for the interferometer provides significant decrease in the aperture, which is important in spaceflight considerations. We have constructed and tested two confocal interferometers. A slow-response thermal-controlled interferometer provides a stable system for laboratory investigation, while a piezoelectric interferometer provides a rapid response for solar observations. In this paper we provide design parameters, show construction details, and report on the laboratory test for these interferometers. The field of view versus aperture for confocal interferometers is compared with other types of spectral imaging filters. We propose a multiple etalon system for observing with these units using existing planar interferometers as pre-filters. The radiometry for these tests established that high spectral resolution profiles can be obtained with imaging confocal interferometers. These sub-picometer spectral data of the photosphere in both the visible and near-infrared can provide important height variation information. However, at the diffraction-limited spatial resolution of the telescope, the spectral data is photon starved due to the decreased spectral passband.

  4. Targeted vertical cross-sectional imaging with handheld near-infrared dual axes confocal fluorescence endomicroscope.

    Science.gov (United States)

    Qiu, Zhen; Liu, Zhongyao; Duan, Xiyu; Khondee, Supang; Joshi, Bishnu; Mandella, Michael J; Oldham, Kenn; Kurabayashi, Katsuo; Wang, Thomas D

    2013-02-01

    We demonstrate vertical cross-sectional (XZ-plane) images of near-infrared (NIR) fluorescence with a handheld dual axes confocal endomicroscope that reveals specific binding of a Cy5.5-labeled peptide to pre-malignant colonic mucosa. This view is perpendicular to the tissue surface, and is similar to that used by pathologists. The scan head is 10 mm in outer diameter (OD), and integrates a one dimensional (1-D) microelectromechanical systems (MEMS) X-axis scanner and a bulky lead zirconate titanate (PZT) based Z-axis actuator. The microscope images in a raster-scanning pattern with a ±6 degrees (mechanical) scan angle at ~3 kHz in the X-axis (fast) and up to 10 Hz (0-400 μm) in the Z-axis (slow). Vertical cross-sectional fluorescence images are collected with a transverse and axial resolution of 4 and 5 μm, respectively, over a field-of-view of 800 μm (width) × 400 μm (depth). NIR vertical cross-sectional fluorescence images of fresh mouse colonic mucosa demonstrate histology-like imaging performance with this miniature instrument.

  5. GPU accelerated real-time confocal fluorescence lifetime imaging microscopy (FLIM) based on the analog mean-delay (AMD) method.

    Science.gov (United States)

    Kim, Byungyeon; Park, Byungjun; Lee, Seungrag; Won, Youngjae

    2016-12-01

    We demonstrated GPU accelerated real-time confocal fluorescence lifetime imaging microscopy (FLIM) based on the analog mean-delay (AMD) method. Our algorithm was verified for various fluorescence lifetimes and photon numbers. The GPU processing time was faster than the physical scanning time for images up to 800 × 800, and more than 149 times faster than a single core CPU. The frame rate of our system was demonstrated to be 13 fps for a 200 × 200 pixel image when observing maize vascular tissue. This system can be utilized for observing dynamic biological reactions, medical diagnosis, and real-time industrial inspection.

  6. Miniature near-infrared dual-axes confocal microscope utilizing a two-dimensional microelectromechanical systems scanner.

    Science.gov (United States)

    Liu, Jonathan T C; Mandella, Michael J; Ra, Hyejun; Wong, Larry K; Solgaard, Olav; Kino, Gordon S; Piyawattanametha, Wibool; Contag, Christopher H; Wang, Thomas D

    2007-02-01

    The first, to our knowledge, miniature dual-axes confocal microscope has been developed, with an outer diameter of 10 mm, for subsurface imaging of biological tissues with 5-7 microm resolution. Depth-resolved en face images are obtained at 30 frames per second, with a field of view of 800 x 100 microm, by employing a two-dimensional scanning microelectromechanical systems mirror. Reflectance and fluorescence images are obtained with a laser source at 785 nm, demonstrating the ability to perform real-time optical biopsy.

  7. Design methodology for a confocal imaging system using an objective microlens array with an increased working distance.

    Science.gov (United States)

    Choi, Woojae; Shin, Ryung; Lim, Jiseok; Kang, Shinill

    2016-09-12

    In this study, a design methodology for a multi-optical probe confocal imaging system was developed. To develop an imaging system that has the required resolving power and imaging area, this study focused on a design methodology to create a scalable and easy-to-implement confocal imaging system. This system overcomes the limitations of the optical complexities of conventional multi-optical probe confocal imaging systems and the short working distance using a micro-objective lens module composed of two microlens arrays and a telecentric relay optical system. The micro-objective lens module was fabricated on a glass substrate using backside alignment photolithography and thermal reflow processes. To test the feasibility of the developed methodology, an optical system with a resolution of 1 μm/pixel using multi-optical probes with an array size of 10 × 10 was designed and constructed. The developed system provides a 1 mm × 1 mm field of view and a sample scanning range of 100 μm. The optical resolution was evaluated by conducting sample tests using a knife-edge detecting method. The measured lateral resolution of the system was 0.98 μm.

  8. The three-dimensional elemental distribution based on the surface topography by confocal 3D-XRF analysis

    Energy Technology Data Exchange (ETDEWEB)

    Yi, Longtao; Qin, Min; Wang, Kai; Peng, Shiqi; Sun, Tianxi; Liu, Zhiguo [Beijing Normal University, College of Nuclear Science and Technology, Beijing (China); Lin, Xue [Northwest University, School of Cultural Heritage, Xi' an (China)

    2016-09-15

    Confocal three-dimensional micro-X-ray fluorescence (3D-XRF) is a good surface analysis technology widely used to analyse elements and elemental distributions. However, it has rarely been applied to analyse surface topography and 3D elemental mapping in surface morphology. In this study, a surface adaptive algorithm using the progressive approximation method was designed to obtain surface topography. A series of 3D elemental mapping analyses in surface morphology were performed in laboratories to analyse painted pottery fragments from the Majiayao Culture (3300-2900 BC). To the best of our knowledge, for the first time, sample surface topography and 3D elemental mapping were simultaneously obtained. Besides, component and depth analyses were also performed using synchrotron radiation confocal 3D-XRF and tabletop confocal 3D-XRF, respectively. The depth profiles showed that the sample has a layered structure. The 3D elemental mapping showed that the red pigment, black pigment, and pottery coat contain a large amount of Fe, Mn, and Ca, respectively. From the 3D elemental mapping analyses at different depths, a 3D rendering was obtained, clearly showing the 3D distributions of the red pigment, black pigment, and pottery coat. Compared with conventional 3D scanning, this method is time-efficient for analysing 3D elemental distributions and hence especially suitable for samples with non-flat surfaces. (orig.)

  9. Brain PET scan

    Science.gov (United States)

    ... have false results on a PET scan. Blood sugar or insulin levels may affect the test results in people with diabetes . PET scans may be done along with a CT scan. This combination scan is called a PET/CT. Alternative Names Brain positron emission tomography; PET scan - brain References Chernecky ...

  10. Coronary Calcium Scan

    Science.gov (United States)

    ... Back To Health Topics / Coronary Calcium Scan Coronary Calcium Scan Also known as Calcium Scan Test A coronary calcium scan is a CT scan of your heart that detects and measures the amount of calcium in the walls of your coronary arteries. Overview ...

  11. Spatial Scan Statistic: Selecting clusters and generating elliptic clusters

    OpenAIRE

    Christiansen, Lasse Engbo; Andersen, Jens Strodl

    2004-01-01

    The spatial scan statistic is widely used to search for clusters. This paper shows that the usually applied elimination of overlapping clusters to find secondary clusters is sensitive to smooth changes in the shape of the clusters. We present an algorithm for generation of set of confocal elliptic clusters. In addition, we propose a new way to present the information in a given set of clusters based on the significance of the clusters.

  12. Spatial Scan Statistic: Selecting clusters and generating elliptic clusters

    DEFF Research Database (Denmark)

    Christiansen, Lasse Engbo; Andersen, Jens Strodl

    2004-01-01

    The spatial scan statistic is widely used to search for clusters. This paper shows that the usually applied elimination of overlapping clusters to find secondary clusters is sensitive to smooth changes in the shape of the clusters. We present an algorithm for generation of set of confocal elliptic...... clusters. In addition, we propose a new way to present the information in a given set of clusters based on the significance of the clusters....

  13. Confocal microscopy on the beamline: novel three-dimensional imaging and sample positioning

    OpenAIRE

    Khan, I.; Gillilan, R; Kriksunov, I.; Williams, R.; Zipfel, W.R.; Englich, U.

    2012-01-01

    Possibilities of applying confocal microscopy on an X-ray beamline have been explored. Confocal microscopy images have the potential to give detailed, on-axis and three-dimensional views of protein crystals on a synchrotron beamline.

  14. Three-Dimensional Visualization of Interfacial Phenomena Using Confocal Microscopy

    Science.gov (United States)

    Shieh, Ian C.

    Surfactants play an integral role in numerous functions ranging from stabilizing the emulsion in a favorite salad dressing to organizing the cellular components that make life possible. We are interested in lung surfactant, which is a mixture of lipids and proteins essential for normal respiration because it modulates the surface tension of the air-liquid interface of the thin fluid lining in the lungs. Through this surface tension modulation, lung surfactant ensures effortless lung expansion and prevents lung collapse during exhalation, thereby effecting proper oxygenation of the bloodstream. The function of lung surfactant, as well as numerous interfacial lipid systems, is not solely dictated by the behavior of materials confined to the two-dimensional interface. Rather, the distributions of materials in the liquid subphase also greatly influence the performance of interfacial films of lung surfactant. Therefore, to better understand the behavior of lung surfactant and other interfacial lipid systems, we require a three-dimensional characterization technique. In this dissertation, we have developed a novel confocal microscopy methodology for investigating the interfacial phenomena of surfactants at the air-liquid interface of a Langmuir trough. Confocal microscopy provides the excellent combination of in situ, fast, three-dimensional visualization of multiple components of the lung surfactant system that other characterization techniques lack. We detail the solutions to the numerous challenges encountered when imaging a dynamic air-liquid interface with a high-resolution technique like confocal microscopy. We then use confocal microscopy to elucidate the distinct mechanisms by which a polyelectrolyte (chitosan) and nonadsorbing polymer (polyethylene glycol) restore the function of lung surfactant under inhibitory conditions mimicking the effects of lung trauma. Beyond this physiological model, we also investigate several one- and two-component interfacial films

  15. Thyroid Scan and Uptake

    Medline Plus

    Full Text Available ... of the Thyroid Scan and Uptake? What is a Thyroid Scan and Uptake? A thyroid scan is ... code: Phone no: Thank you! Do you have a personal story about radiology? Share your patient story ...

  16. Heart PET scan

    Science.gov (United States)

    ... nuclear medicine scan; Heart positron emission tomography; Myocardial PET scan ... A PET scan requires a small amount of radioactive material (tracer). This tracer is given through a vein (IV), ...

  17. Thyroid Scan and Uptake

    Medline Plus

    Full Text Available ... of the Thyroid Scan and Uptake? What is a Thyroid Scan and Uptake? A thyroid scan is ... taking our brief survey: Survey Do you have a personal story about radiology? Share your patient story ...

  18. Clinical feasibility of rapid confocal melanoma feature detection

    Science.gov (United States)

    Hennessy, Ricky; Jacques, Steve; Pellacani, Giovanni; Gareau, Daniel

    2010-02-01

    In vivo reflectance confocal microscopy shows promise for the early detection of malignant melanoma. One diagnostic trait of malignancy is the presence of pagetoid melanocytes in the epidermis. For automated detection of MM, this feature must be identified quantitatively through software. Beginning with in vivo, noninvasive confocal images from 10 unequivocal MMs and benign nevi, we developed a pattern recognition algorithm that automatically identified pagetoid melanocytes in all four MMs and identified none in five benign nevi. One data set was discarded due to artifacts caused by patient movement. With future work to bring the performance of this pattern recognition technique to the level of the clinicians on difficult lesions, melanoma diagnosis could be brought to primary care facilities and save many lives by improving early diagnosis.

  19. Ultrafast superresolution fluorescence imaging with spinning disk confocal microscope optics.

    Science.gov (United States)

    Hayashi, Shinichi; Okada, Yasushi

    2015-05-01

    Most current superresolution (SR) microscope techniques surpass the diffraction limit at the expense of temporal resolution, compromising their applications to live-cell imaging. Here we describe a new SR fluorescence microscope based on confocal microscope optics, which we name the spinning disk superresolution microscope (SDSRM). Theoretically, the SDSRM is equivalent to a structured illumination microscope (SIM) and achieves a spatial resolution of 120 nm, double that of the diffraction limit of wide-field fluorescence microscopy. However, the SDSRM is 10 times faster than a conventional SIM because SR signals are recovered by optical demodulation through the stripe pattern of the disk. Therefore a single SR image requires only a single averaged image through the rotating disk. On the basis of this theory, we modified a commercial spinning disk confocal microscope. The improved resolution around 120 nm was confirmed with biological samples. The rapid dynamics of micro-tubules, mitochondria, lysosomes, and endosomes were observed with temporal resolutions of 30-100 frames/s. Because our method requires only small optical modifications, it will enable an easy upgrade from an existing spinning disk confocal to a SR microscope for live-cell imaging. © 2015 Hayashi and Okada. This article is distributed by The American Society for Cell Biology under license from the author(s). Two months after publication it is available to the public under an Attribution–Noncommercial–Share Alike 3.0 Unported Creative Commons License (http://creativecommons.org/licenses/by-nc-sa/3.0).

  20. Laser confocal cylindrical radius measurement method and its system.

    Science.gov (United States)

    Xiao, Yang; Qiu, Lirong; Zhao, Weiqian

    2017-08-10

    This paper proposes a laser confocal cylindrical radius of the curvature measurement (CCRM) method. The CCRM method precisely identifies the positions of the vertex and curvature center of the test cylindrical surface by using the property so that the maximum point of the laser confocal axial intensity curve precisely corresponds to the focus of the laser confocal measurement system, and the accurate distance of these two positions is obtained by the distance measuring instrument, thus achieving the precise measurement of the cylindrical radius. The quadratic fitting method is used to further improve the measurement accuracy. Compared with existing measurement methods, the CCRM method has high measurement precision, simple structure, and strong environmental interference capability, and it is more suitable for engineering applications. Based on the CCRM, the CCRM system is established, and theoretical analysis and preliminary experiments indicate that the relative uncertainty of cylindrical radius measurement is better than 0.045%. Therefore, the CCRM provides an effective approach for the high-precision measurement of cylindrical radius.

  1. Confocal fluorometer for diffusion tracking in 3D engineered tissue constructs

    Science.gov (United States)

    Daly, D.; Zilioli, A.; Tan, N.; Buttenschoen, K.; Chikkanna, B.; Reynolds, J.; Marsden, B.; Hughes, C.

    2016-03-01

    We present results of the development of a non-contacting instrument, called fScan, based on scanning confocal fluorometry for assessing the diffusion of materials through a tissue matrix. There are many areas in healthcare diagnostics and screening where it is now widely accepted that the need for new quantitative monitoring technologies is a major pinch point in patient diagnostics and in vitro testing. With the increasing need to interpret 3D responses this commonly involves the need to track the diffusion of compounds, pharma-active species and cells through a 3D matrix of tissue. Methods are available but to support the advances that are currently only promised, this monitoring needs to be real-time, non-invasive, and economical. At the moment commercial meters tend to be invasive and usually require a sample of the medium to be removed and processed prior to testing. This methodology clearly has a number of significant disadvantages. fScan combines a fiber based optical arrangement with a compact, free space optical front end that has been integrated so that the sample's diffusion can be measured without interference. This architecture is particularly important due to the "wet" nature of the samples. fScan is designed to measure constructs located within standard well plates and a 2-D motion stage locates the required sample with respect to the measurement system. Results are presented that show how the meter has been used to evaluate movements of samples through collagen constructs in situ without disturbing their kinetic characteristics. These kinetics were little understood prior to these measurements.

  2. Scanning electron microscopy of bone.

    Science.gov (United States)

    Boyde, Alan

    2012-01-01

    This chapter described methods for Scanning Electron Microscopical imaging of bone and bone cells. Backscattered electron (BSE) imaging is by far the most useful in the bone field, followed by secondary electrons (SE) and the energy dispersive X-ray (EDX) analytical modes. This chapter considers preparing and imaging samples of unembedded bone having 3D detail in a 3D surface, topography-free, polished or micromilled, resin-embedded block surfaces, and resin casts of space in bone matrix. The chapter considers methods for fixation, drying, looking at undersides of bone cells, and coating. Maceration with alkaline bacterial pronase, hypochlorite, hydrogen peroxide, and sodium or potassium hydroxide to remove cells and unmineralised matrix is described in detail. Attention is given especially to methods for 3D BSE SEM imaging of bone samples and recommendations for the types of resin embedding of bone for BSE imaging are given. Correlated confocal and SEM imaging of PMMA-embedded bone requires the use of glycerol to coverslip. Cathodoluminescence (CL) mode SEM imaging is an alternative for visualising fluorescent mineralising front labels such as calcein and tetracyclines. Making spatial casts from PMMA or other resin embedded samples is an important use of this material. Correlation with other imaging means, including microradiography and microtomography is important. Shipping wet bone samples between labs is best done in glycerol. Environmental SEM (ESEM, controlled vacuum mode) is valuable in eliminating -"charging" problems which are common with complex, cancellous bone samples.

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

    Science.gov (United States)

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

    2013-01-01

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

  4. Combined AFM and confocal fluorescence microscope for applications in bio-nanotechnology.

    Science.gov (United States)

    Kassies, R; van der Werf, K O; Lenferink, A; Hunter, C N; Olsen, J D; Subramaniam, V; Otto, C

    2005-01-01

    We present a custom-designed atomic force fluorescence microscope (AFFM), which can perform simultaneous optical and topographic measurements with single molecule sensitivity throughout the whole visible to near-infrared spectral region. Integration of atomic force microscopy (AFM) and confocal fluorescence microscopy combines the high-resolution topographical imaging of AFM with the reliable (bio)-chemical identification capability of optical methods. The AFFM is equipped with a spectrograph enabling combined topographic and fluorescence spectral imaging, which significantly enhances discrimination of spectroscopically distinct objects. The modular design allows easy switching between different modes of operation such as tip-scanning, sample-scanning or mechanical manipulation, all of which are combined with synchronous optical detection. We demonstrate that coupling the AFM with the fluorescence microscope does not compromise its ability to image with a high spatial resolution. Examples of several modes of operation of the AFFM are shown using two-dimensional crystals and membranes containing light-harvesting complexes from the photosynthetic bacterium Rhodobacter sphaeroides.

  5. Dimensional metrology of lab-on-a-chip internal structures: a comparison of optical coherence tomography with confocal fluorescence microscopy.

    Science.gov (United States)

    Reyes, D R; Halter, M; Hwang, J

    2015-07-01

    The characterization of internal structures in a polymeric microfluidic device, especially of a final product, will require a different set of optical metrology tools than those traditionally used for microelectronic devices. We demonstrate that optical coherence tomography (OCT) imaging is a promising technique to characterize the internal structures of poly(methyl methacrylate) devices where the subsurface structures often cannot be imaged by conventional wide field optical microscopy. The structural details of channels in the devices were imaged with OCT and analyzed with an in-house written ImageJ macro in an effort to identify the structural details of the channel. The dimensional values obtained with OCT were compared with laser-scanning confocal microscopy images of channels filled with a fluorophore solution. Attempts were also made using confocal reflectance and interferometry microscopy to measure the channel dimensions, but artefacts present in the images precluded quantitative analysis. OCT provided the most accurate estimates for the channel height based on an analysis of optical micrographs obtained after destructively slicing the channel with a microtome. OCT may be a promising technique for the future of three-dimensional metrology of critical internal structures in lab-on-a-chip devices because scans can be performed rapidly and noninvasively prior to their use. © 2015 The Authors Journal of Microscopy © 2015 Royal Microscopical Society.

  6. Ophthalmic applications of confocal microscopy: diagnostics, refractive surgery, and eye banking

    Science.gov (United States)

    Masters, Barry R.

    1990-11-01

    Confocal microscopy of ocular tissue provides two advantages over traditional imaging techniques: increased range and transverse resolution and increased contrast. The semitransparent cornea and ocular lens in the living eye can be optically sectioned and observed by reflected light confocal microscopy. Within the cornea we observed various cell components nerve fibers nerve cell bodies and fibrous networks. The confocal microscopic images from the in-situ ocular lens show the lens capsule the lens epithelium and the individual lens fibrils. All of the reflected light confocal microscopic images have high contrast and high resolution. Some of the applications of confocal imaging in ophthalmology include: diagnostics of the cornea and the ocular lens examination prior to and after refractive surgery examination of intraocular lenses (IOL) and examination of eye bank material. Other ophthalmic uses of confocal imaging include: studies of wound healing therapeutics and the effects of contact lenses on the cornea. The proposed features of a clinical confocal microscope are reviewed. 2.

  7. Multimodal optical workstation for simultaneous linear, nonlinear microscopy and nanomanipulation: upgrading a commercial confocal inverted microscope.

    Science.gov (United States)

    Mathew, Manoj; Santos, Susana I C O; Zalvidea, Dobryna; Loza-Alvarez, Pablo

    2009-07-01

    In this work we propose and build a multimodal optical workstation that extends a commercially available confocal microscope (Nikon Confocal C1-Si) to include nonlinear/multiphoton microscopy and optical manipulation/stimulation tools such as nanosurgery. The setup allows both subsystems (confocal and nonlinear) to work independently and simultaneously. The workstation enables, for instance, nanosurgery along with simultaneous confocal and brightfield imaging. The nonlinear microscopy capabilities are added around the commercial confocal microscope by exploiting all the flexibility offered by this microscope and without need for any mechanical or electronic modification of the confocal microscope systems. As an example, the standard differential interference contrast condenser and diascopic detector in the confocal microscope are readily used as a forward detection mount for second harmonic generation imaging. The various capabilities of this workstation, as applied directly to biology, are demonstrated using the model organism Caenorhabditis elegans.

  8. In vivo confocal microscopy: corneal changes of hydrogel contact lens wearers.

    Science.gov (United States)

    Yagmur, Meltem; Okay, Okan; Sizmaz, Selcuk; Unal, Ilker; Yar, Kemal

    2011-10-01

    To evaluate the corneal findings in hydrogel contact lens wearers by in vivo confocal scanning microscopy. One hundred and forty-two eyes of 71 myopic contact lens wearers (group 1) and 142 eyes of 71 non-contact lens wearers (group 2), whose age, gender and refractive error matched, were enrolled in order to detect the corneal changes by in vivo confocal microscopy through the central cornea. The average age was 25.5 ± 5.7 (16-52) and 25.6 ± 5.6 (17-49) in groups 1 and 2, respectively. The mean duration of contact lens wear was 43.9 ± 15.3 (6-240) months. Anterior keratocyte density was 667.5 ± 128.3 cells/mm(2) in group 1 and 821.4 ± 136.7 cells/mm(2) in group 2 (P = 0.001). Posterior keratocyte densities of groups 1 and 2 were 540.2 ± 87.6 cells/mm(2) and 628.2 ± 72.4 cells/mm(2), respectively (P lenses with a mean Dk/t ratio of 26.5 × 10(-9) ± 5.9 (8.9-32 × 10(-9)). Stromal microdots occurred with contact lenses with a mean Dk/t ratio of 13.2 × 10(-9) ± 17.5 × 10(-9) (8.9-20 × 10(-9)). In vivo examination of the cornea with confocal microscopy revealed a number of changes. These changes can be attributed both to the mechanical and the hypoxic effects of soft contact lenses. In soft contact lenses with a high Dk/t ratio, these changes would be less frequent.

  9. [The ocular surface of severe alkali burns patients on confocal microscopy].

    Science.gov (United States)

    Zhu, Wen-qing; Xu, Jian-jiang; Sun, Xing-huai; Qiu, Ting; Hong, Jia-xu; Wang, Yan; Wang, Wen-tao

    2010-01-01

    To analyze the morphology on the ocular surface of severe alkali burns patients by in vivo laser scanning confocal microscopy. This research was a retrospective observation case series. From February to November 2008 in Eye Ear Nose and Throat Hospital of Fudan University, 39 alkali burns patients who classified as III or IV according to Roper-Hall classification were enrolled in this study. They were divided into four groups according to the course of disease: A (less than 3 months), B (3 - 6 months), C (6 - 12 months) and D (over 12 months). In vivo laser scanning confocal microscopic examinations were performed on the injured cornea, the limbus and the bulbar conjunctiva and the images were recorded. The morphology of the injured cornea, the limbus and the bulbar conjunctiva was analyzed and the densities of the inflammatory cells and dendritic cells in the limbus were calculated. One-way analysis of variance was used to compare the means of the inflammatory cells and dendritic cells. Subsequently the data between two groups were analyzed by least significant difference. The corneal epitheliums of the patients in Group A manifested large irregular features with hyperreflective cytoplasm and hyporeflective nuclei, sometimes losing cell features. There were numerous small hyperreflective inflammatory cells in groups beneath the superficial epitheliums. Shallow corneal stroma was edema, and it was hard to discriminate the morphology of the stromal cells. Deep stromal cells were in the activated state. The images of the endothelial layer were unclear. In Group B and Group C, there were the same manifestation of the superficial epitheliums as the group A and it disappeared in Group D. The inflammatory cells beneath the superficial epitheliums reduced and some residual basal epitheliums and hyperreflective conjunctiva-like epitheliums were visible in Group B and Group C. In Group D, there were small oval tight-arranged cells with punctiform hyperreflective nuclei

  10. Confocal Imaging of Biological Tissues Using Second Harmonic Generation

    Energy Technology Data Exchange (ETDEWEB)

    Kim, B-M.; Stoller, P.; Reiser, K.; Eichler, J.; Yan, M.; Rubenchik, A.; Da Silva, L.

    2000-03-06

    A confocal microscopy imaging system was devised to selectively detect Second harmonic signals generated by biological tissues. Several types of biological tissues were examined using this imaging system, including human teeth, bovine blood vessels, and chicken skin. All these tissues generated strong second harmonic signals. There is considerable evidence that the source of these signals in tissue is collagen. Collagen, the predominant component of most tissues, is known to have second order nonlinear susceptibility. This technique may have diagnostic usefulness in pathophysiological conditions characterized by changes in collagen structure including malignant transformation of nevi, progression of diabetic complications, and abnormalities in wound healing.

  11. 3D confocal imaging in CUBIC-cleared mouse heart

    Energy Technology Data Exchange (ETDEWEB)

    Nehrhoff, I.; Bocancea, D.; Vaquero, J.; Vaquero, J.J.; Lorrio, M.T.; Ripoll, J.; Desco, M.; Gomez-Gaviro, M.V.

    2016-07-01

    Acquiring high resolution 3D images of the heart enables the ability to study heart diseases more in detail. Here, the CUBIC (clear, unobstructed brain imaging cocktails and computational analysis) clearing protocol was adapted for thick mouse heart sections to increase the penetration depth of the confocal microscope lasers into the tissue. The adapted CUBIC clearing of the heart lets the antibody penetrate deeper into the tissue by a factor of five. The here shown protocol enables deep 3D highresolution image acquisition in the heart. This allows a much more accurate assessment of the cellular and structural changes that underlie heart diseases. (Author)

  12. Atherosclerotic plaque detection by confocal Brillouin and Raman microscopies

    Science.gov (United States)

    Meng, Zhaokai; Basagaoglu, Berkay; Yakovlev, Vladislav V.

    2015-02-01

    Atherosclerosis, the development of intraluminal plaque, is a fundamental pathology of cardiovascular system and remains the leading cause of morbidity and mortality worldwide. Biomechanical in nature, plaque rupture occurs when the mechanical properties of the plaque, related to the morphology and viscoelastic properties, are compromised, resulting in intraluminal thrombosis and reduction of coronary blood flow. In this report, we describe the first simultaneous application of confocal Brillouin and Raman microscopies to ex-vivo aortic wall samples. Such a non-invasive, high specific approach allows revealing a direct relationship between the biochemical and mechanical properties of atherosclerotic tissue.

  13. Water-Immersible MEMS scanning mirror designed for wide-field fast-scanning photoacoustic microscopy

    Science.gov (United States)

    Yao, Junjie; Huang, Chih-Hsien; Martel, Catherine; Maslov, Konstantin I.; Wang, Lidai; Yang, Joon-Mo; Gao, Liang; Randolph, Gwendalyn; Zou, Jun; Wang, Lihong V.

    2013-03-01

    By offering images with high spatial resolution and unique optical absorption contrast, optical-resolution photoacoustic microscopy (OR-PAM) has gained increasing attention in biomedical research. Recent developments in OR-PAM have improved its imaging speed, but have sacrificed either the detection sensitivity or field of view or both. We have developed a wide-field fast-scanning OR-PAM by using a water-immersible MEMS scanning mirror (MEMS-ORPAM). Made of silicon with a gold coating, the MEMS mirror plate can reflect both optical and acoustic beams. Because it uses an electromagnetic driving force, the whole MEMS scanning system can be submerged in water. In MEMS-ORPAM, the optical and acoustic beams are confocally configured and simultaneously steered, which ensures uniform detection sensitivity. A B-scan imaging speed as high as 400 Hz can be achieved over a 3 mm scanning range. A diffraction-limited lateral resolution of 2.4 μm in water and a maximum imaging depth of 1.1 mm in soft tissue have been experimentally determined. Using the system, we imaged the flow dynamics of both red blood cells and carbon particles in a mouse ear in vivo. By using Evans blue dye as the contrast agent, we also imaged the flow dynamics of lymphatic vessels in a mouse tail in vivo. The results show that MEMS-OR-PAM could be a powerful tool for studying highly dynamic and time-sensitive biological phenomena.

  14. Thyroid Scan and Uptake

    Medline Plus

    Full Text Available ... concern for you. If you had an intravenous line inserted for the procedure, it will usually be ... procedure that same day that requires an intravenous line. Actual scanning time for a thyroid scan is ...

  15. Thyroid Scan and Uptake

    Medline Plus

    Full Text Available ... the limitations of the Thyroid Scan and Uptake? What is a Thyroid Scan and Uptake? A thyroid ... body converts food to energy. top of page What are some common uses of the procedure? The ...

  16. RBC nuclear scan

    Science.gov (United States)

    ... page: //medlineplus.gov/ency/article/003835.htm RBC nuclear scan To use the sharing features on this page, please enable JavaScript. An RBC nuclear scan uses small amounts of radioactive material to ...

  17. Microscopia confocal in vivo nos depósitos corneanos por amiodarona In vivo confocal microscopy in amiodarone corneal deposits

    Directory of Open Access Journals (Sweden)

    Gustavo Victor

    2007-02-01

    Full Text Available OBJETIVO: Descrever os achados da microscopia confocal in vivo em pacientes nos diversos estágios de ceratopatia induzida por amiodarona, e correlacionar o estadiamento biomicroscópico com o estadiamento confocal. MÉTODOS: Vinte olhos de 10 pacientes (6 homens e 4 mulheres em tratamento com amiodarona, que apresentavam ceratopatia induzida pela droga, foram selecionados para o estudo, com a microscopia confocal (MC. RESULTADOS: A média de idade foi 58 ± 6,2 anos (50-66 anos e o tempo de uso da droga foi de 6 ± 3,2 anos (2-11 anos. Todos pacientes tinham acuidade visual com correção melhor ou igual a 20/40. A biomicroscopia evidenciou ceratopatia por amiodarona: dois pacientes no estágio 1, quatro no estágio 2 e quatro no estágio 3. Todas as córneas apresentaram inclusões intracelulares brilhantes e de alta refletividade na camada epitelial basal. A partir dos estágios 2 e 3, foram encontrados microdepósitos em todas camadas corneanas. Foram observados afilamento e aumento da tortuosidade dos nervos corneanos nos estágios 2 e 3 da ceratopatia. A contagem endotelial média foi de 2.524 ± 150,3 células/mm². CONCLUSÃO: O epitélio basal foi o mais acometido nos diferentes estágios da ceratopatia. Nos pacientes do estágio 1 a biomicroscopia, os microdepósitos subepiteliais são restritos ao epitélio superficial e basal, ao passo que nos pacientes dos estágios 2 e 3, os microdepósitos afetam todas camadas corneanas. À medida que a ceratopatia avança, os nervos corneanos ficam mais afilados e tortuosos.PURPOSE: To describe in vivo confocal microscopy findings in patients with different stages of amiodarone-induced keratopathy, and correlate biomicroscopy stages with confocal stages. METHODS: Twenty eyes of 10 patients (6 men and 4 women, who receive treatment with amiodarone were selected for the study with confocal microscopy (MC. RESULTS: The average age was 58 ± 6.2 years (50-66 years and time of use of the drug was 6

  18. Implementation of Accurate and Fast DNA Cytometry by Confocal Microscopy in 3D

    Directory of Open Access Journals (Sweden)

    Lennert S. Ploeger

    2005-01-01

    Full Text Available Background: DNA cytometry is a powerful method for measuring genomic instability. Standard approaches that measure DNA content of isolated cells may induce selection bias and do not allow interpretation of genomic instability in the context of the tissue. Confocal Laser Scanning Microscopy (CLSM provides the opportunity to perform 3D DNA content measurements on intact cells in thick histological sections. Because the technique is technically challenging and time consuming, only a small number of usually manually selected nuclei were analyzed in different studies, not allowing wide clinical evaluation. The aim of this study was to describe the conditions for accurate and fast 3D CLSM cytometry with a minimum of user interaction to arrive at sufficient throughput for pilot clinical applications. Methods: Nuclear DNA was stained in 14 μm thick tissue sections of normal liver and adrenal stained with either YOYO-1 iodide or TO-PRO-3 iodide. Different pre-treatment strategies were evaluated: boiling in citrate buffer (pH 6.0 followed by RNase application for 1 or 18 hours, or hydrolysis. The image stacks obtained with CLSM at microscope magnifications of ×40 or ×100 were analyzed off-line using in-house developed software for semi-automated 3D fluorescence quantitation. To avoid sectioned nuclei, the top and bottom of the stacks were identified from ZX and YZ projections. As a measure of histogram quality, the coefficient of variation (CV of the diploid peak was assessed. Results: The lowest CV (10.3% was achieved with a protocol without boiling, with 1 hour RNase treatment and TO-PRO-3 iodide staining, and a final image recording at ×60 or ×100 magnifications. A sample size of 300 nuclei was generally achievable. By filtering the set of automatically segmented nuclei based on volume, size and shape, followed by interactive removal of the few remaining faulty objects, a single measurement was completely analyzed in approximately 3 hours

  19. [A new laser scan system for video ophthalmoscopy. Initial clinical experiences also in relation to digital image processing].

    Science.gov (United States)

    Fabian, E; Mertz, M; Hofmann, H; Wertheimer, R; Foos, C

    1990-06-01

    The clinical advantages of a scanning laser ophthalmoscope (SLO) and video imaging of fundus pictures are described. Image quality (contrast, depth of field) and imaging possibilities (confocal stop) are assessed. Imaging with different lasers (argon, He-Ne) and changes in imaging rendered possible by confocal alignment of the imaging optics are discussed. Hard copies from video images are still of inferior quality compared to fundus photographs. Methods of direct processing and retrieval of digitally stored SLO video fundus images are illustrated by examples. Modifications for a definitive laser scanning system - in regard to the field of view and the quality of hard copies - are proposed.

  20. Computerized Reconstruction of Pulpal Blood Vessels Examined under Confocal Microscope

    Directory of Open Access Journals (Sweden)

    Digka Anna

    2015-03-01

    Full Text Available The purpose of this study was the evaluation of 3 different histological methods for studying pulpal blood vessels in combination with 2 types of confocal microscope and computer assisted 3-dimensional reconstruction. 10 human, healthy, free of restorations or caries teeth that were extracted for orthodontic reasons were used. From these teeth, the pulp tissues of 5 were removed, fixed in formalin solution, dehydrated and embedded in paraffin. Serial cross sections 5μm thick were taken from 3 of the above mentioned pulpal tissues and stained with CD34 according to the immunohistochemical ABC technique, while the rest 2 were stained with CD34 and Cy5 by means of immunofluorescence after serial cross sectioning of 10μm. 5 of the 10 teeth were fixed, decalcified, serial cross sectioned (30μm thickness and stained with eosin. The physical sections were examined under 2 types of confocal laser microscope. Serial images were taken for each section, alignment of the images was followed and finally 3-dimensional reconstructions of the pulpal vessels were achieved.

  1. Variety of corneal endothelial cell in glaucoma by confocal microscope

    Directory of Open Access Journals (Sweden)

    Hong-Liang Gao

    2014-10-01

    Full Text Available ATM: To define the causes of corneal endothelial cell damage, to investigate the preventive methods, and to observe the variety of corneal endothelial cell in glaucoma using confocal microscope.METHODS: Totally, 143 eyes of 97 patients with different types of glaucoma, and matched normal people were 20 cases, all 40 eyes. The cell density, cell area and cell variable coefficient were measured used confocal microscope. These indicatives of every kind of glaucoma were compared.RESULTS: The corneal endothelial cell density of normal group was 2 893.88±255.026/mm2, the group of acute angle-closure glaucoma(AACGwas 1 674.11±683.95/mm2, and the group of open angle glaucoma(OAGwas 2 687.22±391.87/mm2, the group of chronic angle-closure glaucoma(CACGwas 2 706.97±351.27/mm2. In all index the average cell density of corneal endothelial and the average area have statistical significance(F=62.950, 8.795; P=0.000, especially the group of AACG. CONCLUSION: The index of corneal endothelial cell in AACG is lower than that of normal. All index in OAG and CACG is difference with that of normal, but the difference has no statistical significance. And the dominant factor of damaged corneal endothelial is the time of intraocular hypertension.

  2. Quantification and confocal imaging of protein specific molecularly imprinted polymers.

    Science.gov (United States)

    Hawkins, Daniel M; Trache, Andreea; Ellis, E Ann; Stevenson, Derek; Holzenburg, Andreas; Meininger, Gerald A; Reddy, Subrayal M

    2006-09-01

    We have employed FITC--albumin as the protein template molecule in an aqueous phase molecular imprinted polymer (HydroMIP) strategy. For the first time, the use of a fluorescently labeled template is reported, with subsequent characterization of the smart material to show that the HydroMIP possesses a significant molecular memory in comparison to that of the nonimprinted control polymer (HydroNIP). The imaging of the FITC--albumin imprinted HydroMIP using confocal microscopy is described, with the in situ removal of the imprinted protein displayed in terms of observed changes in the fluorescence of the imprinted polymer, both before and after template elution (using a 10% SDS/10% AcOH (w/v) solution). We also report the imaging of a bovine hemoglobin (BHb) imprinted HydroMIP using two-photon confocal microscopy and describe the effects of template elution upon protein autofluorescence. The findings further contribute to the understanding of aqueous phase molecular imprinting protocols and document the use of fluorescence as a useful tool in template labeling/detection and novel imaging strategies.

  3. Assessment of corneal alterations by confocal microscopy in vernal keratoconjunctivitis.

    Science.gov (United States)

    Nebbioso, Marcella; Zicari, Anna Maria; Lollobrigida, Valeria; Marenco, Marco; Duse, Marzia

    2015-01-01

    Vernal keratoconjunctivitis (VKC) is a bilateral chronic, seasonally exacerbated inflammation of the ocular surface that especially affects male children and young boys. To evaluate the corneal microscopic features of patients affected by VKC and to assess whether some corneal changes were associated with specific ocular symptoms and/or signs. 20 children aged between 4 and 14 years were enrolled. All patients underwent corneal confocal microscopy by Confoscan CS3 (Nidek). 350 images of the central cornea of each eye were obtained with a ×40 noncontact lens 3,5 micron gap in automode. Some alterations of the sub-basal and stromal corneal nerves were detected. These alterations were more evident in patients with higher severity of photophobia. On the other hand, there were scarce other signs of the anterior segment of the eye. Our preliminary findings show that there is another group of patients affected by VKC, characterized by an intense photophobia caused by corneal damage and without other significant ocular alterations. Therefore confocal microscopy may be useful for an early identification of corneal alterations before the onset of severe ocular symptoms and to set an appropriate therapeutic management.

  4. In vivo confocal microscopy in goldenhar syndrome: a case report.

    Science.gov (United States)

    Triolo, Giacinto; Ferrari, Giulio; Doglioni, Claudio; Rama, Paolo

    2013-10-16

    Goldenhar Syndrome is characterized by malformations of multiple anatomical districts. Between these, bulbar dermoids are common and represent a significant clinical problem as they can affect both ocular function and aesthetic comfort.Histologic characterization of dermoids has been extensively performed; however, no reports exist describing in vivo confocal microscopy (IVCM) of these lesions. We aimed to (i) describe the in vivo confocal morphology of limbal dermoids in Goldenhar syndrome and (ii) compare these findings with standard light microscopy. A 15-year-old Caucasian female affected by Goldenhar Syndrome showed a left, infero-temporal, limbal neoformation, with extension to the left orbital region. Prior to surgical removal, IVCM was performed with the Heidelberg Retina Tomograph II, Cornea Module, using the "section" modality. The IVCM sections showed structures resembling corneal epithelium and vascular structures. Surgical removal of the lesion was decided as it caused poor eyelid closure. After surgical removal, sectioning and standard optical microscopy were performed. The comparison between IVCM imaging and standard microscopy sections were highly correlated in the detection of the pilar and vascular structures. This study showed that IVCM may be a useful technique to study limbal dermoids, given its ability to detect typical microscopic features and its comparability to optical microscopy, which is the current standard.

  5. Application of confocal X-ray fluorescence micro-spectroscopy to the investigation of paint layers.

    Science.gov (United States)

    Sun, Tianxi; Liu, Zhiguo; Wang, Guangfu; Ma, Yongzhong; Peng, Song; Sun, Weiyuan; Li, Fangzuo; Sun, Xuepeng; Ding, Xunliang

    2014-12-01

    A confocal micro X-ray fluorescence (MXRF) spectrometer based on polycapillary X-ray optics was used for the identification of paint layers. The performance of the confocal MXRF was studied. Multilayered paint fragments of a car were analyzed nondestructively to demonstrate that this confocal MXRF instrument could be used in the discrimination of the various layers in multilayer paint systems. Copyright © 2014 Elsevier Ltd. All rights reserved.

  6. Low-power, Confocal Imaging of Protein Localization in Living Cells (7214-150) Project

    Data.gov (United States)

    National Aeronautics and Space Administration — The proposed technology genetically labels intracellular structures and visualizes protein interactions in living cells using a compact, confocal microscope with...

  7. Comparison of two confocal micro-XRF spectrometers with different design aspects.

    Science.gov (United States)

    Smolek, S; Nakazawa, T; Tabe, A; Nakano, K; Tsuji, K; Streli, C; Wobrauschek, P

    2014-03-01

    Two different confocal micro X-ray fluorescence spectrometers have been developed and installed at Osaka City University and the Vienna University of Technology Atominstitut. The Osaka City University system is a high resolution spectrometer operating in air. The Vienna University of Technology Atominstitut spectrometer has a lower spatial resolution but is optimized for light element detection and operates under vacuum condition. The performance of both spectrometers was compared. In order to characterize the spatial resolution, a set of nine specially prepared single element thin film reference samples (500 nm in thickness, Al, Ti, Cr, Fe Ni, Cu, Zr, Mo, and Au) was used. Lower limits of detection were determined using the National Institute of Standards and Technology standard reference material glass standard 1412. A paint layer sample (cultural heritage application) and paint on automotive steel samples were analyzed with both instruments. The depth profile information was acquired by scanning the sample perpendicular to the surface. © 2013 The Authors. X-Ray Spectrometry published by John Wiley & Sons, Ltd.

  8. Confocal microscopy evaluation of the effect of irrigants on Enterococcus faecalis biofilm: An in vitro study.

    Science.gov (United States)

    Flach, Nicole; Böttcher, Daiana Elisabeth; Parolo, Clarissa Cavalcanti Fatturi; Firmino, Luciana Bitello; Malt, Marisa; Lammers, Marcelo Lazzaron; Grecca, Fabiana Soares

    2016-01-01

    The purpose of this study was to evaluate in vitro the effectiveness of two endodontic irrigants and their association against Enterococcus faecalis (E. faecalis) by confocal laser scanning microscope (CLSM). Twenty-four bovine incisors were inoculated in a monoculture of E. faecalis for 21 days. After this period, the teeth were divided into three test groups (n = 5) according to the chemical used. Group 1: 2.5% sodium hypochlorite (NaOCl), group 2: 2% chlorhexidine gel (CHX), group 3: 2.5% NaOCl + 2% CHX gel, and two control groups (n = 3): negative control group (NCG)-sterile and without root canals preparation and positive control group (PCG)-saline. Then, the samples were stained with SYTO9 and propidium iodide and subjected to analysis by CLSM. Bacterial viability was quantitatively analyzed by the proportions of dead and live bacteria in the biofilm remnants. Statistical analysis was performed by the One-way ANOVA test (p = 0.05). No statistical differences were observed to bacterial viability. According to CLSM analysis, none of the tested substances could completely eliminate E. faecalis from the root canal space. Until now, there are no irrigant solutions able to completely eliminate E. faecalis from the root canal. In this regard, the search for irrigants able to intensify the antimicrobial action is of paramount importance. © Wiley Periodicals, Inc.

  9. Antibacterial Efficacy of Pastes Against E Faecalis in Primary Root Dentin: A Confocal Microscope Study.

    Science.gov (United States)

    Verma, R; Sharma, D S; Pathak, A K

    2015-01-01

    Management of abscessed primary teeth often present endodontic failure owing to questioned efficiency of dressings or obturating pastes to eliminate Enterococcus faecalis, a resistant bacterium, residing in depth of dentinal tubules. The present study evaluates the antimicrobial efficacy of two antibacterial and two obturating pastes in dentinal tubules of primary teeth infected with Enterococcus faecalis using viability stain and confocal laser scanning microscope (CLSM). Total 28 samples were prepared.Four groups with 6 samples each were made according to antibacterial pastes i.e. 1% or 2%Chlorhexidine (CHX) + calcium hydroxide (CH), CH + iodoform (Metapex) and Zinc Oxide Eugenol (ZOE). Dentinal tubules from the root canal side were infected with E. faecalis by centrifugation of the bacterial suspension. Two specimens from each group were subjected to 1, 7 and 15 days antibacterial pastes exposure. Viability staining followed by CLSM were used to quantitatively analyze the dead cell count directly inside dentin. Univariate analysis showed that all medicaments were significantly effective (p ZOE(15)> Metapex(15)> 2%CHX+CH(15)> 2%CHX+CH(7)> 2%CHX+CH(1)> 1%CHX+CH(7)> 2%CHX+CH(15)> Metapex(1)> ZOE(1)> ZOE(7). All medicaments were effective against E. faecalis in dentine of primary teeth and their efficacy increased with longer contact with 1%CHX+CH being most effective at day 15. Inclusion of 1% CHX in dressings or obturating pastes might minimize the endodontic relapse and maximize the tooth retention in functional state in pediatric dentistry.

  10. Ex Vivo Confocal Spectroscopy of Autofluorescence in Age-Related Macular Degeneration.

    Directory of Open Access Journals (Sweden)

    Joel Kaluzny

    Full Text Available We investigated the autofluorescence (AF signature of the microscopic features of retina with age-related macular degeneration (AMD using 488 nm excitation.The globes of four donors with AMD and four age-matched controls were embedded in paraffin and sectioned through the macula. Sections were excited using a 488 nm argon laser, and the AF emission was captured using a laser scanning confocal microscope (496-610 nm, 6 nm resolution. The data cubes were then analyzed to compare peak emission spectra between the AMD and the controls. Microscopic features, including individual lipofuscin and melanolipofuscin granules, Bruch's Membrane, as well macroscopic features, were considered.Overall, the AMD eyes showed a trend of blue-shifted emission peaks compared with the controls. These differences were statistically significant when considering the emission of the combined RPE/Bruch's Membrane across all the tissue cross-sections (p = 0.02.The AF signatures of ex vivo AMD RPE/BrM show blue-shifted emission spectra (488 nm excitation compared with the control tissue. The magnitude of these differences is small (~4 nm and highlights the potential challenges of detecting these subtle spectral differences in vivo.

  11. Efficacy of oral exfoliative cytology in diabetes mellitus patients: a light microscopic and confocal microscopic study.

    Science.gov (United States)

    Gopal, Deepika; Malathi, N; Reddy, B Thirupathi

    2015-03-01

    Diabetes mellitus (DM) has become a global problem. By monitoring the health status of these individuals, diabetic complications can be prevented. We aimed to analyze alterations in the morphology and cytomorphometry of buccal epithelial cells of type 2 DM patients using oral exfoliative cytology technique and determine its importance in public health screening, diagnosis and monitoring of diabetes mellitus. The study was carried out in 100 type 2 DM patients and 30 healthy individuals. Smears were taken from the right buccal mucosa and stained by the Papanicolaou technique. Staining with Acridine orange was carried out to view qualitative changes with confocal laser scanning microscope (LSM-510 Meta). The cytomorphometry was evaluated using IMAGE PRO PLUS 5.5 software with Evolution LC camera. All findings were statistically analyzed. The results showed that with increase in fasting plasma glucose levels, there is significant increase in nuclear area, decrease in cytoplasmic area, and increase in nuclear cytoplasmic ratio (p < 0.05) when compared to the control group. Various qualitative changes were noted, such as cell degeneration, micronuclei, binucleation, intracytoplasmic inclusion, candida and keratinization. In the present study, we found significant alterations in the cytomorphometry and cytomorphology of buccal epithelial cells of type 2 DM patients. This study supports and extends the view that these cellular changes can alert the clinician to the possibility of diabetes and aid in monitoring of diabetes throughout the lifetime of the patient.

  12. [Imaging of corneal dystrophies: Correlations between en face anterior segment OCT and in vivo confocal microscopy].

    Science.gov (United States)

    Ghouali, W; Tahiri Joutei Hassani, R; Liang, H; Dupont-Monod, S; Auclin, F; Baudouin, C; Labbé, A

    2015-05-01

    To evaluate the usefulness of en face Optical Coherence Tomography (OCT) for evaluation of corneal dystrophies and to describe correlations with in vivo confocal microscopy (IVCM). Thirty-two eyes of 16 patients with 4 types of corneal dystrophies (epithelial basement membrane dystrophy, Fuchs dystrophy, Reis-Bücklers corneal dystrophy and Crocodile Shagreen dystrophy) were enrolled in this study. Axial and reconstructed en face scans were acquired using OCT. Images were then correlated to IVCM findings. En face OCT provided new insights into the structure, size and depth of corneal tissue alterations in various corneal dystrophies. OCT en face images were well correlated with IVCM features. Despite lower resolution than IVCM, en face OCT offers the advantages of being non-invasive and allowing the analysis of larger corneal areas. En face OCT provides useful new information in corneal dystrophies. This imaging technique will probably increase in popularity in the near future for the assessment of various anterior segment diseases. Copyright © 2015 Elsevier Masson SAS. All rights reserved.

  13. Mapping of normal corneal K-structures by in vivo laser confocal microscopy.

    Science.gov (United States)

    Yokogawa, Hideaki; Kobayashi, Akira; Sugiyama, Kazuhisa

    2008-09-01

    To produce 2-dimensional reconstruction maps of normal human corneal fibrous structures beneath the Bowman layer (K-structures) by in vivo laser confocal microscopy and to show association of structures with the anterior corneal mosaic (ACM). Central corneal regions of 3 healthy volunteers were scanned. Acquired images of K-structures for each eye were arranged and mapped into a subconfluent montage. For each subject, electrical tracings of K-structures were superimposed on a slit-lamp photograph of the ACM produced by rubbing the eyelid. A mean of 677 +/- 211 images of K-structures were obtained for each eye. Mean dimensions of the mapped areas were 5.88 +/- 0.50 (horizontal) and 3.51 +/- 1.37 mm (vertical). In all subjects, K-structures formed a netlike pattern (mean area, 0.082 +/- 0.051 mm), and electrical tracings had good concordance with the ACM. This is the first study, to our knowledge, to elucidate the overall distribution of K-structures in normal human corneas. The netlike pattern of K-structures corresponded well with ACM pattern. These results support the hypothesis that the K-structures are the anterior collagen fiber bundles running at the posterior surface of the Bowman layer and thus are the structural basis for ACM formation.

  14. Grading keratinocyte atypia in actinic keratosis: a correlation of reflectance confocal microscopy and histopathology.

    Science.gov (United States)

    Pellacani, G; Ulrich, M; Casari, A; Prow, T W; Cannillo, F; Benati, E; Losi, A; Cesinaro, A M; Longo, C; Argenziano, G; Soyer, H P

    2015-11-01

    Actinic Keratosis (AK) is the clinical manifestation of cutaneous dysplasia of epidermal keratinocytes, with progressive trend towards squamous cell carcinoma. To evaluate the strength of the correlation between keratinocyte atypia, as detected by Reflectance Confocal Microscopy (RCM) and histopathology, and to develop a more objective atypia grading scale for RCM quantification, through a discrete ranking. A total of 48 AKs and two control areas (photodamaged and non-photodamaged skin) were selected for this study. All these areas were documented by RCM and biopsied for histopathology. One representative image of the epidermis was selected for RCM and for histopathology and used for side-by-side comparison with purpose written software. The assessor chose which of two images displayed more keratinocyte atypia, and an ordered list from the image showing the least to the most keratinocyte atypia was generated. Three evaluations were obtained for RCM and two for histopathology. Good interobserver correlation was obtained for RCM and histopathology grading, with high concordance between RCM and histopathology grading. Expert rater scan consistently distinguish different grades of cytological atypia. Non-invasive RCM data from in vivo imaging can be graded for keratinocyte atypia, comparable to histopathological grading. © 2015 European Academy of Dermatology and Venereology.

  15. Comparison of two confocal micro-XRF spectrometers with different design aspects

    Science.gov (United States)

    Smolek, S; Nakazawa, T; Tabe, A; Nakano, K; Tsuji, K; Streli, C; Wobrauschek, P

    2014-01-01

    Two different confocal micro X-ray fluorescence spectrometers have been developed and installed at Osaka City University and the Vienna University of Technology Atominstitut. The Osaka City University system is a high resolution spectrometer operating in air. The Vienna University of Technology Atominstitut spectrometer has a lower spatial resolution but is optimized for light element detection and operates under vacuum condition. The performance of both spectrometers was compared. In order to characterize the spatial resolution, a set of nine specially prepared single element thin film reference samples (500 nm in thickness, Al, Ti, Cr, Fe Ni, Cu, Zr, Mo, and Au) was used. Lower limits of detection were determined using the National Institute of Standards and Technology standard reference material glass standard 1412. A paint layer sample (cultural heritage application) and paint on automotive steel samples were analyzed with both instruments. The depth profile information was acquired by scanning the sample perpendicular to the surface. © 2013 The Authors. X-Ray Spectrometry published by John Wiley & Sons, Ltd. PMID:26430286

  16. In vivo confocal microscopy of meibomian glands and palpebral conjunctiva in vernal keratoconjunctivitis

    Directory of Open Access Journals (Sweden)

    Qiaoling Wei

    2015-01-01

    Full Text Available Purpose: To investigate the correlations between conjunctival inflammatory status and meibomian gland (MG morphology in vernal keratoconjunctivitis (VKC patients by using in vivo confocal microscopy (CM. Materials and Methods: Nineteen VKC patients (7 limbal, 7 tarsal, and 5 mixed forms and 16 normal volunteers (controls were enrolled. All subjects underwent CM scanning to obtain the images of upper palpebral conjunctiva and MGs. Inflammatory cell (IC density in palpebral conjunctival epithelial and stromal layers, Langerhans cell (LC density at lid margins and the stroma adjacent to the MG, and MG acinar unit density (MGAUD were recorded. The longest and shortest diameters of MG acinar were measured. The Kruskal-Wallis test was used to compare the parameter differences whereas the Spearman′s rank correlation analysis was applied to determine their correlations. Results: Among all groups, no significant statistical differences were found in epithelial and stromal IC densities, mean values of MG acinar unit densities, or longest and shortest diameters. Both LC parameters in the tarsal-mixed groups were significantly higher than those in the limbal and control groups. All LC densities of VKC patients showed a positive correlation with MGAUD and shortest diameter. Conclusions: In VKC patients, the conjunctival inflammatory status could be associated with the MG status. In vivo CM is a noninvasive, efficient tool in the assessment of MG status and ocular surface.

  17. In vivo confocal microscopy of meibomian glands and palpebral conjunctiva in vernal keratoconjunctivitis.

    Science.gov (United States)

    Wei, Qiaoling; Le, Qihua; Hong, Jiaxu; Xiang, Jun; Wei, Anji; Xu, Jianjiang

    2015-04-01

    To investigate the correlations between conjunctival inflammatory status and meibomian gland (MG) morphology in vernal keratoconjunctivitis (VKC) patients by using in vivo confocal microscopy (CM). Nineteen VKC patients (7 limbal, 7 tarsal, and 5 mixed forms) and 16 normal volunteers (controls) were enrolled. All subjects underwent CM scanning to obtain the images of upper palpebral conjunctiva and MGs. Inflammatory cell (IC) density in palpebral conjunctival epithelial and stromal layers, Langerhans cell (LC) density at lid margins and the stroma adjacent to the MG, and MG acinar unit density (MGAUD) were recorded. The longest and shortest diameters of MG acinar were measured. The Kruskal-Wallis test was used to compare the parameter differences whereas the Spearman's rank correlation analysis was applied to determine their correlations. Among all groups, no significant statistical differences were found in epithelial and stromal IC densities, mean values of MG acinar unit densities, or longest and shortest diameters. Both LC parameters in the tarsal-mixed groups were significantly higher than those in the limbal and control groups. All LC densities of VKC patients showed a positive correlation with MGAUD and shortest diameter. In VKC patients, the conjunctival inflammatory status could be associated with the MG status. In vivo CM is a noninvasive, efficient tool in the assessment of MG status and ocular surface.

  18. Measurement of surgically induced corneal deformations using three-dimensional confocal microscopy.

    Science.gov (United States)

    Petroll, W M; Roy, P; Chuong, C J; Hall, B; Cavanagh, H D; Jester, J V

    1996-03-01

    The goal of this study was to develop and apply a new set of experimental techniques for measuring the local deformations induced by partial-thickness corneal incisions in situ. Eight adult cat eyes were enucleated and cannulated, with corneal viability maintained as close to in vivo conditions as possible and intraocular pressure (IOP) carefully controlled. Experimental measurements were made pre/post radial keratotomy (RK) surgery in situ at IOPs of 15, 30, and 45 mm Hg. Incision depth and cross-sectional profiles were measured at the midpoint of selected incisions using three-dimensional (3-D) tandem scanning confocal microscopy (TSCM); central corneal curvature was estimated using a commercial corneal topographical analysis system, and corneal thickness was assessed by both 3-D TSCM and ultrasonic pachymetry. Corneas were then processed for light microscopy and incision depth was measured histologically. Finite element models were developed for comparison with the experimental measurements. There was no significant change in central corneal thickness (-5.3 +/- 3.9%, n = 8) over the course of the experiments, demonstrating that normal endothelial cell function and normal stromal hydration was maintained. The in situ TSCM incision depth measurements were significantly correlated with the histological measurements (slope = 0.95, R = 0.854, p mechanical behavior of the cornea after refractive surgery. These data should provide the foundation for future studies into the relationships between local tissue mechanics and corneal wound healing.

  19. Localization of extracellular matrix components in developing mouse salivary glands by confocal microscopy

    Science.gov (United States)

    Hardman, P.; Spooner, B. S.

    1992-01-01

    The importance of the extracellular matrix (ECM) in epithelial-mesenchymal interactions in developing organisms is well established. Proteoglycans and interstitial collagens are required for the growth, morphogenesis, and differentiation of epithelial organs and the distribution of these molecules has been described. However, much less is known about other ECM macromolecules in developing epithelial organs. We used confocal microscopy to examine the distribution of laminin, heparan sulfate (BM-1) proteoglycan, fibronectin, and collagen types I, IV, and V, in mouse embryonic salivary glands. Organ rudiments were isolated from gestational day 13 mouse embryos and cultured for 24, 48, or 72 hours. Whole mounts were stained by indirect immunofluorescence and then examined using a Zeiss Laser Scan Microscope. We found that each ECM component examined had a distinct distribution and that the distribution of some molecules varied with culture time. Laminin was mainly restricted to the basement membrane. BM-1 proteoglycan was concentrated in the basement membrane and also formed a fine network throughout the mesenchyme. Type IV collagen was mainly located in the basement membrane of the epithelium, but it was also present throughout the mesenchyme. Type V collagen was distributed throughout the mesenchyme at 24 hours, but at 48 hours was principally located in the basement membrane. Type I collagen was distributed throughout the mesenchyme at all culture times, and accumulated in the clefts and particularly at the epithelial-mesenchymal interface as time in culture increased. Fibronectin was observed throughout the mesenchyme at all times.

  20. In situ Observation of Calcium Oxide Treatment of Inclusions in Molten Steel by Confocal Microscopy

    Science.gov (United States)

    Khurana, Bharat; Spooner, Stephen; Rao, M. B. V.; Roy, Gour Gopal; Srirangam, Prakash

    2017-06-01

    Calcium treatment of aluminum killed steel was observed in situ using high-temperature confocal scanning laser microscope (HT-CSLM). This technique along with a novel experimental design enables continuous observation of clustering behavior of inclusions before and after the calcium treatment. Results show that the increase in average inclusion size in non-calcium-treated condition was much faster compared to calcium-treated condition. Results also show that the magnitude of attractive capillary force between inclusion particles in non-treated condition was about 10-15 N for larger particles (10 µm) and 10-16 N for smaller particles (5 µm) and acting length of force was about 30 µm. In the case of calcium-treated condition, the magnitude and acting length of force was reduced to 10-16 N and 10 µm, respectively, for particles of all sizes. This change in attractive capillary attractive force is due to change in inclusion morphology from solid alumina disks to liquid lens particles during calcium treatment.

  1. Efficiency of the confocal method of laser endomicroscopy in complex diagnoses of diseases of common bile duct

    Science.gov (United States)

    Anaskin, S. G.; Panchenkov, D. N.; Chertyuk, V. B.; Sazonov, D. V.; Zabozlayev, F. G.; Danilevskaya, O. V.; Mokshina, N. V.; Korniletsky, I. D.

    2017-01-01

    One of the more frequent manifestations of diseases of the bile ducts are its’ strictures or stenoses that could be of either malignant or benign nature. Current methods of diagnosing this pathology include computer tomography (CT) scan, magnetic resonance cholangiopancreatography (MRCP), endoscopic ultrasound (EUS) and endoscopic retrograde cholangiopancreatography (ERCP). However, these methods are not always informative, which makes this a current and topical problem. A fundamentally new method that broadens the capabilities of ERCP when diagnosing diseases of the bile duct accompanied by the development of strictures or stenoses is probe-based confocal laser endomicroscopy (pCLE). The method is based on the principle of confocal fluorescence microscopy. The most elaborate complications arise with the presence of the pre-existing pancreatobiliary pathology: pseudotumoral chronic pancreatitis, acute cholangitis, etc. Early stage cholangiocarcinoma diagnosis can be difficult (and not always possible) even with the help of modern research methods. For the timely diagnostic it is advantageous to conduct pCLE and targeted biopsy of the zone with most manifested changes. In all instances, the first use of the pCLE method for diagnostic purposes allowed us to clarify and correctly verify the diagnosis. When concerning the diseases of the bile duct, the modern stage of pCLE development can be of critical importance when other methods are not effective.

  2. Living matter observations with a novel hyperspectral supercontinuum confocal microscope for VIS to near-IR reflectance spectroscopy.

    Science.gov (United States)

    Bertani, Francesca R; Ferrari, Luisa; Mussi, Valentina; Botti, Elisabetta; Costanzo, Antonio; Selci, Stefano

    2013-10-25

    A broad range hyper-spectroscopic microscope fed by a supercontinuum laser source and equipped with an almost achromatic optical layout is illustrated with detailed explanations of the design, implementation and data. The real novelty of this instrument, a confocal spectroscopic microscope capable of recording high resolution reflectance data in the VIS-IR spectral range from about 500 nm to 2.5 μm wavelengths, is the possibility of acquiring spectral data at every physical point as defined by lateral coordinates, X and Y, as well as at a depth coordinate, Z, as obtained by the confocal optical sectioning advantage. With this apparatus we collect each single scanning point as a whole spectrum by combining two linear spectral detector arrays, one CCD for the visible range, and one InGaAs infrared array, simultaneously available at the sensor output channel of the home made instrument. This microscope has been developed for biomedical analysis of human skin and other similar applications. Results are shown illustrating the technical performances of the instrument and the capability in extracting information about the composition and the structure of different parts or compartments in biological samples as well as in solid statematter. A complete spectroscopic fingerprinting of samples at microscopic level is shown possible by using statistical analysis on raw data or analytical reflectance models based on Abelés matrix transfer methods.

  3. Anatomical and metabolic small-animal whole-body imaging using ring-shaped confocal photoacoustic computed tomography

    Science.gov (United States)

    Xia, Jun; Chatni, Muhammad; Maslov, Konstantin; Wang, Lihong V.

    2013-03-01

    Due to the wide use of animals for human disease studies, small animal whole-body imaging plays an increasingly important role in biomedical research. Currently, none of the existing imaging modalities can provide both anatomical and glucose metabolic information, leading to higher costs of building dual-modality systems. Even with image coregistration, the spatial resolution of the metabolic imaging modality is not improved. We present a ring-shaped confocal photoacoustic computed tomography (RC-PACT) system that can provide both assessments in a single modality. Utilizing the novel design of confocal full-ring light delivery and ultrasound transducer array detection, RC-PACT provides full-view cross-sectional imaging with high spatial resolution. Scanning along the orthogonal direction provides three-dimensional imaging. While the mouse anatomy was imaged with endogenous hemoglobin contrast, the glucose metabolism was imaged with a near-infrared dye-labeled 2-deoxyglucose. Through mouse tumor models, we demonstrate that RC-PACT may be a paradigm shifting imaging method for preclinical research.

  4. Living Matter Observations with a Novel Hyperspectral Supercontinuum Confocal Microscope for VIS to Near-IR Reflectance Spectroscopy

    Directory of Open Access Journals (Sweden)

    Francesca R. Bertani

    2013-10-01

    Full Text Available A broad range hyper-spectroscopic microscope fed by a supercontinuum laser source and equipped with an almost achromatic optical layout is illustrated with detailed explanations of the design, implementation and data. The real novelty of this instrument, a confocal spectroscopic microscope capable of recording high resolution reflectance data in the VIS-IR spectral range from about 500 nm to 2.5 μm wavelengths, is the possibility of acquiring spectral data at every physical point as defined by lateral coordinates, X and Y, as well as at a depth coordinate, Z, as obtained by the confocal optical sectioning advantage. With this apparatus we collect each single scanning point as a whole spectrum by combining two linear spectral detector arrays, one CCD for the visible range, and one InGaAs infrared array, simultaneously available at the sensor output channel of the home made instrument. This microscope has been developed for biomedical analysis of human skin and other similar applications. Results are shown illustrating the technical performances of the instrument and the capability in extracting information about the composition and the structure of different parts or compartments in biological samples as well as in solid statematter. A complete spectroscopic fingerprinting of samples at microscopic level is shown possible by using statistical analysis on raw data or analytical reflectance models based on Abelés matrix transfer methods.

  5. Cellular-level near-wall unsteadiness of high-hematocrit erythrocyte flow using confocal μPIV

    Science.gov (United States)

    Patrick, Michael J.; Chen, Chia-Yuan; Frakes, David H.; Dur, Onur; Pekkan, Kerem

    2011-04-01

    In hemodynamics, the inherent intermittency of two-phase cellular-level flow has received little attention. Unsteadiness is reported and quantified for the first time in the literature using a combination of fluorescent dye labeling, time-resolved scanning confocal microscopy, and micro-particle image velocimetry (μPIV). The near-wall red blood cell (RBC) motion of physiologic high-hematocrit blood in a rectangular microchannel was investigated under pressure-driven flow. Intermittent flow was associated with (1) the stretching of RBCs as they passed through RBC clusters with twisting motions; (2) external flow through local obstacles; and (3) transitionary rouleaux formations. Velocity profiles are presented for these cases. Unsteady flow clustered in local regions. Extra-cellular fluid flow generated by individual RBCs was examined using submicron fluorescent microspheres. The capabilities of confocal μPIV post-processing were verified using synthetic raw PIV data for validation. Cellular interactions and oscillating velocity profiles are presented, and 3D data are made available for computational model validation.

  6. Cellular-level near-wall unsteadiness of high-hematocrit erythrocyte flow using confocal {mu}PIV

    Energy Technology Data Exchange (ETDEWEB)

    Patrick, Michael J. [Carnegie Mellon University, Molecular Biosensor and Imaging Center (MBIC), Pittsburgh, PA (United States); Chen, Chia-Yuan; Dur, Onur; Pekkan, Kerem [Carnegie Mellon University, Department of Biomedical and Mechanical Engineering, Pittsburgh, PA (United States); Frakes, David H. [Arizona State University, School of Biological and Health Systems Engineering and School of Electrical, Computer, and Energy Engineering, Tempe, AZ (United States)

    2011-04-15

    In hemodynamics, the inherent intermittency of two-phase cellular-level flow has received little attention. Unsteadiness is reported and quantified for the first time in the literature using a combination of fluorescent dye labeling, time-resolved scanning confocal microscopy, and micro-particle image velocimetry ({mu}PIV). The near-wall red blood cell (RBC) motion of physiologic high-hematocrit blood in a rectangular microchannel was investigated under pressure-driven flow. Intermittent flow was associated with (1) the stretching of RBCs as they passed through RBC clusters with twisting motions; (2) external flow through local obstacles; and (3) transitionary rouleaux formations. Velocity profiles are presented for these cases. Unsteady flow clustered in local regions. Extra-cellular fluid flow generated by individual RBCs was examined using submicron fluorescent microspheres. The capabilities of confocal {mu}PIV post-processing were verified using synthetic raw PIV data for validation. Cellular interactions and oscillating velocity profiles are presented, and 3D data are made available for computational model validation. (orig.)

  7. Materials and corrosion characterization using the confocal resonator

    Energy Technology Data Exchange (ETDEWEB)

    Tigges, C.P.; Sorensen, N.R.; Hietala, V.M.; Plut, T.A. [and others

    1997-05-01

    Improved characterization and process control is important to many Sandia and DOE programs related to manufacturing. Many processes/structures are currently under-characterized including thin film growth, corrosion and semiconductor structures, such as implant profiles. A sensitive tool is required that is able to provide lateral and vertical imaging of the electromagnetic properties of a sample. The confocal resonator is able to characterize the surface and near-surface impedance of materials. This device may be applied to a broad range of applications including in situ evaluation of thin film processes, physical defect detection/characterization, the characterization of semiconductor devices and corrosion studies. In all of these cases, the technology should work as a real-time process diagnostic or as a feedback mechanism regarding the quality of a manufacturing process. This report summarizes the development and exploration of several diagnostic applications.

  8. Automatic, high-accuracy image registration in confocal microscopy.

    Science.gov (United States)

    Liu, Jian; Li, Yong; Wang, Weibo; Wang, Yuhang; Zhang, He; Tan, Jiubin

    2017-11-10

    We proposed a high-accuracy image registration method of confocal microscopy for a large field of view and high resolution. The spatial information (edge information) and the entropy correlation coefficient have been both taken into account for higher accuracy of registration. The edge information is introduced to calculate the normalization correlation coefficient of the image. Then the normalization correlation coefficient and the entropy correlation coefficient of the original image have been used to improve the proposed similarity measures, the normalized mutual information with edge information (called EMI). Meanwhile, a parallel particle swarm optimization (pa-PSO) with the idea of conditional initialization and parallel cooperation is developed to speed up the convergence rate and further reduce the mismatch. Experiments verified that the registration accuracy can be up to 0.2 pixel and has better robustness to the noise.

  9. Reflectance confocal microscopy for cutaneous infections and infestations.

    Science.gov (United States)

    Cinotti, E; Perrot, J L; Labeille, B; Cambazard, F

    2016-05-01

    Reflectance confocal microscopy (RCM) is a high-resolution emerging imaging technique that allows non-invasive diagnosis of several cutaneous disorders. A systematic review of the literature on the use of RCM for the study of infections and infestations has been performed to evaluate the current use of this technique and its possible future applications in this field. RCM is particularly suitable for the identification of Sarcoptes scabies, Demodex folliculorum, Ixodes, Dermatophytes and Candida species in the clinical practice and for the follow-up after treatment. The cytopathic effect of herpes simplex virus, varicella zoster virus and molluscipoxvirus is also detectable by this imaging technique even in a pre-vesicular stage. In addition, thanks to its non-invasiveness, RCM allows pathophysiological studies. © 2015 European Academy of Dermatology and Venereology.

  10. Endocrine and metabolic disease: Confocal microscopy as a diagnostic aid

    Directory of Open Access Journals (Sweden)

    Jaikrit Bhutani

    2015-01-01

    Full Text Available Diabetes is a systemic disease associated with many complications. These can be prevented and managed effectively if detected promptly. Confocal microscopy (CFM is a diagnostic tool which has the potential to help in early detection of disease and timely management. CFM has the potential to serve as an excellent noninvasive modality for in vivo imaging and morphological analysis, which can aid us in assessing and monitoring various infectious and pathological diseases at the cellular level. Besides ophthalmological indications, CFM has shown good sensitivity and specificity for identifying those at risk of neuropathy and foot ulceration, monitoring evolution and therapeutic response in a wide range of neuropathies apart from diabetic neuropathy. Through this communication, we aim to sensitize the endocrinologists towards cerebral cavernous malformation as a biomarker to evaluate potential outcomes and therapies in human diabetic neuropathy.

  11. Straightness measurements by use of a reflection confocal optical system.

    Science.gov (United States)

    Matsuda, K; Roy, M; O'Byrne, J W; Fekete, P W; Eiju, T; Sheppard, C J

    1999-09-01

    Straightness measurement is a very important technique in the field of mechanical engineering. A particular application for straightness measurement is high-accuracy machining on a diamond-turning lathe. We propose a novel, to our knowledge, optical method for measuring the straightness of motion, and its mathematical analysis is outlined. The technique is based on measurement of the lateral displacement of point images by use of reflection confocal optical systems. The advantages of this method are that (i) the lateral displacements in the direction of the two axes perpendicular to the optical axis can be measured, (ii) the rotation angles around all three axes can be measured, and (iii) reflection optical systems are more compact in length than are transmission optical systems.

  12. The Reliability and Reproducibility of Corneal Confocal Microscopy in Children.

    Science.gov (United States)

    Pacaud, Danièle; Romanchuk, Kenneth G; Tavakoli, Mitra; Gougeon, Claire; Virtanen, Heidi; Ferdousi, Maryam; Nettel-Aguirre, Alberto; Mah, Jean K; Malik, Rayaz A

    2015-08-01

    To assess the image and patient level interrater agreement and repeatability within 1 month for corneal nerve fiber length (CNFL) measured using in vivo corneal confocal microscopy (IVCCM) in children. Seventy-one subjects (mean [SD] age 14.3 [2.6] years, range 8-18 years; 44 with type 1 diabetes and 27 controls; 36 males and 35 females) were included. 547 images (∼6 images per subject) were analyzed manually by two independent and masked observers. One-month repeat visit images were analyzed by a single masked observer in 21 patients. Automated image analysis was then performed using a specialized computerized software (ACCMetrics). For CNFL, the ICC (95% CI) were 0.94 (0.93-0.95) for image-level, 0.86 (0.78-0.91) for patient-level, and 0.88 (0.72-0.95) for the 1-month repeat assessment, and the Bland-Altman plots showed minimal bias between observers. Although there was excellent agreement between manual and automated analysis according to an ICC 0.89 (0.82-0.93), the Bland-Altman plot showed a consistent bias with manual measurements providing higher readings. In vivo corneal confocal microscopy image analysis shows good reproducibility with excellent intraindividual and interindividual variability in pediatric subjects. Since the image-level reproducibility is stronger than the patient-level reproducibility, refinement of the method for image selection will likely further increase the robustness of this novel, rapid, and noninvasive approach to detect early neuropathy in children with diabetes. Further studies on the use of IVCCM to identify early subclinical neuropathy in children are indicated.

  13. Reflectance confocal microscopy for the evaluation of sensitive skin.

    Science.gov (United States)

    Ma, Y-F; Yuan, C; Jiang, W-C; Wang, X-L; Humbert, P

    2017-05-01

    Nowadays, the diagnosis for sensitive skin relies on subjective assessment or on the combination of subjective and objective evaluation. No quantitative evaluation is available. It could be expected that confocal microscopy imaging could be of interest to better define the condition. Total 166 healthy female subjects were recruited in this study. Firstly, all subjects completed the sensitive questionnaire. Then, the cutaneous structures were measured by the reflectance confocal microscopy (RCM) on the face and fossa cubitalia. The lactic acid sting test was conducted finally. According to the results of self-perception sensitive skin questionnaire and lactic acid stinging test to evaluate facial skin sensitivity the both positive subjects were regarded as sensitive skin group and both negative group as healthy control group. The results of RCM indicating that the proportion of 'disarranged honeycomb pattern' and 'spongiform edema' in the sensitive group and healthy control group were statistically different (P 0.05). The epidermal thickness was 38.88 ± 6.81 μm, healthy control group was 40.31 ± 9.37 μm in, respectively, sensitive skin group and healthy control group, there was no significant statistical difference between the two groups (P > 0.05). The honeycomb structure depth of sensitive group was 20.57 ± 4.86 μm. It was for 23.27 ± 6.38 μm, healthy control group the difference being statistically different between the two groups (P skin signs of RCM evaluation of sensitive skin effectively. Indeed, sensitive skin honeycomb structure depth was thinner compared with healthy control group. Such a specific pattern has good clinical and monitoring value for the further exploration. RCM could provide new data and patterns for the evaluation of sensitive skin. © 2016 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

  14. Quantification of confocal images of human corneal endothelium

    Science.gov (United States)

    Laird, Jeffery A.; Beuerman, Roger W.; Kaufman, Stephen C.

    1996-05-01

    Real-time, in vivo, confocal microscopic examination permits visualization of human corneal endothelium cells as bright bodies organized into a densely packed hexagonal arrangement. Quantification of endothelial cell number would be useful in assessing the condition of this cell layer in various disease states. In this study, we sought to use an image analysis method developed in this laboratory that utilizes digital filtering techniques and morphological operations to determine the boundaries of each cell. Images were corrected to establish a uniform luminance level, and then convolved by various matrices until distinct peaks in luminance value were identified. These peaks were used as seed points from which cell boundaries were recursively expanded until they collided with other cell boundaries. This method automatically counts the number of cells and determines the size and position of each cell. The resulting histograms of cell size are readily indicative of changes in cellular density, cell loss, and deviation from uniform arrangement. The numbers of cells counted by this method are consistently within 3% of the numbers counted manually. Results relating cell counts obtained by manual and computerized methods are as follows: 200/184; 276/262; 87/87; 234/232; 236/232; 299/297; 145/147; 119/122; 237/243; 119/119; 245/253; 189/193. Thus, confocal microscopy coupled with these image analysis and statistical procedures provides an accurate quantitative approach to monitoring the endothelium under normal, pathological, and experimental conditions, such as those following surgery and trauma or in the evaluation of the efficacy of topical therapeutic agents.

  15. Diagnosis of thalassemia and iron deficiency anemia using confocal and atomic force microscopy

    Science.gov (United States)

    Tariq, Saira; Bilal, Muhammad; Shahzad, Shaheen; Firdous, Shamaraz; Aziz, Uzma; Ahmed, Mushtaq

    2017-11-01

    Anemia is the most prevalent blood disorder, categorized into thalassemia and iron deficiency anemia. In anemia, the morphology of erythrocytes is disturbed, thus leading to abnormal functioning of the erythrocytes. Globally, thalassemia affects 1.3% of individuals and is one of the most widespread monogenic disorders in Pakistan. All over the World, women and children are most frequently affected by a type of nutritional deficiency known as iron deficiency anemia. The morphological changes that occur in erythrocytes due to these diseases are investigated in this study at the nano-scale level. Fifty samples of blood from individuals suffering from thalassemia or iron deficiency anemia were obtained from different hospitals in Rawalpindi and Islamabad. The blood samples were scanned using atomic force microscopy (AFM) and laser scanning confocal microscopy (LSCM) to check the morphological changes in both types of anemia. According to the present study, thalassemia is most prevalent in females in the age group between 5 and 15 years old, and iron deficiency is most prevalent in females in the age groups of 16–25 and 36–45 years old. Erythrocyte morphology is the significant determinant for diagnosing and discriminating between these two types of diseases. The study reports deformed erythrocytes in anemic patients, which were different from the ones that existed in the control. Thalassemia erythrocytes showed a crenated shape, iron deficiency anemia erythrocytes showed an elliptocyte shape and healthy erythrocytes showed a biconcave disk shape when using AFM and LSCM. These techniques seem to be very promising, cheap and less time consuming in determining the structure–function relationship of erythrocytes of thalassemic and iron deficiency anemic patients. The results of LSCM and AFM are quite useful in determining the morphological changes in erythrocytes and to study the disease at the molecular level within short period of time. Hence, we encourage

  16. Estudio del endotelio corneal en el queratocono por microscopia confocal Study of the corneal endothelium confocal microscopy in keratoconus

    Directory of Open Access Journals (Sweden)

    María del Carmen Benítez Merino

    2011-12-01

    Full Text Available Objetivo: Describir los hallazgos morfométricos del endotelio corneal por microscopia confocal con CONFOSCAN S-4. Métodos: Estudio descriptivo transversal de 102 ojos con queratocono en el período de septiembre de 2008 a septiembre 2009. A estos pacientes se les realizó microscopia confocal con CosfoscanS-4 para el estudio del endotelio corneal atendiendo el grado de queratocono. Se analizó el comportamiento de la evolución del queratocono según edad y sexo. Las imágenes fueron analizadas y procesadas mediante un programa informático diseñado específicamente para esto. Resultados: Fueron semejantes las edades de los pacientes con queratocono grado I y II, (35,2 y 34,7 años, los grado III presentaron una edad promedio mayor (38,4 años, sin diferencias significativas (p= 0,279. El sexo femenino predominó en 80,4 % de los pacientes. El 100 % de los queratoconos grado III tuvieron endotelios patológicos. Los valores promedios de la densidad celular en los queratoconos grado III (2585,9 células/mm² resultó no significativo (p= 0,339. El polimegatismo en los queratoconos grado III para un 48,69 % fue significativo (p= 0,002. En el pleomorfismo resultó significativo las diferencias observadas entre los tres grados (p= 0,002. Conclusión: Predominó el queratocono grado II para las mujeres y el grado I para los hombres. Los hallazgos morfológicos se manifestaron en la forma y tamaño de las células endoteliales. En córneas con queratocono grado II y III confluyeron células de mediano y gran tamaño con pérdida de su hexagonalidad. La densidad celular se mantuvo dentro del rango de valores normales para cualquier grado de queratocono.Objective: To describe the morphometric findings of the corneal endothelium confocal microscopy with CONFOSCAN S-4 Methods: Descriptive cross-sectional study of 102 eyes with keratoconus performed from September 2008 to September 2009. The study patients had undergone confocal microscopy with

  17. Microscopia confocal en córneas de cien ojos sanos Confocal microscopy results of one hundred healthy eye corneas

    Directory of Open Access Journals (Sweden)

    Zulema Gómez Castillo

    2012-06-01

    Full Text Available Objetivo: Analizar las estructuras celulares por microscopia confocal, Confoscan 4, en córneas sanas en nuestro medio. Métodos: Se realizó un estudio prospectivo longitudinal a 100 ojos sanos de médicos que trabajan en nuestra institución, y pacientes que asistieron al servicio de córnea. Esta investigación fue desde mayo de 2007 a mayo 2008, en el Instituto Cubano de Oftalmología "Ramón Pando Ferrer", La Habana. En los médicos se examinaron ambos ojos y en los pacientes el ojo no afectado. Se recopilaron un total de 50 casos sin afección corneal. Resultados: De los 100 ojos estudiados, 64 tenían paquimetrías por encima del valor medio. Estuvieron presentes los tres tipos de células epiteliales en casi la totalidad de los pacientes; así como los queratocitos en las diferentes profundidades del estroma corneal. La mayoría de los ojos tenían un conteo celular endotelial por encima de 2 500, cifra comprendida dentro de los valores normales. Se encontraron fibras nerviosas en cada una de sus capas. Conclusiones: La microscopia confocal se presenta como una nueva herramienta que permite observar en vivo la histología corneal y complementar las observaciones de la biomicroscopia convencional. Esto constituye un reto para el mejor entendimiento de la histopatología corneal. De esta manera podemos actuar de forma profiláctica y terapéutica, en el seguimiento y evolución de patologías corneales.Objective: This paper is aimed at analyzing the corneal cellular structures through Confoscan S4-aided confocal microscopy in apparently healthy corneas. Methods: A prospective longitudinal study of 100 healthy eyes from practicing doctors, and from patients who had attended the corneal service at “Ramón Pando Ferrer” Cuban Institute of Ophthalmology in Havana since May 2007 was conducted. Both eyes of participating doctors were examined whereas the non-affected eye was examined in the patients. A total of 50 cases with no corneal

  18. [In vivo reflectance-mode confocal laser microscopy: basic principles and clinical and research employments in dermatology].

    Science.gov (United States)

    Levi, Assi; Ingber, Arieh; Enk, David Claes

    2012-10-01

    Reflectance-mode confocal scanning laser microscopy is a novel, non-invasive imaging technique which permits real time visualization of cellular components in the skin at a resolution close to that of conventional histology. It has been widely used in the diagnosis of both benign and malignant tumors of the skin. In recent years it was also employed in the investigation of a variety of inflammatory and infectious skin conditions. The non-invasive nature of the procedure allows examination of multiple lesions and/ or repetitive sampling of one lesion over time, making it an excellent tool for followup and for monitoring treatment outcome in medical and cosmetic dermatology. This review summarizes the main indications for the use of this novel technique in clinical and experimental dermatology.

  19. Nanoparticle uptake and their co-localization with cell compartments - a confocal Raman microscopy study at single cell level

    Science.gov (United States)

    Estrela-Lopis, I.; Romero, G.; Rojas, E.; Moya, S. E.; Donath, E.

    2011-07-01

    Confocal Raman Microscopy, a non-invasive, non-destructive and label-free technique, was employed to study the uptake and localization of nanoparticles (NPs) in the Hepatocarcinoma human cell line HepG2 at the level of single cells. Cells were exposed to carbon nanotubes (CNTs) the surface of which was engineered with polyelectrolytes and lipid layers, aluminium oxide and cerium dioxide nanoparticles. Raman spectra deconvolution was applied to obtain the spatial distributions of NPs together with lipids/proteins in cells. The colocalization of the NPs with different intracellular environments, lipid bodies, protein and DNA, was inferred. Lipid coated CNTs associated preferentially with lipid rich regions, whereas polyelectrolyte coated CNTs were excluded from lipid rich regions. Al2O3 NPs were found in the cytoplasm. CeO2 NPs were readily taken up and have been observed all over the cell. Raman z-scans proved the intracellular distribution of the respective NPs.

  20. Effect of Different Irrigating Solutions on Depth of Penetration of Sealer into Dentinal Tubules: A Confocal Microscopic Study.

    Science.gov (United States)

    Thota, Murali Mohan; Sudha, Kakollu; Malini, D L; Madhavi, Singiri Bindhu

    2017-01-01

    The aim of the study was to evaluate the effect of different irrigating solutions used in final irrigation on depth of sealer penetration into dentinal tubules. Thirty recently extracted, human mandibular premolar teeth with single canals were randomly divided into two groups, and one of the two irrigants was used in each group - Group A (Chitosan) and Group B (Ethylenediaminetetraacetic acid). All the teeth were obturated with gutta-percha and AH 26(®) sealer labeled with fluorescent dye. The teeth were sectioned at distances 2, 5, and 8 mm from the root apex. Maximum depth of sealer penetration was measured using confocal laser scanning microscopy. Statistical analysis used One-way analysis of variance and t-test. At coronal third depth, the sealer penetration was greater in ethylenediaminetetraacetic acid (EDTA) group; however, depth of sealer penetration was greater at apical third in chitosan group. Final irrigation with EDTA and chitosan after the use of sodium hypochlorite affected sealer penetration.

  1. In-Situ Observation of Crystallization and Growth in High-Temperature Melts Using the Confocal Laser Microscope

    Science.gov (United States)

    Sohn, Il; Dippenaar, Rian

    2016-08-01

    This review discusses the innovative efforts initiated by Emi and co-workers for in-situ observation of phase transformations at high temperatures for materials. By using the high-temperature confocal laser-scanning microscope (CLSM), a robust database of the phase transformation behavior during heating and cooling of slags, fluxes, and steel can be developed. The rate of solidification and the progression of solid-state phase transformations can be readily investigated under a variety of atmospheric conditions and be correlated with theoretical predictions. The various research efforts following the work of Emi and co-workers have allowed a deeper fundamental understanding of the elusive solidification and phase transformation mechanisms in materials beyond the ambit of steels. This technique continues to evolve in terms of its methodology, application to other materials, and its contribution to technology.

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

    OpenAIRE

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

    2013-01-01

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

  3. Ca(2+ release events in cardiac myocytes up close: insights from fast confocal imaging.

    Directory of Open Access Journals (Sweden)

    Vyacheslav M Shkryl

    Full Text Available The spatio-temporal properties of Ca(2+ transients during excitation-contraction coupling and elementary Ca(2+ release events (Ca(2+ sparks were studied in atrial and ventricular myocytes with ultra-fast confocal microscopy using a Zeiss LSM 5 LIVE system that allows sampling rates of up to 60 kHz. Ca(2+ sparks which originated from subsarcolemmal junctional sarcoplasmic reticulum (j-SR release sites in atrial myocytes were anisotropic and elongated in the longitudinal direction of the cell. Ca(2+ sparks in atrial cells originating from non-junctional SR and in ventricular myocytes were symmetrical. Ca(2+ spark recording in line scan mode at 40,000 lines/s uncovered step-like increases of [Ca(2+]i. 2-D imaging of Ca(2+ transients revealed an asynchronous activation of release sites and allowed the sequential recording of Ca(2+ entry through surface membrane Ca(2+ channels and subsequent activation of Ca(2+-induced Ca(2+ release. With a latency of 2.5 ms after application of an electrical stimulus, Ca(2+ entry could be detected that was followed by SR Ca(2+ release after an additional 3 ms delay. Maximum Ca(2+ release was observed 4 ms after the beginning of release. The timing of Ca(2+ entry and release was confirmed by simultaneous [Ca(2+]i and membrane current measurements using the whole cell voltage-clamp technique. In atrial cells activation of discrete individual release sites of the j-SR led to spatially restricted Ca(2+ release events that fused into a peripheral ring of elevated [Ca(2+]i that subsequently propagated in a wave-like fashion towards the center of the cell. In ventricular myocytes asynchronous Ca(2+ release signals from discrete sites with no preferential subcellular location preceded the whole-cell Ca(2+ transient. In summary, ultra-fast confocal imaging allows investigation of Ca(2+ signals with a time resolution similar to patch clamp technique, however in a less invasive fashion.

  4. Influences of edges and steep slopes in 3D interference and confocal microscopy

    Science.gov (United States)

    Xie, Weichang; Hagemeier, Sebastian; Woidt, Carsten; Hillmer, Harmut; Lehmann, Peter

    2016-04-01

    Optical measurement techniques are widely applied in high-resolution contour, topography and roughness measurement. In this context vertical scanning white-light interferometers and confocal microscopes have become mature instruments over the last decades. The accuracy of measurement results is highly related not only to the type and physical properties of the measuring instruments, but also to the measurement object itself. This contribution focuses on measurement effects occurring at edges and height steps using white-light interferometers of different numerical apertures. If the edge is perfectly perpendicular, batwing effects appear at height steps. These batwings show maximum height if the height-to-wavelength-ratio (HWR) is about one forth or three forth, and they disappear if the HWR value is about an integer multiple of one half. The wavelength that is relevant in this context is the effective wavelength, i.e. the center wavelength of the illuminating light multiplied by a correction factor known as the numerical aperture correction. However, in practice the edges are usually not perfectly perpendicular. In this case, the measurement results depend also on the derivative of the surface height function and they may differ from theory and the prediction according to the HWR value. Measurements of such steps show systematical effects depending on the lateral resolution of the instrument. In this context, a Linnik interferometer with a magnification of 100x and NA = 0.9 is used to characterize the three dimensional topography of more or less rectangular calibration specimens and quasi-perpendicular structures produced by the nanoimprint technology. The Linnik interferometer is equipped with LED light sources emitting at different wavelengths, so that the HWR value can be changed. This is possible since the high NA objective lenses show a rather limited depth of focus such that the temporal coherence gating may be replaced by focal gating in this particular

  5. Ca(2+) release events in cardiac myocytes up close: insights from fast confocal imaging.

    Science.gov (United States)

    Shkryl, Vyacheslav M; Blatter, Lothar A

    2013-01-01

    The spatio-temporal properties of Ca(2+) transients during excitation-contraction coupling and elementary Ca(2+) release events (Ca(2+) sparks) were studied in atrial and ventricular myocytes with ultra-fast confocal microscopy using a Zeiss LSM 5 LIVE system that allows sampling rates of up to 60 kHz. Ca(2+) sparks which originated from subsarcolemmal junctional sarcoplasmic reticulum (j-SR) release sites in atrial myocytes were anisotropic and elongated in the longitudinal direction of the cell. Ca(2+) sparks in atrial cells originating from non-junctional SR and in ventricular myocytes were symmetrical. Ca(2+) spark recording in line scan mode at 40,000 lines/s uncovered step-like increases of [Ca(2+)]i. 2-D imaging of Ca(2+) transients revealed an asynchronous activation of release sites and allowed the sequential recording of Ca(2+) entry through surface membrane Ca(2+) channels and subsequent activation of Ca(2+)-induced Ca(2+) release. With a latency of 2.5 ms after application of an electrical stimulus, Ca(2+) entry could be detected that was followed by SR Ca(2+) release after an additional 3 ms delay. Maximum Ca(2+) release was observed 4 ms after the beginning of release. The timing of Ca(2+) entry and release was confirmed by simultaneous [Ca(2+)]i and membrane current measurements using the whole cell voltage-clamp technique. In atrial cells activation of discrete individual release sites of the j-SR led to spatially restricted Ca(2+) release events that fused into a peripheral ring of elevated [Ca(2+)]i that subsequently propagated in a wave-like fashion towards the center of the cell. In ventricular myocytes asynchronous Ca(2+) release signals from discrete sites with no preferential subcellular location preceded the whole-cell Ca(2+) transient. In summary, ultra-fast confocal imaging allows investigation of Ca(2+) signals with a time resolution similar to patch clamp technique, however in a less invasive fashion.

  6. Influence of the hydrostatic pulpal pressure on droplets formation in current etch-and-rinse and self-etch adhesives: a video rate/TSM microscopy and fluid filtration study.

    Science.gov (United States)

    Sauro, Salvatore; Mannocci, Francesco; Toledano, Manuel; Osorio, Raquel; Thompson, Ian; Watson, Timothy F

    2009-11-01

    The aim of this study was to investigate the droplet formation using a real-time/confocal microscopy technique, when different self-etching and etch-and-rinse adhesives were applied in the presence or absence of pulpal pressure. Resin-dentin permeability (%P) was also evaluated. Optibond FL, Silorane adhesive, Scotchbond 1XT, G-Bond and DC-Bond were bonded in the presence or in absence of simulated pulpal pressure. A fluid-transport model was used to measure the water permeability through resin-bonded dentin. Half of the specimens bonded in the presence of the hydrostatic pulpal pressure (20 cm H2O) were light cured, whereas the remnant half received no light curing. The same was done with the half of the specimens bonded under no pulpal pressure. The specimens were investigated under a confocal TSM. Optibond FL and G-Bond had the lowest dentin permeability. Optibond FL adhesive showed few water droplets on the polymerized external surface and within the resin-dentin interface. G-Bond showed static interfacial globular-like droplet formation. DC-Bond and Scotchbond 1XT were the most water permeable adhesives both in the presence and in absence of pulpal pressure. A dynamic interfacial non-globular-like droplet formation was observed. Severe droplet formation was observed on the polymerized external surface. The presence of the pulpal pressure may cause increasing in fluid filtration and droplet formation in simplified adhesives containing HEMA. The adhesives containing 4-META (G-Bond) may be affected by static phase separation but by very low osmotic droplets formation and water permeability. The three-step adhesives are less affected by these problems.

  7. Scanning laser Doppler vibrometry

    DEFF Research Database (Denmark)

    Brøns, Marie; Thomsen, Jon Juel

    With a Scanning Laser Doppler Vibrometer (SLDV) a vibrating surface is automatically scanned over predefined grid points, and data processed for displaying vibration properties like mode shapes, natural frequencies, damping ratios, and operational deflection shapes. Our SLDV – a PSV-500H from...

  8. Frequency scanning microstrip antennas

    DEFF Research Database (Denmark)

    Danielsen, Magnus; Jørgensen, Rolf

    1979-01-01

    The principles of using radiating microstrip resonators as elements in a frequency scanning antenna array are described. The resonators are cascade-coupled. This gives a scan of the main lobe due to the phase-shift in the resonator in addition to that created by the transmission line phase...

  9. Stromal wound healing explains refractive instability and haze development after photorefractive keratectomy: a 1-year confocal microscopic study.

    Science.gov (United States)

    Moller-Pedersen, T; Cavanagh, H D; Petroll, W M; Jester, J V

    2000-07-01

    To evaluate the mechanism(s) producing refractive instability and corneal haze development after photorefractive keratectomy (PRK). Prospective, nonrandomized, comparative case series, self-controlled. Seventeen eyes of 17 patients with low- to moderate-grade myopia (-2.88 to -9.13 diopters [D]) were included. Surgical intervention was a standardized, 6-mm diameter PRK procedure using the Meditec MEL 60 excimer laser (Aesculap-Meditec, Heroldsberg, Germany). The photoablation center was evaluated before surgery and at 1, 3, 6, 9, and 12 months after PRK using rapid, continuous z-scans of confocal images, termed confocal microscopy through focusing (CMTF). Simultaneous epithelial and stromal thickness analysis and objective assessment of corneal light backscattering were obtained from digital image analysis of the CMTF scans. Corneal reinnervation and anterior stromal keratocyte density and wound healing morphologic features were evaluated on high resolution, in vivo confocal images. Manifest refraction was measured and corneal clarity was graded by slit-lamp biomicroscopy. Epithelial thickness averaged 45+/-10 microm at 1 month, 50+/-8 microm at 3 months, and 52+/-6 microm at 12 months after PRK, as compared with 51+/-4 microm before surgery, demonstrating complete restoration of the preoperative thickness without compensatory hyperplasia. Interestingly, epithelial rethickening had no significant correlation with refractive regression. By contrast, stromal regrowth (from 1-12 months) averaged 6+/-12 microm (range, 27 microm thinning-22 microm rethickening) and correlated closely (r = 0.84, Pwound healing mechanisms. In agreement with these findings, all "hazy" corneas showed increased numbers of anterior stromal wound healing keratocytes with increased reflectivity of both nuclei and cell bodies, suggesting that cellular-based reflections, as opposed to extracellular matrix deposition, are the major origin of increased corneal light scattering after PRK. Taken

  10. Diagnostic value of probe-based confocal laser endomicroscopy and high-definition virtual chromoendoscopy in early esophageal squamous neoplasia.

    Science.gov (United States)

    Guo, Jing; Li, Chang-Qing; Li, Ming; Zuo, Xiu-Li; Yu, Tao; Liu, Jian-Wei; Liu, Jing; Kou, Guan-Jun; Li, Yan-Qing

    2015-01-01

    Detection and differentiation of esophageal squamous neoplasia (ESN) are of value in improving patient outcomes. Probe-based confocal laser endomicroscopy (pCLE) can serve in targeted biopsies in the diagnosis of GI neoplasia. However, its performance in ESN has not yet been reported. To investigate the diagnostic value of pCLE for early ESN screened by high-definition virtual chromoendoscopy (I-Scan) and verified by Lugol chromoendoscopy and histopathology. Prospective and noninferiority trial. Single center in China. Patients were enrolled who (1) previously had histologically verified early ESN or (2) were about to undergo screening endoscopy and were 50 to 80 years of age between February 2013 and February 2014. The esophagus was investigated sequentially by white-light endoscopy, I-Scan, then pCLE and iodine chromoendoscopy. The results were interpreted and compared with histopathologic results. Diagnostic characteristics of pCLE and I-Scan. In total, 356 patients were enrolled. In all, 42 patients were histologically proven to have 47 neoplasias. The diagnostic value of pCLE for ESN during ongoing endoscopy has a sensitivity, specificity, and accuracy of 94.6%, 90.7%, and 92.3%, respectively. The interobserver and intraobserver agreement was good and excellent, with κ values of 0.699 and 0.895, respectively. The detection rate by using I-Scan and Lugol chromoendoscopy was 10.4% and 12.9%, respectively (P<.01 for noninferiority). Single center. pCLE shows promise in diagnosing and differentiating ESN in vivo. The screening performance of I-Scan in the detection of ESN is noninferior to that of iodine chromoendoscopy. Copyright © 2015. Published by Elsevier Inc.

  11. Characterization of the stem anatomy of the Eocene fern Dennstaedtiopsis aerenchymata (Dennstaedtiaceae) by use of confocal laser scanning microscopy.

    Science.gov (United States)

    Shi, Chris S; Schopf, J William; Kudryavtsev, Anatoliy B

    2013-08-01

    Permineralization provides the most faithful known mode of three-dimensional preservation of the morphology and cellular anatomy of fossil plants. Standard optical microscopic documentation of such structures can provide only an approximation of their true three-dimensional form and is incapable of revealing fine-structural (anatomy of permineralized fossil plants can provide accurate data in two and three dimensions at high spatial resolution, information that can be critically important to taxonomic, taphonomic, and developmental interpretations. Results presented here from this first detailed CLSM-based study of permineralized plant axes indicate that this nonintrusive, nondestructive technique should be widely applicable in paleobotany.

  12. Real-Time Demonstration of Split Skin Graft Inosculation and Integra Dermal Matrix Neovascularization Using Confocal Laser Scanning Microscopy

    OpenAIRE

    Greenwood, John; Amjadi, Mahyar; Dearman, Bronwyn; Mackie, Ian

    2009-01-01

    Objectives: During the first 48 hours after placement, an autograft ?drinks? nutrients and dissolved oxygen from fluid exuding from the underlying recipient bed (?plasmatic imbibition?). The theory of inosculation (that skin grafts subsequently obtain nourishment via blood vessel ?anastomosis? between new vessels invading from the wound bed and existing graft vessels) was hotly debated from the late 19th to mid-20th century. This study aimed to noninvasively observe blood flow in split skin g...

  13. Contribution to the age determination of fingerprint constituents using laser fluorescence spectroscopy and confocal laser scanning microscopy

    Science.gov (United States)

    Barcikowski, Stephan; Bunte, Jens; Ostendorf, Andreas; Aehnlich, Joerg; Herrmann, Rainer

    2004-12-01

    The development of an instrument measuring the age of a fingerprint could raise the effectiveness and reliability of the dactyloscopy and lower the expenses of unnecessary fingerprint analysis. The purpose of the presented investigation is to determine the compounds in a fingerprint residue, which can be used as a time relevant indicator. The identification of the information carrier inside the secreted substance shall determine the main components, which allow age-related statements. A multiphoton excitation is used to determine the age without degenerating or changing samples in any way, so that the evidence is usable for further examination and documentation. Non-invasive fluorescence analysis on the basis of the surface concentration of fingerprint components is presented.

  14. Influence of nail varnish on the remineralization of enamel single sections assessed by microradiography and confocal laser scanning microscopy

    NARCIS (Netherlands)

    Iijima, Y; Takagi, O; Duschner, H; Ruben, J; Arends, J

    1998-01-01

    Single-section techniques are attractive in enamel de- and remineralization investigations because they allow longitudinal studies in which mineral changes can be assessed by microradiography (TMR), Nail Varnish (NV) is in general applied to coat the cut thin-section sides, The aims of this study

  15. The simplicity of males: Dwarf males of four species of Osedax (Siboglinidae; Annelida) investigated by confocal laser scanning microscopy

    DEFF Research Database (Denmark)

    Worsaae, Katrine; Rouse, Greg W

    2010-01-01

    Dwarf males of the bone-eating worms Osedax (Siboglinidae, Annelida) have been proposed to develop from larvae that settle on females rather than on bone. The apparent arrest in somatic development and resemblance of the males to trochophore larvae has been posited as an example of paedomorphosis...

  16. Musculature of Notholca acuminata (Rotifera: Ploima: Brachionidae) revealed by confocal scanning laser microscopy

    DEFF Research Database (Denmark)

    Sørensen, M.V.; Funch, P.; Hooge, M.

    2003-01-01

    , stomach, gut, and cloaca, including thin longitudinal gut fibers and viscero-cloacal fibers, never before reported in other species of rotifers. The dorsal, lateral, and ventral retractor muscles and the incomplete circular muscles associated with the body wall appear to be apomorphies for the Rotifera....... Muscle-revealing staining shows promise for providing additional information on previously unrecognized complexity in rotifer musculature that will be useful in functional morphology and phylogenetic analyses...

  17. Musculature of Notholca acuminata (Rotifera : Ploima : Brachionidae) revealed by confocal scanning laser microscopy

    DEFF Research Database (Denmark)

    Sørensen, M.V.; Funch, P.; Hooge, M.

    2003-01-01

    , stomach, gut, and cloaca, including thin longitudinal gut fibers and viscero-cloacal fibers, never before reported in other species of rotifers. The dorsal, lateral, and ventral retractor muscles and the incomplete circular muscles associated with the body wall appear to be apomorphies for the Rotifera....... Muscle-revealing staining shows promise for providing additional information on previously unrecognized complexity in rotifer musculature that will be useful in functional morphology and phylogenetic analyses...

  18. Imunolocalization of delta-giardin within the ventral disc of Giardia duodenalis using laser scanning confocal microscopy

    Science.gov (United States)

    Giardia duodenalis is a ubiquitous protozoan parasite that colonizes the upper small intestine of humans and animals causing diarrheal disease. To maintain infection within the small intestine, trophozoites (the replicative stage of the parasite) attach to the epithelial layer of the gut and resist ...

  19. The Use of Intravital Two-Photon and Thick Section Confocal Imaging to Analyze B Lymphocyte Trafficking in Lymph Nodes and Spleen.

    Science.gov (United States)

    Park, Chung; Hwang, Il-Young; Kehrl, John H

    2018-01-01

    Intravital two-photon laser scanning microscopy (TP-LSM) has allowed the direct observation of immune cells in intact organs of living animals. In the B cell biology field TP-LSM has detailed the movement of B cells in high endothelial venules and during their transmigration into lymph organs; described the movement and positioning of B cells within lymphoid organs; outlined the mechanisms by which antigen is delivered to B cells; observed B cell interacting with T cells, other cell types, and even with pathogens; and delineated the egress of B cells from the lymph node (LN) parenchyma into the efferent lymphatics. As the quality of TP-LSM improves and as new fluorescent probes become available additional insights into B cell behavior and function await new investigations. Yet intravital TP-LSM has some disadvantages including a lower resolution than standard confocal microscopy, a narrow imaging window, and a shallow depth of imaging. We have found that supplementing intravital TP-LSM with conventional confocal microscopy using thick LN sections helps to overcome some of these shortcomings. Here, we describe procedures for visualizing the behavior and trafficking of fluorescently labeled, adoptively transferred antigen-activated B cells within the inguinal LN of live mice using two-photon microscopy. Also, we introduce procedures for fixed thick section imaging using standard confocal microscopy, which allows imaging of fluorescently labeled cells deep in the LN cortex and in the spleen with high resolution.

  20. Confocal Imaging of Neuropeptide Y-pHluorin: A Technique to Visualize Insulin Granule Exocytosis in Intact Murine and Human Islets.

    Science.gov (United States)

    Makhmutova, Madina; Liang, Tao; Gaisano, Herbert; Caicedo, Alejandro; Almaça, Joana

    2017-09-13

    Insulin secretion plays a central role in glucose homeostasis under normal physiological conditions as well as in disease. Current approaches to study insulin granule exocytosis either use electrophysiology or microscopy coupled to the expression of fluorescent reporters. However most of these techniques have been optimized for clonal cell lines or require dissociating pancreatic islets. In contrast, the method presented here allows for real time visualization of insulin granule exocytosis in intact pancreatic islets. In this protocol, we first describe the viral infection of isolated pancreatic islets with adenovirus that encodes a pH-sensitive green fluorescent protein (GFP), pHluorin, coupled to neuropeptide Y (NPY). Second, we describe the confocal imaging of islets five days after viral infection and how to monitor the insulin granule secretion. Briefly, the infected islets are placed on a coverslip on an imaging chamber and imaged under an upright laser-scanning confocal microscope while being continuously perfused with extracellular solution containing various stimuli. Confocal images spanning 50 µm of the islet are acquired as time-lapse recordings using a fast-resonant scanner. The fusion of insulin granules with the plasma membrane can be followed over time. This procedure also allows for testing a battery of stimuli in a single experiment, is compatible with both mouse and human islets, and can be combined with various dyes for functional imaging (e.g., membrane potential or cytosolic calcium dyes).

  1. Laser Scanning in Forests

    Directory of Open Access Journals (Sweden)

    Håkan Olsson

    2012-09-01

    Full Text Available The introduction of Airborne Laser Scanning (ALS to forests has been revolutionary during the last decade. This development was facilitated by combining earlier ranging lidar discoveries [1–5], with experience obtained from full-waveform ranging radar [6,7] to new airborne laser scanning systems which had components such as a GNSS receiver (Global Navigation Satellite System, IMU (Inertial Measurement Unit and a scanning mechanism. Since the first commercial ALS in 1994, new ALS-based forest inventory approaches have been reported feasible for operational activities [8–12]. ALS is currently operationally applied for stand level forest inventories, for example, in Nordic countries. In Finland alone, the adoption of ALS for forest data collection has led to an annual savings of around 20 M€/year, and the work is mainly done by companies instead of governmental organizations. In spite of the long implementation times and there being a limited tradition of making changes in the forest sector, laser scanning was commercially and operationally applied after about only one decade of research. When analyzing high-ranked journal papers from ISI Web of Science, the topic of laser scanning of forests has been the driving force for the whole laser scanning research society over the last decade. Thus, the topic “laser scanning in forests” has provided a significant industrial, societal and scientific impact. [...

  2. Confocal reader for biochip screening and fluorescence microscopy.

    Science.gov (United States)

    Ruckstuhl, Thomas; Walser, Andreas; Verdes, Dorinel; Seeger, Stefan

    2005-03-15

    We developed a fluorescence reader for the sensitive detection of surface-generated fluorescence. The system is applicable for high resolution imaging as well as for the readout of large biochips. The surface of a microscope coverslip is scanned with a laser beam focused to a waist diameter of 500 nm (FWHM) by means of a single aspheric lens. Scanning large areas with a focused beam usually evokes the need of automatic control elements to adjust the laser spot to the designated position at the surface. Due to the special design of the reader, the focus keeps at the plane of the surface even when scanning large areas, obviating the requirement of any real time control. Thus the instrument is straightforward and inexpensive. Nevertheless it features a high sensitivity and high optical resolution. The versatility of the instrument is demonstrated by imaging cells and reading out a DNA-chip. The excellent sensitivity is shown by detecting single fluorescently labeled antibodies.

  3. CRAFT: Multimodality confocal skin imaging for early cancer diagnosis.

    Science.gov (United States)

    Peng, Tong; Xie, Hao; Ding, Yichen; Wang, Weichao; Li, Zhiming; Jin, Dayong; Tang, Yuanhe; Ren, Qiushi; Xi, Peng

    2012-05-01

    Although histological analysis serves as a gold standard to cancer diagnosis, its application on skin cancer detection is largely prohibited due to its invasive nature. To obtain both the structural and pathological information in situ, a Confocal Reflectance/Auto-Fluorescence Tomography (CRAFT) system was established to examine the skin sites in vivo with both reflectance and autofluorescence modes simultaneously. Nude mice skin with cancerous sites and normal skin sites were imaged and compared with the system. The cellular density and reflective intensity in cancerous sites reflects the structural change of the tissue. With the decay coefficient analysis, the corresponding NAD(P)H decay index for cancerous sites is 1.65-fold that of normal sites, leading to a 97.8% of sensitivity and specificity for early cancer diagnosis. The results are verified by the followed histological analysis. Therefore, CRAFT may provide a novel method for the in vivo, non-invasive diagnosis of early cancer. Copyright © 2012 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  4. Confocal imaging to quantify passive transport across biomimetic lipid membranes.

    Science.gov (United States)

    Li, Su; Hu, Peichi; Malmstadt, Noah

    2010-09-15

    The ability of a molecule to pass through the plasma membrane without the aid of any active cellular mechanisms is central to that molecule's pharmaceutical characteristics. Passive transport has been understood in the context of Overton's rule, which states that more lipophilic molecules cross membrane lipid bilayers more readily. Existing techniques for measuring passive transport lack reproducibility and are hampered by the presence of an unstirred layer (USL) that dominates transport across the bilayer. This report describes assays based on spinning-disk confocal microscopy (SDCM) of giant unilamellar vesicles (GUVs) that allow for the detailed investigation of passive transport processes and mechanisms. This approach allows the concentration field to be directly observed, allowing membrane permeability to be determined easily from the transient concentration profile data. A series of molecules of increasing hydrophilicity was constructed, and the transport of these molecules into GUVs was observed. The observed permeability trend is consistent with Overton's rule. However, the values measured depart from the simple partition-diffusion proportionality model of passive transport. This technique is easy to implement and has great promise as an approach to measure membrane transport. It is optimally suited to precise quantitative measurements of the dependence of passive transport on membrane properties.

  5. A histologic evaluation of retrieved hydroxyapatite-coated blade-form implants using scanning electron, light, and confocal laser scanning microscopies.

    Science.gov (United States)

    Takeshita, F; Ayukawa, Y; Iyama, S; Suetsugu, T; Kido, M A

    1996-10-01

    We histologically examined seven hydroxyapatite-coated (HA) blade implants removed from patients. Four of them radiologically showed severe bone loss and were easily removed with an elevator. Three radiologically showed vertical bone loss and were removed by surgical procedure. Our histological evaluation indicated that coating separation from the HA implants had occurred, and HA coating resorption by bone tissues was suspected in an implant left in situ for 8 years. Several multinucleated giant cells were seen with a few released particles of HA coating at the point lacking bone contact with the HA coating. The presence of microorganisms on and in the HA coating layer was also noted.

  6. Evaluation of Enterococcus faecalis adhesion, penetration, and method to prevent the penetration of Enterococcus faecalis into root cementum: Confocal laser scanning microscope and scanning electron microscope analysis

    OpenAIRE

    Rahul S Halkai; Hegde, Mithra N; Halkai, Kiran R.

    2016-01-01

    Aim: To ascertain the role of Enterococcus faecalis in persistent infection and a possible method to prevent the penetration of E. faecalis into root cementum. Methodology: One hundred and twenty human single-rooted extracted teeth divided into five groups. Group I (control): intact teeth, Group II: no apical treatment done, Group III divided into two subgroups. In Groups IIIa and IIIb, root apex treated with lactic acid of acidic and neutral pH, respectively. Group IV: apical root cementu...

  7. Feasibility and reliability of pancreatic cancer staging using a new confocal non-fluorescent microscopy probe: a double-blind study in rats.

    Science.gov (United States)

    Akladios, Cherif; De Ruijter, Vivian; Perretta, Sylvana; Aprahamian, Marc; Ignat, Mihaela; Lindner, Veronique; Averous, Gerlinde; Dallemagne, Bernard; Marescaux, Jacques

    2017-02-01

    Surgical management of pancreatic cancer depends on tumor resectability and staging. Lymph node (LN) metastases represent an important decision-making factor when it comes to surgical treatment. To evaluate a new in vivo, endoscopic confocal microscopy (CM) system not requiring fluorescence markers, for detection and staging of pancreatic cancer in rats. A confocal system consisting of a confocal scanning laser operating in reflection mode and a dedicated rigid Hopkins rod-lens endoscope were used for in vivo imaging in a rat model of pancreatic ductal adenocarcinoma. A double-blind study compared CM to standard histology in (1) the detection of tumors in rat bearing cancer (n = 11) and controls (n = 6), and (2) in the detection of local nodal involvement at 3 and 6 weeks after tumor induction. CM detected all pancreatic tumors with 100 % sensitivity and specificity and identified 15 metastatic LNs with an average adenocarcinoma nodule diameter of 2.3 mm (range from 1 to 4.2 mm) out of the 66 examined. CM demonstrated a sensitivity of 87.5 % and a specificity of 98 % in LN detection. The Spearman's rank correlation/rho calculator was of 0.87. CM demonstrated a negative predictive value of 96.1 % and a positive predictive value of 93.3 % in the detection of metastatic LNs. Interpretation of confocal images has a high concurrence rate with histopathology examination for primary tumor and lymphatic involvement detection making it a promising technique for in vivo real-time detection and staging of pancreatic cancer. Larger studies are warranted to confirm these preliminary results.

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

    Science.gov (United States)

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

    2014-10-01

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

  9. Investigations in optoelectronic image processing in scanning laser microscopy

    Science.gov (United States)

    Chaliha, Hiranya Kumar

    A considerable amount of work has been done on scann-ing laser microscopy since its applications were first pointed out by Roberts and Young[1], Minsky [2] and Davidovits et al [3]. The advent of laser has made it possible to focus an intense beam of laser light in a scanning optical microscope (SOM) [4, 5] and hence explore regions of microscopy[6] uncovered by conven-tional microscopy. In the simple SOM [7, 8, 9], the upper spatial frequency in amplitude transmittance or reflectance of an object for which transfer function is nonzero is same as that in a conventional optical microscope. However, in Type II SOM [7] or confocal SOM that employs a coherent or a point detector, the spatial frequency bandwidth is twice that obtained in a conventional microscope. Besides this confocal set-up is found to be very useful in optical sectioning and consequently in 3-D image processing[10, 11, 12] specially of biological specimens. Such systems are also suitable for studies of semiconductor materials [13], super-resolution [14] and various imaginative ways of image processing[15, 16, 17] including phase imaging[18]. A brief survey of related advances in scanning optical microscopy has been covered in the chapter 1 of the thesis. The performance of SOM may be investigated by concent-rating also on signal derived by one dimensional scan of the object specimen. This simplified mode may also be adapted to give wealth of information for biological and semiconductor specimens. Hence we have investigated the design of a scanning laser system suited specifically for studies of line scan image signals of microscopic specimens when probed through a focused laser spot. An electro-mechanical method of scanning of the object specimen has been designed with this aim in mind. Chapter 2, Part A of the thesis deals with the design consider-ations of such a system. For analysis of scan signals at a later instant of time so as to facilitate further processing, an arrangement of microprocessor

  10. Thyroid Scan and Uptake

    Medline Plus

    Full Text Available ... energy. top of page What are some common uses of the procedure? The thyroid scan is used ... gland evaluate changes in the gland following medication use, surgery, radiotherapy or chemotherapy top of page How ...

  11. Thyroid Scan and Uptake

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    Full Text Available ... nuclear medicine procedures are able to pinpoint molecular activity within the body, they offer the potential to ... tells you otherwise, you may resume your normal activities after your nuclear medicine scan. If any special ...

  12. Body CT (CAT Scan)

    Science.gov (United States)

    ... Professions Site Index A-Z Computed Tomography (CT) - Body Computed tomography (CT) of the body uses special ... the Body? What is CT Scanning of the Body? Computed tomography, more commonly known as a CT ...

  13. Thyroid Scan and Uptake

    Medline Plus

    Full Text Available ... scan and uptake uses small amounts of radioactive materials called radiotracers, a special camera and a computer ... last two months that used iodine-based contrast material. Your doctor will instruct you on how to ...

  14. Thyroid Scan and Uptake

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    Full Text Available ... evaluate changes in the gland following medication use, surgery, radiotherapy or chemotherapy top of page How should ... such as an x-ray or CT scan, surgeries or treatments using iodinated contrast material within the ...

  15. Thyroid Scan and Uptake

    Medline Plus

    Full Text Available ... imaging procedures. For many diseases, nuclear medicine scans yield the most useful information needed to make a ... any. Nuclear medicine is less expensive and may yield more precise information than exploratory surgery. Risks Because ...

  16. Thyroid Scan and Uptake

    Medline Plus

    Full Text Available ... Actual scanning time for each thyroid uptake is five minutes or less. top of page What will ... diagnostic procedures have been used for more than five decades, and there are no known long-term ...

  17. Thyroid Scan and Uptake

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    Full Text Available ... A thyroid scan is a type of nuclear medicine imaging. The radioactive iodine uptake test (RAIU) is ... thyroid function, but does not involve imaging. Nuclear medicine is a branch of medical imaging that uses ...

  18. Thyroid Scan and Uptake

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    Full Text Available ... several hours before your exam because eating can affect the accuracy of the uptake measurement. Jewelry and ... often unattainable using other imaging procedures. For many diseases, nuclear medicine scans yield the most useful information ...

  19. Thyroid Scan and Uptake

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    Full Text Available ... iodine, medications and anesthetics. are breastfeeding. In the days prior to your examination, blood tests may be ... are scheduled for an additional procedure that same day that requires an intravenous line. Actual scanning time ...

  20. Thyroid Scan and Uptake

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    Full Text Available ... gland in the neck that controls metabolism , a chemical process that regulates the rate at which the body converts food to energy. top of page What are some common uses of the procedure? The thyroid scan is ...

  1. Thyroid Scan and Uptake

    Medline Plus

    Full Text Available ... liquid or capsule form, it is typically swallowed up to 24 hours before the scan. The radiotracer given by intravenous injection is usually given up to 30 minutes prior to the test. When ...

  2. Thyroid Scan and Uptake

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    Full Text Available ... regulates the rate at which the body converts food to energy. top of page What are some ... often unattainable using other imaging procedures. For many diseases, nuclear medicine scans yield the most useful information ...

  3. Thyroid Scan and Uptake

    Medline Plus

    Full Text Available ... Because nuclear medicine procedures are able to pinpoint molecular activity within the body, they offer the potential ... or imaging device that produces pictures and provides molecular information. The thyroid scan and thyroid uptake provide ...

  4. Thyroid Scan and Uptake

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    Full Text Available ... body. top of page How does the procedure work? With ordinary x-ray examinations, an image is ... with other imaging techniques, such as CT or MRI. However, nuclear medicine scans are more sensitive than ...

  5. Thyroid Scan and Uptake

    Medline Plus

    Full Text Available ... a special camera or imaging device that produces pictures and provides molecular information. The thyroid scan and ... and with the help of a computer, create pictures offering details on both the structure and function ...

  6. Pediatric CT Scans

    Science.gov (United States)

    The Radiation Epidemiology Branch and collaborators have initiated a retrospective cohort study to evaluate the relationship between radiation exposure from CT scans conducted during childhood and adolescence and the subsequent development of cancer.

  7. Thyroid Scan and Uptake

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    Full Text Available ... abnormal was found, and should not be a cause of concern for you. If you had an ... abnormal was found, and should not be a cause of concern for you. Actual scanning time for ...

  8. Scanning Auger Electron Microscope

    Data.gov (United States)

    Federal Laboratory Consortium — A JEOL model 7830F field emission source, scanning Auger microscope.Specifications / Capabilities:Ultra-high vacuum (UHV), electron gun range from 0.1 kV to 25 kV,...

  9. Thyroid Scan and Uptake

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    Full Text Available ... scan and thyroid uptake provide information about the structure and function of the thyroid. The thyroid is ... computer, create pictures offering details on both the structure and function of organs and tissues in your ...

  10. The Scanning Optical Microscope.

    Science.gov (United States)

    Sheppard, C. J. R.

    1978-01-01

    Describes the principle of the scanning optical microscope and explains its advantages over the conventional microscope in the improvement of resolution and contrast, as well as the possibility of producing a picture from optical harmonies generated within the specimen.

  11. Thyroid Scan and Uptake

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    Full Text Available ... scan you are undergoing. top of page What does the equipment look like? The special camera and ... area of your body. top of page How does the procedure work? With ordinary x-ray examinations, ...

  12. Thyroid Scan and Uptake

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    Full Text Available ... eat for several hours before your exam because eating can affect the accuracy of the uptake measurement. ... often unattainable using other imaging procedures. For many diseases, nuclear medicine scans yield the most useful information ...

  13. Thyroid Scan and Uptake

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    Full Text Available ... found, and should not be a cause of concern for you. If you had an intravenous line ... found, and should not be a cause of concern for you. Actual scanning time for each thyroid ...

  14. Slow Scan Telemedicine

    Science.gov (United States)

    1984-01-01

    Originally developed under contract for NASA by Ball Bros. Research Corporation for acquiring visual information from lunar and planetary spacecraft, system uses standard closed circuit camera connected to a device called a scan converter, which slows the stream of images to match an audio circuit, such as a telephone line. Transmitted to its destination, the image is reconverted by another scan converter and displayed on a monitor. In addition to assist scans, technique allows transmission of x-rays, nuclear scans, ultrasonic imagery, thermograms, electrocardiograms or live views of patient. Also allows conferencing and consultation among medical centers, general practitioners, specialists and disease control centers. Commercialized by Colorado Video, Inc., major employment is in business and industry for teleconferencing, cable TV news, transmission of scientific/engineering data, security, information retrieval, insurance claim adjustment, instructional programs, and remote viewing of advertising layouts, real estate, construction sites or products.

  15. Thyroid Scan and Uptake

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    Full Text Available ... of page What are some common uses of the procedure? The thyroid scan is used to determine ... you are undergoing. top of page What does the equipment look like? The special camera and imaging ...

  16. Thyroid Scan and Uptake

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    Full Text Available ... to identify disease in its earliest stages as well as a patient’s immediate response to therapeutic interventions. ... but is often performed on hospitalized patients as well. Thyroid Scan You will be positioned on an ...

  17. Thyroid Scan and Uptake

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    Full Text Available ... Uptake? A thyroid scan is a type of nuclear medicine imaging. The radioactive iodine uptake test (RAIU) ... of thyroid function, but does not involve imaging. Nuclear medicine is a branch of medical imaging that ...

  18. Thyroid Scan and Uptake

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    Full Text Available ... medical tests that help physicians diagnose and evaluate medical conditions. These imaging scans use radioactive materials called radiopharmaceuticals or radiotracers . Depending on the type of nuclear medicine exam, the radiotracer is either injected into the body, ...

  19. Thyroid Scan and Uptake

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    Full Text Available ... many types of cancers, heart disease, gastrointestinal, endocrine, neurological disorders and other abnormalities within the body. Because ... with other imaging techniques, such as CT or MRI. However, nuclear medicine scans are more sensitive than ...

  20. Thyroid Scan and Uptake

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    Full Text Available ... are noninvasive and, with the exception of intravenous injections, are usually painless medical tests that help physicians ... before the scan. The radiotracer given by intravenous injection is usually given up to 30 minutes prior ...