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Sample records for combined confocal laser

  1. Extended Field Laser Confocal Microscopy (EFLCM): Combining automated Gigapixel image capture with in silico virtual microscopy

    PubMed Central

    Flaberg, Emilie; Sabelström, Per; Strandh, Christer; Szekely, Laszlo

    2008-01-01

    Background Confocal laser scanning microscopy has revolutionized cell biology. However, the technique has major limitations in speed and sensitivity due to the fact that a single laser beam scans the sample, allowing only a few microseconds signal collection for each pixel. This limitation has been overcome by the introduction of parallel beam illumination techniques in combination with cold CCD camera based image capture. Methods Using the combination of microlens enhanced Nipkow spinning disc confocal illumination together with fully automated image capture and large scale in silico image processing we have developed a system allowing the acquisition, presentation and analysis of maximum resolution confocal panorama images of several Gigapixel size. We call the method Extended Field Laser Confocal Microscopy (EFLCM). Results We show using the EFLCM technique that it is possible to create a continuous confocal multi-colour mosaic from thousands of individually captured images. EFLCM can digitize and analyze histological slides, sections of entire rodent organ and full size embryos. It can also record hundreds of thousands cultured cells at multiple wavelength in single event or time-lapse fashion on fixed slides, in live cell imaging chambers or microtiter plates. Conclusion The observer independent image capture of EFLCM allows quantitative measurements of fluorescence intensities and morphological parameters on a large number of cells. EFLCM therefore bridges the gap between the mainly illustrative fluorescence microscopy and purely quantitative flow cytometry. EFLCM can also be used as high content analysis (HCA) instrument for automated screening processes. PMID:18627634

  2. Combining microtomy and confocal laser scanning microscopy for structural analyses of plant-fungus associations.

    PubMed

    Rath, Magnus; Grolig, Franz; Haueisen, Janine; Imhof, Stephan

    2014-05-01

    The serious problem of extended tissue thickness in the analysis of plant-fungus associations was overcome using a new method that combines physical and optical sectioning of the resin-embedded sample by microtomy and confocal microscopy. Improved tissue infiltration of the fungal-specific, high molecular weight fluorescent probe wheat germ agglutinin conjugated to Alexa Fluor® 633 resulted in high fungus-specific fluorescence even in deeper tissue sections. If autofluorescence was insufficient, additional counterstaining with Calcofluor White M2R or propidium iodide was applied in order to visualise the host plant tissues. Alternatively, the non-specific fluorochrome acid fuchsine was used for rapid staining of both, the plant and the fungal cells. The intricate spatial arrangements of the plant and fungal cells were preserved by immobilization in the hydrophilic resin Unicryl™. Microtomy was used to section the resin-embedded roots or leaves until the desired plane was reached. The data sets generated by confocal laser scanning microscopy of the remaining resin stubs allowed the precise spatial reconstruction of complex structures in the plant-fungus associations of interest. This approach was successfully tested on tissues from ectomycorrhiza (Betula pendula), arbuscular mycorrhiza (Galium aparine; Polygala paniculata, Polygala rupestris), ericoid mycorrhiza (Calluna vulgaris), orchid mycorrhiza (Limodorum abortivum, Serapias parviflora) and on one leaf-fungus association (Zymoseptoria tritici on Triticum aestivum). The method provides an efficient visualisation protocol applicable with a wide range of plant-fungus symbioses. PMID:24249491

  3. Mobile connected dermatoscope and confocal laser scanning microscope: a useful combination applied in facial simple sensitive skin.

    PubMed

    Zha, W F; Song, W M; Ai, J J; Xu, A E

    2012-08-01

    Little is known as the effects of mobile connected dermatoscope services on diagnostic accuracy for sensitive skin. Confocal laser scanning microscope (CLSM) can non-invasively measure the thickness of epidermis. Combination of the two devices to observe sensitive skin may receive unexpected effects. To evaluate the application effect on sensitive skin with the combination of Handyscope and confocal laser scanning microscope. Twenty simple sensitive-skinned patients and 20 volunteers participated in the study. Cheek, typically, dermoscopic images were obtained from patients, and the changes in the skin texture were observed. Their epidermis thicknesses as well as the volunteers' were measured so that the thicknesses of the two groups were compared. Dermoscopic pictures of the skin texture obviously showed that dilated capillaries looked like earthworms with pigmented patches more or less floating above, and skin roughness as well as deepened dermatoglyph were also conspicuously present in some patients. The mean epidermal thickness of the patients was 79.01 μm and the volunteers' was 85.78 μm. The difference between the two groups reached 6.77 μm. There was a statistical significance (P = 0.001). Mobile connected dermatoscope and confocal laser scanning microscope might be the choice for simple sensitive skin investigation. PMID:22515509

  4. Development of a confocal laser scanning fluorescence microscope using two-photon excitation in combination with time-gated detection

    NASA Astrophysics Data System (ADS)

    Sytsma, Joost; Vroom, Jurrien; Gerritsen, Hans C.; Levine, Yehudi K.

    1995-03-01

    Fluorescent molecules having single-photon absorption in the blue and the UV can be excited with infra-red light via a process known as two-photon excitation. The combination of this technique with scanning techniques can be exploited for 3D microscopic imaging. The two- photon process is confined to a restricted volume in the sample determined by the laser focus, resulting in inherent confocality. Other advantages are reduced photo-bleaching of the samples and a larger penetration depth of the excitation light. The implementation of time-gated detection techniques allows fluorescent lifetime imaging. This drastically improves the selectivity and contrast of the images.

  5. Laser differential confocal radius measurement.

    PubMed

    Zhao, Weiqian; Sun, Ruoduan; Qiu, Lirong; Sha, Dingguo

    2010-02-01

    A new laser differential confocal radius measurement (DCRM) is proposed for high precision measurement of radius. Based on the property of an axial intensity curve that the absolute zero precisely corresponds to the focus of the objective in a differential confocal system (DCS), DCRM uses the zero point of the DCS axial intensity curve to precisely identify the cat's-eye and confocal positions of the test lens, and measures the accurate distance between the two positions to achieve the high-precision measurement of radius of curvature (ROC). In comparison with the existing measurement methods, DCRM proposed has a high measurement precision, a strong environmental anti-interference capability and a low cost. The theoretical analyses and preliminary experimental results indicate that DCRM has a relative measurement error of better than 5 ppm. PMID:20174065

  6. Spectrally encoded confocal scanning laser ophthalmoscope

    NASA Astrophysics Data System (ADS)

    Tao, Yuankai K.; Izatt, Joseph A.

    2010-02-01

    Fundus imaging has become an essential clinical diagnostic tool in ophthalmology. Current generation scanning laser ophthalmoscopes (SLO) offer advantages over conventional fundus photography and indirect ophthalmoscopy in terms of light efficiency and contrast. As a result of the ability of SLO to provide rapid, continuous imaging of retinal structures and its versatility in accommodating a variety of illumination wavelengths, allowing for imaging of both endogenous and exogenous fluorescent contrast agents, SLO has become a powerful tool for the characterization of retinal pathologies. However, common implementations of SLO, such as the confocal scanning laser ophthalmoscope (CSLO) and line-scanning laser ophthalmoscope (LSLO), require imaging or multidimensional scanning elements which are typically implemented in bulk optics placed close to the subject eye. Here, we apply a spectral encoding technique in one dimension combined with single-axis lateral scanning to create a spectrally encoded confocal scanning laser ophthalmoscope (SECSLO) which is fully confocal. This novel implementation of the SLO allows for high contrast, high resolution in vivo human retinal imaging with image transmission through a single-mode optical fiber. Furthermore, the scanning optics are similar and the detection engine is identical to that of current-generation spectral domain optical coherence tomography (SDOCT) systems, potentially allowing for a simplistic implementation of a joint SECSLO-SDOCT imaging system.

  7. A handheld laser scanning confocal reflectance imaging–confocal Raman microspectroscopy system

    PubMed Central

    Patil, Chetan A.; Arrasmith, Christopher L.; Mackanos, Mark A.; Dickensheets, David L.; Mahadevan-Jansen, Anita

    2012-01-01

    Confocal reflectance microscopy and confocal Raman spectroscopy have shown potential for non-destructive analysis of samples at micron-scale resolutions. Current studies utilizing these techniques often employ large bench-top microscopes, and are not suited for use outside of laboratory settings. We have developed a microscope which combines laser scanning confocal reflectance imaging and confocal Raman spectroscopy into a compact handheld probe that is capable of high-resolution imaging and spectroscopy in a variety of settings. The compact size of the probe is largely due to the use of a MEMS mirror for beam scanning. The probe is capable of axial resolutions of up to 4 μm for the confocal imaging channel and 10 μm for the confocal Raman spectroscopy channel. Here, we report instrument design, characterize optical performance, and provide images and spectra from normal skin to demonstrate the instrument’s capabilities for clinical diagnostics. PMID:22435097

  8. CONFOCAL MICROSCOPY SYSTEM PERFORMANCE: LASER POWER MEASUREMENTS

    EPA Science Inventory

    Laser power abstract
    The reliability of the confocal laser-scanning microscope (CLSM) to obtain intensity measurements and quantify fluorescence data is dependent on using a correctly aligned machine that contains a stable laser power. The laser power test appears to be one ...

  9. Confocal unstable-resonator semiconductor laser

    NASA Technical Reports Server (NTRS)

    Salzman, J.; Lang, R.; Yariv, A.; Larson, A.

    1986-01-01

    GaAs/GaAlAs heterostructure lasers with a monolithic confocal unstable resonator were demonstrated. The curved mirrors satisfying the confocal condition were fabricated by etching. Close to threshold, the lasers operate in a single lateral mode with a nearly collimated output beam. A single-lobe far-field intensity distribution as narrow as 1.9-deg full width at half maximum was measured.

  10. Confocal scanning beam laser microscope/macroscope: applications in fluorescence

    NASA Astrophysics Data System (ADS)

    Dixon, Arthur E.; Damaskinos, Savvas; Ribes, Alfonso

    1996-03-01

    A new confocal scanning beam laser microscope/macroscope is described that combines the rapid scan of a scanning beam laser microscope with the large specimen capability of a scanning stage microscope. This instrument combines an infinity-corrected confocal scanning laser microscope with a scanning laser macroscope that uses a telecentric f*(Theta) laser scan lens to produce a confocal imaging system with a resolution of 0.25 microns at a field of view of 25 microns and 5 microns at a field of view of 75,000 microns. The frame rate is 5 seconds per frame for a 512 by 512 pixel image, and 25 seconds for a 2048 by 2048 pixel image. Applications in fluorescence are discussed that focus on two important advantages of the instrument over a confocal scanning laser microscope: an extremely wide range of magnification, and the ability to image very large specimens. Examples are presented of fluorescence and reflected-light images of high quality printing, fluorescence images of latent fingerprints, packaging foam, and confocal autofluorescence images of a cricket.

  11. Confocal Laser Induced Fluorescence of Argon Plasmas

    NASA Astrophysics Data System (ADS)

    Scime, Earl; Soderholm, Mark

    2015-11-01

    Laser Induced Fluorescence (LIF) provides measurements of flow speed, temperature and when absolutely calibrated, density of ions or neutrals in a plasma. Traditionally, laser induced fluorescence requires two ports on a plasma device. One port is used for laser injection and the other is used for fluorescence emission collection. Traditional LIF is tedious and time consuming to align. These difficulties motivate the development of an optical configuration that requires a single port and remains fully aligned at all times; confocal LIF. Our confocal optical design employs a single two inch diameter lens to both inject the laser light and collect the stimulated emission from an argon plasma. A pair of axicon lenses create an annular beam path for the emission collection and the pump laser light is confined inside the annulus of the collection beam. The measurement location is scanned radially by manually adjusting the final focusing lens position. Here we present optical modeling of and initial results from the axicon based confocal optical system. The confocal measurements are compared to traditional, two-port, LIF measurements over the same radial range. This work is supported by US National Science Foundation grant number PHY-1360278.

  12. Combined Confocal and Magnetic Resonance Microscopy

    SciTech Connect

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

    2002-05-12

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

  13. Characterization of Nanoscale Transformations in Polyelectrolyte Multilayers Fabricated from Plasmid DNA Using Laser Scanning Confocal Microscopy in Combination with Atomic Force Microscopy

    PubMed Central

    Fredin, Nathaniel J.; Flessner, Ryan M.; Jewell, Christopher M.; Bechler, Shane L.; Buck, Maren E.; Lynn, David M.

    2010-01-01

    Laser scanning confocal microscopy (LSCM) and atomic force microscopy (AFM) were used to characterize changes in nanoscale structure that occur when ultrathin polyelectrolyte multilayers (PEMs) are incubated in aqueous media. The PEMs investigated here were fabricated by the deposition of alternating layers of plasmid DNA and a hydrolytically degradable polyamine onto a precursor film composed of alternating layers of linear poly(ethylene imine) (LPEI) and sodium poly(styrene sulfonate) (SPS). Past studies of these materials in the context of gene delivery revealed transformations from a morphology that is smooth and uniform to one characterized by the formation of nanometer-scale particulate structures. We demonstrate that in-plane registration of LSCM and AFM images acquired from the same locations of films fabricated using fluorescently labeled polyelectrolytes allows the spatial distribution of individual polyelectrolyte species to be determined relative to the locations of topographic features that form during this transformation. Our results suggest that this physical transformation leads to a morphology consisting of a relatively less disturbed portion of film composed of polyamine and DNA juxtaposed over an array of particulate structures composed predominantly of LPEI and SPS. Characterization by scanning electron microscopy (SEM) and energy-dispersive X-ray (EDX) microanalysis provides additional support for this interpretation. The combination of these different microscopy techniques provides insight into the structures and dynamics of these multicomponent thin films that cannot be achieved using any one method alone, and that could prove useful for the further development of these assemblies as platforms for the surface-mediated delivery of DNA. PMID:20155860

  14. CONFOCAL LASER SCANNING MICROSCOPY OF RAT FOLLICLE DEVELOPMENT

    EPA Science Inventory

    This study used confocal laser scanning microscopy (CLSM) to study follicular development in millimeter pieces of rat ovary. To use this technology, it is essential to stain the tissue before laser excitation with the confocal microscope. Various fluorescent stains (Yo-Pro, Bo-Pr...

  15. Needle-based confocal laser endomicroscopy

    PubMed Central

    Giovannini, Marc

    2015-01-01

    New applications of confocal laser endomicroscopy were developed as pCLE in the bile duct and nCLE for pancreatic cystic tumors, pancreatic masses and lymph nodes. The aim of this paper would be to give you an update in this new technology and to try to define its place in the diagnosis of cystic and solid pancreatic masses. The material used was a 19G EUS-needle in which the stylet was replaced by the Confocal mini-probe. The mini-probe (0.632 mm of diameter) is pre-loaded and screwed by a locking device in the EUS-Needle and guided endosonographically in the target. Regarding pancreatic cystic lesion, the presence of epithelial villous structures based on nCLE was associated with pancreatic cystic neoplasm (IPMN) (P = 0.004) and provided a sensitivity of 59%, specificity of 100%, positive predictive value of 100%, and negative predictive value of 50%. A superficial vascular network pattern visualized on nCLE was identified in serous cystadenomas. It corresponded on pathological specimen to a dense and subepithelial capillary vascularization. The accuracy, sensitivity, specificity, positive predictive value, and negative predictive value of this sign for the diagnosis of SCA were 87%, 69%, 100%, 100%, and 82%, respectively. In pancreatic adenocarcinomas, nCLE found vascular leakage with irregular vessels with leakage of fluorescein into the tumor, large dark clumps which correspond to humps of malignant cells. These criteria correlate with the histological structure of those tumors which are characterized by tumoral glands, surrounded by fibrosis in case of fibrous stroma tumor. Neuroendocrine tumors showed a dense network of small vessels on a dark background, which fits with the histological structure based on cord of cells surrounded by vessels and by fibrosis. nCLE is feasible during a EUS examination; these preliminary results are very encouraging and may be used in the future in case of inconclusive EUS-FNA. PMID:26643694

  16. Optimization of confocal scanning laser ophthalmoscope design

    PubMed Central

    Dhalla, Al-Hafeez; Kelly, Michael P.; Farsiu, Sina; Izatt, Joseph A.

    2013-01-01

    Abstract. Confocal scanning laser ophthalmoscopy (cSLO) enables high-resolution and high-contrast imaging of the retina by employing spatial filtering for scattered light rejection. However, to obtain optimized image quality, one must design the cSLO around scanner technology limitations and minimize the effects of ocular aberrations and imaging artifacts. We describe a cSLO design methodology resulting in a simple, relatively inexpensive, and compact lens-based cSLO design optimized to balance resolution and throughput for a 20-deg field of view (FOV) with minimal imaging artifacts. We tested the imaging capabilities of our cSLO design with an experimental setup from which we obtained fast and high signal-to-noise ratio (SNR) retinal images. At lower FOVs, we were able to visualize parafoveal cone photoreceptors and nerve fiber bundles even without the use of adaptive optics. Through an experiment comparing our optimized cSLO design to a commercial cSLO system, we show that our design demonstrates a significant improvement in both image quality and resolution. PMID:23864013

  17. Probe based confocal laser endomicroscopy of the pancreatobiliary system

    PubMed Central

    Almadi, Majid A; Neumann, Helmut

    2015-01-01

    AIM: To review applications of confocal laser endomicroscopy (CLE) in pancreatobiliary lesions and studies that assessed training and interpretation of images. METHODS: A computerized literature search was performed using OVID MEDLINE, EMBASE, Cochrane library, and the ISI Web of Knowledge from 1980 to October 2014. We also searched abstracts from major meetings that included the Digestive Disease Week, Canadian Digestive Disease Week and the United European Gastroenterology Week using a combination of controlled vocabulary and text words related to pCLE, confocal, endomicroscopy, probe-based confocal laser endomicroscopy, and bile duct to identify reports of trials. In addition, recursive searches and cross-referencing was performed, and manual searches of articles identified after the initial search was also completed. We included fully published articles and those in abstract form. Given the relatively recent introduction of CLE we included randomized trials and cohort studies. RESULTS: In the evaluation of indeterminate pancreatobiliary strictures CLE with ERCP compared to ERCP alone can increase the detection of cancerous strictures with a sensitivity of (98% vs 45%) and has a negative predictive value (97% vs 69%), but decreased the specificity (67% vs 100%) and the positive predictive value (71% vs 100%) when compared to index pathology. Modifications in the classification systems in indeterminate biliary strictures have increased the specificity of pCLE from 67% to 73%. In pancreatic cystic lesions there is a need to develop similar systems to interpret and characterize lesions based on CLE images obtained. The presence of superficial vascular network predicts serous cystadenomas accurately. Also training in acquiring and interpretation of images is feasible in those without any prior knowledge in CLE in a relatively simple manner and computer-aided diagnosis software is a promising innovation. CONCLUSION: The role of pCLE in the evaluation of

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

    PubMed

    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. PMID:26917278

  19. Laser confocal feedback tomography and nano-step height measurement

    PubMed Central

    Tan, Yidong; Wang, Weiping; Xu, Chunxin; Zhang, Shulian

    2013-01-01

    A promising method for tomography and step height measurement is proposed, which combines the high sensitivity of the frequency-shifted feedback laser and the axial positioning ability of confocal microscopy. By demodulating the feedback-induced intensity modulation signals, the obtained amplitude and phase information are used to respectively determine the coarse and fine measurement of the samples. Imaging the micro devices and biological samples by the demodulated amplitude, this approach is proved to be able to achieve the cross-sectional image in highly scattered mediums. And then the successful height measurement of nano-step on a glass-substrate grating by combination of both amplitude and phase information indicates its axial high resolution (better than 2 nm) in a non-ambiguous range of about ten microns. PMID:24145717

  20. Morphological and ultrastructural characterization of ionoregulatory cells in the teleost Oreochromis niloticus following salinity challenge combining complementary confocal scanning laser microscopy and transmission electron microscopy using a novel prefixation immunogold labeling technique.

    PubMed

    Fridman, Sophie; Rana, Krishen J; Bron, James E

    2013-10-01

    Aspects of ionoregulatory or mitochondria-rich cell (MRC) differentiation and adaptation in Nile tilapia yolk-sac larvae following transfer from freshwater to elevated salinities, that is, 12.5 and 20 ppt are described. Investigations using immunohistochemistry on whole-mount Nile tilapia larvae using anti- Na⁺/K⁺-ATPase as a primary antibody and Fluoronanogold™ (Nanoprobes) as a secondary immunoprobe allowed fluorescent labeling with the high resolution of confocal scanning laser microscopy combined with the detection of immunolabeled target molecules at an ultrastructural level using transmission electron microscopy (TEM). It reports, for the first time, various developmental stages of MRCs within the epithelial layer of the tail of yolk-sac larvae, corresponding to immature, developing, and mature MRCs, identifiable by their own characteristic ultrastructure and form. Following transfer to hyperosmotic salinities the density of immunogold particles and well as the intricacy of the tubular system appeared to increase. In addition, complementary confocal scanning laser microscopy allowed identification of immunopositive ramifying extensions that appeared to emanate from the basolateral portion of the cell that appeared to be correlated with the localization of subsurface tubular areas displaying immunogold labeled Na⁺/K⁺-ATPase. This integrated approach describes a reliable and repeatable prefixation immunogold labeling technique allowing precise visualization of NaK within target cells combined with a 3D imaging that offers valuable insights into MRC dynamics at an ultrastructural level. PMID:23873584

  1. Laser confocal radius measurement method for unpolished spheres.

    PubMed

    Wang, Xu; Zhao, Weiqian; Qiu, Lirong; Yang, Shuai; Wang, Zhongyu

    2016-06-10

    A laser confocal radius measurement method for unpolished spheres (CRMUS) is proposed for measuring the radius of an unpolished sphere during optical sphere processing. CRMUS uses the laser confocal focusing technique to accurately identify the cat's eye and confocal positions of the unpolished sphere, and then uses the distance between the cat's eye and confocal positions measured by a distance measurement interferometer to derive the radius. The partially coherent optical theoretical model of the CRMUS derived indicates that the CRMUS is able to measure the radius of the unpolished sphere with a roughness of less than 0.15 μm. Using an unpolished sphere made of Schott BK7 as the test sphere, experimental results indicate that the CRMUS has a relative expanded uncertainty of less than 20 ppm. The CRMUS could greatly increase processing efficiency. PMID:27409012

  2. A New Multichannel Spectral Imaging Laser Scanning Confocal Microscope

    PubMed Central

    Zhang, Yunhai; Hu, Bian; Dai, Yakang; Yang, Haomin; Huang, Wei; Xue, Xiaojun; Li, Fazhi; Zhang, Xin; Jiang, Chenyu; Gao, Fei; Chang, Jian

    2013-01-01

    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. PMID:23585775

  3. [Combined endoscopic diagnostics with catheter confocal endomicroscopy for gastric neoplasia detection].

    PubMed

    Shuleshova, A G; Zav'ialov, M O; Ul'ianov, D N; Kanareĭtseva, T D

    2014-01-01

    The analysis of combined endoscopic diagnostics with catheter confocal laser endomicroscopy (CCLE) for detection of gastric neoplasia in 103 patients is presented in the article. It was described the main principles of catheter confocal laser endomicroscopy by using of Cellvizio-system ("Mauna Kea Technologies", France). All patients underwent esophagogastroduodenoscopy before catheter confocal laser endomicroscopy. Such modes as HRE-endoscopy, NBI-endoscopy and Zoom-endoscopy were used. It was revealed different neoplastic changes of stomach mucous coat and early cancer forms of stomach in 185 cases. It was noted expediency and high informational content of CCLE which leads to detect the foci of intestinal metaplasia by colonic type, foci of dysplasia and early cancer of stomach mucous coat. The role of conventional morphological study for verification of changes detected with CCLE was shown. PMID:25327669

  4. Confocal laser scanning microscopy with spatiotemporal structured illumination.

    PubMed

    Gao, Peng; Nienhaus, G Ulrich

    2016-03-15

    Confocal laser scanning microscopy (CLSM), which is widely utilized in the biological and biomedical sciences, is limited in spatial resolution due to diffraction to about half the light wavelength. Here we have combined structured illumination with CLSM to enhance its spatial resolution. To this end, we have used a spatial light modulator (SLM) to generate fringe patterns of different orientations and phase shifts in the excitation spot without any mechanical movement. We have achieved 1.8 and 1.7 times enhanced lateral and axial resolutions, respectively, by synthesizing the object spectrum along different illumination directions. This technique is thus a promising tool for high-resolution morphological or fluorescence imaging, especially in deep tissue. PMID:26977667

  5. Surface microstructure profilometry based on laser confocal feedback

    NASA Astrophysics Data System (ADS)

    Wang, Weiping; Zhang, Shulian; Li, Yan

    2015-10-01

    We demonstrate a surface microstructure profile measurement method, which utilizes the positioning ability of confocal technology and the high sensitivity of frequency-shift feedback of a microchip laser. The surface profile is measured by combination of the amplitude and phase information of the feedback light reflected by the sample. The amplitude information is used for coarse measurement and to determine the integral number of half lasing wavelengths contained in the sample profile variation. The phase information is used for fine measurement and to determine the fractional number. The measurement realizes both a large axial measuring range of tens of microns and a high axial resolution of ˜2 nm. Meanwhile, a heterodyne phase measurement approach is introduced to compensate for environmental disturbance and to realize high axial resolution measurement under common room conditions. The surface profile of a grating is measured and proves the feasibility of the method.

  6. Three-dimensional scanning confocal laser microscope

    DOEpatents

    Anderson, R. Rox; Webb, Robert H.; Rajadhyaksha, Milind

    1999-01-01

    A confocal microscope for generating an image of a sample includes a first scanning element for scanning a light beam along a first axis, and a second scanning element for scanning the light beam at a predetermined amplitude along a second axis perpendicular to the first axis. A third scanning element scans the light beam at a predetermined amplitude along a third axis perpendicular to an imaging plane defined by the first and second axes. The second and third scanning element are synchronized to scan at the same frequency. The second and third predetermined amplitudes are percentages of their maximum amplitudes. A selector determines the second and third predetermined amplitudes such that the sum of the percentages is equal to one-hundred percent.

  7. FOOD SURFACE TEXTURE MEASUREMENT USING REFLECTIVE CONFOCAL LASER SCANNING MICROSCOPY

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Confocal laser scanning microscopy (CLSM) was used in the reflection mode to characterize the surface texture (roughness) of sliced food surfaces. Sandpapers of grit size between 150 and 600 were used as the height reference to standardize the CLSM hardware settings. Sandpaper particle sizes were v...

  8. Confocal laser scanning microscopy of apoptosis in organogenesis-stage mouse embryos

    EPA Science Inventory

    Confocal laser scanning microscopy combined with a vital stain has been used to study apoptosis in organogenesis-stage mouse embryos. In order to achieve optical sectioning through embryos, it was necessary to use low power objectives and to prepare the sample appropriately. Mous...

  9. Confocal Fabry-Perot interferometer for frequency stabilization of laser

    NASA Astrophysics Data System (ADS)

    Pan, H.-J.; Ruan, P.; Wang, H.-W.; Li, F.

    2011-02-01

    The frequency shift of laser source of Doppler lidar is required in the range of a few megahertzs. To satisfy this demand, a confocal Fabry-Perot (F-P) interferometer was manufactured as the frequency standard for frequency stabilization. After analyzing and contrasting the center frequency shift of confocal Fabry-Perot interferometers that are made of three different types of material with the change of temperature, the zerodur material was selected to fabricate the interferometer, and the cavity mirrors were optically contacted onto the end of spacer. The confocal Fabry-Perot interferometer was situated within a double-walled chamber, and the change of temperature in the chamber was less than 0.01 K. The experimental results indicate that the free spectral range is 500 MHz, the full-width at half maximum is 3.33 MHz, and the finesse is 150.

  10. Automatic analysis for neuron by confocal laser scanning microscope

    NASA Astrophysics Data System (ADS)

    Satou, Kouhei; Aoki, Yoshimitsu; Mataga, Nobuko; Hensh, Takao K.; Taki, Katuhiko

    2005-12-01

    The aim of this study is to develop a system that recognizes both the macro- and microscopic configurations of nerve cells and automatically performs the necessary 3-D measurements and functional classification of spines. The acquisition of 3-D images of cranial nerves has been enabled by the use of a confocal laser scanning microscope, although the highly accurate 3-D measurements of the microscopic structures of cranial nerves and their classification based on their configurations have not yet been accomplished. In this study, in order to obtain highly accurate measurements of the microscopic structures of cranial nerves, existing positions of spines were predicted by the 2-D image processing of tomographic images. Next, based on the positions that were predicted on the 2-D images, the positions and configurations of the spines were determined more accurately by 3-D image processing of the volume data. We report the successful construction of an automatic analysis system that uses a coarse-to-fine technique to analyze the microscopic structures of cranial nerves with high speed and accuracy by combining 2-D and 3-D image analyses.

  11. Managing multiple image stacks from confocal laser scanning microscopy

    NASA Astrophysics Data System (ADS)

    Zerbe, Joerg; Goetze, Christian H.; Zuschratter, Werner

    1999-05-01

    A major goal in neuroanatomy is to obtain precise information about the functional organization of neuronal assemblies and their interconnections. Therefore, the analysis of histological sections frequently requires high resolution images in combination with an overview about the structure. To overcome this conflict we have previously introduced a software for the automatic acquisition of multiple image stacks (3D-MISA) in confocal laser scanning microscopy. Here, we describe a Windows NT based software for fast and easy navigation through the multiple images stacks (MIS-browser), the visualization of individual channels and layers and the selection of user defined subregions. In addition, the MIS browser provides useful tools for the visualization and evaluation of the datavolume, as for instance brightness and contrast corrections of individual layers and channels. Moreover, it includes a maximum intensity projection, panning and zoom in/out functions within selected channels or focal planes (x/y) and tracking along the z-axis. The import module accepts any tiff-format and reconstructs the original image arrangement after the user has defined the sequence of images in x/y and z and the number of channels. The implemented export module allows storage of user defined subregions (new single image stacks) for further 3D-reconstruction and evaluation.

  12. Imaging retinal densitometry with a confocal Scanning Laser Ophthalmoscope.

    PubMed

    van Norren, D; van de Kraats, J

    1989-01-01

    We describe a novel use of the Scanning Laser Ophthalmoscope (SLO), viz. as an imaging retinal densitometer. In our SLO a helium-neon or an argon laser beam is moved in a raster pattern over the retina; the reflected light is descanned (confocal SLO) and collected by a photomultiplier. Images of the fundus subtending 22 by 18 deg are displayed on a TV monitor. Single frames taken with 514 nm light were stored in a computer in arrays of 256 by 256 pixels and density differences between dark adapted and bleached images were calculated. With a full bleach density differences of about 0.35 were found in the center of the fovea; at retinal eccentricities of 15-20 deg we found 0.15. After selective bleaching with 633 nm light substantial density differences were only seen in the foveal area. We conclude that the confocal SLO is a very suitable instrument for imaging fundus reflectometry. PMID:2631402

  13. Laser ablation of basal cell carcinomas guided by confocal microscopy

    NASA Astrophysics Data System (ADS)

    Sierra, Heidy; Cordova, Miguel; Nehal, Kishwer; Rossi, Anthony; Chen, Chih-Shan Jason; Rajadhyaksha, Milind

    2016-02-01

    Laser ablation offers precise and fast removal of superficial and early nodular types of basal cell carcinomas (BCCs). Nevertheless, the lack of histological confirmation has been a limitation. Reflectance confocal microscopy (RCM) imaging combined with a contrast agent can offer cellular-level histology-like feedback to detect the presence (or absence) of residual BCC directly on the patient. We conducted an ex vivo bench-top study to provide a set of effective ablation parameters (fluence, number of passes) to remove superficial BCCs while also controlling thermal coagulation post-ablation to allow uptake of contrast agent. The results for an Er:YAG laser (2.9 um and pulse duration 250us) show that with 6 passes of 25 J/cm2, thermal coagulation can be effectively controlled, to allow both the uptake of acetic acid (contrast agent) and detection of residual (or absence) BCCs. Confirmation was provided with histological examination. An initial in vivo study on 35 patients shows that the uptake of contrast agent aluminum chloride) and imaging quality is similar to that observed in the ex vivo study. The detection of the presence of residual tumor or complete clearance was confirmed in 10 wounds with (additional) histology and in 25 lesions with follow-up imaging. Our results indicate that resolution is sufficient but further development and use of appropriate contrast agent are necessary to improve sensitivity and specificity. Advances in RCM technology for imaging of lateral and deep margins directly on the patient may provide less invasive, faster and less expensive image-guided approaches for treatment of BCCs.

  14. Imaging System With Confocally Self-Detecting Laser.

    DOEpatents

    Webb, Robert H.; Rogomentich, Fran J.

    1996-10-08

    The invention relates to a confocal laser imaging system and method. The system includes a laser source, a beam splitter, focusing elements, and a photosensitive detector. The laser source projects a laser beam along a first optical path at an object to be imaged, and modulates the intensity of the projected laser beam in response to light reflected from the object. A beam splitter directs a portion of the projected laser beam onto a photodetector. The photodetector monitors the intensity of laser output. The laser source can be an electrically scannable array, with a lens or objective assembly for focusing light generated by the array onto the object of interest. As the array is energized, its laser beams scan over the object, and light reflected at each point is returned by the lens to the element of the array from which it originated. A single photosensitive detector element can generate an intensity-representative signal for all lasers of the array. The intensity-representative signal from the photosensitive detector can be processed to provide an image of the object of interest.

  15. Laser-excited confocal-fluorescence gel scanner

    SciTech Connect

    Mathies, R.A.; Scherer, J.R.; Quesada, M.A. ); Rye, H.S.; Glazer, A.N. )

    1994-04-01

    A high-sensitivity, laser-excited, confocal-fluorescence scanner has been developed for the detection of fluorescently labeled nucleic acids separated on slab gels. The gel is placed on a motor-driven, two-dimensional scan stage and raster scanned past the optical detection system. The 488-nm argon ion laser beam is introduced into the confocal optical system at a long-pass dichroic beam splitter and focused within the gel to an [similar to]2 [mu]m diameter spot by a high-numerical aperture microscope objective. The resulting fluorescence is gathered by the objective, passed back through the first long-pass beam splitter, and relayed to a second dichroic beam splitter that separates the red and green emissions. The fluorescence is then focused on confocal spatial filters to reduce stray and scattered light, passed through spectral filters, and detected with photomultipliers. The resulting signals are amplified, filtered, and digitized for display on a computer. This system can detect as little as 5[times]10[sup [minus]12] M fluorescein, the resolution as operated is 160 [mu]m, and it can scan a 6 cm[times]6 cm gel using a scan rate of 4 cm/s in 12 min. The detection of DNA on slab gels, two-color DNA fragment sizing, and microtiter plate scanning are presented to illustrate some of the possible applications of this apparatus.

  16. [Calibration Procedure of Laser Confocal Micro-Raman Spectrometer].

    PubMed

    Zhao, Ying-chun; Ren, Ling-ling; Wei, Wei-sheng; Yao, Ya-xuan

    2015-09-01

    As a common spectral characterization technique, Raman spectroscopy is widely used and has a specified calibration procedure. Based on laser confocal micro-Raman spectrometer, in this paper, we briefly introduced the principle, configuration and main components of Raman spectrometer. In addition, the calibration procedures were also presented, with an emphasis on the calibration of spectrometer (spectrograph) and that of excitation laser wavelength. On the basis of conventional calibration method, a novel and more accurate method was proposed to obtain the actual excitation wavelength, that is, calibration at the point of Raman shift Δν=0. Using this novel calibration method of excitation wavelength, Raman frequency shift values of sulfur were measured, and compared with the standard values from American Society Testing and Materials (ASTM). As a result, the measured values after calibration were consistent with those ASTM values, which indicated that the calibration method is accurate. Thus, a more reasonable calibration procedure of the laser confocal micro-Raman spectrometer was provided. PMID:26669164

  17. Confocal scanning laser ophthalmoscopic imaging resolution of secondary retinal effects induced by laser radiation

    NASA Astrophysics Data System (ADS)

    Zwick, Harry; Lund, David J.; Stuck, Bruce E.; Zuclich, Joseph A.; Elliot, Rowe; Schuschereba, Steven T.; Gagliano, Donald A.; Belkin, M.; Glickman, Randolph D.

    1996-02-01

    We have evaluated secondary laser induced retinal effects in non-human primates with a Rodenstock confocal scanning laser ophthalmoscope. A small eye animal model, the Garter snake, was employed to evaluate confocal numerical aperture effects in imaging laser retinal damage in small eyes vs. large eyes. Results demonstrate that the confocal image resolution in the Rhesus monkey eye is sufficient to differentiate deep retinal scar formation from retinal nerve fiber layer (NFL) damage and to estimate the depth of the NFL damage. The best comparison with histological depth was obtained for the snake retina, yielding a ratio close to 1:1 compared to 2:1 for the Rhesus. Resolution in the Garter snake allows imaging the photoreceptor matrix and therefore, evaluation of the interrelationship between the primary damage site (posterior retina), the photoreceptor matrix, and secondary sites in the anterior retina such as the NFL and the epiretinal vascular system. Alterations in both the retinal NFL and epiretinal blood flow rate were observed within several minutes post Argon laser exposure. Unique aspects of the snake eye such as high tissue transparency and inherently high contrast cellular structures, contribute to the confocal image quality. Such factors may be nearly comparable in primate eyes suggesting that depth of resolution can be improved by smaller confocal apertures and more sensitive signal processing techniques.

  18. Atomic force microscopy and confocal laser scanning microscopy on the cytoskeleton of permeabilised and embedded cells.

    PubMed

    Meller, Karl; Theiss, Carsten

    2006-03-01

    We describe a technical method of cell permeabilisation and embedding to study the organisation and distribution of intracellular proteins with aid of atomic force microscopy and confocal laser scanning microscopy in identical areas. While confocal laser scanning microscopy is useful for the identification of certain proteins subsequent labelling with markers or antibodies, atomic force microscopy allows the observation of macromolecular structures in fixed and living cells. To demonstrate the field of application of this preparatory technique, cells were permeabilised, fixed, and the actin cytoskeleton was stained with phalloidin-rhodamine. Confocal laser scanning microscopy was used to show the organisation of these microfilaments, e.g. geodesic dome structures. Thereafter, cells were embedded in Durcupan water-soluble resin, followed by UV-polymerisation of resin at 4 degrees C. This procedure allowed intracellular visualisation of the cell nucleus or cytoskeletal elements by atomic force microscopy, for instance to analyse the globular organisation of actin filaments. Therefore, this method offers a great potential to combine both microscopy techniques in order to understand and interpret intracellular protein relations, for example, the biochemical and morphological interaction of the cytoskeleton. PMID:16360280

  19. Visualization and quantification of dentin structure using confocal laser scanning microscopy

    NASA Astrophysics Data System (ADS)

    Kimura, Yuichi; Wilder-Smith, Petra B.; Krasieva, Tatiana B.; Arrastia-Jitosho, Anna-Marie A.; Liaw, Lih-Huei L.; Matsumoto, Koukichi

    1997-07-01

    Dentin was visualized using a new fluorescence technique and confocal laser scanning microscopy. Thirty extracted human teeth showing no clinical signs of caries were investigated. All teeth were horizontally sectioned to approximately 200 micrometers thickness and sections were subjected to different pretreatment conditions as follows: vacuum only, ultrasonication only, sodium hypochlorite only, sodium hypochlorite and vacuum, sodium hypochlorite and ultrasonication, and a combination of sodium hypochlorite, vacuum, and ultrasonication. Some samples were left untreated to serve as control. Following pretreatment, rhodamine 123 fluorescent dye was used for staining at concentrations ranging from 10-3 to 10-7 M for 1 to 24 h at pH 6.0, 6.5, or 7.4. Optical staining occurred at pH 7.4 and concentrations >= 10-5 M over 3 h or longer. Surface images obtained using confocal laser scanning microscopy were similar to those observed by scanning electron microscopy without the need for sample- altering conventional scanning electron microscope preparation techniques. Subsurface imaging to a depth of approximately 60 micrometers was achieved using confocal laser microscope techniques. This fluorescence technique offers a useful new alternative for visualization and quantification of dentin.

  20. Laser multi-reflection confocal long focal-length measurement

    NASA Astrophysics Data System (ADS)

    Li, Zhigang; Qiu, Lirong; Zhao, Weiqian; Xiao, Yang

    2016-06-01

    We propose a new laser multi-reflection confocal focal-length measurement (MCFM) method to meet the requirements of a high-precision measurement for a long focal-length more than 2 m. It places an optical flat and a reflector behind the test lens for reflecting the measuring beam repeatedly, and then, uses the property that the peak points of confocal response curves precisely corresponds to the convergence points of a multi-reflected measuring beam to exactly identify the positions of the convergence points. Subsequently, it obtains the position variation of the reflector with a different number of reflections by a distance measuring instrument, and thereby achieving the high precise long focal-length measurement. The theoretical analyses and preliminary experimental results indicate that MCFM has a relative standard uncertainty of 0.066% for a test lens with the focal-length of 9.76 m. MCFM can provide a novel approach for the high-precision focal-length measurement.

  1. Diffusion of photoacid generators by laser scanning confocal microscopy

    NASA Astrophysics Data System (ADS)

    Zhang, Ping L.; Webber, Stephen E.; Mendenhall, J.; Byers, Jeffrey D.; Chao, Keith K.

    1998-06-01

    Diffusion of the photogenerated acid during the period of time between exposure and development can cause contrast loss and ultimately loss of the latent image. This is especially relevant for chemically amplified photoresists that require a post-exposure baking step, which in turn facilitates acid diffusion due to the high temperature normally employed. It is thus important to develop techniques with good spatial resolution to monitor the photogeneration of acid. More precisely, we need techniques that provide two distinct types of information: spatial resolution on various length scales within the surface layer and also sufficient depth resolution so that one can observe the transition from very surface layer to bulk structure in the polymer blend coated on silicon substrate. Herein laser scanning confocal microscopy is used to evaluate the resist for the first time. We report the use of the confocal microscopy to map the pag/dye distribution in PHS matrices, with both reflectance images and fluorescence images. A laser beam is focused onto a small 3D volume element, termed a voxel. It is typically 200 nm X 200 nm laterally and 800 nm axially. The illuminated voxel is viewed such that only signals emanating from this voxel are detected, i.e., signal from outside the probed voxel is not detected. By adjusting the vertical position of the laser focal point, the voxel can be moved to the designated lateral plane to produce an image. Contrast caused by topology difference between the exposed and unexposed area can be eliminated. Bis-p-butylphenyl iodonium triflat (7% of polyhydroxystyrene) is used as photoacid generators. 5% - 18% (by weight, PHS Mn equals 13 k) resist in PGMEA solution is spin cast onto the treated quartz disk with thickness of 1.4 micrometers , 5 micrometers space/10 micrometers pitch chrome mask is used to generate the pattern with mercury DUV illumination. Fluoresceinamine, the pH-sensitive dye, is also used to enhance the contrast of

  2. Photobleaching property of confocal laser scanning microscopy with masked illumination

    NASA Astrophysics Data System (ADS)

    Kim, DongUk; Moon, Sucbei; Song, Hoseong; Yang, Wenzhong; Kim, Dug Y.

    2010-02-01

    Confocal laser scanning microscopy (CLSM) has become the tool of choice for high-contrast fluorescence imaging in the study of the three-dimensional and dynamic properties of biological system. However, the high cost and complexity of commercial CLSMs urges many researchers to individually develop low cost and flexible confocal microscopy systems. The high speed scanner is an influential factor in terms of cost and system complexity. Resonant galvo scanners at several kHz have been commonly used in custom-built CLSMs. However, during the repeated illumination for live cell imaging or 3D image formation, photobleaching and image distortion occurred at the edges of the scan field may be more serious than the center due to an inherent property (e.g. sinusoidal angular velocity) of the scan mirror. Usually, no data is acquired at the edges due to large image distortion but the excitation beam is still illuminated. Here, we present the photobleaching property of CLSM with masked illumination, a simple and low cost method, to exclude the unintended excitation illumination at the edges. The mask with a square hole in its center is disposed at the image plane between the scan lens and the tube lens in order to decrease photobleaching and image distortion at the edges. The excluded illumination section is used as the black level of the detected signals for a signal quantizing step. Finally, we demonstrated the reduced photobleaching at the edges on a single layer of fluorescent beads and real-time image acquisition without a standard composite video signal by using a frame grabber.

  3. The use of laser scanning confocal microscopy (LSCM) in materials science.

    PubMed

    Hovis, D B; Heuer, A H

    2010-12-01

    Laser scanning confocal microscopes are essential and ubiquitous tools in the biological, biochemical and biomedical sciences, and play a similar role to scanning electron microscopes in materials science. However, modern laser scanning confocal microscopes have a number of advantages for the study of materials, in addition to their obvious uses for high resolution reflected and transmitted light optical microscopy. In this paper, we provide several examples that exploit the laser scanning confocal microscope's capabilities of pseudo-infinite depth of field imaging, topographic imaging, photo-stimulated luminescence imaging and Raman spectroscopic imaging. PMID:21077878

  4. Probe-based confocal laser endomicroscopy in head and neck malignancies: early preclinical experience

    NASA Astrophysics Data System (ADS)

    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.

  5. Confocal laser endomicroscopy in gastrointestinal and pancreatobiliary diseases.

    PubMed

    Nakai, Yousuke; Isayama, Hiroyuki; Shinoura, Susumu; Iwashita, Takuji; Samarasena, Jason B; Chang, Kenneth J; Koike, Kazuhiko

    2014-01-01

    Confocal laser endomicroscopy (CLE) is an emerging diagnostic procedure that enables in vivo pathological evaluation during ongoing endoscopy. There are two types of CLE: endoscope-based CLE (eCLE), which is integrated in the tip of the endoscope, and probe-based CLE (pCLE), which goes through the accessory channel of the endoscope. Clinical data of CLE have been reported mainly in gastrointestinal (GI) diseases including Barrett's esophagus, gastric neoplasms, and colon polyps, but, recently, a smaller pCLE, which goes through a catheter or a fine-needle aspiration needle, was developed and clinical data in the diagnosis of biliary stricture or pancreatic cysts have been increasingly reported. The future application of this novel technique expands beyond the pathological diagnosis to functional or molecular imaging. Despite these promising data, the generalizability of the procedure should be confirmed especially in Japan and other Asian countries, where the current diagnostic yield for GI luminal diseases is high. Given the high cost of CLE devices, cost-benefit analysis should also be considered. PMID:24033351

  6. Confocal laser scanning microscopy in study of bone calcification

    NASA Astrophysics Data System (ADS)

    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.

  7. Confocal laser endomicroscopy features of sessile serrated adenomas/polyps

    PubMed Central

    Parikh, Neil D; Gibson, Joanna; Nagar, Anil; Ahmed, Ali A

    2015-01-01

    Background and aims Sessile serrated adenomas/polyps (SSA/Ps) are difficult to differentiate from non-neoplastic tissue on white-light endoscopy. Confocal laser endomicroscopy (CLE) provides subcellular imaging and real-time “optical biopsy”. The aim of this study was to prospectively describe CLE features of SSA/Ps. Patients and methods Consecutive patients with SSA/Ps were prospectively evaluated with probe-based CLE imaging. CLE images and polyp histology were independently reviewed by three endoscopists and an expert gastrointestinal (GI) pathologist. Distinguishing CLE features of SSA/Ps were identified in conjunction with pathologic correlation. Results In total, 260 CLE images were generated from nine SSA/Ps evaluated in seven patients. Four consensus CLE features of SSA/P were identified: (1) a mucus cap with a bright, cloud-like appearance; (2) thin, branching crypts; (3) increased number of goblet cells and microvesicular mucin-containing cells; and (4) architectural disarray, with dystrophic goblet cells and lack of regular circular crypts Conclusion This is a novel description of characteristic CLE features of SSA/Ps. The four features we identified are easy to detect and may allow for CLE to serve as a diagnostic modality. PMID:27536371

  8. Laparoscopic Manipulation of a Probe-based Confocal Laser Endomicroscope Using a Steerable Intravascular Catheter

    PubMed Central

    Desjardins, Adrien E.; Gurusamy, Kurinchi; Hawkes, David J.; Davidson, Brian R.

    2015-01-01

    Probe-based confocal laser endomicroscopy is an emerging imaging modality that enables visualization of histologic details during endoscopy and surgery. A method of guiding the probe with millimeter accuracy is required to enable imaging in all regions of the abdomen accessed during laparoscopy. On the basis of a porcine model of laparoscopic liver resection, we report our experience of using a steerable intravascular catheter to guide a probe-based confocal laser endomicroscope. PMID:25807277

  9. Confocal laser endomicroscopy to monitor the colonic mucosa of mice.

    PubMed

    Mielke, Lisa; Preaudet, Adele; Belz, Gabrielle; Putoczki, Tracy

    2015-06-01

    The gastrointestinal tract is a unique organ system that provides an epithelial barrier between our underlying immune system and luminal pathogens. Disruption of gastrointestinal homeostasis, as a result of impaired barrier function, is associated with numerous pathologies including inflammatory bowel disease and colorectal cancer. In parallel to the clinical development of endoscopy technologies to monitor and diagnose these pathologies in humans, advanced mouse colonoscopy techniques are being developed. When these technologies are coupled with model systems of human disease, which are essential to our understanding of the pathophysiology of gastrointestinal diseases, the requirement for euthanasia of multiple cohorts of mice is eliminated. Here we highlight the suitability of white light endoscopy to monitor the progression of colitis in mice. We further outline the experimental power of combined standard endoscopy with confocal microendoscopy, which permits visualization of fluorescent markers in a single animal in real-time. Together, these technologies will enhance our understanding of the interplay between components of the gastrointestinal microenvironment and their role in disease. PMID:25960174

  10. Imaging Single ZnO Vertical Nanowire Laser Cavities using UV-Laser Scanning Confocal Microscopy

    SciTech Connect

    Gargas, D.J.; Toimil-Molares, M.E.; Yang, P.

    2008-11-17

    We report the fabrication and optical characterization of individual ZnO vertical nanowire laser cavities. Dilute nanowire arrays with interwire spacing>10 ?m were produced by a modified chemical vapor transport (CVT) method yielding an ideal platform for single nanowire imaging and spectroscopy. Lasing characteristics of a single vertical nanowire are presented, as well as high-resolution photoluminescence imaging by UV-laser scanning confocal microscopy. In addition, three-dimensional (3D) mapping of the photoluminescence emission performed in both planar and vertical dimensions demonstrates height-selective imaging useful for vertical nanowires and heteronanostructures emerging in the field of optoelectronics and nanophotonics.

  11. Use of a white light supercontinuum laser for confocal interference-reflection microscopy

    PubMed Central

    Chiu, L-D; Su, L; Reichelt, S; Amos, WB

    2012-01-01

    Shortly after its development, the white light supercontinuum laser was applied to confocal scanning microscopy as a more versatile substitute for the multiple monochromatic lasers normally used for the excitation of fluorescence. This light source is now available coupled to commercial confocal fluorescence microscopes. We have evaluated a supercontinuum laser as a source for a different purpose: confocal interferometric imaging of living cells and artificial models by interference reflection. We used light in the range 460–700 nm where this source provides a reasonably flat spectrum, and obtained images free from fringe artefacts caused by the longer coherence length of conventional lasers. We have also obtained images of cytoskeletal detail that is difficult to see with a monochromatic laser. PMID:22432542

  12. Interobserver Agreement of Confocal Laser Endomicroscopy for Bladder Cancer

    PubMed Central

    Chang, Timothy C.; Liu, Jen-Jane; Hsiao, Shelly T.; Pan, Ying; Mach, Kathleen E.; Leppert, John T.; McKenney, Jesse K.; Rouse, Robert V.

    2013-01-01

    Abstract Background and Purpose Emerging optical imaging technologies such as confocal laser endomicroscopy (CLE) hold promise in improving bladder cancer diagnosis. The purpose of this study was to determine the interobserver agreement of image interpretation using CLE for bladder cancer. Methods Experienced CLE urologists (n=2), novice CLE urologists (n=6), pathologists (n=4), and nonclinical researchers (n=5) were recruited to participate in a 2-hour computer-based training consisting of a teaching and validation set of intraoperative white light cystoscopy (WLC) and CLE video sequences from patients undergoing transurethral resection of bladder tumor. Interobserver agreement was determined using the κ statistic. Results Of the 31 bladder regions analyzed, 19 were cancer and 12 were benign. For cancer diagnosis, experienced CLE urologists had substantial agreement for both CLE and WLC+CLE (90%, κ 0.80) compared with moderate agreement for WLC alone (74%, κ 0.46), while novice CLE urologists had moderate agreement for CLE (77%, κ 0.55), WLC (78%, κ 0.54), and WLC+CLE (80%, κ 0.59). Pathologists had substantial agreement for CLE (81%, κ 0.61), and nonclinical researchers had moderate agreement (77%, κ 0.49) in cancer diagnosis. For cancer grading, experienced CLE urologists had fair to moderate agreement for CLE (68%, κ 0.64), WLC (74%, κ 0.67), and WLC+CLE (53%, κ 0.33), as did novice CLE urologists for CLE (53%, κ 0.39), WLC (66%, κ 0.50), and WLC+CLE (61%, κ 0.49). Pathologists (65%, κ 0.55) and nonclinical researchers (61%, κ 0.56) both had moderate agreement for CLE in cancer grading. Conclusions CLE is an adoptable technology for cancer diagnosis in novice CLE observers after a short training with moderate interobserver agreement and diagnostic accuracy similar to WLC alone. Experienced CLE observers may be capable of achieving substantial levels of agreement for cancer diagnosis that is higher than with WLC alone. PMID:23072435

  13. Clinical applications of in vivo fluorescence confocal laser scanning microscopy

    NASA Astrophysics Data System (ADS)

    Oh, Chilhwan; Park, Sangyong; Kim, Junhyung; Ha, Seunghan; Park, Gyuman; Lee, Gunwoo; Lee, Onseok; Chun, Byungseon; Gweon, Daegab

    2008-02-01

    Living skin for basic and clinical research can be evaluated by Confocal Laser Scanning Microscope (CLSM) non-invasively. CLSM imaging system can achieve skin image its native state either "in vivo" or "fresh biopsy (ex vivo)" without fixation, sectioning and staining that is necessary for routine histology. This study examines the potential fluorescent CLSM with a various exogenous fluorescent contrast agent, to provide with more resolution images in skin. In addition, in vivo fluorescent CLSM researchers will be extended a range of potential clinical application. The prototype of our CLSM system has been developed by Prof. Gweon's group. The operating parameters are composed of some units, such as illuminated wavelength 488 nm, argon illumination power up to 20mW on the skin, objective lens, 0.9NA oil immersion, axial resolution 1.0μm, field of view 200μm x 100μm (lateral resolution , 0.3μm). In human volunteer, fluorescein sodium was administrated topically and intradermally. Animal studies were done in GFP transgenic mouse, IRC mouse and pig skin. For imaging of animal skin, fluorescein sodium, acridine orange, and curcumine were used for fluorescein contrast agent. We also used the GFP transgenic mouse for fluorescein CLSM imaging. In intact skin, absorption of fluorescein sodium by individual corneocyte and hair. Intradermal administrated the fluorescein sodium, distinct outline of keratinocyte cell border could be seen. Curcumin is a yellow food dye that has similar fluorescent properties to fluorescein sodium. Acridin Orange can be highlight nuclei in viable keratinocyte. In vivo CLSM of transgenic GFP mouse enable on in vivo, high resolution view of GFP expressing skin tissue. GFP signals are brightest in corneocyte, kertinocyte, hair and eccrine gland. In intact skin, absorption of fluorescein sodium by individual corneocyte and hair. Intradermal administrated the fluorescein sodium, distinct outline of keratinocyte cell border could be seen. In

  14. Combined FLIM and reflectance confocal microscopy for epithelial imaging

    NASA Astrophysics Data System (ADS)

    Jabbour, Joey M.; Cheng, Shuna; Shrestha, Sebina; Malik, Bilal; Jo, Javier A.; Applegate, Brian; Maitland, Kristen C.

    2012-03-01

    Current methods for detection of oral cancer lack the ability to delineate between normal and precancerous tissue with adequate sensitivity and specificity. The usual diagnostic mechanism involves visual inspection and palpation followed by tissue biopsy and histopathology, a process both invasive and time-intensive. A more sensitive and objective screening method can greatly facilitate the overall process of detection of early cancer. To this end, we present a multimodal imaging system with fluorescence lifetime imaging (FLIM) for wide field of view guidance and reflectance confocal microscopy for sub-cellular resolution imaging of epithelial tissue. Moving from a 12 x 12 mm2 field of view with 157 ìm lateral resolution using FLIM to 275 x 200 μm2 with lateral resolution of 2.2 μm using confocal microscopy, hamster cheek pouch model is imaged both in vivo and ex vivo. The results indicate that our dual modality imaging system can identify and distinguish between different tissue features, and, therefore, can potentially serve as a guide in early oral cancer detection..

  15. Laser scanning confocal microscopy: history, applications, and related optical sectioning techniques.

    PubMed

    Paddock, Stephen W; Eliceiri, Kevin W

    2014-01-01

    Confocal microscopy is an established light microscopical technique for imaging fluorescently labeled specimens with significant three-dimensional structure. Applications of confocal microscopy in the biomedical sciences include the imaging of the spatial distribution of macromolecules in either fixed or living cells, the automated collection of 3D data, the imaging of multiple labeled specimens and the measurement of physiological events in living cells. The laser scanning confocal microscope continues to be chosen for most routine work although a number of instruments have been developed for more specific applications. Significant improvements have been made to all areas of the confocal approach, not only to the instruments themselves, but also to the protocols of specimen preparation, to the analysis, the display, the reproduction, sharing and management of confocal images using bioinformatics techniques. PMID:24052346

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

    PubMed Central

    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. PMID:24639675

  17. Laser excited confocal microscope fluorescence scanner and method

    DOEpatents

    Mathies, Richard A.; Peck, Konan

    1992-01-01

    A fluorescent scanner for scanning the fluorescence from a fluorescence labeled separated sample on a sample carrier including a confocal microscope for illuminating a predetermined volume of the sample carrier and/or receiving and processing fluorescence emissions from said volume to provide a display of the separated sample.

  18. Laser excited confocal microscope fluorescence scanner and method

    DOEpatents

    Mathies, R.A.; Peck, K.

    1992-02-25

    A fluorescent scanner is designed for scanning the fluorescence from a fluorescence labeled separated sample on a sample carrier. The scanner includes a confocal microscope for illuminating a predetermined volume of the sample carrier and/or receiving and processing fluorescence emissions from the volume to provide a display of the separated sample. 8 figs.

  19. Further study of trichosanthin's effect on mouse embryos with confocal laser scanning microscopy

    NASA Astrophysics Data System (ADS)

    Xu, Hui; Zhang, Chunyang; Ma, Hui; Chen, Die Yan

    2001-09-01

    Trichosanthin(TCS), a ribosome inactivating protein extracted from the root tuber of a traditional Chinese medicine herb Tian Huo Fen(THF), possessed abortifacient, anti-tumor and anti-human immunodeficiency virus(HIV) activities. For centuries in China, THF has been used as an effective folk medicine to terminate early and midtrimester pregnancies and to treat ectopic pregnancies, hydatidiform moles and trophoblastic tumor. We observed the changes in reactive oxygen species and intracellular calcium in mouse embryos induced by TCS with confocal laser scanning microscopy in combination with the fluorescene diacetate (DCFHDA) and Fluo-3-AM. The results indicated that TCS induced increase in intracellular calcium and production of reactive oxygen species in mouse embryos , and TCS inhibited the development of mouse embryos effectively. Mouse embryos of different developmental stages before implantation are used in the experiments. This provides new insight into mechanism for abortifacient activity of TCS.

  20. Scanning microphotolysis: a new photobleaching technique based on fast intensity modulation of a scanned laser beam and confocal imaging.

    PubMed

    Wedekind, P; Kubitscheck, U; Peters, R

    1994-10-01

    The fluorescence photobleaching method has been widely used to study molecular transport in single living cells and other microsystems while confocal microscopy has opened new avenues to high-resolution, three-dimensional imaging. A new technique, scanning microphotolysis (Scamp), combines the potential of photobleaching, beam scanning and confocal imaging. A confocal scanning laser microscope was equipped with a sufficiently powerful laser and a novel device, the 'Scamper'. This consisted essentially of a filter changer, an acousto-optical modulator (AOM) and a computer. The computer was programmed to activate the AOM during scanning according to a freely defined image mask. As a result, almost any desired pattern could be bleached ('written') into fluorescent samples at high definition and then imaged ('read') at non-bleaching conditions, employing full confocal resolution. Furthermore, molecular transport could be followed by imaging the dissipation of bleach patterns. Experiments with living cells concerning dynamic processes in cytoskeletal filaments and the lateral mobility of membrane lipids suggest a wide range of potential biological applications. Thus, Scamp offers new possibilities for the optical manipulation and analysis of both technical and biological microsystems. PMID:7799426

  1. Spectral imaging technique for retinal perfusion detection using confocal scanning laser ophthalmoscopy

    NASA Astrophysics Data System (ADS)

    Rasta, Seyed Hossein; Manivannan, Ayyakkannu; Sharp, Peter F.

    2012-11-01

    To evaluate retinal perfusion in the human eye, a dual-wavelength confocal scanning laser ophthalmoscope (cSLO) was developed that provides spectral imaging of the fundus using a combination of red (670 nm) and near-infrared (810 nm) wavelengths. The image of the ocular fundus was analyzed to find out if quantitative measurements of the reflectivity of tissue permit assessment of the oxygen perfusion of tissue. We explored problems that affect the reproducibility of patient measurements such as non-uniformity errors on the image. For the first time, an image processing technique was designed and used to minimize the errors of oxygen saturation measurements by illumination correction in retina wide field by increasing SNR. Retinal images were taken from healthy and diabetic retinopathy eyes using the cSLO with a confocal aperture of 100 μm. The ratio image (RI) of red/IR, as oxygen saturation (SO2) index, was calculated for normal eyes. The image correction technique improved the reproducibility of the measurements. Average RI intensity variation of healthy retina tissue was determined within a range of about 5.5%. The capability of the new technique to discriminate oxygenation levels of retinal artery and vein was successfully demonstrated and showed good promise in the diagnosis of the perfused retina.

  2. Fabrication of microgrooves on a curved surface by the confocal measurement system using pulse laser and continuous laser

    NASA Astrophysics Data System (ADS)

    Noh, Jiwhan; Cho, Ilhwan; Lee, Seungwoo; Na, Suckjoo; Lee, Jae-Hoon

    2012-03-01

    In order to fabricate microgrooves on a curved surface, the curved surface was measured with a confocal system and then it was used for laser microprocessing. This paper proposes a new method of using a pulse laser for the confocal system to measure the curved surface. It also compares the conventional way of using a continuous laser and a new way of using the pulse laser with the confocal system. Using the data measured with the pulse laser for fabrication, microgrooves were fabricated on a curved surface. The width of the fabricated microgroove was 10 μm and the depth was 27 μm. The microgroove fabricated on a curved surface as a part of this study can be used in injection molding to manufacture a micropatterned plastic surface at a low cost. This plastic surface can be applied for a superhydrophobic surface, a self-cleaning surface, or a biochip.

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

    PubMed

    Mowla, Alireza; Taimre, Thomas; Lim, Yah Leng; Bertling, Karl; Wilson, Stephen J; Prow, Tarl W; Soyer, H Peter; Rakić, Aleksandar D

    2016-01-01

    Optical interrogation of suspicious skin lesions is standard care in the management of skin cancer worldwide. Morphological and functional markers of malignancy are often combined to improve expert human diagnostic power. We propose the evaluation of the combination of two independent optical biomarkers of skin tumours concurrently. The morphological modality of reflectance confocal microscopy (RCM) is combined with the functional modality of laser Doppler flowmetry, which is capable of quantifying tissue perfusion. To realize the idea, we propose laser feedback interferometry as an implementation of RCM, which is able to detect the Doppler signal in addition to the confocal reflectance signal. Based on the proposed technique, we study numerical models of skin tissue incorporating two optical biomarkers of malignancy: (i) abnormal red blood cell velocities and concentrations and (ii) anomalous optical properties manifested through tissue confocal reflectance, using Monte Carlo simulation. We also conduct a laboratory experiment on a microfluidic channel containing a dynamic turbid medium, to validate the efficacy of the technique. We quantify the performance of the technique by examining a signal to background ratio (SBR) in both the numerical and experimental models, and it is shown that both simulated and experimental SBRs improve consistently using this technique. This work indicates the feasibility of an optical instrument, which may have a role in enhanced imaging of skin malignancies. PMID:27598157

  4. Effects of Fluorescein Staining on Laser In Vivo Confocal Microscopy Images of the Cornea

    PubMed Central

    Sindt, Christine W.; Critser, D. Brice; Grout, Trudy K.; Kern, Jami R.

    2012-01-01

    This study was designed to identify whether topical fluorescein, a common ophthalmic tool, affects laser in vivo confocal microscopy of the cornea, a tool with growing applications. Twenty-five eye care specialists were asked to identify presence or absence of fluorescein in 99 confocal micrographs of healthy corneas. Responses were statistically similar to guessing for the epithelium (48% ± 14% of respondents correct per image) and the subbasal nerve plexus (49% ± 11% correct), but results were less clear for the stroma. Dendritic immune cells were quantified in bilateral images from subjects who had been unilaterally stained with fluorescein. Density of dendritic immune cells was statistically similar between the unstained and contralateral stained eyes of 24 contact lens wearers (P = .72) and of 10 nonwearers (P = .53). Overall, the results indicated that fluorescein staining did not interfere with laser confocal microscopy of corneal epithelium, subbasal nerves, or dendritic immune cells. PMID:22363837

  5. Probe-based confocal laser endomicroscopy of the urinary tract: the technique.

    PubMed

    Chang, Timothy C; Liu, Jen-Jane; Liao, Joseph C

    2013-01-01

    Probe-based confocal laser endomicroscopy (CLE) is an emerging optical imaging technology that enables real-time in vivo microscopy of mucosal surfaces during standard endoscopy. With applications currently in the respiratory and gastrointestinal tracts, CLE has also been explored in the urinary tract for bladder cancer diagnosis. Cellular morphology and tissue microarchitecture can be resolved with micron scale resolution in real time, in addition to dynamic imaging of the normal and pathological vasculature. The probe-based CLE system (Cellvizio, Mauna Kea Technologies, France) consists of a reusable fiberoptic imaging probe coupled to a 488 nm laser scanning unit. The imaging probe is inserted in the working channels of standard flexible and rigid endoscopes. An endoscope-based CLE system (Optiscan, Australia), in which the confocal endomicroscopy functionality is integrated onto the endoscope, is also used in the gastrointestinal tract. Given the larger scope diameter, however, application in the urinary tract is currently limited to ex vivo use. Confocal image acquisition is done through direct contact of the imaging probe with the target tissue and recorded as video sequences. As in the gastrointestinal tract, endomicroscopy of the urinary tract requires an exogenenous contrast agent-most commonly fluorescein, which can be administered intravenously or intravesically. Intravesical administration is a well-established method to introduce pharmacological agents locally with minimal systemic toxicity that is unique to the urinary tract. Fluorescein rapidly stains the extracellular matrix and has an established safety profile. Imaging probes of various diameters enable compatibility with different caliber endoscopes. To date, 1.4 and 2.6 mm probes have been evaluated with flexible and rigid cystoscopy. Recent availability of a < 1 mm imaging probe opens up the possibility of CLE in the upper urinary tract during ureteroscopy. Fluorescence cystoscopy (i

  6. MAMMALIAN APOPTOSIS IN WHOLE NEONATAL OVARIES USING CONFOCAL LASER SCANNING MICROSCOPY

    EPA Science Inventory

    MAMMALIAN APOPTOSIS IN WHOLE NEONATAL OVARIES USING CONFOCAL LASER SCANNING MICROSCOPY

    Robert M. Zucker Susan C. Jeffery and Sally D. Perreault

    Reproductive Toxicology Division, National Health and Environmental Effects Research Laboratory, U.S. Environmental Prot...

  7. Detailed three-dimensional visualization of resilin in the exoskeleton of arthropods using confocal laser scanning microscopy.

    PubMed

    Michels, J; Gorb, S N

    2012-01-01

    Resilin is a rubber-like protein found in the exoskeleton of arthropods. It often contributes large proportions to the material of certain structures in movement systems. Accordingly, the knowledge of the presence and distribution of resilin is essential for the understanding of the functional morphology of these systems. Because of its specific autofluorescence, resilin can be effectively visualized using fluorescence microscopy. However, the respective excitation maximum is in the UV range, which is not covered by the lasers available in most of the modern commercial confocal laser scanning microscopes. The goal of this study was to test the potential of confocal laser scanning microscopy (CLSM) in combination with a 405 nm laser to visualize and analyse the presence and distribution of resilin in arthropod exoskeletons. The results clearly show that all resilin-dominated structures, which were visualized successfully using wide-field fluorescence microscopy (WFM) and a 'classical' UV excitation, could also be visualized efficiently with the proposed CLSM method. Furthermore, with the application of additional laser lines CLSM turned out to be very appropriate for studying differences in the material composition within arthropod exoskeletons in great detail. As CLSM has several advantages over WFM with respect to detailed morphological imaging, the application of the proposed CLSM method may reveal new information about the micromorphology and material composition of resilin-dominated exoskeleton structures leading to new insights into the functional morphology and biomechanics of arthropods. PMID:22142031

  8. Correlated Biofilm Imaging, Transport and Metabolism Measurements via Combined Nuclear Magnetic Resonance and Confocal Microscopy

    SciTech Connect

    Mclean, Jeffrey S.; Ona, Ositadinma; Majors, Paul D.

    2008-02-18

    Bacterial biofilms are complex, three-dimensional, communities that are found nearly everywhere in nature1 and are being recognized as the cause of treatment-resistant infections1 2. Advanced methods are required to characterize their collective and spatial patterns of metabolism however most techniques are invasive or destructive. Here we describe the use of a combined confocal laser scanning microscopy (CLSM) and nuclear magnetic resonance (NMR) microscopy system to monitor structure, mass transport, and metabolism in active biofilms. Non-invasive NMR methods provide macroscopic structure along with spatially-resolved metabolite profiles and diffusion measurements. CLSM enables monitoring of cells by fluorescent protein reporters to investigate biofilm structure and gene expression concurrently. A planar sample chamber design facilitates depth-resolved measurements on 140 nL sample volumes under laminar flow conditions. The techniques and approaches described here are applicable to environmental and medically relevant microbial communities, thus providing key metabolic information for promoting beneficial biofilms and treating associated diseases.

  9. Evaluation of human sclera after femtosecond laser ablation using two photon and confocal microscopy

    NASA Astrophysics Data System (ADS)

    Sun, Hui; Kurtz, Ronald; Juhasz, Tibor

    2012-08-01

    Glaucoma is the second-leading cause of blindness worldwide and is often associated with elevated intraocular pressure (IOP). Partial thickness intrascleral channels can be created with a femtosecond laser operating at a wavelength of 1700 nm. Such channels have the potential to increase outflow facility and reduce elevated IOP. Analysis of the dimensions and location of these channels is important in understanding their effects. We describe the application of two-photon microscopy and confocal microscopy for noninvasive imaging of the femtosecond laser created partial-thickness scleral channels in human cadaver eyes. High-resolution images, hundreds of microns deep in the sclera, were obtained to allow determination of the shape and dimension of such channels. This demonstrates that concept of integrating femtosecond laser surgery, and two-photon and confocal imaging has the future potential for image-guided high-precision surgery in transparent and translucent tissue.

  10. Multiplex fluorescence in situ hybridization (M-FISH) and confocal laser scanning microscopy (CLSM) to analyze multispecies oral biofilms.

    PubMed

    Karygianni, Lamprini; Hellwig, Elmar; Al-Ahmad, Ali

    2014-01-01

    Multiplex fluorescence in situ hybridization (M-FISH) constitutes a favorable microbiological method for the analysis of spatial distribution of highly variable phenotypes found in multispecies oral biofilms. The combined use of confocal laser scanning microscopy (CLSM) produces high-resolution three-dimensional (3D) images of individual bacteria in their natural environment. Here, we describe the application of M-FISH on early (Streptococcus spp., Actinomyces naeslundii) and late colonizers (Fusobacterium nucleatum, Veillonella spp.) of in situ-formed oral biofilms, the acquisition of CLSM images, as well as the qualitative and quantitative analysis of these digitally obtained and processed images. PMID:24664826

  11. Modeling and simulation of protein uptake in cation exchanger visualized by confocal laser scanning microscopy.

    PubMed

    Yang, Kun; Shi, Qing-Hong; Sun, Yan

    2006-12-01

    Confocal laser scanning microscopy (CLSM) has been extensively applied in the area of protein chromatography to investigate the uptake mechanism of protein in adsorbents. However, due to the light attenuation in the deeper layers of a specimen, quantitative analysis using CLSM data is still far from reality. In this work, an attenuation equation for describing the darkening of the CLSM image in the deeper scanning layers was developed. Bovine serum albumin (BSA) adsorption to SP Sepharose FF was performed by batch adsorption and micro-column chromatography on which protein concentration in single absorbents were visualized by CLSM. The parameters in the equation were estimated by fitting it to the fluorescence intensity profiles obtained at adsorption equilibrium, and then the equation was used to simulate the effect caused by the light scattering and absorption. CLSM analysis demonstrated that BSA adsorption to SP Sepharose FF followed the shrinking core pattern and was predicted reasonably well by the pore diffusion model in combination with the attenuation equation. By comparison of the CLSM data with the simulations, it shows that the attenuation equation was useful to demonstrate the validity of an intraparticle mass transport model for the estimation of intraparticle protein concentration profiles. PMID:17034803

  12. Handheld histology-equivalent sectioning laser-scanning confocal optical microscope for interventional imaging.

    PubMed

    Kumar, Karthik; Avritscher, Rony; Wang, Youmin; Lane, Nancy; Madoff, David C; Yu, Tse-Kuan; Uhr, Jonathan W; Zhang, Xiaojing

    2010-04-01

    A handheld, forward-imaging, laser-scanning confocal microscope (LSCM) demonstrating optical sectioning comparable with microtome slice thicknesses in conventional histology, targeted towards interventional imaging, is reported. Fast raster scanning (approximately 2.5 kHz line scan rate, 3.0-5.0 frames per second) was provided by a 2-axis microelectromechanical system (MEMS) scanning mirror fabricated by a method compatible with complementary metal-oxide-semiconductor (CMOS) processing. Cost-effective rapid-prototyped packaging combined the MEMS mirror with micro-optical components into a probe with 18 mm outer diameter and 54 mm rigid length. ZEMAX optical design simulations indicate the ability of the handheld optical system to obtain lateral resolution of 0.31 and axial resolution of 2.85 microm. Lateral and axial resolutions are experimentally measured at 0.5 microm and 4.2 microm respectively, with field of view of 200 x 125 microm. Results of reflectance imaging of ex vivo swine liver, and fluorescence imaging of the expression of cytokeratin and mammaglobin tumor biomarkers in epithelial human breast tissue from metastatic breast cancer patients are presented. The results indicate that inexpensive, portable handheld optical microscopy tools based on silicon micromirror technologies could be important in interventional imaging, complementing existing coarse-resolution techniques to improve the efficacy of disease diagnosis, image-guided excisional microsurgery, and monitored photodynamic therapy. PMID:20012209

  13. Appraisal of needle-based confocal laser endomicroscopy in the diagnosis of pancreatic cysts

    PubMed Central

    Krishna, Somashekar G; Lee, Jeffery H

    2016-01-01

    Nearly 2.5% of cross-sectional imaging studies will report a finding of a cystic pancreatic lesion. Even though most of these are incidental findings, it remains very concerning for both patients and treating clinicians. Differentiating and predicting malignant transformation in pancreatic cystic lesions is clinically challenging. Current evaluation of suspicious cystic lesions includes a combination of radiologic imaging, endoscopic ultrasound (EUS) and cyst fluid analyses. Despite these attempts, precise diagnostic stratification among non-mucinous, mucinous, and malignant cystic lesions is often not possible until surgical resection. EUS-guided needle based confocal laser endomicroscopy (nCLE) for evaluation of pancreatic cysts is emerging as a powerful technique with remarkable potential. Though limited imaging data from 3 large clinical trials (INSPECT, DETECT and CONTACT) are currently the reference standard for nCLE imaging, nonetheless these have not been validated in large studies. The aim of this review article is to review the evolving role of EUS-guided nCLE in management of pancreatic cystic lesions in terms of its significance, adverse events, limitations, and implications. PMID:26819534

  14. Detection of Gold Nanoparticles Aggregation Growth Induced by Nucleic Acid through Laser Scanning Confocal Microscopy

    PubMed Central

    Gary, Ramla; Carbone, Giovani; Petriashvili, Gia; De Santo, Maria Penelope; Barberi, Riccardo

    2016-01-01

    The gold nanoparticle (GNP) aggregation growth induced by deoxyribonucleic acid (DNA) is studied by laser scanning confocal and environmental scanning electron microscopies. As in the investigated case the direct light scattering analysis is not suitable, we observe the behavior of the fluorescence produced by a dye and we detect the aggregation by the shift and the broadening of the fluorescence peak. Results of laser scanning confocal microscopy images and the fluorescence emission spectra from lambda scan mode suggest, in fact, that the intruding of the hydrophobic moiety of the probe within the cationic surfactants bilayer film coating GNPs results in a Förster resonance energy transfer. The environmental scanning electron microscopy images show that DNA molecules act as template to assemble GNPs into three-dimensional structures which are reminiscent of the DNA helix. This study is useful to design better nanobiotechnological devices using GNPs and DNA. PMID:26907286

  15. Detection of Gold Nanoparticles Aggregation Growth Induced by Nucleic Acid through Laser Scanning Confocal Microscopy.

    PubMed

    Gary, Ramla; Carbone, Giovani; Petriashvili, Gia; De Santo, Maria Penelope; Barberi, Riccardo

    2016-01-01

    The gold nanoparticle (GNP) aggregation growth induced by deoxyribonucleic acid (DNA) is studied by laser scanning confocal and environmental scanning electron microscopies. As in the investigated case the direct light scattering analysis is not suitable, we observe the behavior of the fluorescence produced by a dye and we detect the aggregation by the shift and the broadening of the fluorescence peak. Results of laser scanning confocal microscopy images and the fluorescence emission spectra from lambda scan mode suggest, in fact, that the intruding of the hydrophobic moiety of the probe within the cationic surfactants bilayer film coating GNPs results in a Förster resonance energy transfer. The environmental scanning electron microscopy images show that DNA molecules act as template to assemble GNPs into three-dimensional structures which are reminiscent of the DNA helix. This study is useful to design better nanobiotechnological devices using GNPs and DNA. PMID:26907286

  16. Application of laser differential confocal technique in back vertex power measurement for phoropters

    NASA Astrophysics Data System (ADS)

    Li, Fei; Li, Lin; Ding, Xiang; Liu, Wenli

    2012-10-01

    A phoropter is one of the most popular ophthalmic instruments used in optometry and the back vertex power (BVP) is one of the most important parameters to evaluate the refraction characteristics of a phoropter. In this paper, a new laser differential confocal vertex-power measurement method which takes advantage of outstanding focusing ability of laser differential confocal (LDC) system is proposed for measuring the BVP of phoropters. A vertex power measurement system is built up. Experimental results are presented and some influence factor is analyzed. It is demonstrated that the method based on LDC technique has higher measurement precision and stronger environmental anti-interference capability compared to existing methods. Theoretical analysis and experimental results indicate that the measurement error of the method is about 0.02m-1.

  17. Confocal, two-photon laser-induced fluorescence technique for the detection of nitric oxide.

    PubMed

    Reeves, M; Musculus, M; Farrell, P

    1998-10-01

    We describe a confocal two-photon laser-induced fluorescence scheme for the detection of gaseous NO. Excitation from a simple YAG-pumped Coumarin 450 dye system near 452.6 nm was used to promote the two-photon NO(A (2)?(+), nu? = 0 ? X (2)?, nu? = 0) transition in the gamma(0, 0) band. Subsequent fluorescence detection in the range 200-300 nm permitted almost total rejection of elastic and geometric scatter of laser radiation for excellent signal/noise ratio characteristics. The goal of the research was to apply NO fluorescence to a relatively realistic limited optical access combustion environment. A confocal optical arrangement was demonstrated for single-point measurements of NO concentration in gas samples and in atmospheric-pressure flames. The technique is suitable for applications that offer only a single direction for optical access and when significant elastic scatter is present. PMID:18301470

  18. Correlated biofilm imaging, transport and metabolism measurements via combined nuclear magnetic resonance and confocal microscopy.

    PubMed

    McLean, Jeffrey S; Ona, Ositadinma N; Majors, Paul D

    2008-02-01

    Bacterial biofilms are complex, three-dimensional communities found nearly everywhere in nature and are also associated with many human diseases. Detailed metabolic information is critical to understand and exploit beneficial biofilms as well as combat antibiotic-resistant, disease-associated forms. However, most current techniques used to measure temporal and spatial metabolite profiles in these delicate structures are invasive or destructive. Here, we describe imaging, transport and metabolite measurement methods and their correlation for live, non-invasive monitoring of biofilm processes. This novel combination of measurements is enabled by the use of an integrated nuclear magnetic resonance (NMR) and confocal laser scanning microscope (CLSM). NMR methods provide macroscopic structure, metabolic pathway and rate data, spatially resolved metabolite concentrations and water diffusion profiles within the biofilm. In particular, current depth-resolved spectroscopy methods are applied to detect metabolites in 140-190 nl volumes within biofilms of the dissimilatory metal-reducing bacterium Shewanella oneidensis strain MR-1 and the oral bacterium implicated in caries disease, Streptococcus mutans strain UA159. The perfused sample chamber also contains a transparent optical window allowing for the collection of complementary fluorescence information using a unique, in-magnet CLSM. In this example, the entire three-dimensional biofilm structure was imaged using magnetic resonance imaging. This was then correlated to a fluorescent CLSM image by employing a green fluorescent protein reporter construct of S. oneidensis. Non-invasive techniques such as described here, which enable measurements of dynamic metabolic processes, especially in a depth-resolved fashion, are expected to advance our understanding of processes occurring within biofilm communities. PMID:18253132

  19. Correlated biofilm imaging, transport and metabolism measurements via combined nuclear magnetic resonance and confocal microscopy

    PubMed Central

    McLean, Jeffrey S; Ona, Ositadinma N; Majors, Paul D

    2015-01-01

    Bacterial biofilms are complex, three-dimensional communities found nearly everywhere in nature and are also associated with many human diseases. Detailed metabolic information is critical to understand and exploit beneficial biofilms as well as combat antibiotic-resistant, disease-associated forms. However, most current techniques used to measure temporal and spatial metabolite profiles in these delicate structures are invasive or destructive. Here, we describe imaging, transport and metabolite measurement methods and their correlation for live, non-invasive monitoring of biofilm processes. This novel combination of measurements is enabled by the use of an integrated nuclear magnetic resonance (NMR) and confocal laser scanning microscope (CLSM). NMR methods provide macroscopic structure, metabolic pathway and rate data, spatially resolved metabolite concentrations and water diffusion profiles within the biofilm. In particular, current depth-resolved spectroscopy methods are applied to detect metabolites in 140–190 nl volumes within biofilms of the dissimilatory metal-reducing bacterium Shewanella oneidensis strain MR-1 and the oral bacterium implicated in caries disease, Streptococcus mutans strain UA159. The perfused sample chamber also contains a transparent optical window allowing for the collection of complementary fluorescence information using a unique, in-magnet CLSM. In this example, the entire three-dimensional biofilm structure was imaged using magnetic resonance imaging. This was then correlated to a fluorescent CLSM image by employing a green fluorescent protein reporter construct of S. oneidensis. Non-invasive techniques such as described here, which enable measurements of dynamic metabolic processes, especially in a depth-resolved fashion, are expected to advance our understanding of processes occurring within biofilm communities. PMID:18253132

  20. UNDERSTANDING THE EFFECTS OF SURFACTANT ADDITION ON RHEOLOGY USING LASER SCANNING CONFOCAL MICROSCOPY

    SciTech Connect

    White, T

    2007-05-08

    The effectiveness of three dispersants to modify rheology was examined using rheology measurements and laser scanning confocal microscopy (LSCM) in simulated waste solutions. All of the dispersants lowered the yield stress of the slurries below the baseline samples. The rheology curves were fitted reasonably to a Bingham Plastic model. The three-dimensional LSCM images of simulants showed distinct aggregates were greatly reduced after the addition of dispersants leading to a lowering of the yield stress of the simulated waste slurry solutions.

  1. Three-dimensional imaging of monogenoidean sclerites by laser scanning confocal fluorescence microscopy.

    PubMed

    Galli, Paolo; Strona, Giovanni; Villa, Anna Maria; Benzoni, Francesca; Fabrizio, Stefani; Doglia, Silvia Maria; Kritsky, Delane C

    2006-04-01

    A nondestructive protocol for preparing specimens of Monogenoidea for both alpha-taxonomic studies and reconstruction of 3-dimensional structure is presented. Gomori's trichrome, a stain commonly used to prepare whole-mount specimens of monogenoids for taxonomic purposes, is used to provide fluorescence of genital spines, the copulatory organ, accessory piece, squamodisc, anchors, hooks, bars, and clamps under laser scanning confocal microscopy. PMID:16729702

  2. Application of confocal laser microscopy for monitoring mesh implants in herniology

    SciTech Connect

    Zakharov, V P; Belokonev, V I; Bratchenko, I A; Timchenko, P E; Vavilov, A V; Volova, L T

    2011-04-30

    The state of the surface of mesh implants and their encapsulation region in herniology is investigated by laser confocal microscopy. A correlation between the probability of developing relapses and the size and density of implant microdefects is experimentally shown. The applicability limits of differential reverse scattering for monitoring the post-operation state of implant and adjacent tissues are established based on model numerical experiments. (optical technologies in biophysics and medicine)

  3. Three-dimensional functional imaging of lung parenchyma using optical coherence tomography combined with confocal fluorescence microscopy

    NASA Astrophysics Data System (ADS)

    Gaertner, Maria; Cimalla, Peter; Knels, Lilla; Meissner, Sven; Koch, Edmund

    2011-03-01

    Optical coherence tomography (OCT), as a non-invasive technique for studying tissue morphology, is widely used in in vivo studies, requiring high resolution and fast three-dimensional imaging. Based on light scattering it reveals micrometer sized substructures of the samples due to changes in their optical properties and therefore allows quantification of the specimen's geometry. Utilizing fluorescence microscopy further information can be obtained from molecular compositions embedded in the investigated object. Fluorescent markers, specifically binding to the substance of interest, reveal the sample's chemical structure and give rise to functional studies. This research presents the application of a combined OCT and laser scanning confocal microscopy (LSCM) system to investigate structural details in lung tissue. OCT reveals the three-dimensional morphology of the alveoli whereas fluorescence detection, arising from the fluorophore Sulforhodamin B (SRB) which is binding to elastin, shows the elastic meshwork of the organs extracellular matrix. Different plains of fluorescence can be obtained by using a piezo driven objective and exploiting the confocal functionality of the setup. Both techniques combined in one optical system not only ease the experimental procedure but also contribute to a thorough description of tissue's morphology and chemical composition.

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

    PubMed

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

    2015-01-27

    Understanding surface and interfacial lateral organization in material and biological systems is critical in nearly every field of science. The continued development of tools and techniques viable for elucidation of interfacial and surface information is therefore necessary to address new questions and further current investigations. Sum frequency spectroscopy (SFS) is a label-free, nonlinear optical technique with inherent surface specificity that can yield critical organizational information on interfacial species. Unfortunately, SFS provides no spatial information on a surface; small scale heterogeneities that may exist are averaged over the large areas typically probed. Over the past decade, this has begun to be addressed with the advent of SFS microscopy. Here we detail the construction and function of a total internal reflection (TIR) SFS spectral and confocal fluorescence imaging microscope directly amenable to surface investigations. This instrument combines, for the first time, sample scanning TIR-SFS imaging with confocal fluorescence microscopy. PMID:25506739

  5. 3-D reconstruction of neurons from multichannel confocal laser scanning image series.

    PubMed

    Wouterlood, Floris 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 to 3-D reconstruct structures of interest. The operator first configures separate channels (e.g., laser, filters, and detector settings) for each applied fluorochrome and then acquires Z-series of confocal images: one series per channel. Channel signal separation is extremely important. Measures to avoid bleaching are vital. Post-acquisition deconvolution of the image series is often performed to increase resolution before 3-D reconstruction takes place. In the 3-D reconstruction programs described in this unit, reconstructions can be inspected in real time from any viewing angle. By altering viewing angles and by switching channels off and on, the spatial relationships of 3-D-reconstructed structures with respect to structures visualized in other channels can be studied. Since each brand of CLSM, computer program, and 3-D reconstruction package has its own proprietary set of procedures, a general approach is provided in this protocol wherever possible. PMID:24723320

  6. Laser-induced cartilage damage: an ex-vivo model using confocal microscopy

    NASA Astrophysics Data System (ADS)

    Frenz, Martin; Zueger, Benno J.; Monin, D.; Weiler, C.; Mainil-Varlet, P. M.; Weber, Heinz P.; Schaffner, Thomas

    1999-06-01

    Although there is an increasing popularity of lasers in orthopedic surgery, there is a growing concern about negative side effects of this therapy e.g. prolonged restitution time, radiation damage to adjacent cartilage or depth effects like bone necrosis. Despite case reports and experimental investigations over the last few years little is known about the extent of acute cartilage damage induced by different lasers types and energies. Histological examination offers only limited insights in cell viability and metabolism. Ho:YAG and Er:YAG lasers emitting at 2.1 micrometer and 2.94 micrometer, respectively, are ideally suited for tissue treatment because these wavelengths are strongly absorbed in water. The Purpose of the present study is to evaluate the effect of laser type and energy on chondrocyte viability in an ex vivo model. Free running Er:YAG (E equals 100 and 150 mJ) and Ho:YAG (E equals 500 and 800 mJ) lasers were used at different energy levels using a fixed pulse length of 400 microseconds. The energy was delivered at 8 Hz through optical fibers. Fresh bovine hyaline cartilage samples were mounted in a water bath at room temperature and the fiber was positioned at 30 degree and 180 degree angles relative to the tissue surface. After laser irradiation the samples were assessed by a life-dead cell viability test using a confocal microscope and by standard histology. Thermal damage was much deeper with Ho:YAG (up to 1800 micrometer) than with the Er:YAG laser (up to 70 micrometer). The cell viability test revealed a damage zone about twice the one determined by standard histology. Confocal microscopy is a powerful tool for assessing changes in tissue structure after laser treatment. In addition this technique allows to quantify these alterations without necessitating time consuming and expensive animal experiments.

  7. A Monolithic Confocal Laser Coupler For an Optical Pick-up

    NASA Astrophysics Data System (ADS)

    Mizuno, Takeshi; Doi, Masato; YoshinobuHiguchi, YoshinobuHiguchi; Taniguchi, Takehiro; NobukataOkano, NobukataOkano; Nakao, Takashi; Narui, Hironobu; Matsuda, Osamu

    1999-04-01

    We propose a novel optical pick-up using a confocal lasercoupler for an optical disk player. The laser coupler consists of aglass window and a monolithic optical element which includes a laserdiode, 8 photodiodes, and a pyramid-shaped prism mirror positionednear the confocal plane which acts as a knife edge and aphoto-coupler. The focusing-error signal is detected using theconfocal knife edge (CKE) method and the tracking-error signal isdetected using the CKE push-pull method. The jitter of Compact Disc(CD) readout was 6.7 ns at a line velocity of 1.2 m/s, and the DCoffsets of the tracking-error signal were sppressed to less than 1/3for a radial lens displacement of ±400 µm compared to theconventional push-pull method.

  8. Investigation of phosphatidylcholine enhancing FITC-insulin across buccal mucosa by confocal laser scanning microscopy

    NASA Astrophysics Data System (ADS)

    Tian, Weiqun; Su, Li; Zeng, Shaoqun; Luo, Qingming; Gao, Qiuhua; Xu, Huibi

    2002-04-01

    The aim was to characterize the transport of fluorescein isothiocyanate (FITC)-labeled dextran and insulin with different resoluble compounds for peptides and proteins through buccal mucosa. The penetration rate of insulin molecules through porcine buccal mucosa (a nonkeratinized epithelium, comparable to human buccal mucosa) was investigated by measuring transbuccal fluxes and by analyzing the distribution of the fluorescent probe in the rabbit buccal mucosa epithelium, using confocal laser scanning microscopy for visualizing permeation pathways. The confocal images of the distribution pattern of FITC-dextran and FITC-insulin showed that the paracellular route is the major pathway of FITC-dextran through buccal mucosa epithelium, the intra-cellular route is the major pathway of FITC-insulin through buccal mucosa epithelium. The permeation rate can be increased by co-administration of soybean phosphatidylcholine (SPC).

  9. Investigation of metallurgical phenomena related to process and product development by means of High Temperature Confocal Scanning Laser Microscopy

    NASA Astrophysics Data System (ADS)

    Diéguez-Salgado, U.; Michelic, S.; Bernhard, C.

    2016-03-01

    An increased interest for high temperature metallurgical processes appeared during the last decades, in order to achieve the high quality requirements in steel products. A defined steel cleanness and microstructure essentially influence the final product quality. The high temperatures involved in metallurgical processes and the lack of in situ observations do not only complicate the verification of simulation model predictions but also make significant conclusions regarding the industrial processes difficult. For that reason, new tools and techniques are necessary to develop. By combining the advances of a laser, confocal optics and an infrared image furnace, the High Temperature Confocal Scanning Laser Microscopy (HTCSLM) is a strong tool which enables high temperature in situ observations of different metallurgical phenomena. Next to solidification processes and phase transformations also the behavior of inclusions at different interfaces in the system steel-slag-refractory can be observed. The present study focuses on the aspects of inclusion agglomeration in the liquid steel and the inclusion behavior at the steel/refractory interface in two different steel grades. Out of the obtained experimental data, attraction forces are calculated and compared. This information provides an important basis for a better understanding of inclusion behavior in industrial processes and the therewith related process optimization, like for example the clogging phenomenon during continuous casting.

  10. Combining confocal Raman microscopy and freeze-drying for quantification of substance penetration into human skin.

    PubMed

    Franzen, Lutz; Anderski, Juliane; Planz, Viktoria; Kostka, Karl-Heinz; Windbergs, Maike

    2014-12-01

    In the area of dermatological research, the knowledge of rate and extent of substance penetration into the human skin is essential not only for evaluation of therapeutics, but also for risk assessment of chemicals and cosmetic ingredients. Recently, confocal Raman microscopy emerged as a novel analytical technique for analysis of substance skin penetration. In contrast to destructive drug extraction and quantification, the technique is non-destructive and provides high spatial resolution in three dimensions. However, the generation of time-resolved concentration depth profiles is restrained by ongoing diffusion of the penetrating substance during analysis. To prevent that, substance diffusion in excised human skin can instantly be stopped at defined time points by freeze-drying the sample. Thus, combining sample preparation by freeze-drying with drug quantification by confocal Raman microscopy yields a novel analytical platform for non-invasive and quantitative in vitro analysis of substance skin penetration. This work presents the first proof-of-concept study for non-invasive quantitative substance depth profiling in freeze-dried excised human stratum corneum by confocal Raman microscopy. PMID:25219950

  11. Monitoring Pancreatic Carcinogenesis by the Molecular Imaging of Cathepsin E In Vivo Using Confocal Laser Endomicroscopy

    PubMed Central

    Cui, Lei; Wang, Biyuan; Cui, Wenli; Li, Minghua; Cheng, Yingsheng

    2014-01-01

    The monitoring of pancreatic ductal adenocarcinoma (PDAC) in high-risk populations is essential. Cathepsin E (CTSE) is specifically and highly expressed in PDAC and pancreatic intraepithelial neoplasias (PanINs), and its expression gradually increases along with disease progression. In this study, we first established an in situ 7,12-dimethyl-1,2-benzanthracene (DMBA)-induced rat model for PanINs and PDAC and then confirmed that tumorigenesis properties in this model were consistent with those of human PDAC in that CTSE expression gradually increased with tumor development using histology and immunohistochemistry. Then, using in vivo imaging of heterotopically implanted tumors generated from CTSE- overexpressing cells (PANC-1-CTSE) in nude mice and in vitro imaging of PanINs and PDAC in DMBA-induced rats, the specificity of the synthesized CTSE-activatable probe was verified. Quantitative determination identified that the fluorescence signal ratio of pancreatic tumor to normal pancreas gradually increased in association with progressive pathological grades, with the exception of no significant difference between PanIN-II and PanIN-III grades. Finally, we monitored pancreatic carcinogenesis in vivo using confocal laser endomicroscopy (CLE) in combination with the CTSE-activatable probe. A prospective double-blind control study was performed to evaluate the accuracy of this method in diagnosing PDAC and PanINs of all grades (>82.7%). This allowed us to establish effective diagnostic criteria for CLE in PDAC and PanINs to facilitate the monitoring of PDAC in high-risk populations. PMID:25184278

  12. Nondestructive 3D confocal laser imaging with deconvolution of seven whole stardust tracks with complementary XRF and quantitative analysis

    SciTech Connect

    Greenberg, M.; Ebel, D.S.

    2009-03-19

    We present a nondestructive 3D system for analysis of whole Stardust tracks, using a combination of Laser Confocal Scanning Microscopy and synchrotron XRF. 3D deconvolution is used for optical corrections, and results of quantitative analyses of several tracks are presented. The Stardust mission to comet Wild 2 trapped many cometary and ISM particles in aerogel, leaving behind 'tracks' of melted silica aerogel on both sides of the collector. Collected particles and their tracks range in size from submicron to millimeter scale. Interstellar dust collected on the obverse of the aerogel collector is thought to have an average track length of {approx}15 {micro}m. It has been our goal to perform a total non-destructive 3D textural and XRF chemical analysis on both types of tracks. To that end, we use a combination of Laser Confocal Scanning Microscopy (LCSM) and X Ray Florescence (XRF) spectrometry. Utilized properly, the combination of 3D optical data and chemical data provides total nondestructive characterization of full tracks, prior to flattening or other destructive analysis methods. Our LCSM techniques allow imaging at 0.075 {micro}m/pixel, without the use of oil-based lenses. A full textural analysis on track No.82 is presented here as well as analysis of 6 additional tracks contained within 3 keystones (No.128, No.129 and No.140). We present a method of removing the axial distortion inherent in LCSM images, by means of a computational 3D Deconvolution algorithm, and present some preliminary experiments with computed point spread functions. The combination of 3D LCSM data and XRF data provides invaluable information, while preserving the integrity of the samples for further analysis. It is imperative that these samples, the first extraterrestrial solids returned since the Apollo era, be fully mapped nondestructively in 3D, to preserve the maximum amount of information prior to other, destructive analysis.

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

    NASA Astrophysics Data System (ADS)

    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.

  14. Next generation of optical diagnostics for bladder cancer using probe-based confocal laser endomicroscopy

    NASA Astrophysics Data System (ADS)

    Liu, Jen-Jane; Chang, Timothy C.; Pan, Ying; Hsiao, Shelly T.; Mach, Kathleen E.; Jensen, Kristin C.; Liao, Joseph C.

    2012-02-01

    Real-time imaging with confocal laser endomicroscopy (CLE) probes that fit in standard endoscopes has emerged as a clinically feasible technology for optical biopsy of bladder cancer. Confocal images of normal, inflammatory, and neoplastic urothelium obtained with intravesical fluorescein can be differentiated by morphologic characteristics. We compiled a confocal atlas of the urinary tract using these diagnostic criteria to be used in a prospective diagnostic accuracy study. Patients scheduled to undergo transurethral resection of bladder tumor underwent white light cystoscopy (WLC), followed by CLE, and histologic confirmation of resected tissue. Areas that appeared normal by WLC were imaged and biopsied as controls. We imaged and prospectively analyzed 135 areas in 57 patients. We show that CLE improves the diagnostic accuracy of WLC for diagnosing benign tissue, low and high grade cancer. Interobserver studies showed a moderate level of agreement by urologists and nonclinical researchers. Despite morphologic differences between inflammation and cancer, real-time differentiation can still be challenging. Identification of bladder cancer-specific contrast agents could provide molecular specificity to CLE. By using fluorescently-labeled antibodies or peptides that bind to proteins expressed in bladder cancer, we have identified putative molecular contrast agents for targeted imaging with CLE. We describe one candidate agent - anti-CD47 - that was instilled into bladder specimens. The tumor and normal urothelium were imaged with CLE, with increased fluorescent signal demonstrated in areas of tumor compared to normal areas. Thus, cancer-specificity can be achieved using molecular contrast agents ex vivo in conjunction with CLE.

  15. Experimental research on radius of curvature measurement of spherical lenses based on laser differential confocal technique

    NASA Astrophysics Data System (ADS)

    Ding, Xiang; Sun, Ruoduan; Li, Fei; Zhao, Weiqian; Liu, Wenli

    2011-11-01

    A new approach based on laser differential confocal technique is potential to achieve high accuracy in radius of curvature (ROC) measurement. It utilizes two digital microscopes with virtual pinholes on the CCD detectors to precisely locate the cat's-eye and the confocal positions, which can enhance the focus-identification resolution. An instrumental system was established and experimental research was carried out to determine how error sources contribute to the uncertainty of ROC measurement, such as optical axis misalignment, dead path of the interferometer, surface figure error of tested lenses and temperature fluctuation, etc. Suggestions were also proposed on how these factors could be avoided or suppressed. The system performance was tested by employing four pairs of template lenses with a serial of ROC values. The relative expanded uncertainty was analyzed and calculated based on theoretical analysis and experimental determination, which was smaller than 2x10-5 (k=2). The results were supported by comparison measurement between the differential confocal radius measurement (DCRM) system and an ultra-high accuracy three-dimensional profilometer, showing good consistency. It demonstrated that the DCRM system was capable of high-accuracy ROC measurement.

  16. Spectrally encoded slit confocal microscopy using a wavelength-swept laser

    NASA Astrophysics Data System (ADS)

    Kim, Soocheol; Hwang, Jaehyun; Heo, Jung; Ryu, Suho; Lee, Donghak; Kim, Sang-Hoon; Oh, Seung Jae; Joo, Chulmin

    2015-03-01

    We present an implementation of spectrally encoded slit confocal microscopy. The method employs a rapid wavelength-swept laser as the light source and illuminates a specimen with a line focus that scans through the specimen as the wavelength sweeps. The reflected light from the specimen is imaged with a stationary line scan camera, in which the finite pixel height serves as a slit aperture. This scanner-free operation enables a simple and cost-effective implementation in a small form factor, while allowing for the three-dimensional imaging of biological samples.

  17. Use of the confocal laser scanning microscope in studies on the developmental biology of marine crustaceans.

    PubMed

    Buttino, Isabella; Ianora, Adrianna; Carotenuto, Ylenia; Zupo, Valerio; Miralto, Antonio

    2003-03-01

    Confocal Laser Scanning Microscope techniques have been applied to study the developmental biology of marine copepods and decapod larvae. The lipophylic probes DiI and DiOC(6) were used to study both the external and internal morphology of these crustaceans, whereas the same DiOC(6) and the specific nuclear probe Hoechst 33342 were used to study embryonic development of copepods in vivo. To distinguish viable from non-viable copepod embryos, the vital dye dichlorodihydrofluorescein diacetate (H(2)DCFDA) was used. Major advantages and difficulties in the use of these non-invasive techniques in studies of the reproductive biology of marine crustaceans are discussed. PMID:12567403

  18. Ti-6Al-4V electron beam weld qualification using laser scanning confocal microscopy

    SciTech Connect

    Wanjara, P. . E-mail: priti.wanjara@cnrc-nrc.gc.ca; Brochu, M.; Jahazi, M.

    2005-03-15

    Processing conditions for manufacturing Ti-6Al-4V components by welding using an electron beam source are known to influence the transformation microstructure in the narrow fusion and heat-affected zones of the weld region. This work examined the effect of multiple-sequence welding on the characteristics of the transformed beta microstructure, using laser scanning confocal microscopy to resolve the Widmanstaetten alpha-beta structure in the fusion zone. The evolution in the alpha interlamellar spacing and plate thickness with processing was then related to microhardness measurements in the weld region.

  19. Probing the compressibility of tumor cell nuclei by combined atomic force-confocal microscopy

    NASA Astrophysics Data System (ADS)

    Krause, Marina; te Riet, Joost; Wolf, Katarina

    2013-12-01

    The cell nucleus is the largest and stiffest organelle rendering it the limiting compartment during migration of invasive tumor cells through dense connective tissue. We here describe a combined atomic force microscopy (AFM)-confocal microscopy approach for measurement of bulk nuclear stiffness together with simultaneous visualization of the cantilever-nucleus contact and the fate of the cell. Using cantilevers functionalized with either tips or beads and spring constants ranging from 0.06-10 N m-1, force-deformation curves were generated from nuclear positions of adherent HT1080 fibrosarcoma cell populations at unchallenged integrity, and a nuclear stiffness range of 0.2 to 2.5 kPa was identified depending on cantilever type and the use of extended fitting models. Chromatin-decondensating agent trichostatin A (TSA) induced nuclear softening of up to 50%, demonstrating the feasibility of our approach. Finally, using a stiff bead-functionalized cantilever pushing at maximal system-intrinsic force, the nucleus was deformed to 20% of its original height which after TSA treatment reduced further to 5% remaining height confirming chromatin organization as an important determinant of nuclear stiffness. Thus, combined AFM-confocal microscopy is a feasible approach to study nuclear compressibility to complement concepts of limiting nuclear deformation in cancer cell invasion and other biological processes.

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

    PubMed Central

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

    2015-01-01

    The staining of neurons with silver began in the 1800s, but until now the great resolving power of the laser scanning confocal microscope has not been utilized to capture the in-focus and three-dimensional cytoarchitecture of metal-impregnated cells. Here, we demonstrate how spectral confocal microscopy, typically reserved for fluorescent imaging, can be used to visualize metal-labeled tissues. This imaging does not involve the reflectance of metal particles, but rather the excitation of silver (or gold) nanoparticles and their putative surface plasmon resonance. To induce such resonance, silver or gold particles were excited with visible-wavelength laser lines (561 or 640 nm), and the maximal emission signal was collected at a shorter wavelength (i.e., higher energy state). Because the surface plasmon resonances of noble metal nanoparticles offer a superior optical signal and do not photobleach, our novel protocol holds enormous promise of a rebirth and further development of silver- and gold-based cell labeling protocols. DOI: http://dx.doi.org/10.7554/eLife.09388.001 PMID:26670545

  1. Confocal Laser Microscope Scanning Applied To Three-Dimensional Studies Of Biological Specimens.

    NASA Astrophysics Data System (ADS)

    Franksson, Olof; Liljeborg, Anders; Carlsson, Kjell; Forsgren, Per-Ola

    1987-08-01

    The depth-discriminating property of confocal laser microscope scanners can be used to record the three-dimensional structure of specimens. A number of thin sections (approx. 1 μm thick) can be recorded by a repeated process of image scanning and refocusing of the microscope. We have used a confocal microscope scanner in a number of feasibility studies to investigate its possibilities and limitations. It has proved to be well suited for examining fluorescent specimens with a complicated three-dimensional structure, such as nerve cells. It has also been used to study orchid seeds, as well as cell colonies, greatly facilitating evaluation of such specimens. Scanning of the specimens is performed by a focused laser beam that is deflected by rotating mirrors, and the reflected or fluorescent light from the specimen is detected. The specimen thus remains stationary during image scanning, and is only moved stepwise in the vertical direction for refocusing between successive sections. The scanned images consist of 256*256 or 512*512 pixels, each pixel containing 8 bits of data. After a scanning session a large number of digital images, representing consecutive sections of the specimen, are stored on a disk memory. In a typical case 200 such 256*256 images are stored. To display and process this information in a meaningful way requires both appropriate software and a powerful computer. The computer used is a 32-bits minicomputer equipped with an array processor (FPS 100). The necessary software was developed at our department.

  2. Technique of laser confocal and Raman spectroscopy for living cell analysis

    NASA Astrophysics Data System (ADS)

    Meng, Xiaochen; Zhu, Lianqing

    2013-10-01

    Because of the shortcomings of the main methods used to analysis single cell, the need of single living cell analysis with no damage, unmarked and in situ dynamic multi-parameter measurement is urgent in the life sciences and biomedical advanced research field. And the method of for living cells analysis is proposed. The spectral pretreatment technology of living cell is the key work of laser confocal Raman spectroscopy. To study the spectrum processing methods for Raman spectrum on single living cell and develop the pre-process techniques to enhance the signal-to-noise ratio, sensitivity, and decrease the influence of fluorescence, elimination the cosmic rays was used to improve the spectrum. The classification, average and filtration of spectrum were applied to enhance signal-to-noise ratio. The fluorescence was depressed for quantity analysis or utilized for analysis by comparing the background and the spectrum. The results show that the proposed technique for laser confocal Raman spectrum of single cell can perform the sensitive and weak intensity peaks and reflect the information of molecules structures very well.

  3. Observation of dendritic cell morphology under light, phase-contrast or confocal laser scanning microscopy.

    PubMed

    Tan, Yuen-Fen; Leong, Chooi-Fun; Cheong, Soon-Keng

    2010-12-01

    Dendritic cells (DCs) are professional antigen presenting cells of the immune system. They can be generated in vitro from peripheral blood monocytes supplemented with GM-CSF, IL-4 and TNF alpha. During induction, DCs will increase in size and acquire multiple cytoplasmic projections when compared to their precursor cells such as monocytes or haematopoietic stem cells which are usually round or spherical. Morphology of DCs can be visualized by conventional light microscopy after staining or phase-contrast inverted microscopy or confocal laser scanning microscopy. In this report, we described the morphological appearances of DCs captured using the above-mentioned techniques. We found that confocal laser scanning microscopy yielded DCs images with greater details but the operating cost for such a technique is high. On the other hand, the images obtained through light microscopy after appropriate staining or phase contrast microscopy were acceptable for identification purpose. Besides, these equipments are readily available in most laboratories and the cost of operation is affordable. Nevertheless, morphological identification is just one of the methods to characterise DCs. Other methods such as phenotypic expression markers and mixed leukocyte reactions are additional tools used in the characterisation of DCs. PMID:21329180

  4. The method of axial drift compensation of laser differential confocal microscopy based on zero-tracking

    NASA Astrophysics Data System (ADS)

    Wang, Yajie; Cui, Han; Wang, Yun; Qiu, Lirong; Zhao, Weiqian

    2015-08-01

    Laser differential confocal microscopy (DCM) has advantages of high axial resolution and strong ability of focus identification. However, the imaging mechanism of point scanning needs long measurement time, in the process due to itself mechanical instability and the influence of environment vibration the axial drift of object position is inevitable, which will reduce lateral resolution of the DCM. To ensure the lateral resolution we propose an axial drift compensation method based on zero-tracking in this paper. The method takes advantage of the linear region of differential confocal axial response curve, gets axial drift by detecting the laser intensity; uses grating sensor to monitor the real-time axial drift of lifting stage and realizes closed-loop control; uses capacitive sensor of objective driver to measure its position. After getting the axial drift of object, the lifting stage and objective driver will be driven to compensate position according to the axial drift. This method is realized by using Visual Studio 2010, and the experiment demonstrates that the compensation precision of the proposed method can reach 6 nm. It is not only easy to implement, but also can compensate the axial drift actively and real-timely. Above all, this method improves the system stability of DCM effectively.

  5. Laser Scanning In Vivo Confocal Microscopy of Clear Grafts after Penetrating Keratoplasty

    PubMed Central

    Wang, Dai; Song, Peng; Wang, Shuting; Sun, Dapeng; Wang, Yuexin; Zhang, Yangyang

    2016-01-01

    Purpose. To evaluate the changes of keratocytes and dendritic cells in the central clear graft by laser scanning in vivo confocal microscopy after penetrating keratoplasty (PK). Methods. Thirty adult subjects receiving PK at Shandong Eye Institute and with clear grafts and no sign of immune rejection after surgery were recruited into this study, and 10 healthy adults were controls. The keratocytes and dendritic cells in the central graft were evaluated by laser scanning confocal microscopy, as well as epithelium cells, keratocytes, corneal endothelium cells, and corneal nerves (especially subepithelial plexus nerves). Results. Median density of subepithelial plexus nerves, keratocyte density in each layer of the stroma, and density of corneal endothelium cells were all lower in clear grafts than in controls. The dendritic cells of five (16.7%) patients were active in Bowman's membrane and stromal membrane of the graft after PK. Conclusions. Activated dendritic cells and Langerhans cells could be detected in some of the clear grafts, which indicated that the subclinical stress of immune reaction took part in the chronic injury of the clear graft after PK, even when there was no clinical rejection episode. PMID:27034940

  6. EUS-Guided Needle-Based Confocal Laser Endomicroscopy: A Novel Technique With Emerging Applications

    PubMed Central

    Koduru, Pramoda; Joshi, Virendra; Karstensen, John G.; Saftoiu, Adrian; Vilmann, Peter; Giovannini, Marc

    2015-01-01

    Endoscopic ultrasound (EUS) has emerged as an excellent tool for imaging the gastrointestinal tract, as well as surrounding structures. EUS-guided fine-needle aspiration (EUS-FNA) has become the standard of care for the tissue sampling of a variety of masses and lymph nodes within and around 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 the diagnostic accuracy of EUS-FNA. The device has been studied in animals as well as in humans, and the results so far have been promising. Recently, this method has also been used for the visualization of regulatory proteins and receptors in the pancreas, setting a cornerstone for nCLE in molecular imaging. The aim of this article is to review the role of EUS-guided nCLE in modern endoscopy and its implications in molecular imaging. PMID:27099595

  7. Confocal Laser Endomicroscopy of Bladder and Upper Tract Urothelial Carcinoma: A New Era of Optical Diagnosis?

    PubMed Central

    Chen, Stephanie P.; Liao, Joseph C.

    2014-01-01

    Urothelial carcinoma of the bladder and upper tract pose significant diagnostic and therapeutic challenges. White light endoscopy plays a central role in the management of urothelial carcinoma but has several well-recognized shortcomings. New optical imaging technologies may improve diagnostic accuracy, enhance local cancer control, and better stratify treatment options. Confocal laser endomicroscopy enables dynamic imaging of the cellular structures below the mucosal surface and holds promise in providing real time optical diagnosis and grading of urothelial carcinoma. A variety of imaging probes are available that are compatible with the full spectrum of cystoscopes and ureteroscopes. We review the underlying principles and technique of confocal laser endomicroscopy in the urinary tract, with emphasis on specific application towards urothelial carcinoma. While the available data are largely related to urothelial carcinoma of the bladder, the lessons learned are directly applicable to the upper tract, where the clinical needs are significant. Ongoing efforts to optimize this technology offer an exciting glimpse into future advances in optical imaging and intraoperative image guidance. PMID:25002073

  8. Development of confocal laser microscope system for examination of microscopic characteristics of radiophotoluminescence glass dosemeters.

    PubMed

    Maki, Daisuke; Ishii, Tetsuya; Sato, Fuminobu; Kato, Yushi; Yamamoto, Takayoshi; Iida, Toshiyuki

    2011-03-01

    A confocal laser microscope system was developed for the measurement of radiophotoluminescence (RPL) photons emitted from a minute alpha-ray-irradiated area in an RPL glass dosemeter. The system was composed mainly of an inverted-type microscope, an ultraviolet laser, an XY movable stage and photon-counting circuits. The photon-counting circuits were effective in the reduction of the background noise level in the measurement of RPL photons. The performance of this microscope system was examined by the observation of standard RPL glass samples irradiated using (241)Am alpha rays. The spatial resolution of this system was ∼ 3 μm, and with regard to the sensitivity of this system, a hit of more than four to five alpha rays in unit area produced enough amount of RPL photons to construct the image. PMID:21212081

  9. Diagnosis of gastric intraepithelial neoplasia by narrow-band imaging and confocal laser endomicroscopy

    PubMed Central

    Wang, Shu-Fang; Yang, Yun-Sheng; Wei, Li-Xin; Lu, Zhong-Sheng; Guo, Ming-Zhou; Huang, Jin; Peng, Li-Hua; Sun, Gang; Ling-Hu, En-Qiang; Meng, Jiang-Yun

    2012-01-01

    AIM: To evaluate the diagnosis of different differentiated gastric intraepithelial neoplasia (IN) by magnification endoscopy combined with narrow-band imaging (ME-NBI) and confocal laser endomicroscopy (CLE). METHODS: Eligible patients with suspected gastric IN lesions previously diagnosed by endoscopy in secondary hospitals and scheduled for further diagnosis and treatment were recruited for this study. Excluded from the study were patients who had liver cirrhosis, impaired renal function, acute gastrointestinal (GI) bleeding, coagulopathy, esophageal varices, jaundice, and GI post-surgery. Also excluded were those who were pregnant, breastfeeding, were younger than 18 years old, or were unable to provide informed consent. All patients had all mucus and bile cleared from their stomachs. They then received upper GI endoscopy. When a mucosal lesion is found during observation with white-light imaging, the lesion is visualized using maximal magnification, employing gradual movement of the tip of the endoscope to bring the image into focus. Saved images are analyzed. Confocal images were evaluated by two endoscopists (Huang J and Li MY), who were familiar with CLE, blinded to the related information about the lesions, and asked to classify each lesion as either a low grade dysplasia (LGD) or high grade dysplasia (HGD) according to given criteria. The results were compared with the final histopathologic diagnosis. ME-NBI images were evaluated by two endoscopists (Lu ZS and Ling-Hu EQ) who were familiar with NBI, blinded to the related information about the lesions and CLE images, and were asked to classify each lesion as a LGD or HGD according to the “microvascular pattern and surface pattern” classification system. The results were compared with the final histopathologic diagnosis. RESULTS: The study included 32 pathology-proven low grade gastric IN and 26 pathology-proven high grade gastric IN that were detected with any of the modalities. CLE and ME-NBI enabled

  10. Detection of fluorescent organic nanoparticles by confocal laser endomicroscopy in a rat model of Barrett’s esophageal adenocarcinoma

    PubMed Central

    Dassie, Elisa; Arcidiacono, Diletta; Wasiak, Iga; Damiano, Nunzio; Dall’Olmo, Luigi; Giacometti, Cinzia; Facchin, Sonia; Cassaro, Mauro; Guido, Ennio; De Lazzari, Franca; Marin, Oriano; Ciach, Tomasz; Fery-Forgues, Suzanne; Alberti, Alfredo; Battaglia, Giorgio; Realdon, Stefano

    2015-01-01

    For many years, novel strategies for cancer detection and treatment using nanoparticles (NPs) have been developed. Esophageal adenocarcinoma is the sixth leading cause of cancer-related deaths in Western countries, and despite recent advances in early detection and treatment, its prognosis is still very poor. This study investigated the use of fluorescent organic NPs as potential diagnostic tool in an experimental in vivo model of Barrett’s esophageal adenocarcinoma. NPs were made of modified polysaccharides loaded with [4-(dicyanomethylene)-2-methyl-6-(4-dimethylaminostyryl)-4H-pyran] (DCM), a well-known fluorescent dye. The NP periphery might or might not be decorated with ASYNYDA peptide that has an affinity for esophageal cancer cells. Non-operated and operated rats in which gastroesophageal reflux was surgically induced received both types of NPs (NP-DCM and NP-DCM-ASYNYDA) by intravenous route. Localization of mucosal NPs was assessed in vivo by confocal laser endomicroscopy, a technique which enables a “real time” and in situ visualization of the tissue at a cellular level. After injection of NP-DCM and NP-DCM-ASYNYDA, fluorescence was observed in rats affected by esophageal cancer, whereas no signal was observed in control non-operated rats, or in rats with simple esophagitis or Barrett’s esophagus mucosa. Fluorescence was observable in vivo 30 minutes after the administration of NPs. Interestingly, NP-DCM-ASYNYDA induced strong fluorescence intensity 24 hours after administration. These observations suggested that NPs could reach the tumor cells, likely by enhanced permeability and retention effect, and the peptide ASYNYDA gave them high specificity for esophageal cancer cells. Thus, the combination of NP platform and confocal laser endomicroscopy could play an important role for highlighting esophageal cancer conditions. This result supports the potential of this strategy as a targeted carrier for photoactive and bioactive molecules in esophageal

  11. Detection of fluorescent organic nanoparticles by confocal laser endomicroscopy in a rat model of Barrett's esophageal adenocarcinoma.

    PubMed

    Dassie, Elisa; Arcidiacono, Diletta; Wasiak, Iga; Damiano, Nunzio; Dall'Olmo, Luigi; Giacometti, Cinzia; Facchin, Sonia; Cassaro, Mauro; Guido, Ennio; De Lazzari, Franca; Marin, Oriano; Ciach, Tomasz; Fery-Forgues, Suzanne; Alberti, Alfredo; Battaglia, Giorgio; Realdon, Stefano

    2015-01-01

    For many years, novel strategies for cancer detection and treatment using nanoparticles (NPs) have been developed. Esophageal adenocarcinoma is the sixth leading cause of cancer-related deaths in Western countries, and despite recent advances in early detection and treatment, its prognosis is still very poor. This study investigated the use of fluorescent organic NPs as potential diagnostic tool in an experimental in vivo model of Barrett's esophageal adenocarcinoma. NPs were made of modified polysaccharides loaded with [4-(dicyanomethylene)-2-methyl-6-(4-dimethylaminostyryl)-4H-pyran] (DCM), a well-known fluorescent dye. The NP periphery might or might not be decorated with ASYNYDA peptide that has an affinity for esophageal cancer cells. Non-operated and operated rats in which gastroesophageal reflux was surgically induced received both types of NPs (NP-DCM and NP-DCM-ASYNYDA) by intravenous route. Localization of mucosal NPs was assessed in vivo by confocal laser endomicroscopy, a technique which enables a "real time" and in situ visualization of the tissue at a cellular level. After injection of NP-DCM and NP-DCM-ASYNYDA, fluorescence was observed in rats affected by esophageal cancer, whereas no signal was observed in control non-operated rats, or in rats with simple esophagitis or Barrett's esophagus mucosa. Fluorescence was observable in vivo 30 minutes after the administration of NPs. Interestingly, NP-DCM-ASYNYDA induced strong fluorescence intensity 24 hours after administration. These observations suggested that NPs could reach the tumor cells, likely by enhanced permeability and retention effect, and the peptide ASYNYDA gave them high specificity for esophageal cancer cells. Thus, the combination of NP platform and confocal laser endomicroscopy could play an important role for highlighting esophageal cancer conditions. This result supports the potential of this strategy as a targeted carrier for photoactive and bioactive molecules in esophageal cancer

  12. Roughness of biopores and cracks in Bt-horizons by confocal laser scanning microscopy

    NASA Astrophysics Data System (ADS)

    Leue, Martin; Gerke, Horst H.

    2016-04-01

    During preferential flow events in structured soils, the movement of water and reactive solutes is mostly restricted to larger inter-aggregate pores, cracks, and biopores. The micro-topography of such macropores in terms of pore shapes, geometry, and roughness is crucial for describing the exchange of water and solutes between macropores and the soil matrix. The objective of this study was to determine the surface roughness of intact structural surfaces from the Bt-horizon of Luvisols by confocal laser scanning microscopy. For this purpose, samples with the structural surface types including cracks with and without clay-organic coatings from Bt-horizons developed on loess and glacial till were compared. The surface roughness of these structures was calculated in terms of three parameters from selected surface regions of 0.36 mm² determined with a confocal laser scanning microscope of the type Keyence VK-X100K. These data were evaluated in terms of the root-mean-squared roughness, Rq, the curvature, Rku, and the ratio between surface area and base area, RA. Values of Rq and RA were smaller for coated as compared to uncoated cracks and earthworm burrows of the Bt-horizons from both parent materials. The results indicated that the illuviation of clayey material led to a "smoothing" of the crack surfaces, which was similar for the coarser textured till-Bt and the finer-textured loess-Bt surfaces. The roughness indicated by Rq and RA values was only slightly smaller and that indicated by Rku slightly higher for the structural surfaces from the loess as compared to those from the glacial till. These results suggest a minor importance of the parent material on the roughness of structural surfaces in the Bt-horizon. The similarity of Rq, RA, and Rku values between surfaces of earthworm burrows and uncoated cracks did not confirm an expected smoothing effect of the burrow walls by the earthworm. In contrast to burrow walls, root channels from the loess-Bt were smoother

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

    PubMed Central

    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

  14. Laser multi-reflection differential confocal long focal-length measurement.

    PubMed

    Li, Zhigang; Qiu, Lirong; Zhao, Weiqian; Zhao, Qi

    2016-06-20

    We propose a new laser multi-reflection differential confocal focal-length measurement (LDCFM) method to meet the requirements of high-precision measurements of long focal lengths. An optical flat and a reflector are placed behind a test lens for reflecting the measuring beam repeatedly. Then, LDCFM uses the property that the null points of differential confocal response curves precisely correspond to the convergence points of the multi-reflected measuring beam to exactly determine the positions of the convergence points accurately. Subsequently, the position variation of the reflector is measured with different reflection times by using a distance-measuring instrument, and thereby the long focal length is measured precisely. Theoretical analyses and preliminary experimental results indicate that the LDCFM method has a relative expanded standard uncertainty (k=2) of 0.04% for the test lens with a focal length of 9.76 m. The LDCFM method can provide a novel approach for high-precision focal-length measurements. PMID:27409117

  15. Aperture combined Raman laser

    SciTech Connect

    Woods, C.; Tang, K.; Howton, C.; Muller, D.; Hunter, R.O. Jr.

    1983-01-01

    Excimer lasers, while able to produce large powers and energies, are limited to a few discrete wavelengths. Efficient Raman shifting promises the availability of a much broader wavelength range. A method was developed which both Raman shifts and allows for multiple pump beams.

  16. Parallel deconvolution of large 3D images obtained by confocal laser scanning microscopy.

    PubMed

    Pawliczek, Piotr; Romanowska-Pawliczek, Anna; Soltys, Zbigniew

    2010-03-01

    Various deconvolution algorithms are often used for restoration of digital images. Image deconvolution is especially needed for the correction of three-dimensional images obtained by confocal laser scanning microscopy. Such images suffer from distortions, particularly in the Z dimension. As a result, reliable automatic segmentation of these images may be difficult or even impossible. Effective deconvolution algorithms are memory-intensive and time-consuming. In this work, we propose a parallel version of the well-known Richardson-Lucy deconvolution algorithm developed for a system with distributed memory and implemented with the use of Message Passing Interface (MPI). It enables significantly more rapid deconvolution of two-dimensional and three-dimensional images by efficiently splitting the computation across multiple computers. The implementation of this algorithm can be used on professional clusters provided by computing centers as well as on simple networks of ordinary PC machines. PMID:19725070

  17. Evaluation of the Cytotoxic Behavior of Fungal Extracellular Synthesized Ag Nanoparticles Using Confocal Laser Scanning Microscope

    PubMed Central

    Salaheldin, Taher A.; Husseiny, Sherif M.; Al-Enizi, Abdullah M.; Elzatahry, Ahmed; Cowley, Alan H.

    2016-01-01

    Silver nanoparticles have been synthesized by subjecting a reaction medium to a Fusarium oxysporum biomass at 28 °C for 96 h. The biosynthesized Ag nanoparticles were characterized on the basis of their anticipated peak at 405 nm using UV-Vis-NIR spectroscopy. Structural confirmation was evident from the characteristic X-ray diffraction (XRD) pattern, high-resolution transmission electron Microscopy (HRTEM) and the particle size analyzer. The Ag nanoparticles were of dimension 40 ± 5 nm and spherical in shape. The study mainly focused on using the confocal laser scanning microscope (CLSM) to examine the cytotoxic activities of fungal synthesized Ag nanoparticles on a human breast carcinoma cell line MCF7 cell, which featured remarkable vacuolation, thus indicating a potent cytotoxic activity. PMID:26950118

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

    PubMed

    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. PMID:24274517

  19. An alternative method of promoter assessment by confocal laser scanning microscopy.

    PubMed

    Sahoo, Dipak K; Ranjan, Rajiv; Kumar, Deepak; Kumar, Alok; Sahoo, Bhabani S; Raha, Sumita; Maiti, Indu B; Dey, Nrisingha

    2009-10-01

    A rapid and useful method of promoter activity analysis using techniques of confocal laser scanning microscopy (CLSM) is described in the present study. The activities of some pararetroviral promoters such as CaMV35S (Cauliflower mosaic virus), FMVSgt3 (Figwort mosaic virus sub-genomic transcript) and MMVFLt12 (Mirabilis mosaic virus full-length transcript) coupled to GFP (green fluorescent protein) and GUS (beta-glucuronidase) reporter genes were determined simultaneously by the CLSM technique and other available conventional methods for reporter gene assay based on relevant biochemical and molecular approaches. Consistent and comparable results obtained by CLSM as well as by other conventional assay methods confirm the effectiveness of the CLSM approach for assessment of promoter activity. Hence the CLSM method can be suggested as an alternative way for promoter analysis on the basis of high throughput. PMID:19540268

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

    NASA Technical Reports Server (NTRS)

    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. Characterization of acoustic lenses with the Foucault test by confocal laser scanning microscopy

    NASA Astrophysics Data System (ADS)

    Ahmed Mohamed, E. T.; Abdelrahman, A.; Pluta, M.; Grill, W.

    2010-03-01

    In this work, the Foucault knife-edge test, which has traditionally been known as the classic test for optical imaging devices, is used to characterize an acoustic lens for operation at 1.2 GHz. A confocal laser scanning microscope (CLSM) was used as the illumination and detection device utilizing its pinhole instead of the classical knife edge that is normally employed in the Foucault test. Information about the geometrical characteristics, such as the half opening angle of the acoustic lens, were determined as well as the quality of the calotte of the lens used for focusing. The smallest focal spot size that could be achieved with the examined lens employed as a spherical reflector was found to be about 1 μm. By comparison to the idealized resolution a degradation of about a factor of 2 can be deduced. This limits the actual quality of the acoustic focus.

  2. Visualization of microcrack anisotropy in granite affected by afault zone, using confocal laser scanning microscope

    SciTech Connect

    Onishi, Celia T.; Shimizu, Ichiko

    2004-01-02

    Brittle deformation in granite can generate a fracture system with different patterns. Detailed fracture analyses at both macroscopic and microscopic scales, together with physical property data from a drill-core, are used to classify the effects of reverse fault deformation in four domains: (1) undeformed granite, (2) fractured granite with cataclastic seams, (3) fractured granite from the damage zone, and (4) foliated cataclasite from the core of the fault. Intact samples from two orthogonal directions, horizontal (H) and vertical (V), from the four domains indicate a developing fracture anisotropy toward the fault, which is highly developed in the damage zone. As a specific illustration of this phenomenon, resin impregnation, using a confocal laser scanning microscope (CLSM) technique is applied to visualize the fracture anisotropy developed in the Toki Granite, Japan. As a result, microcrack networks have been observed to develop in H sections and elongate open cracks in V sections, suggesting that flow pathways can be determined by deformation.

  3. Evaluation of the Cytotoxic Behavior of Fungal Extracellular Synthesized Ag Nanoparticles Using Confocal Laser Scanning Microscope.

    PubMed

    Salaheldin, Taher A; Husseiny, Sherif M; Al-Enizi, Abdullah M; Elzatahry, Ahmed; Cowley, Alan H

    2016-01-01

    Silver nanoparticles have been synthesized by subjecting a reaction medium to a Fusarium oxysporum biomass at 28 °C for 96 h. The biosynthesized Ag nanoparticles were characterized on the basis of their anticipated peak at 405 nm using UV-Vis-NIR spectroscopy. Structural confirmation was evident from the characteristic X-ray diffraction (XRD) pattern, high-resolution transmission electron Microscopy (HRTEM) and the particle size analyzer. The Ag nanoparticles were of dimension 40 ± 5 nm and spherical in shape. The study mainly focused on using the confocal laser scanning microscope (CLSM) to examine the cytotoxic activities of fungal synthesized Ag nanoparticles on a human breast carcinoma cell line MCF7 cell, which featured remarkable vacuolation, thus indicating a potent cytotoxic activity. PMID:26950118

  4. Is Seeing Really Believing? Probe-based Confocal Laser Endomicroscopy in the Evaluation of Pancreaticobiliary Disease.

    PubMed

    Storm, Andrew C; Lee, Linda S

    2016-01-01

    Confocal laser endomicroscopy for real-time diagnosis during endoscopic procedures has now been in the mainstream clinical arena for a decade. Indeterminate biliary strictures and pancreatic cysts remain 2 difficult diagnostic challenges for the gastroenterologist, and the role this technology will play in the approach to these problems is still evolving. There is now a body of literature to guide the endoscopist in the use of this imaging tool, including how it may be useful in excluding biliary malignancy, and how miniaturization has allowed for endoscopic ultrasound-guided application of the probe within cysts. Interobserver variability remains a weakness of the system. Tips for use of this tool and interpretation of the imaging data it provides are discussed. PMID:26927493

  5. Three-dimensional reconstruction of paramecium primaurelia oral apparatus through confocal laser scanning optical microscopy

    NASA Astrophysics Data System (ADS)

    Beltrame, Francesco; Ramoino, Paola; Fato, Marco; Delmonte Corrado, Maria U.; Marcenaro, Giampiero; Crippa Franceschi, Tina

    1992-06-01

    Studies on the complementary mating types of Paramecium primaurelia (Protozoa, Ciliates) have shown that cell lines which differ from each other in mating type expression are characterized by different cell contents, organization, and physiology. Referring to these differences and to the differential rates of food vacuole formation, oral apparatuses of the two mating type cells are assumed to possibly differ from each other in some traits, such as, for instance, in their lengths. In our work, the highly organized oral structures are analyzed by means of a laser scanning confocal optical microscope (CLSM), which provides their 3-D visualization and measurement. The extraction of the 3-D intrinsic information related to the biological objects under investigation can be in turn related to their functional state, according to the classical paradigm of structure to function relationships identification. In our experiments, we acquired different data sets. These are optical slices of the biological sample under investigation, acquired in a confocal situation, through epi-illumination, in reflection, and, for comparison with conventional microscopy, 2-D images acquired via a standard TV camera coupled to the microscope itself. Our CLSM system is equipped with a laser beam at 488 and 514 nm and the data have been acquired with various steps of optical slicing, ranging from .04 to .25 micrometers. The volumes obtained by piling-up the slices are rendered through different techniques, some of them directly implemented on the workstation controlling the CLSM system, some of them on a SUN SPARC station 1, where the original data were transferred via an Ethernet link. In this last instance, original software has been developed for the visualization and animation of the 3-D structures, running under UNIX and X-Window, according to a ray-tracing algorithm.

  6. Real time confocal laser scanning microscopy: Potential applications in space medicine and cell biology

    NASA Astrophysics Data System (ADS)

    Rollan, Ana; Ward, Thelma; McHale, Anthony P.

    Photodynamic therapy (PDT), in which tissues may be rendered fatally light-sensitive represents a relatively novel treatment for cancer and other disorders such as cardiovascular disease. It offers significant application to disease control in an isolated environment such as space flight. In studying PDT in the laboratory, low energy lasers such as HeNe lasers are used to activate the photosensitized cellular target. A major problem associated with these studies is that events occurring during actual exposure of the target cells to the system cannot be examined in real time. In this study HeLa cells were photosensitized and photodynamic activation was accomplished using the scanning microbeam from a confocal laser scanning microscope. This form of activation allowed for simultaneous photoactivation and observation and facilitated the recording of events at a microscopic level during photoactivation. Effects of photodynamic activation on the target cells were monitored using the fluorophores rhodamine 123 and ethidium homodimer-1. Potential applications of these forms of analyses to space medicine and cell biology are discussed.

  7. Three-dimensional measurements with a novel technique combination of confocal and focus variation with a simultaneous scan

    NASA Astrophysics Data System (ADS)

    Matilla, A.; Mariné, J.; Pérez, J.; Cadevall, C.; Artigas, R.

    2016-04-01

    The most common optical measurement technologies used today for the three dimensional measurement of technical surfaces are Coherence Scanning Interferometry (CSI), Imaging Confocal Microscopy (IC), and Focus Variation (FV). Each one has its benefits and its drawbacks. FV will be the ideal technology for the measurement of those regions where the slopes are high and where the surface is very rough, while CSI and IC will provide better results for smoother and flatter surface regions. In this work we investigated the benefits and drawbacks of combining Interferometry, Confocal and focus variation to get better measurement of technical surfaces. We investigated a way of using Microdisplay Scanning type of Confocal Microscope to acquire on a simultaneous scan confocal and focus Variation information to reconstruct a three dimensional measurement. Several methods are presented to fuse the optical sectioning properties of both techniques as well as the topographical information. This work shows the benefit of this combination technique on several industrial samples where neither confocal nor focus variation is able to provide optimal results.

  8. Femtosecond laser subsurface scleral treatment in cadaver human sclera and evaluation using two-photon and confocal microscopy

    NASA Astrophysics Data System (ADS)

    Sun, Hui; Fan, Zhongwei; Yan, Ying; Lian, Fuqiang; Kurtz, Ron; Juhasz, Tibor

    2016-03-01

    Glaucoma is the second-leading cause of blindness worldwide and is often associated with elevated intraocular pressure (IOP). Partial-thickness drainage channels can be created with femtosecond laser in the translucent sclera for the potential treatment of glaucoma. We demonstrate the creation of partial-thickness subsurface drainage channels with the femtosecond laser in the cadaver human eyeballs and describe the application of two-photon microscopy and confocal microscopy for noninvasive imaging of the femtosecond laser created partial-thickness scleral channels in cadaver human eyes. A femtosecond laser operating at a wavelength of 1700 nm was scanned along a rectangular raster pattern to create the partial thickness subsurface drainage channels in the sclera of cadaver human eyes. Analysis of the dimensions and location of these channels is important in understanding their effects. We describe the application of two-photon microscopy and confocal microscopy for noninvasive imaging of the femtosecond laser created partial-thickness scleral channels in cadaver human eyes. High-resolution images, hundreds of microns deep in the sclera, were obtained to allow determination of the shape and dimension of such partial thickness subsurface scleral channels. Our studies suggest that the confocal and two-photon microscopy can be used to investigate femtosecond-laser created partial-thickness drainage channels in the sclera of cadaver human eyes.

  9. Assessment of Nd:YAG laser-induced injury to rabbit nasal septal cartilage using confocal microscopy and Live/Dead assay in an ex vivo model

    NASA Astrophysics Data System (ADS)

    Li, Chao; Krasieva, Tatiana B.; Zorin, Roman; Sun, Chung-Ho; Lam, Anthony; Gardiner, David M.; Wong, Brian J.

    2004-07-01

    Identification of proliferating chondrocytes along the periphery of laser ablation sites in irradiated cartilage has led to interest in studying the use of laser heating alone to stimulate chondrocyte growth. However, excessive heat produced by a laser can also cause chondrocyte necrosis and apoptosis. The objective of this study was to evaluate acute injury to cartilage following irradiation by an Nd:YAG (λ=1.32μm) laser in intact ex-vivo tissue specimens. Rabbit nasal septal cartilage was irradiated using an Nd:YAG laser using pulse durations (4, 6, and 8 seconds) and power (4, 6, and 8 watts) settings previously determined to produce cell division. Immediately after laser irradiation, the extent of thermal injury to the cartilage samples was evaluated using a Live/Dead cell viability assay combined with confocal microscopy. Thermal injury was assessed with respect to distribution of live and dead cells surrounding the laser spot where regeneration was previously observed. The cell viability assay identified necrotic tissue within and immediately around the laser spot. Moving away from the center of the laser spot, a mixed population of necrotic and live chondrocytes was observed. As expected, a correlation between irradiation time, power and degree of injury was found. The results of this experiment will be used to determine the threshold required to produce regeneration while minimizing thermal injury.

  10. 3-D laser confocal microscopy study of the oxidation of NdFeB magnets in atmospheric conditions

    NASA Astrophysics Data System (ADS)

    Meakin, J. P.; Speight, J. D.; Sheridan, R. S.; Bradshaw, A.; Harris, I. R.; Williams, A. J.; Walton, A.

    2016-08-01

    Neodymium iron boron (NdFeB) magnets are used in a number of important applications, such as generators in gearless wind turbines, motors in electric vehicles and electronic goods (e.g.- computer hard disk drives, HDD). Hydrogen can be used as a processing gas to separate and recycle scrap sintered Nd-Fe-B magnets from end-of-life products to form a powder suitable for recycling. However, the magnets are likely to have been exposed to atmospheric conditions prior to processing, and any oxidation could lead to activation problems for the hydrogen decrepitation reaction. Many previous studies on the oxidation of NdFeB magnets have been performed at elevated temperatures; however, few studies have been formed under atmospheric conditions. In this paper a combination of 3-D laser confocal microscopy and Raman spectroscopy have been used to assess the composition, morphology and rate of oxidation/corrosion on scrap sintered NdFeB magnets. Confocal microscopy has been employed to measure the growth of surface reaction products at room temperature, immediately after exposure to air. The results showed that there was a significant height increase at the triple junctions of the Nd-rich grain boundaries. Using Raman spectroscopy, the product was shown to consist of Nd2O3 and formed only on the Nd-rich triple junctions. The diffusion coefficient of the triple junction reaction product growth at 20 °C was determined to be approximately 4 × 10-13 cm2/sec. This value is several orders of magnitude larger than values derived from the diffusion controlled oxide growth observations at elevated temperatures in the literature. This indicates that the growth of the room temperature oxidation products are likely defect enhanced processes at the NdFeB triple junctions.

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

    NASA Astrophysics Data System (ADS)

    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.

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

    PubMed

    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. PMID:26256640

  13. Evaluation of confocal laser scanning microscopy for enumeration of virus-like particles in aquatic systems

    PubMed Central

    Agis, Martin; Luef, Birgit

    2016-01-01

    Abstract Abundances of virus-like particles (VLPs, mostly bacteriophages) are high in aquatic environments; therefore, techniques for precise enumeration are essential in ecological monitoring. VLPs were determined after staining with SYBR Gold by conventional epifluorescence microscopy and compared to enumerations performed by confocal laser scanning microscopy (CLSM). In order to assess the potential of CLSM for viral direct counts (VDCs), we processed samples from different freshwater and marine systems. Optical sectioning by CLSM and production of an overlay picture of multiple scans enables the often uneven whole investigated filter area to be brought to the plane of focus. This allows for subsequent image analysis of digitally created high-quality images. Another advantage using the CLSM was that the short spot excitation of the stain via laser beam minimized fading of the stain. The VDC results show that there is no significant difference between the two methods. Regarding the known difficulties of viral abundance estimates on particulate material, CLSM was further applied to enumerate VLPs on a small set of marine transparent exopolymeric particles sampled from the Atlantic Ocean. Our data suggest that CLSM is a useful tool to count viruses in water samples as well as attached to certain types of aquatic aggregates. PMID:23108709

  14. Real time diagnosis of bladder cancer with probe-based confocal laser endomicroscopy

    NASA Astrophysics Data System (ADS)

    Liu, Jen-Jane; Wu, Katherine; Adams, Winifred; Hsiao, Shelly T.; Mach, Kathleen E.; Beck, Andrew H.; Jensen, Kristin C.; Liao, Joseph C.

    2011-02-01

    Probe-based confocal laser endomicroscopy (pCLE) is an emerging technology for in vivo optical imaging of the urinary tract. Particularly for bladder cancer, real time optical biopsy of suspected lesions will likely lead to improved management of bladder cancer. With pCLE, micron scale resolution is achieved with sterilizable imaging probes (1.4 or 2.6 mm diameter), which are compatible with standard cystoscopes and resectoscopes. Based on our initial experience to date (n = 66 patients), we have demonstrated the safety profile of intravesical fluorescein administration and established objective diagnostic criteria to differentiate between normal, benign, and neoplastic urothelium. Confocal images of normal bladder showed organized layers of umbrella cells, intermediate cells, and lamina propria. Low grade bladder cancer is characterized by densely packed monomorphic cells with central fibrovascular cores, whereas high grade cancer consists of highly disorganized microarchitecture and pleomorphic cells with indistinct cell borders. Currently, we are conducting a diagnostic accuracy study of pCLE for bladder cancer diagnosis. Patients scheduled to undergo transurethral resection of bladder tumor are recruited. Patients undergo first white light cystocopy (WLC), followed by pCLE, and finally histologic confirmation of the resected tissues. The diagnostic accuracy is determined both in real time by the operative surgeon and offline after additional image processing. Using histology as the standard, the sensitivity, specificity, positive and negative predictive value of WLC and WLC + pCLE are calculated. With additional validation, pCLE may prove to be a valuable adjunct to WLC for real time diagnosis of bladder cancer.

  15. Utilizing confocal laser endomicroscopy for evaluating the adequacy of laparoscopic liver ablation

    PubMed Central

    Johnson, Sean P.; Walker‐Samuel, Simon; Gurusamy, Kurinchi; Clarkson, Matthew J.; Thompson, Stephen; Song, Yi; Totz, Johannes; Cook, Richard J.; Desjardins, Adrien E.; Hawkes, David J.; Davidson, Brian R.

    2015-01-01

    Background Laparoscopic liver ablation therapy can be used for the treatment of primary and secondary liver malignancy. The increased incidence of cancer recurrence associated with this approach, has been attributed to the inability of monitoring the extent of ablated liver tissue. Methods The feasibility of assessing liver ablation with probe‐based confocal laser endomicroscopy (CLE) was studied in a porcine model of laparoscopic microwave liver ablation. Following the intravenous injection of the fluorophores fluorescein and indocyanine green, CLE images were recorded at 488 nm and 660 nm wavelength and compared to liver histology. Statistical analysis was performed to assess if fluorescence intensity change can predict the presence of ablated liver tissue. Results CLE imaging of fluorescein at 488 nm provided good visualization of the hepatic microvasculature; whereas, CLE imaging of indocyanine green at 660 nm enabled detailed visualization of hepatic sinusoid architecture and interlobular septations. Fluorescence intensity as measured in relative fluorescence units was found to be 75–100% lower in ablated compared to healthy liver regions. General linear mixed modeling and ROC analysis found the decrease in fluorescence to be statistically significant. Conclusion Laparoscopic, dual wavelength CLE imaging using two different fluorophores enables clinically useful visualization of multiple liver tissue compartments, in greater detail than is possible at a single wavelength. CLE imaging may provide valuable intraoperative information on the extent of laparoscopic liver ablation. Lasers Surg. Med. 48:299–310, 2016. © 2015 The Authors. Lasers in Surgery and Medicine Published by Wiley Periodicals, Inc. PMID:26718623

  16. Evaluation of hypericin-mediated photodynamic therapy in combination with angiogenesis inhibitor bevacizumab using in vivo fluorescence confocal endomicroscopy

    NASA Astrophysics Data System (ADS)

    Bhuvaneswari, Ramaswamy; Thong, Patricia S. P.; Gan, Yik-Yuen; Soo, Khee; Olivo, Malini

    2010-01-01

    Photodynamic therapy (PDT) is an alternative cancer treatment modality that offers localized treatment using a photosensitizer and light. However, tumor angiogenesis is a major concern following PDT-induced hypoxia as it promotes recurrence. Bevacizumab is a monoclonal antibody that targets vascular endothelial growth factor (VEGF), thus preventing angiogenesis. The combination of PDT with antiangiogenic agents such as bevacizumab has shown promise in preclinical studies. We use confocal endomicroscopy to study the antiangiogenic effects of PDT in combination with bevacizumab. This technique offers in vivo surface and subsurface fluorescence imaging of tissue. Mice bearing xenograft bladder carcinoma tumors were treated with PDT, bevacizumab, or PDT and bevacizumab combination therapy. In tumor regression experiments, combination therapy treated tumors show the most regression. Confocal fluorescence endomicroscopy enables visualization of tumor blood vessels following treatment. Combination therapy treated tumors show the most posttreatment damage with reduced cross-sectional area of vessels. Immunohistochemistry and immunofluorescence studies show that VEGF expression is significantly downregulated in the tumors treated by combination therapy. Overall, combining PDT and bevacizumab is a promising cancer treatment approach. We also demonstrate that confocal endomicroscopy is useful for visualization of vasculature and evaluation of angiogenic response following therapeutic intervention.

  17. Nonlinear combining of laser beams.

    PubMed

    Lushnikov, Pavel M; Vladimirova, Natalia

    2014-06-15

    We propose to combine multiple laser beams into a single diffraction-limited beam by beam self-focusing (collapse) in a Kerr medium. Beams with total power above critical are first combined in the near field and then propagated in the optical fiber/waveguide with Kerr nonlinearity. Random fluctuations during propagation eventually trigger a strong self-focusing event and produce a diffraction-limited beam carrying the critical power. PMID:24978503

  18. Toward real-time virtual biopsy of oral lesions using confocal laser endomicroscopy interfaced with embedded computing

    NASA Astrophysics Data System (ADS)

    Thong, Patricia S. P.; Tandjung, Stephanus S.; Movania, Muhammad Mobeen; Chiew, Wei-Ming; Olivo, Malini; Bhuvaneswari, Ramaswamy; Seah, Hock-Soon; Lin, Feng; Qian, Kemao; Soo, Khee-Chee

    2012-05-01

    Oral lesions are conventionally diagnosed using white light endoscopy and histopathology. This can pose a challenge because the lesions may be difficult to visualise under white light illumination. Confocal laser endomicroscopy can be used for confocal fluorescence imaging of surface and subsurface cellular and tissue structures. To move toward real-time "virtual" biopsy of oral lesions, we interfaced an embedded computing system to a confocal laser endomicroscope to achieve a prototype three-dimensional (3-D) fluorescence imaging system. A field-programmable gated array computing platform was programmed to enable synchronization of cross-sectional image grabbing and Z-depth scanning, automate the acquisition of confocal image stacks and perform volume rendering. Fluorescence imaging of the human and murine oral cavities was carried out using the fluorescent dyes fluorescein sodium and hypericin. Volume rendering of cellular and tissue structures from the oral cavity demonstrate the potential of the system for 3-D fluorescence visualization of the oral cavity in real-time. We aim toward achieving a real-time virtual biopsy technique that can complement current diagnostic techniques and aid in targeted biopsy for better clinical outcomes.

  19. Oral biofilm analysis of palatal expanders by fluorescence in-situ hybridization and confocal laser scanning microscopy.

    PubMed

    Klug, Barbara; Rodler, Claudia; Koller, Martin; Wimmer, Gernot; Kessler, Harald H; Grube, Martin; Santigli, Elisabeth

    2011-01-01

    Confocal laser scanning microscopy (CLSM) of natural heterogeneous biofilm is today facilitated by a comprehensive range of staining techniques, one of them being fluorescence in situ hybridization (FISH). We performed a pilot study in which oral biofilm samples collected from fixed orthodontic appliances (palatal expanders) were stained by FISH, the objective being to assess the three-dimensional organization of natural biofilm and plaque accumulation. FISH creates an opportunity to stain cells in their native biofilm environment by the use of fluorescently labeled 16S rRNA-targeting probes. Compared to alternative techniques like immunofluorescent labeling, this is an inexpensive, precise and straightforward labeling technique to investigate different bacterial groups in mixed biofilm consortia. General probes were used that bind to Eubacteria (EUB338 + EUB338II + EUB338III; hereafter EUBmix), Firmicutes (LGC354 A-C; hereafter LGCmix), and Bacteroidetes (Bac303). In addition, specific probes binding to Streptococcus mutans (MUT590) and Porphyromonas gingivalis (POGI) were used. The extreme hardness of the surface materials involved (stainless steel and acrylic resin) compelled us to find new ways of preparing the biofilm. As these surface materials could not be readily cut with a cryotome, various sampling methods were explored to obtain intact oral biofilm. The most workable of these approaches is presented in this communication. Small flakes of the biofilm-carrying acrylic resin were scraped off with a sterile scalpel, taking care not to damage the biofilm structure. Forceps were used to collect biofilm from the steel surfaces. Once collected, the samples were fixed and placed directly on polysine coated glass slides. FISH was performed directly on these slides with the probes mentioned above. Various FISH protocols were combined and modified to create a new protocol that was easy to handle. Subsequently the samples were analyzed by confocal laser scanning

  20. Observation of the early stage of insulin crystallization by confocal laser scanning microscopy

    NASA Astrophysics Data System (ADS)

    Mühlig, P.; Klupsch, Th.; Schell, U.; Hilgenfeld, R.

    2001-11-01

    It is demonstrated that high resolution confocal laser scanning microscopy (CLSM) is a powerful tool for in situ observation and analysis of protein crystal growth. CLSM is used to study the early crystallization stage of Des-ThrB30 human insulin in aqueous solution, under conditions known to lead to monoclinic crystals. A modified batch crystallization method for CLSM purposes is applied which allows the growth behavior of crystallites to be studied in reflected light. A few hours after the start of the experiment, microcrystallites of characteristic shapes (mainly prismatic and pyramidal) are observed, the number of which strongly depends on the concentration of higher insulin aggregates in the initial solution. From direct observation as well as from model calculations we conclude that for solute concentrations up to about 3.5-times the saturation value, growth starts from few active insulin precipitate particles while 3D nucleation is neglegible for observation times up to 24 h. The anisotropic growth rates of monoclinic, prismatic crystallites are measured along the long edge of the cover face and perpendicular to the latter. A simultaneous crossover to signifcantly higher growth rates is found when the crystallite size reaches about 2 μm. The higher growth rates are connected with the appearence of striations. We argue that this growth rate crossover is caused by an increased 2D nucleation rate at the edges and corners, which finally results in bunching of steps simultaneously spreading over adjacent crystallite faces.

  1. Confocal Laser Endomicroscopy in Gastrointestinal and Pancreatobiliary Diseases: A Systematic Review and Meta-Analysis.

    PubMed

    Fugazza, Alessandro; Gaiani, Federica; Carra, Maria Clotilde; Brunetti, Francesco; Lévy, Michaël; Sobhani, Iradj; Azoulay, Daniel; Catena, Fausto; de'Angelis, Gian Luigi; de'Angelis, Nicola

    2016-01-01

    Confocal laser endomicroscopy (CLE) is an endoscopic-assisted technique developed to obtain histopathological diagnoses of gastrointestinal and pancreatobiliary diseases in real time. The objective of this systematic review is to analyze the current literature on CLE and to evaluate the applicability and diagnostic yield of CLE in patients with gastrointestinal and pancreatobiliary diseases. A literature search was performed on MEDLINE, EMBASE, Scopus, and Cochrane Oral Health Group Specialized Register, using pertinent keywords without time limitations. Both prospective and retrospective clinical studies that evaluated the sensitivity, specificity, or accuracy of CLE were eligible for inclusion. Of 662 articles identified, 102 studies were included in the systematic review. The studies were conducted between 2004 and 2015 in 16 different countries. CLE demonstrated high sensitivity and specificity in the detection of dysplasia in Barrett's esophagus, gastric neoplasms and polyps, colorectal cancers in inflammatory bowel disease, malignant pancreatobiliary strictures, and pancreatic cysts. Although CLE has several promising applications, its use has been limited by its low availability, high cost, and need of specific operator training. Further clinical trials with a particular focus on cost-effectiveness and medicoeconomic analyses, as well as standardized institutional training, are advocated to implement CLE in routine clinical practice. PMID:26989684

  2. Spatial Gradients in Particle Reinforced Polymers Characterized by X-Ray Attenuation and Laser Confocal Microscopy

    SciTech Connect

    LAGASSE,ROBERT R.; THOMPSON,KYLE R.

    2000-06-12

    The goal of this work is to develop techniques for measuring gradients in particle concentration within filled polymers, such as encapsulant. A high concentration of filler particles is added to such materials to tailor physical properties such as thermal expansion coefficient. Sedimentation and flow-induced migration of particles can produce concentration gradients that are most severe near material boundaries. Therefore, techniques for measuring local particle concentration should be accurate near boundaries. Particle gradients in an alumina-filled epoxy resin are measured with a spatial resolution of 0.2 mm using an x-ray beam attenuation technique, but an artifact related to the finite diameter of the beam reduces accuracy near the specimen's edge. Local particle concentration near an edge can be measured more reliably using microscopy coupled with image analysis. This is illustrated by measuring concentration profiles of glass particles having 40 {micro}m median diameter using images acquired by a confocal laser fluorescence microscope. The mean of the measured profiles of volume fraction agrees to better than 3% with the expected value, and the shape of the profiles agrees qualitatively with simple theory for sedimentation of monodisperse particles. Extending this microscopy technique to smaller, micron-scale filler particles used in encapsulant for microelectronic devices is illustrated by measuring the local concentration of an epoxy resin containing 0.41 volume fraction of silica.

  3. Confocal Laser Endomicroscopy for In Vivo Diagnosis of Clostridium difficile Associated Colitis — A Pilot Study

    PubMed Central

    Neumann, Helmut; Günther, Claudia; Vieth, Michael; Grauer, Martin; Wittkopf, Nadine; Mudter, Jonas; Becker, Christoph; Schoerner, Christoph; Atreya, Raja; Neurath, Markus F.

    2013-01-01

    Background Clostridium difficile infection (CDI) is one of the most dreaded causes of hospital-acquired diarrhea. Main objective was to investigate whether confocal laser endomicroscopy (CLE) has the capability for in vivo diagnosis of C. difficile associated histological changes. Second objective was to prove the presence of intramucosal bacteria using CLE. Methods 80 patients were prospectively included, 10 patients were diagnosed with CDI based on toxigenic culture. To validate the presence of intramucosal bacteria ex vivo, CLE was performed in pure C. difficile culture; additionally fluorescence in situ hybridization (FISH) was performed. Finally, CLE with fluorescence labelled oligonucleotide probe specific for C. difficile was performed ex vivo in order to prove the presence of bacteria. Results CLE identified CDI-associated histological changes in vivo (sensitivity and accuracy of 88.9% and 96.3%). In addition, intramucosal bacteria were visualized. The presence of these bacteria could be proven by CLE with labeled, specific molecular C. difficile probe and FISH-technique. Based on comparison between CLE and FISH analyses, sensitivity and specificity for the presence of intramucosal bacteria were 100%. Conclusion CLE has the potential for in vivo diagnosis of CDI associated colitis. In addition, CLE allowed the detection of intramucosal bacteria in vivo. PMID:23527018

  4. Three-dimensional imaging of the intact mouse cochlea by fluorescent laser scanning confocal microscopy.

    PubMed

    MacDonald, Glen H; Rubel, Edwin W

    2008-09-01

    The complex anatomy of the mammalian cochlea is most readily understood by representation in three-dimensions. However, the cochlea is often sectioned to minimize the effects of its anatomic complexity and optical properties on image acquisition by light microscopy. We have found that optical aberrations present in the decalcified cochlea can be greatly reduced by dehydration through graded ethanols followed by clearing with a mixture of five parts methyl salicylate and three parts benzyl benzoate (MSBB). Clearing the cochlea with MSBB enables acquisition of high-resolution images with multiple fluorescent labels, through the full volume of the cochlea by laser scanning confocal microscopy. The resulting images are readily applicable to three-dimensional morphometric analysis and volumetric visualizations. This method promises to be particularly useful for three-dimensional characterization of anatomy, innervation and expression of genes or proteins in the many new animal models of hearing and balance generated by genetic manipulation. Furthermore, the MSBB is compatible with most non-protein fluorophores used for histological labeling, and may be removed with traditional transitional solvents to allow subsequent epoxy embedding for sectioning. PMID:18573326

  5. In Vivo Laser Scanning Confocal Microscopy of Human Meibomian Glands in Aging and Ocular Surface Diseases

    PubMed Central

    Fasanella, Vincenzo; Mastropasqua, Rodolfo; Brescia, Lorenza; Di Staso, Federico; Ciancaglini, Marco; Mastropasqua, Leonardo

    2016-01-01

    Meibomian glands (MGs) play a crucial role in the ocular surface homeostasis by providing lipids to the superficial tear film. Their dysfunction destabilizes the tear film leading to a progressive loss of the ocular surface equilibrium and increasing the risk for dry eye. In fact, nowadays, the meibomian gland dysfunction is one of the leading causes of dry eye. Over the past decades, MGs have been mainly studied by using meibography, which, however, cannot image the glandular structure at a cellular level. The diffusion of the in vivo laser scanning confocal microscopy (LSCM) provided a new approach for the structural assessment of MGs permitting a major step in the noninvasive evaluation of these structures. LSCM is capable of showing MGs modifications during aging and in the most diffuse ocular surface diseases such as dry eye, allergy, and autoimmune conditions and in the drug-induced ocular surface disease. On the other hand, LSCM may help clinicians in monitoring the tissue response to therapy. In this review, we summarized the current knowledge about the role of in vivo LSCM in the assessment of MGs during aging and in the most diffuse ocular surface diseases. PMID:27047965

  6. In Vivo Laser Scanning Confocal Microscopy of Human Meibomian Glands in Aging and Ocular Surface Diseases.

    PubMed

    Fasanella, Vincenzo; Agnifili, Luca; Mastropasqua, Rodolfo; Brescia, Lorenza; Di Staso, Federico; Ciancaglini, Marco; Mastropasqua, Leonardo

    2016-01-01

    Meibomian glands (MGs) play a crucial role in the ocular surface homeostasis by providing lipids to the superficial tear film. Their dysfunction destabilizes the tear film leading to a progressive loss of the ocular surface equilibrium and increasing the risk for dry eye. In fact, nowadays, the meibomian gland dysfunction is one of the leading causes of dry eye. Over the past decades, MGs have been mainly studied by using meibography, which, however, cannot image the glandular structure at a cellular level. The diffusion of the in vivo laser scanning confocal microscopy (LSCM) provided a new approach for the structural assessment of MGs permitting a major step in the noninvasive evaluation of these structures. LSCM is capable of showing MGs modifications during aging and in the most diffuse ocular surface diseases such as dry eye, allergy, and autoimmune conditions and in the drug-induced ocular surface disease. On the other hand, LSCM may help clinicians in monitoring the tissue response to therapy. In this review, we summarized the current knowledge about the role of in vivo LSCM in the assessment of MGs during aging and in the most diffuse ocular surface diseases. PMID:27047965

  7. Multipass cell based on confocal mirrors for sensitive broadband laser spectroscopy in the near infrared.

    PubMed

    Mohamed, T; Zhu, F; Chen, S; Strohaber, J; Kolomenskii, A A; Bengali, A A; Schuessler, H A

    2013-10-10

    We report on broadband absorption spectroscopy in the near IR using a multipass cell design based on highly reflecting mirrors in a confocal arrangement having the particular aim of achieving long optical paths. We demonstrate a path length of 314 m in a cell consisting of two sets of highly reflecting mirrors with identical focal length, spaced 0.5 m apart. The multipass cell covers this path length in a relatively small volume of 1.25 l with the light beam sampling the whole volume. In a first application, the absorption spectra of the greenhouse gases CO(2), CH(4), and CO were measured. In these measurements we used a femtosecond fiber laser with a broadband spectral range spanning the near IR from 1.5 to 1.7 μm. The absorption spectra show a high signal-to-noise ratio, from which we derive a sensitivity limit of 6 ppmv for methane observed in a mixture with air. PMID:24217732

  8. Optical Biopsy of Peripheral Nerve Using Confocal Laser Endomicroscopy: A New Tool for Nerve Surgeons?

    PubMed

    Crowe, Christopher S; Liao, Joseph C; Curtin, Catherine M

    2015-09-01

    Peripheral nerve injuries remain a challenge for reconstructive surgeons with many patients obtaining suboptimal results. Understanding the level of injury is imperative for successful repair. Current methods for distinguishing healthy from damaged nerve are time consuming and possess limited efficacy. Confocal laser endomicroscopy (CLE) is an emerging optical biopsy technology that enables dynamic, high resolution, sub-surface imaging of live tissue. Porcine sciatic nerve was either left undamaged or briefly clamped to simulate injury. Diluted fluorescein was applied topically to the nerve. CLE imaging was performed by direct contact of the probe with nerve tissue. Images representative of both damaged and undamaged nerve fibers were collected and compared to routine H&E histology. Optical biopsy of undamaged nerve revealed bands of longitudinal nerve fibers, distinct from surrounding adipose and connective tissue. When damaged, these bands appear truncated and terminate in blebs of opacity. H&E staining revealed similar features in damaged nerve fibers. These results prompt development of a protocol for imaging peripheral nerves intraoperatively. To this end, improving surgeons' ability to understand the level of injury through real-time imaging will allow for faster and more informed operative decisions than the current standard permits. PMID:26430636

  9. Usefulness and Future Prospects of Confocal Laser Endomicroscopy for Gastric Premalignant and Malignant Lesions.

    PubMed

    Lee, Sang Kil

    2015-11-01

    Confocal laser endomicroscopy (CLE) is a new technology enabling endoscopists to visualize tissue at the cellular level. CLE has the fundamental potential to provide a histologic diagnosis, and may theoretically replace or reduce the need for performing biopsy for histology. The clinical benefits of CLE are more obvious in esophageal disease, including Barrett's esophagus. Currently, this technology has been adapted to the diagnosis and surveillance of Barrett's esophagus and related neoplasia. Standard white light endoscopy is the primary tool for gastric cancer screening. Currently, the only method available to precisely diagnose these lesions is upper endoscopy with an appropriate biopsy. A recent study showed that CLE could characterize dysplasia or cancer and identify the risk factors for gastric cancer, such as intestinal metaplasia and the presence of Helicobacter pylori in vivo, although fewer studies on CLE were performed on the stomach than on Barrett's esophagus and other esophageal diseases. However, the application of CLE to routine clinical endoscopy continues to be refined. This review focused on the usefulness and future prospects of CLE for gastric premalignant and malignant lesions. PMID:26668797

  10. Collagen and Elastic Fibers in Odontogenic Entities: Analysis Using Light and Confocal Laser Microscopic Methods

    PubMed Central

    Moure, Sabrina P; Carrard, Vinicius C; Lauxen, Isabel S; Manso, Pedro Paulo A; Oliveira, Marcia G; Martins, Manoela D; Sant´Ana Filho, Manoel

    2011-01-01

    Dentigerous cyst (DC) and keratocystic odontogenic tumor (KOT) are odontogenic lesions arising from epithelial elements, such as those observed in dental follicles (DF), that have been part of the tooth forming apparatus. These lesions show different clinical and histological characteristics, as well as distinct biological behavior. This study aimed to qualify and quantify collagen and elastic fibers by means of histochemical techniques with light and confocal laser microscopic methods in three odontogenic entities. Eleven DF, 13 DC (n=10 with inflammation, n=3 without inflammation) and 13 KOT were processed to the following techniques: Hematoxylin and Eosin, Masson’s Trichrome, Picrosirius, Direct Blue, and Orcein. DF and DC without inflammation exhibited collagen with similar characteristics: no parallel pattern of fiber orientation, thick fibers with dense arrangement, and absence of distinct layers. A comparison between DC with inflammation and KOT revealed similar collagen organization, showing distinct layers: thin collagen fibers with loose arrangement near the epithelium and thick fibers with dense arrangement in distant areas. The only difference found was that KOT exhibited a parallel collagen orientation in relation to the odontogenic epithelia. It may be suggested that the connective tissue of DC is a reactive tissue, inducing an expansive growth associated with fluid accumulation and inflammatory process, which in turn may be present as part of the lesion itself. In KOT, loosely arranged collagen may be associated with the behavior of the neoplastic epithelium. PMID:21760864

  11. Visualization and quantification of healthy and carious dentin structure using confocal laser scanning microscopy

    NASA Astrophysics Data System (ADS)

    Kimura, Yuichi; Wilder-Smith, Petra B. B.; Krasieva, Tatiana B.; Arrastia-Jitosho, Anna-Marie A.; Liaw, Lih-Huei L.; Matsumoto, Koukichi; Berns, Michael W.

    1996-04-01

    In this study, a fluorescence technique was developed for visualization of dentin using confocal laser scanning microscopy (CLSM). Eighteen extracted human teeth were used: 13 showing no clinical signs of caries and 5 with visually apparent decay. Preliminary study: All teeth were horizontally sectioned to approx. 200 micrometers thickness and pre-treated as follows: no pretreatment; vacuum only; ultrasonication only; sodium hypochlorite (NaOCl) only; vacuum and NaOCl; ultrasonication and NaOCl; or vacuum, ultrasonication and NaOCl. Samples were stained with Rhodamine 123 fluorescent dye at a concentration of 10-5 M in phosphate buffer saline for 1 to 24 hours. Caries study: Dentin surfaces, some with pre-existing caries, were visualized using CLSM. Most dentin tubules in sound dentin appeared open using CLSM, but most dentin tubules in carious dentin appeared closed or narrowed. Surface images obtained using CLSM were similar to those seen by SEM, but additional subsurface imaging was possible using CLSM at depth intervals of 1 micrometers to a depth of 30 - 50 micrometers . This technique shows good potential for non-invasive surface and subsurface imaging of dentin structures.

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

    PubMed

    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. PMID:25959794

  13. In vivo assessment of the structure of skin microcirculation by reflectance confocal-laser-scanning microscopy

    NASA Astrophysics Data System (ADS)

    Sugata, Keiichi; Osanai, Osamu; Kawada, Hiromitsu

    2012-02-01

    One of the major roles of the skin microcirculation is to supply oxygen and nutrition to the surrounding tissue. Regardless of the close relationship between the microcirculation and the surrounding tissue, there are few non-invasive methods that can evaluate both the microcirculation and its surrounding tissue at the same site. We visualized microcapillary plexus structures in human skin using in vivo reflectance confocal-laser-scanning microscopy (CLSM), Vivascope 3000® (Lucid Inc., USA) and Image J software (National Institutes of Health, USA) for video image processing. CLSM is a non-invasive technique that can visualize the internal structure of the skin at the cellular level. In addition to internal morphological information such as the extracellular matrix, our method reveals capillary structures up to the depth of the subpapillary plexus at the same site without the need for additional optical systems. Video images at specific depths of the inner forearm skin were recorded. By creating frame-to-frame difference images from the video images using off-line video image processing, we obtained images that emphasize the brightness depending on changes of intensity coming from the movement of blood cells. Merging images from different depths of the skin elucidates the 3-dimensional fine line-structure of the microcirculation. Overall our results show the feasibility of a non-invasive, high-resolution imaging technique to characterize the skin microcirculation and the surrounding tissue.

  14. Applicability of confocal laser scanning microscopy for evaluation and monitoring of cutaneous wound healing

    NASA Astrophysics Data System (ADS)

    Lange-Asschenfeldt, Susanne; Bob, Adrienne; Terhorst, Dorothea; Ulrich, Martina; Fluhr, Joachim; Mendez, Gil; Roewert-Huber, Hans-Joachim; Stockfleth, Eggert; Lange-Asschenfeldt, Bernhard

    2012-07-01

    There is a high demand for noninvasive imaging techniques for wound assessment. In vivo reflectance confocal laser scanning microscopy (CLSM) represents an innovative optical technique for noninvasive evaluation of normal and diseased skin in vivo at near cellular resolution. This study was designed to test the feasibility of CLSM for noninvasive analysis of cutaneous wound healing in 15 patients (7 male/8 female), including acute and chronic, superficial and deep dermal skin wounds. A commercially available CLSM system was used for the assessment of wound bed and wound margins in order to obtain descriptive cellular and morphological parameters of cutaneous wound repair noninvasively and over time. CLSM was able to visualize features of cutaneous wound repair in epidermal and superficial dermal wounds, including aspects of inflammation, neovascularisation, and tissue remodelling in vivo. Limitations include the lack of mechanic fixation of the optical system on moist surfaces restricting the analysis of chronic skin wounds to the wound margins, as well as a limited optical resolution in areas of significant slough formation. By describing CLSM features of cutaneous inflammation, vascularisation, and epithelialisation, the findings of this study support the role of CLSM in modern wound research and management.

  15. Application of Laser Scanning Confocal Microscopy to Heat and Mass Transport Modeling in Porous Microstructures

    NASA Technical Reports Server (NTRS)

    Marshall, Jochen; Milos, Frank; Fredrich, Joanne; Rasky, Daniel J. (Technical Monitor)

    1997-01-01

    Laser Scanning Confocal Microscopy (LSCM) has been used to obtain digital images of the complicated 3-D (three-dimensional) microstructures of rigid, fibrous thermal protection system (TPS) materials. These orthotropic materials are comprised of refractory ceramic fibers with diameters in the range of 1 to 10 microns and have open porosities of 0.8 or more. Algorithms are being constructed to extract quantitative microstructural information from the digital data so that it may be applied to specific heat and mass transport modeling efforts; such information includes, for example, the solid and pore volume fractions, the internal surface area per volume, fiber diameter distributions, and fiber orientation distributions. This type of information is difficult to obtain in general, yet it is directly relevant to many computational efforts which seek to model macroscopic thermophysical phenomena in terms of microscopic mechanisms or interactions. Two such computational efforts for fibrous TPS materials are: i) the calculation of radiative transport properties; ii) the modeling of gas permeabilities.

  16. Retinal Vasculature of Adult Zebrafish: In Vivo Imaging Using Confocal Scanning Laser Ophthalmoscopy

    PubMed Central

    Bell, Brent A.; Xie, Jing; Yuan, Alex; Kaul, Charles; Hollyfield, Joe G.; Anand-Apte, Bela

    2014-01-01

    Over the past 3 decades the zebrafish (Danio rerio) has become an important biomedical research species. As their use continues to grow additional techniques and tools will be required to keep pace with ongoing research using this species. In this paper we describe a novel method for in vivo imaging of the retinal vasculature in adult animals using a commercially available confocal scanning laser ophthalmoscope (SLO). With this instrumentation, we demonstrate the ability to distinguish diverse vascular phenotypes in different transgenic GFP lines. In addition this technology allows repeated visualization of the vasculature in individual zebrafish over time to document vascular leakage progression and recovery induced by intraocular delivery of proteins that induce vascular permeability. SLO of the retinal vasculature was found to be highly informative, providing images of high contrast and resolution that were capable of resolving individual vascular endothelial cells. Finally, the procedures required to acquire SLO images from zebrafish are non-invasive, simple to perform and can be achieved with low animal mortality, allowing repeated imaging of individual fish. PMID:25447564

  17. A statistical pixel intensity model for segmentation of confocal laser scanning microscopy images.

    PubMed

    Calapez, Alexandre; Rosa, Agostinho

    2010-09-01

    Confocal laser scanning microscopy (CLSM) has been widely used in the life sciences for the characterization of cell processes because it allows the recording of the distribution of fluorescence-tagged macromolecules on a section of the living cell. It is in fact the cornerstone of many molecular transport and interaction quantification techniques where the identification of regions of interest through image segmentation is usually a required step. In many situations, because of the complexity of the recorded cellular structures or because of the amounts of data involved, image segmentation either is too difficult or inefficient to be done by hand and automated segmentation procedures have to be considered. Given the nature of CLSM images, statistical segmentation methodologies appear as natural candidates. In this work we propose a model to be used for statistical unsupervised CLSM image segmentation. The model is derived from the CLSM image formation mechanics and its performance is compared to the existing alternatives. Results show that it provides a much better description of the data on classes characterized by their mean intensity, making it suitable not only for segmentation methodologies with known number of classes but also for use with schemes aiming at the estimation of the number of classes through the application of cluster selection criteria. PMID:20363677

  18. Confocal Laser Endomicroscopy in Gastrointestinal and Pancreatobiliary Diseases: A Systematic Review and Meta-Analysis

    PubMed Central

    Fugazza, Alessandro; Gaiani, Federica; Carra, Maria Clotilde; Brunetti, Francesco; Lévy, Michaël; Sobhani, Iradj; Azoulay, Daniel; Catena, Fausto; de'Angelis, Gian Luigi; de'Angelis, Nicola

    2016-01-01

    Confocal laser endomicroscopy (CLE) is an endoscopic-assisted technique developed to obtain histopathological diagnoses of gastrointestinal and pancreatobiliary diseases in real time. The objective of this systematic review is to analyze the current literature on CLE and to evaluate the applicability and diagnostic yield of CLE in patients with gastrointestinal and pancreatobiliary diseases. A literature search was performed on MEDLINE, EMBASE, Scopus, and Cochrane Oral Health Group Specialized Register, using pertinent keywords without time limitations. Both prospective and retrospective clinical studies that evaluated the sensitivity, specificity, or accuracy of CLE were eligible for inclusion. Of 662 articles identified, 102 studies were included in the systematic review. The studies were conducted between 2004 and 2015 in 16 different countries. CLE demonstrated high sensitivity and specificity in the detection of dysplasia in Barrett's esophagus, gastric neoplasms and polyps, colorectal cancers in inflammatory bowel disease, malignant pancreatobiliary strictures, and pancreatic cysts. Although CLE has several promising applications, its use has been limited by its low availability, high cost, and need of specific operator training. Further clinical trials with a particular focus on cost-effectiveness and medicoeconomic analyses, as well as standardized institutional training, are advocated to implement CLE in routine clinical practice. PMID:26989684

  19. Usefulness and Future Prospects of Confocal Laser Endomicroscopy for Gastric Premalignant and Malignant Lesions

    PubMed Central

    Lee, Sang Kil

    2015-01-01

    Confocal laser endomicroscopy (CLE) is a new technology enabling endoscopists to visualize tissue at the cellular level. CLE has the fundamental potential to provide a histologic diagnosis, and may theoretically replace or reduce the need for performing biopsy for histology. The clinical benefits of CLE are more obvious in esophageal disease, including Barrett’s esophagus. Currently, this technology has been adapted to the diagnosis and surveillance of Barrett’s esophagus and related neoplasia. Standard white light endoscopy is the primary tool for gastric cancer screening. Currently, the only method available to precisely diagnose these lesions is upper endoscopy with an appropriate biopsy. A recent study showed that CLE could characterize dysplasia or cancer and identify the risk factors for gastric cancer, such as intestinal metaplasia and the presence of Helicobacter pylori in vivo, although fewer studies on CLE were performed on the stomach than on Barrett’s esophagus and other esophageal diseases. However, the application of CLE to routine clinical endoscopy continues to be refined. This review focused on the usefulness and future prospects of CLE for gastric premalignant and malignant lesions. PMID:26668797

  20. Optical Biopsy of Peripheral Nerve Using Confocal Laser Endomicroscopy: A New Tool for Nerve Surgeons?

    PubMed Central

    Liao, Joseph C; Curtin, Catherine M

    2015-01-01

    Peripheral nerve injuries remain a challenge for reconstructive surgeons with many patients obtaining suboptimal results. Understanding the level of injury is imperative for successful repair. Current methods for distinguishing healthy from damaged nerve are time consuming and possess limited efficacy. Confocal laser endomicroscopy (CLE) is an emerging optical biopsy technology that enables dynamic, high resolution, sub-surface imaging of live tissue. Porcine sciatic nerve was either left undamaged or briefly clamped to simulate injury. Diluted fluorescein was applied topically to the nerve. CLE imaging was performed by direct contact of the probe with nerve tissue. Images representative of both damaged and undamaged nerve fibers were collected and compared to routine H&E histology. Optical biopsy of undamaged nerve revealed bands of longitudinal nerve fibers, distinct from surrounding adipose and connective tissue. When damaged, these bands appear truncated and terminate in blebs of opacity. H&E staining revealed similar features in damaged nerve fibers. These results prompt development of a protocol for imaging peripheral nerves intraoperatively. To this end, improving surgeons' ability to understand the level of injury through real-time imaging will allow for faster and more informed operative decisions than the current standard permits. PMID:26430636

  1. Probe-Based Confocal Laser Endomicroscopy for Indeterminate Biliary Strictures: Refinement of the Image Interpretation Classification

    PubMed Central

    Giovannini, Marc; Jamidar, Priya; Gan, S. Ian; Cesaro, Paola; Caillol, Fabrice; Filoche, Bernard; Karia, Kunal; Smith, Ioana; Slivka, Adam

    2015-01-01

    Background. Accurate diagnosis and clinical management of indeterminate biliary strictures are often a challenge. Tissue confirmation modalities during Endoscopic Retrograde Cholangiopancreatography (ERCP) suffer from low sensitivity and poor diagnostic accuracy. Probe-based confocal laser endomicroscopy (pCLE) has been shown to be sensitive for malignant strictures characterization (98%) but lacks specificity (67%) due to inflammatory conditions inducing false positives. Methods. Six pCLE experts validated the Paris Classification, designed for diagnosing inflammatory biliary strictures, using a set of 40 pCLE sequences obtained during the prospective registry (19 inflammatory, 6 benign, and 15 malignant). The 4 criteria used included (1) multiple thin white bands, (2) dark granular pattern with scales, (3) increased space between scales, and (4) thickened reticular structures. Interobserver agreement was further calculated on a separate set of 18 pCLE sequences. Results. Overall accuracy was 82.5% (n = 40 retrospectively diagnosed) versus 81% (n = 89 prospectively collected) for the registry, resulting in a sensitivity of 81.2% (versus 98% for the prospective study) and a specificity of 83.3% (versus 67% for the prospective study). The corresponding interobserver agreement for 18 pCLE clips was fair (k = 0.37). Conclusion. Specificity of pCLE using the Paris Classification for the characterization of indeterminate bile duct stricture was increased, without impacting the overall accuracy. PMID:25866506

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

    NASA Astrophysics Data System (ADS)

    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.

  3. Elastomeric photo-actuators and their investigation by confocal laser scanning microscopy

    NASA Astrophysics Data System (ADS)

    Czaniková, Klaudia; Ilčíková, Markéta; Krupa, Igor; Mičušík, Matej; Kasák, Peter; Pavlova, Ewa; Mosnáček, Jaroslav; Chorvát, Dušan, Jr.; Omastová, Mária

    2013-10-01

    The photo-actuation behavior of nanocomposites based on ethylene-vinylacetate copolymer (EVA) and styrene-isoprene-styrene (SIS) block copolymer filled with well-dispersed and modified multiwalled carbon nanotubes (MWCNTs) is discussed in this paper. The nanocomposites were prepared by casting from solution. To improve the dispersion of the MWCNTs in EVA, the MWCNT surface was modified with a non-covalent surfactant, cholesteryl 1-pyrenecarboxylate (PyChol). To prepare SIS nanocomposites, the MWCNT surface was covalently modified with polystyrene chains. The good dispersion of the filler was confirmed by transmission electron microscopy (TEM). Special, custom-made punch/die molds were used to create a Braille element (BE)-like shape, which under shear forces induces a uniaxial orientation of the MWCNTs within the matrix. The uniaxial orientation of MWCNTs is an essential precondition to ensure the photo-actuating behavior of MWCNTs in polymeric matrices. The orientation of the MWCNTs within the matrices was examined by scanning electron microscopy (SEM). Nanocomposite BEs were illuminated from the bottom by a red light-emitting diode (LED), and the photo-actuation was investigated by confocal laser scanning microscopy (CLSM). When the BEs were exposed to light, a temporary increase in the height of the element was detected. This process was observed to be reversible: after switching off the light, the BEs returned to their original shape and height.

  4. Distribution of ALA metabolic products in esophageal carcinoma cells using spectrally resolved confocal laser microscopy

    NASA Astrophysics Data System (ADS)

    Smolka, Jozef; Mateasik, Anton

    2006-08-01

    Aminolevulinic acid (ALA) is an efficient substance used in photodynamic therapy (PDT). It is a precursor of light-sensitive products that can selectively accumulate in malignant cells following the altered activity of the heme biosynthetic pathway enzymes in such cells. These products are synthesized in mitochondria and distributed to various cellular structures [1]. The localization of ALA products in subcellular structures depends on their chemical characteristics as well as on the properties of the intracellular environment [2]. Characterization of such properties is possible by means of fluorescent probes like JC-1 and carboxy SNARF-1. However, the emission spectra of these probes are overlapped with spectral pattern of typical ALA product -protoporphyrin IX (PpIX). Spectral overlap of fluorescence signals prevents to clearly separate a distribution of probes from PpIX distribution what can completely mess the applicability of these probes in characterization of cell properties. The spectrally resolved confocal laser microscopy can be used to overcome this problem. In this study, a distribution of ALA metabolic products in relation to the mitochondrial membrane potential and intracellular pH was examined. Human cell lines (KYSE-450, KYSE-70) from esophageal squamous cell carcinoma were used. Cells were incubated with 1mM solution of ALA for four hours. Two fluorescent probes, carboxy SNARF-1 and JC-1 , were used to monitor intracellular pH levels and to determine membrane potential changes, respectively. The samples were scanned by spectrally resolved laser scanning microscope. Spectral linear unmixing method was used to discriminate and separate regions of accumulation of ALA metabolic products of JC-1 and carboxy SNARF-1.

  5. Imaging of calcium wave propagation in guinea-pig ventricular cell pairs by confocal laser scanning microscopy.

    PubMed

    Takamatsu, T; Minamikawa, T; Kawachi, H; Fujita, S

    1991-08-01

    We describe here the use of a confocal laser scanning microscope for imaging fast dynamic changes of the intracellular calcium ion concentration ([Ca2+]i) in isolated ventricular cell pairs. The scanning apparatus of our system, paired galvanometer mirrors, can perform narrow band scanning of an area of interest at a high temporal resolution of less than 70 msec per image. The actual [Ca2+]i is obtained directly through the fluorescence intensity of injected fluo-3, which responds to changes of [Ca2+]i in optically sectioned unit volumes of the cell. Images of the calcium wave obtained during propagation between paired cells revealed that the wavefront is constant in shape and propagates at constant velocity without any delay at the cell-to-cell junction. The confocal laser scanning microscope with depth-discriminating ability is a valuable tool for taking pictures of the sequence of biological events in living cells. PMID:1782671

  6. Dermoscopy, confocal laser microscopy, and hi-tech evaluation of vascular skin lesions: diagnostic and therapeutic perspectives.

    PubMed

    Grazzini, Marta; Stanganelli, Ignazio; Rossari, Susanna; Gori, Alessia; Oranges, Teresa; Longo, Anna Sara; Lotti, Torello; Bencini, Pier Luca; De Giorgi, Vincenzo

    2012-01-01

    Vascular skin lesions comprise a wide and heterogeneous group of malformations and tumors that can be correctly diagnosed based on natural history and physical examination. However, considering the high incidence of such lesions, a great number of them can be misdiagnosed. In addition, it is not so rare that an aggressive amelanotic melanoma can be misdiagnosed as a vascular lesion. In this regard, dermoscopy and confocal laser microscopy examination can play a central role in increasing the specificity of the diagnosis of such lesions. In fact, the superiority of these tools over clinical examination has encouraged dermatologists to adopt these devices for routine clinical practice, with a progressive spread of their use. In this review, we will go through the dermoscopic and the confocal laser microscopy of diagnosis of most frequent vascular lesions (i.e., hemangiomas angiokeratoma, pyogenic granuloma, angiosarcoma) taking into particular consideration the differential diagnosis with amelanotic melanoma. PMID:22950556

  7. A confocal microscope position sensor for micron-scale target alignment in ultra-intense laser-matter experiments

    NASA Astrophysics Data System (ADS)

    Willis, Christopher; Poole, Patrick L.; Akli, Kramer U.; Schumacher, Douglass W.; Freeman, Richard R.

    2015-05-01

    A diagnostic tool for precise alignment of targets in laser-matter interactions based on confocal microscopy is presented. This device permits precision alignment of targets within the Rayleigh range of tight focusing geometries for a wide variety of target surface morphologies. This confocal high-intensity positioner achieves micron-scale target alignment by selectively accepting light reflected from a narrow range of target focal planes. Additionally, the design of the device is such that its footprint and sensitivity can be tuned for the desired chamber and experiment. The device has been demonstrated to position targets repeatably within the Rayleigh range of the Scarlet laser system at The Ohio State University, where use of the device has provided a marked increase in ion yield and maximum energy.

  8. Confocal laser scanning microscopy measurement of the morphology of vanadium pentoxide nanorods grown by electron beam irradiation or thermal oxidation

    NASA Astrophysics Data System (ADS)

    Kang, Manil; Hong, Donghyuk; Kim, Taesung; Chu, Minwoo; Kim, Sok Won

    2013-01-01

    In order to observe the morphology of nanostructures at the submicroscale, we use a confocal laser scanning (CLS) microscope built in our laboratory. The theoretical resolution of the hand-made CLS microscope is 150 nm and the performance of the microscope is evaluated by observing a USAF target. Vanadium pentoxide nanorods grown by electron beam irradiation and thermal oxidation methods are used as nanostructures and the morphologies of the nanorods observed by confocal laser scanning microscopy (CLSM) are compared with those obtained by scanning electron microscopy. The magnification and resolution of the CLSM were estimated to be approximately 1500 and 800 nm, respectively. From the results, we confirm that the CLSM can be used to measure nanostructures at the sub-micro-scale without a preconditioning process.

  9. Cytosolic pH gradients in cultured neuronal cell lines studied by laser scanning confocal microscopy, real-time confocal microscopy, and spectral imaging microscopy

    NASA Astrophysics Data System (ADS)

    Sanchez-Armass, Sergio; Sennoune, Souad; Martinez, Gloria M.; Ortega, Filiberta; Martinez-Zaguilan, Raul

    2002-06-01

    Changes in intracellular pH are important for the regulation of many physiological processes including: cell growth and differentiation, exocytosis, synaptic transmission, cell motility and invasion, to name a few. In pathological states such as cancer and diabetes, pH regulation is known to be altered. Nevertheless the physiological and pathological significance of this ion, there are still many gaps in our knowledge. The advent of fluorescent pH probes to monitor this ion, has substantially accelerated its study. New advances in the methods of detection of this ion by fluorescence-based approaches have also helped us to understand more about the regulation of cytosolic pH. This study evaluates the usefulness of real time confocal imaging microscopy, laser scanning confocal microscopy, and spectral imaging microscopy to the study of pH. These approaches exhibit unsurpassed temporal, spatial, and spectral resolution and are complementary. We employed cell lines derived from the brain exhibiting soma and dendrites. The existence of cell polarity suggests that the different protein composition/micro environment in discrete subcellular domains may affect the properties of fluorescent ion indicators. We performed in situ calibration of pH probes in discrete cellular regions of the neuronal cell lines to eliminate any bias in data interpretation because of differences in cell thickness/micro environment. We show that there are distinct in situ calibration parameters in different cellular domains. These indicate that in situ titrations in discrete cellular domains are needed to assign pH values. We concluded that there are distinct pH micro domains in discrete cellular regions of neuronal cell lines.

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

    PubMed

    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. PMID:19274580

  11. Multicolor probe-based confocal laser endomicroscopy: a new world for in vivo and real-time cellular imaging

    NASA Astrophysics Data System (ADS)

    Vercauteren, Tom; Doussoux, François; Cazaux, Matthieu; Schmid, Guillaume; Linard, Nicolas; Durin, Marie-Amélie; Gharbi, Hédi; Lacombe, François

    2013-03-01

    Since its inception in the field of in vivo imaging, endomicroscopy through optical fiber bundles, or probe-based Confocal Laser Endomicroscopy (pCLE), has extensively proven the benefit of in situ and real-time examination of living tissues at the microscopic scale. By continuously increasing image quality, reducing invasiveness and improving system ergonomics, Mauna Kea Technologies has turned pCLE not only into an irreplaceable research instrument for small animal imaging, but also into an accurate clinical decision making tool with applications as diverse as gastrointestinal endoscopy, pulmonology and urology. The current implementation of pCLE relies on a single fluorescence spectral band making different sources of in vivo information challenging to distinguish. Extending the pCLE approach to multi-color endomicroscopy therefore appears as a natural plan. Coupling simultaneous multi-laser excitation with minimally invasive, microscopic resolution, thin and flexible optics, allows the fusion of complementary and valuable biological information, thus paving the way to a combination of morphological and functional imaging. This paper will detail the architecture of a new system, Cellvizio Dual Band, capable of video rate in vivo and in situ multi-spectral fluorescence imaging with a microscopic resolution. In its standard configuration, the system simultaneously operates at 488 and 660 nm, where it automatically performs the necessary spectral, photometric and geometric calibrations to provide unambiguously co-registered images in real-time. The main hardware and software features, including calibration procedures and sub-micron registration algorithms, will be presented as well as a panorama of its current applications, illustrated with recent results in the field of pre-clinical imaging.

  12. Homonymous Hemianopic Hyporeflective Retinal Abnormality on Infrared Confocal Scanning Laser Photography: A Novel Sign of Optic Tract Lesion.

    PubMed

    Monteiro, Mario L R; Araújo, Rafael B; Suzuki, Ana C F; Cunha, Leonardo P; Preti, Rony C

    2016-03-01

    Infrared confocal scanning laser photography of a patient with long-standing optic tract lesion revealed a homonymous hemianopic hyporeflective image contralateral to the visual field defect. Spectral domain optical coherence tomography showed thinning of the retinal nerve fiber and retinal ganglion cell layer and thickening of the inner nuclear layer (with microcystic degeneration) in the macular area, matching the infrared image. Hyporeflective image on infrared laser photography is associated with retinal degeneration secondary to anterior visual pathway disease and, when located in homonymous hemianopic retinas, may represent a new sign of an optic tract lesion. PMID:26172159

  13. 3D Imaging of Porous Media Using Laser Scanning Confocal Microscopy with Application to Microscale Transport Processes

    SciTech Connect

    Fredrich, J.T.

    1999-02-10

    We present advances in the application of laser scanning confocal microscopy (LSCM) to image, reconstruct, and characterize statistically the microgeometry of porous geologic and engineering materials. We discuss technical and practical aspects of this imaging technique, including both its advantages and limitations. Confocal imaging can be used to optically section a material, with sub-micron resolution possible in the lateral and axial planes. The resultant volumetric image data, consisting of fluorescence intensities for typically {approximately}50 million voxels in XYZ space, can be used to reconstruct the three-dimensional structure of the two-phase medium. We present several examples of this application, including studying pore geometry in sandstone, characterizing brittle failure processes in low-porosity rock deformed under triaxial loading conditions in the laboratory, and analyzing the microstructure of porous ceramic insulations. We then describe approaches to extract statistical microgeometric descriptions from volumetric image data, and present results derived from confocal volumetric data sets. Finally, we develop the use of confocal image data to automatically generate a three-dimensional mesh for numerical pore-scale flow simulations.

  14. Advances in combined endoscopic fluorescence confocal microscopy and optical coherence tomography

    NASA Astrophysics Data System (ADS)

    Risi, Matthew D.

    Confocal microendoscopy provides real-time high resolution cellular level images via a minimally invasive procedure. Results from an ongoing clinical study to detect ovarian cancer with a novel confocal fluorescent microendoscope are presented. As an imaging modality, confocal fluorescence microendoscopy typically requires exogenous fluorophores, has a relatively limited penetration depth (100 μm), and often employs specialized aperture configurations to achieve real-time imaging in vivo. Two primary research directions designed to overcome these limitations and improve diagnostic capability are presented. Ideal confocal imaging performance is obtained with a scanning point illumination and confocal aperture, but this approach is often unsuitable for real-time, in vivo biomedical imaging. By scanning a slit aperture in one direction, image acquisition speeds are greatly increased, but at the cost of a reduction in image quality. The design, implementation, and experimental verification of a custom multi-point-scanning modification to a slit-scanning multi-spectral confocal microendoscope is presented. This new design improves the axial resolution while maintaining real-time imaging rates. In addition, the multi-point aperture geometry greatly reduces the effects of tissue scatter on imaging performance. Optical coherence tomography (OCT) has seen wide acceptance and FDA approval as a technique for ophthalmic retinal imaging, and has been adapted for endoscopic use. As a minimally invasive imaging technique, it provides morphological characteristics of tissues at a cellular level without requiring the use of exogenous fluorophores. OCT is capable of imaging deeper into biological tissue (˜1-2 mm) than confocal fluorescence microscopy. A theoretical analysis of the use of a fiber-bundle in spectral-domain OCT systems is presented. The fiber-bundle enables a flexible endoscopic design and provides fast, parallelized acquisition of the optical coherence tomography

  15. In vivo probe-based confocal laser endomicroscopy in amiodarone-related pneumonia.

    PubMed

    Salaün, Mathieu; Roussel, Francis; Bourg-Heckly, Geneviève; Vever-Bizet, Christine; Dominique, Stéphane; Genevois, Anne; Jounieaux, Vincent; Zalcman, Gérard; Bergot, Emmanuel; Vergnon, Jean-Michel; Thiberville, Luc

    2013-12-01

    Probe-based confocal laser endomicroscopy (pCLE) allows microscopic imaging of the alveoli during bronchoscopy. The objective of the study was to assess the diagnostic accuracy of pCLE for amiodarone-related pneumonia (AMR-IP). Alveolar pCLE was performed in 36 nonsmoking patients, including 33 consecutive patients with acute or subacute interstitial lung disease (ILD), of which 17 were undergoing treatment with amiodarone, and three were amiodarone-treated patients without ILD. Nine out of 17 patients were diagnosed with high-probability AMR-IP (HP-AMR-IP) by four experts, and three separate observers. Bronchoalveolar lavage findings did not differ between HP-AMR-IP and low-probability AMR-IP (LP-AMR-IP) patients. In HP-AMR-IP patients, pCLE showed large (>20 μm) and strongly fluorescent cells in 32 out of 38 alveolar areas. In contrast, these cells were observed in only two out of 39 areas from LP-AMR-IP patients, in one out of 59 areas from ILD patients not receiving amiodarone and in none of the 10 areas from amiodarone-treated patients without ILD (p<0.001; HP-AMR-IP versus other groups). The presence of at least one alveolar area with large and fluorescent cells had a sensitivity, specificity, negative predictive value and positive predictive value for the diagnosis of AMR-IP of 100%, 88%, 100% and 90%, respectively. In conclusion, pCLE appears to be a valuable tool for the in vivo diagnosis of AMR-IP in subacute ILD patients. PMID:23018901

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

    USGS Publications Warehouse

    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.

  17. Detection of superficial esophageal squamous cell neoplasia by chromoendoscopy-guided confocal laser endomicroscopy

    PubMed Central

    Huang, Jin; Yang, Yun-Sheng; Lu, Zhong-Sheng; Wang, Shuang-Fang; Yang, Jing; Yuan, Jing

    2015-01-01

    AIM: To evaluate the diagnostic potential of Lugol’s chromoendoscopy-guided confocal laser endomicroscopy (CLE) in detecting superficial esophageal squamous cell neoplasia (ESCN). METHODS: Between December 2008 and September 2010, a total of 52 patients were enrolled at the Chinese PLA General Hospital in Beijing, China. First, Lugol’s chromoendoscopy-guided CLE was performed in these patients and the CLE in vivo histological diagnosis was recorded. Then, chromoendoscopy-guided biopsy was performed in the same patients by another endoscopist who was blinded to the CLE findings. Based on the biopsy and CLE diagnosis, en bloc endoscopic resection was performed. The CLE in vivo diagnosis and the histological diagnosis of biopsy of ESCN were compared, using a histological examination of the endoscopic resection specimens as the standard reference. RESULTS: A total of 152 chromoendoscopy-guided biopsies were obtained from 56 lesions. In the 56 lesions of 52 patients, a total of 679 CLE images were obtained vs 152 corresponding biopsies. The sensitivity, specificity, negative predictive value and positive predictive value of chromoendoscopy-guided CLE compared with biopsy were 95.7% vs 82% (P < 0.05), 90% vs 70% (P < 0.05), 81.8% vs 46.7% (P < 0.05), and 97.8% vs 92.7% (P > 0.05), respectively. There was a significant improvement in sensitivity, specificity, negative predictive value, and accuracy when comparing chromoendoscopy-guided CLE with biopsy. CONCLUSION: Lugol’s chromoendoscopy-guided CLE is a real-time, non-invasive endoscopic diagnostic technology; the accuracy of the detection of superficial ESCN is equivalent to or may be superior to biopsy histology. PMID:26078575

  18. Automatic classification of small bowel mucosa alterations in celiac disease for confocal laser endomicroscopy

    NASA Astrophysics Data System (ADS)

    Boschetto, Davide; Di Claudio, Gianluca; Mirzaei, Hadis; Leong, Rupert; Grisan, Enrico

    2016-03-01

    Celiac disease (CD) is an immune-mediated enteropathy triggered by exposure to gluten and similar proteins, affecting genetically susceptible persons, increasing their risk of different complications. Small bowels mucosa damage due to CD involves various degrees of endoscopically relevant lesions, which are not easily recognized: their overall sensitivity and positive predictive values are poor even when zoom-endoscopy is used. Confocal Laser Endomicroscopy (CLE) allows skilled and trained experts to qualitative evaluate mucosa alteration such as a decrease in goblet cells density, presence of villous atrophy or crypt hypertrophy. We present a method for automatically classifying CLE images into three different classes: normal regions, villous atrophy and crypt hypertrophy. This classification is performed after a features selection process, in which four features are extracted from each image, through the application of homomorphic filtering and border identification through Canny and Sobel operators. Three different classifiers have been tested on a dataset of 67 different images labeled by experts in three classes (normal, VA and CH): linear approach, Naive-Bayes quadratic approach and a standard quadratic analysis, all validated with a ten-fold cross validation. Linear classification achieves 82.09% accuracy (class accuracies: 90.32% for normal villi, 82.35% for VA and 68.42% for CH, sensitivity: 0.68, specificity 1.00), Naive Bayes analysis returns 83.58% accuracy (90.32% for normal villi, 70.59% for VA and 84.21% for CH, sensitivity: 0.84 specificity: 0.92), while the quadratic analysis achieves a final accuracy of 94.03% (96.77% accuracy for normal villi, 94.12% for VA and 89.47% for CH, sensitivity: 0.89, specificity: 0.98).

  19. Confocal laser-scanning microscopy for determining the structure of and keratinocyte infiltration through collagen sponges.

    PubMed

    Hanthamrongwit, M; Wilkinson, R; Osborne, C; Reid, W H; Grant, M H

    1996-03-01

    The development of artificial skin substitutes based on cultured cells and biomaterials such as collagen requires an understanding of cellular interactions with the substrate. In this study, human keratinocytes were cultured on the surface of collagen sponges, and confocal laser-scanning microscopy (CLSM) was used to assess both the microstructure of the sponge, and the cell morphology and distribution throughout the sponge. It was found that the pore size increased with increasing depth into the sponge. Both pore size and fiber thickness increased during incubation for up to 10 days at 37 degrees C in culture medium in the absence of cells. This latter effect was not observed when the sponges were incubated in distilled water. Keratinocytes penetrated into the sponge even after only 3 days in culture. By 10 days in culture, the cells had penetrated to the maximum depth that could be examined (120 microns from the sponge surface). In the presence of cells, the inner structure of the collagen sponge had altered after 10 days in culture, with the collagen fibers becoming thicker, and pore geometry less regular. The mechanism responsible for this is unknown at present. Although the presence of the keratinocytes increases distortion of the sponge structure, factors from the medium itself also contribute to this effect. CLSM is a powerful tool for assessing cellular interactions with bioimplants, providing both qualitative and quantitative information. It offers many advantages over scanning electron microscopy (SEM) and histological techniques. CLSM minimizes the time-consuming, extensive preparation of samples required with the latter two methods, and allows noninvasive serial optical sectioning of intact samples. PMID:8698696

  20. Confocal Laser Endomicroscopy for Diagnosis and Monitoring of Pulmonary Alveolar Proteinosis

    PubMed Central

    Averyanov, Alexander; Lesnyak, Viktor; Chernyaev, Andrey; Sorokina, Anastasia

    2015-01-01

    Background: The diagnosis of pulmonary alveolar proteinosis (PAP) is based on computed tomography, histology, and antibodies to granulocyte-macrophage colony-stimulating factor. The role of a novel technique for imaging cells and elastin during endoscopy, probe-based confocal laser endomicroscopy (pCLE), has not yet been investigated in PAP patients. The aim of the present study was to estimate the value of pCLE in the PAP diagnosis and treatment in comparison with the findings of high-resolution computed tomography (HRCT) before and after whole-lung lavage. Methods: In vivo pCLE was performed during bronchoscopy in 6 male patients with PAP before and after whole-lung lavage. In certain lung segments, pCLE was followed by HRCT. Results: During the in vivo pCLE, we found characteristic signs of PAP: a fluorescent floating amorphous substance in the alveoli lumen sticking to conglomerates along with alveolar macrophages. These features were present to a lesser extent after a whole-lung lavage. pCLE revealed specific PAP features not only in segments with crazy-paving and ground-glass opacity, but also in segments without HRCT findings. Conclusions: The alveolar imaging in PAP patients is able to reveal characteristic changes, both in the presence and in the absence of HRCT findings. Therefore, pCLE may be a helpful tool for the diagnosis and whole-lung lavage therapy. Our data prove that accumulation of lipoproteinaceous substances within the alveoli at PAP is a diffuse but not a patchy process. PMID:25590481

  1. Short fatigue crack characterization and detection using confocal scanning laser microscopy (CSLM)

    SciTech Connect

    Varvani-Farahani, A.; Topper, T.H.

    1997-12-31

    This paper presents a new technique for studying the growth and morphology of fatigue cracks. The technique allows short fatigue crack growth, crack depth, aspect ratio (crack depth/half crack length), and crack front configuration to be measured using a Confocal Scanning Laser Microscope (CSLM). CSLM measurements of the initial stage of crack growth in Al 2024-T351 revealed that microstructurally short fatigue cracks grew initially along a plane inclined to the applied stress. The angle of the inclined plane (Stage I crack growth) was found to be about 45 degrees to the axis of the applied tensile load. Aspect ratio and the angle of maximum shear plane (Mode II), obtained using the CSLM technique, showed a good agreement with those obtained using a Surface Removal (SR) technique. The aspect ratios obtained using the CSLM technique were found to remain constant with increasing crack length in Al 2024-T351 and SAE 1045 Steel at 0.83 and 0.80, respectively. Optical sectioning along the length of a crack revealed that the crack front in the interior of the materials has a semi-elliptical shape. These results are in good agreement with results obtained using the SR technique. The CSLM technique was employed to characterize the fracture surface of fatigue cracks in an SAE 1045 Steel. CSLM image processing of the fracture surface near the crack tip constructed a three dimensional profile of fracture surface asperities. The heights of asperities were obtained from this profile. Optical sectioning from a post-image-processed crack provided crack depth and crack mouth width at every point along the crack length for each load level. The crack opening stress was taken as the stress level at which the crack depth stopped increasing with increases in a lied stress. 6 refs., 9 figs., 1 tab.

  2. A prospective cohort study: probe based confocal laser endomicroscopy for peripheral pulmonary lesions (Conference Presentation)

    NASA Astrophysics Data System (ADS)

    Matsumoto, Yuji; Izumo, Takehiro; Hiraishi, Yoshihisa; Tsuchida, Takaaki

    2016-03-01

    Introduction: The diagnostic value of bronchoscopy for peripheral pulmonary lesions (PPLs) has improved since the application of radial endobronchial ultrasound (R-EBUS). Though R-EBUS indicates the position of the PPL, there is often a discrepancy between the obtained R-EBUS image and the diagnostic outcome. Meanwhile, probe based confocal laser endomicroscopy (pCLE) is a novel technique which provides in vivo real-time image of the contacted surface structures. However, its findings have not been established yet. Methods: Consecutive patients who have underwent bronchoscopy for PPLs were prospectively enrolled. R-EBUS with a guide sheath (GS) was inserted to the target PPL under X-ray fluoroscopic guidance. When an adequate R-EBUS image (within or adjacent to) was obtained, pCLE was sequentially inserted through the GS. Then pCLE image was scanned and biopsy was performed where an abnormal finding was estimated. The pCLE findings of PPLs and the background were recorded and analyzed exploratorily. Results: We analyzed 19 cases that we could get appropriate tissues. In all cases, bronchial walls showed longitudinal elastic fibers whereas alveolar walls formed grid-like elastic fiber networks. Conversely, discontinuous, crushed or aggregated alveolar structures accompanied by thickened and distorted fibers were detected in PPLs. Some cases showed dark hollow with fragmented or granular fluorescence. On the other hand, 11 cases (57.9%) indicated normal elastic fibers and needed the position change (3 cases; approached other bronchus, 6 cases; adjusted the position, 2 cases; penetrated the covered bronchial wall). Conclusion: The pCLE has a potential to improve the efficacy of diagnostic bronchoscopy for PPLs.

  3. Toward Automated Analysis of Biofilm Architecture: Bias Caused by Extraneous Confocal Laser Scanning Microscopy Images▿

    PubMed Central

    Merod, Robin T.; Warren, Jennifer E.; McCaslin, Hope; Wuertz, Stefan

    2007-01-01

    An increasing number of studies utilize confocal laser scanning microscopy (CLSM) for in situ visualization of biofilms and rely on the use of image analysis programs to extract quantitative descriptors of architecture. Recently, designed programs have begun incorporating procedures to automatically determine threshold values for three-dimensional CLSM image stacks. We have found that the automated threshold calculation is biased when a stack contains images lacking pixels of biological significance. Consequently, we have created the novel program Auto PHLIP-ML to resolve this bias by iteratively excluding extraneous images based on their area coverage of biomass. A procedure was developed to identify the optimal percent area coverage value used for extraneous image removal (PACVEIR). The optimal PACVEIR was defined to occur when the standard deviation of mean thickness, determined from replicate image stacks, was at a maximum, because it more accurately reflected inherent structural variation. Ten monoculture biofilms of either Ralstonia eutropha JMP228n::gfp or Acinetobacter sp. strain BD413 were tested to verify the routine. All biofilms exhibited an optimal PACVEIR between 0 and 1%. Prior to the exclusion of extraneous images, JMP228n::gfp appeared to develop more homogeneous biofilms than BD413. However, after the removal of extraneous images, JMP228n::gfp biofilms were found to form more heterogeneous biofilms. Similarly, JMP228n::gfp biofilms grown on glass surfaces vis-à-vis polyethylene membranes produced significantly different architectures after extraneous images had been removed but not when such images were included in threshold calculations. This study shows that the failure to remove extraneous images skewed a seemingly objective analysis of biofilm architecture and significantly altered statistically derived conclusions. PMID:17545329

  4. Adhesion of rice flour-based batter to chicken drumsticks evaluated by laser scanning confocal microscopy and texture analysis.

    PubMed

    Mukprasirt, A; Herald, T J; Boyle, D L; Rausch, K D

    2000-09-01

    The convenience and appeal of battered or breaded products have resulted in a sales increase of 100% since 1980. Because of the rapid growth of the Asian-American population and increasing consumption of rice and rice products, rice flour is a logical alternative for wheat flour in traditional batter formulation. The effects of ingredients used in rice flour-based batters on adhesion characteristic for deep-fat fried chicken drumsticks were studied by laser scanning confocal microscopy (LSCM) and texture analysis. Raw chicken drumsticks were predusted with egg albumin powder before dipping into batters prepared from combinations of rice flour, yellow corn flour, oxidized cornstarch, methylcellulose, or xanthan gum. The drumsticks were fried at 175+/-5 C until the internal temperature reached at least 71 C. For LSCM, samples were fixed overnight and were sectioned by vibratome (200 microm) before viewing. Batter adhesion was determined using an attachment specifically designed for chicken drumsticks. Microstructural analysis showed that batter formulated with a 50:50 mixture of rice and corn flours adhered better to drumsticks than batter with other rice flour ratios. Xanthan gum (0.2%) or methylcellulose (0.3%) alone had poor adhesion to chicken skin. However, when combined with other ingredients, xanthan gum increased the amount of batter pick-up before frying by increasing viscosity. Egg albumin significantly facilitated batter adhesion. The results from texture analysis supported the microstructural studies. As rice flour ratio increased from 50 to 70%, the binding force decreased. Rice flour showed potential as an alternative to wheat flour for batter formulas when the appropriate levels of oxidized starch, xanthan gum, and methylcellulose were included in the formulation. PMID:11020085

  5. 3D Quantitative Confocal Laser Microscopy of Ilmenite Volume Distribution in Alpe Arami Olivine

    NASA Astrophysics Data System (ADS)

    Bozhilov, K. N.

    2001-12-01

    The deep origin of the Alpe Arami garnet lherzolite massif in the Swiss Alps proposed by Dobrzhinetskaya et al. (Science, 1996) has been a focus of heated debate. One of the lines of evidence supporting an exhumation from more than 200 km depth includes the abundance, distribution, and orientation of magnesian ilmenite rods in the oldest generation of olivine. This argument has been disputed in terms of the abundance of ilmenite and consequently the maximum TiO2 content in the discussed olivine. In order to address this issue, we have directly measured the volume fraction of ilmenite of the oldest generation of olivine by applying confocal laser scanning microscopy (CLSM). CLSM is a method which allows for three-dimensional imaging and quantitative volume determination by optical sectioning of the objects. The images for 3D reconstruction and measurements were acquired from petrographic thin sections in reflected laser light with 488 nm wavelength. Measurements of more than 80 olivine grains in six thin sections of our material yielded an average volume fraction of 0.31% ilmenite in the oldest generation of olivine from Alpe Arami. This translates into 0.23 wt.% TiO2 in olivine with error in determination of ±0.097 wt.%, a value significantly different from that of 0.02 to 0.03 wt.% TiO2 determined by Hacker et al. (Science, 1997) by a broad-beam microanalysis technique. During the complex geological history of the Alpe Arami massif, several events of metamorphism are recorded which all could have caused increased mobility of the mineral components. Evidence for loss of TiO2 from olivine is the tendency for high densities of ilmenite to be restricted to cores of old grains, the complete absence of ilmenite inclusions from the younger, recrystallized, generation of olivine, and reduction in ilmenite size and abundance in more serpentinized specimens. These observations suggest that only olivine grains with the highest concentrations of ilmenite are close to the

  6. Confocal and Atomic Force Microscopies of Color Centers Produced by Ultrashort Laser Irradiation in LiF Crystals

    NASA Astrophysics Data System (ADS)

    Courrol, Lilia Coronato; Martinez, Oscar; Samad, Ricardo Elgul; Gomes, Laércio; Ranieri, Izilda Márcia; Baldochi, Sonia Licia; de Freitas, Anderson Zanardi; Junior, Nilson Dias Vieira

    2008-04-01

    We report properties of the spatial and spectral distribution of color centers produced in LiF single crystals by ultrashort high intensity laser pulses (60 fs, 10 GW) using confocal spectral microscopy and atomic force microscopy. We could identify a large amount of F centers that gave rise to aggregates such as F2, F4, F2+ and F3+ distributed in cracked shape brownish areas. We have taken a 3D image using confocal microscopy of the sample (luminescent image) and no difference is observed in the different planes. The atomic force microscopy image clearly shows the presence of defects on the modified surface. The formation of micrometer or sub-micrometer voids, filaments and void strings was observed and related to filamentation process.

  7. Detection of a fluorescent-labeled avidin-nucleic acid nanoassembly by confocal laser endomicroscopy in the microvasculature of chronically inflamed intestinal mucosa

    PubMed Central

    Buda, Andrea; Facchin, Sonia; Dassie, Elisa; Casarin, Elisabetta; Jepson, Mark A; Neumann, Helmut; Hatem, Giorgia; Realdon, Stefano; D’Incà, Renata; Sturniolo, Giacomo Carlo; Morpurgo, Margherita

    2015-01-01

    Inflammatory bowel diseases are chronic gastrointestinal pathologies causing great discomfort in both children and adults. The pathogenesis of inflammatory bowel diseases is not yet fully understood and their diagnosis and treatment are often challenging. Nanoparticle-based strategies have been tested in local drug delivery to the inflamed colon. Here, we have investigated the use of the novel avidin-nucleic acid nanoassembly (ANANAS) platform as a potential diagnostic carrier in an experimental model of inflammatory bowel diseases. Fluorescent- labeled ANANAS nanoparticles were administered to mice with chemically induced chronic inflammation of the large intestine. Localization of mucosal nanoparticles was assessed in vivo by dual-band confocal laser endomicroscopy. This technique enables characterization of the mucosal microvasculature and crypt architecture at subcellular resolution. Intravascular nanoparticle distribution was observed in the inflamed mucosa but not in healthy controls, demonstrating the utility of the combination of ANANAS and confocal laser endomicroscopy for highlighting intestinal inflammatory conditions. The specific localization of ANANAS in inflamed tissues supports the potential of this platform as a targeted carrier for bioactive moieties in the treatment of inflammatory bowel disease. PMID:25609952

  8. Hybrid Laser Would Combine Power With Efficiency

    NASA Technical Reports Server (NTRS)

    Sipes, Donald L., Jr

    1986-01-01

    Efficient laser system constructed by using two semiconductor lasers to pump neodymium yttrium aluminum garnet (Nd:YAG) device. Hybrid concept allows digital transmission at data rates of several megabits per second with reasonably sized optical aperture of 20 cm. Beams from two GaAs lasers efficiently coupled for pumping Nd:YAG crystal. Combination of lasers exploits best features of each.

  9. Analysis of micro-structural relaxation phenomena in laser-modified fused silica using confocal Raman microscopy

    SciTech Connect

    Matthews, M; Vignes, R; Cooke, J; Yang, S; Stolken, J

    2009-12-15

    Fused silica micro-structural changes associated with localized 10.6 {micro}m CO{sub 2} laser heating are reported. Spatially-resolved shifts in the high-frequency asymmetric stretch transverse-optic (TO) phonon mode of SiO{sub 2} were measured using confocal Raman microscopy, allowing construction of axial fictive temperature (T{sub f}) maps for various laser heating conditions. A Fourier conduction-based finite element model was employed to compute on-axis temperature-time histories, and, in conjunction with a Tool-Narayanaswamy form for structural relaxation, used to fit T{sub f}(z) profiles to extract relaxation parameters. Good agreement between the calculated and measured T{sub f} was found, yielding reasonable values for relaxation time and activation enthalpy in the laser-modified silica.

  10. Computer Aided Diagnosis for Confocal Laser Endomicroscopy in Advanced Colorectal Adenocarcinoma

    PubMed Central

    Ştefănescu, Daniela; Streba, Costin; Cârţână, Elena Tatiana; Săftoiu, Adrian; Gruionu, Gabriel; Gruionu, Lucian Gheorghe

    2016-01-01

    Introduction Confocal laser endomicroscopy (CLE) is becoming a popular method for optical biopsy of digestive mucosa for both diagnostic and therapeutic procedures. Computer aided diagnosis of CLE images, using image processing and fractal analysis can be used to quantify the histological structures in the CLE generated images. The aim of this study is to develop an automatic diagnosis algorithm of colorectal cancer (CRC), based on fractal analysis and neural network modeling of the CLE-generated colon mucosa images. Materials and Methods We retrospectively analyzed a series of 1035 artifact-free endomicroscopy images, obtained during CLE examinations from normal mucosa (356 images) and tumor regions (679 images). The images were processed using a computer aided diagnosis (CAD) medical imaging system in order to obtain an automatic diagnosis. The CAD application includes image reading and processing functions, a module for fractal analysis, grey-level co-occurrence matrix (GLCM) computation module, and a feature identification module based on the Marching Squares and linear interpolation methods. A two-layer neural network was trained to automatically interpret the imaging data and diagnose the pathological samples based on the fractal dimension and the characteristic features of the biological tissues. Results Normal colon mucosa is characterized by regular polyhedral crypt structures whereas malignant colon mucosa is characterized by irregular and interrupted crypts, which can be diagnosed by CAD. For this purpose, seven geometric parameters were defined for each image: fractal dimension, lacunarity, contrast correlation, energy, homogeneity, and feature number. Of the seven parameters only contrast, homogeneity and feature number were significantly different between normal and cancer samples. Next, a two-layer feed forward neural network was used to train and automatically diagnose the malignant samples, based on the seven parameters tested. The neural network

  11. Study of hydroxyl carbonate apatite formation on bioactive glass coated dental ceramics by confocal laser scanning microscopy (CLSM)

    NASA Astrophysics Data System (ADS)

    Stanciu, G. A.; Savu, B.; Sandulescu, I.; Paraskevopoulos, K.; Koidis, P.

    2007-03-01

    Some dental ceramics were coated with a bioactive glass and resulted the formation of a stable and well bonded with the ceramic substrate thin layer. After immersion in a solution with ion concentrations similar to those of human blood plasma the development of hydroxy carbonate apatite layer on the surface of bioactive glass may be observed. The objective of this study was to investigate structural surface changes of bioactive glass, after exposure in a simulated body fluid for a different number of days. The roughness and topography of the hydroxyapatite surface were investigated by Confocal Scanning Laser Microscopy. The chemical composition was analyzed by Energy Dispersive Spectroscopy measurements.

  12. Endoscopic Ultrasound-Guided Needle-Based Probe Confocal Laser Endomicroscopy (nCLE) of Intrapancreatic Ectopic Spleen.

    PubMed

    Bastidas, Amanda B; Holloman, David; Lankarani, Ali; Nieto, Jose M

    2016-04-01

    Accessory spleens and splenosis represent the congenital and acquired type of ectopic splenic tissue. Generally, they are asymptomatic entities posing as solid hypervascular masses at the splenic hilum or in other organs, such as the pancreas. Intrapancreatic ectopic spleen mimics pancreatic neoplasms on imaging studies, and due to the lack of radiological diagnostic criteria, patients undergo unnecessary distal pancreatectomy. We present the first case of intrapancreatic ectopic spleen in which the concomitant use of needle-based probe confocal laser endomicroscopy and fine-needle aspiration supported the final diagnosis. PMID:27144203

  13. Inverse image alignment method for image mosaicing and video stabilization in fundus indocyanine green angiography under confocal scanning laser ophthalmoscope.

    PubMed

    Zhou, Yongjin; Xue, Hui; Wan, Mingxi

    2003-01-01

    An efficient image registration algorithm, the Inverse Compositional image alignment method based on minimization of Sum of Squared Differences of images, is applied in fundus blood vessel angiography under confocal scanning laser ophthalmoscope, to build image mosaics which have larger field of view without loss of resolution to assist diagnosis. Furthermore, based on similar technique, the angiography video stabilization algorithm is implemented for fundus documenting. The actual underlying models of motion between images and corresponding convergence criteria are also discussed. The experiment results in fundus images demonstrate the effectiveness of the registration scheme. PMID:14575786

  14. Endoscopic Ultrasound-Guided Needle-Based Probe Confocal Laser Endomicroscopy (nCLE) of Intrapancreatic Ectopic Spleen

    PubMed Central

    Bastidas, Amanda B.; Holloman, David; Lankarani, Ali

    2016-01-01

    Accessory spleens and splenosis represent the congenital and acquired type of ectopic splenic tissue. Generally, they are asymptomatic entities posing as solid hypervascular masses at the splenic hilum or in other organs, such as the pancreas. Intrapancreatic ectopic spleen mimics pancreatic neoplasms on imaging studies, and due to the lack of radiological diagnostic criteria, patients undergo unnecessary distal pancreatectomy. We present the first case of intrapancreatic ectopic spleen in which the concomitant use of needle-based probe confocal laser endomicroscopy and fine-needle aspiration supported the final diagnosis. PMID:27144203

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

    NASA Astrophysics Data System (ADS)

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

    2016-01-01

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

  16. Studies of porphyrin-containing specimens using an optical spectrometer connected to a confocal scanning laser microscope.

    PubMed

    Trepte, O; Rokahr, I; Andersson-Engels, S; Carlsson, K

    1994-12-01

    A spectrometer has been developed for use with a confocal scanning laser microscope. With this unit, spectral information from a single point or a user-defined region within the microscope specimen can be recorded. A glass prism is used to disperse the spectral components of the recorded light over a linear CCD photodiode array with 256 elements. A regulated cooling unit keeps the detector at 277 K, thereby allowing integration times of up to 60 s. The spectral resolving power, lambda/delta lambda, ranges from 350 at lambda = 400 nm to 100 at lambda = 700 nm. Since the entrance aperture of the spectrometer has the same size as the detector pinhole used during normal confocal scanning, the three-dimensional spatial resolution is equivalent to that of normal confocal scanning. Light from the specimen is deflected to the spectrometer by a solenoid controlled mirror, allowing fast and easy switching between normal confocal scanning and spectrometer readings. With this equipment, studies of rodent liver specimens containing porphyrins have been made. The subcellular localization is of interest for the mechanisms of photodynamic therapy (PDT) of malignant tumours. Spectroscopic detection is necessary to distinguish the porphyrin signal from other fluorescent components in the specimen. Two different substances were administered to the tissue, Photofrin, a haematoporphyrin derivative (HPD) and delta-amino levulinic acid (ALA), a precursor to protoporphyrin IX and haem in the haem cycle. Both are substances under clinical trials for PDT of malignant tumours. Following administration of these compounds to the tissue, the potent photosensitizer and fluorescent compound Photofrin, or protoporphyrin IX, respectively, is accumulated.(ABSTRACT TRUNCATED AT 250 WORDS) PMID:7869364

  17. Use of endoscopic distal attachment cap to enhance image stabilization in probe-based confocal laser endomicroscopy in colorectal lesions*

    PubMed Central

    Ussui, Vivian; Xu, Can; Crook, Julia E.; Diehl, Nancy N.; Hardee, Joy; Staggs, Estela G.; Shahid, Muhammad W.; Wallace, Michael B.

    2015-01-01

    Background and study aims: Colorectal cancer can be prevented through the use of colonoscopy with polypectomy. Most colon polyps are benign or low grade adenomas. However, currently all lesions need histopathologic analysis, which increases diagnostic costs and delays the final diagnosis. Confocal laser endomicroscopy (CLE) is a new technology that enables real-time endomicroscopy. However, there are challenges to maintaining a stable image with currently available systems. We conducted a small study to obtain a preliminary assessment of whether the use of an endoscopic distal attachment cap may enhance image quality of CLE in comparison with images obtained with free-hand acquisition. Patients and methods: Forty outpatients underwent colonoscopy for evaluation of colon polyps in a single academic medical center. Patients were assigned randomly to 1 of 2 study arms on the basis of whether an endoscopic distal attachment cap was used (n = 21, Cap Used) or not used (n = 19, No Cap) in the procedure. The quality of confocal images and probe stabilization was summarized. Results: A total of 81 polyps were identified. The proportion of polyps with images of high quality was 74 % (28/38) in the Cap Used group and 79 % (30/38) in the No Cap arm. Image stability was also similar with and without a cap. Diagnostic accuracy was estimated to be slightly higher in the Cap Used group for probe-based confocal laser endomicroscopy (pCLE; 78 % vs 70 %). This was also true for white-light and narrow-band imaging. Conclusions: This preliminary study did not yield any evidence to support that the use of an endoscopic distal attachment cap improves the quality of images obtained during CLE. PMID:26528511

  18. Raman confocal microscopy and AFM combined studies of cancerous cells treated with Paclitaxel

    NASA Astrophysics Data System (ADS)

    Derely, L.; Collart Dutilleul, P.-Y.; Michotte de Welle, Sylvain; Szabo, V.; Gergely, C.; Cuisinier, F. J. G.

    2011-03-01

    Paclitaxel interferes with the normal function of microtubule breakdown, induces apoptosis in cancer cells and sequesters free tubulin. As this drug acts also on other cell mechanisms it is important to monitor its accumulation in the cell compartments. The intracellular spreading of the drug was followed using a WITEC 300R confocal Raman microscope equipped with a CCD camera. Hence Atomic force microscopy (an MFP3D- Asylum Research AFM) in imaging and force mode was used to determine the morphological and mechanical modifications induced on living cells. These studies were performed on living epithelial MCF-7 breast cancer cells. Paclitaxel was added to cell culture media for 3, 6 and 9 hours. Among the specific paclitaxel Raman bands we selected the one at 1670 cm-1 because it is not superposed by the spectrum of the cells. Confocal Raman images are formed by monitoring this band, the NH2 and the PO4 band. Paclitaxel slightly accumulates in the nucleus forming patches. The drug is also concentrated in the vicinity of the cell membrane and in an area close to the nucleus where proteins accumulate. Our AFM images reveal that the treated cancerous MCF-7 cells keep the same size as the non treated ones, but their shape becomes more oval. Cell's elasticity is also modified: a difference of 2 kPa in the Young Modulus characterizes the treated MCF-7 mammary cancerous cell. Our observations demonstrate that paclitaxel acts not only on microtubules but accumulates also in other cell compartments (nucleus) where microtubules are absent.

  19. Assessment of possibilities of ceramic biomaterial fracture surface reconstruction using laser confocal microscopy and long working distance objective lenses.

    PubMed

    Stach, Sebastian; Sapota, Wiktoria; Wróbel, Zygmunt; Ţălu, Ştefan

    2016-05-01

    A numerical description of fracture is an important step in the search of the correlation between specific micromechanisms of decohesion and material characteristics designated with the use of fracture mechanics methods. This issue is essential for the proper orientation of the search for basic relationships between chemical composition, technology, structure, and properties of materials. It often happens that fracture surfaces are well developed, which can significantly hinder or even prevent the measurement and reconstruction of the tested material surface geometry. In this article, comparative measurements of a biomaterial surface were performed using laser confocal microscopy. To this end, short working distance lenses dedicated to a focused UV laser beam and long working distance objective lenses were used. The article includes a quantitative comparative analysis and interpretation of the obtained results. Microsc. Res. Tech. 79:385-392, 2016. © 2016 Wiley Periodicals, Inc. PMID:26918261

  20. Confocal microscopy to guide Erbium:yttrium aluminum garnet laser ablation of basal cell carcinoma: an ex vivo feasibility study

    PubMed Central

    Larson, Bjorg A.; Chen, Chih-Shan Jason; Rajadhyaksha, Milind

    2013-01-01

    Abstract. For the removal of superficial and nodular basal cell carcinomas (BCCs), laser ablation provides certain advantages relative to other treatment modalities. However, efficacy and reliability tend to be variable because tissue is vaporized such that none is available for subsequent histopathological examination for residual BCC (and to confirm complete removal of tumor). Intra-operative reflectance confocal microscopy (RCM) may provide a means to detect residual tumor directly on the patient and guide ablation. However, optimization of ablation parameters will be necessary to control collateral thermal damage and preserve sufficient viability in the underlying layer of tissue, so as to subsequently allow labeling of nuclear morphology with a contrast agent and imaging of residual BCC. We report the results of a preliminary study of two key parameters (fluence, number of passes) vis-à-vis the feasibility of labeling and RCM imaging in human skin ex vivo, following ablation with an erbium:yttrium aluminum garnet laser. PMID:24045654

  1. Visualising fouling of a chromatographic matrix using confocal scanning laser microscopy.

    PubMed

    Siu, Sun Chau; Boushaba, Rihab; Topoyassakul, Vithaya; Graham, Alex; Choudhury, Sorwar; Moss, Guy; Titchener-Hooker, Nigel J

    2006-11-01

    Confocal scanning laser microscopy (CSLM) was used to visualise the spatial location of foulants during the fouling of Q Sepharose FF matrix in finite batch experiments and for examining the subsequent effectiveness of clean-in-place (CIP) treatments in cleaning the heavily fouled beads. Beads were severely fouled with partially clarified E. coli homogenate by contacting the beads with the foulant for contact times of 5 min, 1 or 12 h. The use of two different fluorescent dyes, PicoGreen and Cy5.5, for labelling genomic PicoGreen-labelled dsDNA and protein respectively, allowed the direct observation of the chromatographic beads. The extent of fouling was assessed by measuring the subsequent adsorption of Cy5.5-labelled BSA to the beads. Control studies established that the labelling of BSA did not affect significantly the protein properties. In the control case of contacting the unfouled matrix with Cy5.5-labelled BSA, protein was able to penetrate the entire matrix volume. After fouling, Cy5.5-labelled BSA was unable to penetrate the bead but only to bind near the bead surface where it slowly displaced PicoGreen-conjugated dsDNA, which bound only at the exterior of the beads. Labelled host cell proteins bound throughout the bead interior but considerably less at the core; suggesting that other species might have occupied that space. The gross levels of fouling achieved drastically reduced the binding capacity and maximum Cy5.5-labelled BSA uptake rate. The capacity of the resin was reduced by 2.5-fold when incubated with foulant for up to 1 h. However, when the resin was fouled for a prolonged time of 12 h a further sixfold decrease in capacity was seen. The uptake rate of Cy5.5-labelled BSA decreased with increased fouling time of the resin. Incubating the fouled beads in 1 M NaCl dissociated PicoGreen-labelled dsDNA from the bead exterior within 15 min of incubation but proved ineffective in removing all the foulant protein. Cy5.5-labelled BSA was still unable

  2. Grating rhomb diode laser power combiner

    NASA Technical Reports Server (NTRS)

    Minott, Peter O.; Abshire, James B.

    1987-01-01

    A compact device for spectrally combining many laser-diode beams into a single multi-wavelength beam has been developed for use in NASA's intersatellite communications programs. The prototype device combines seven 30 milliwatt beams into a single beam with 70 percent efficiency producing an output of approximately 150 milliwatts. All beams are coaxial and can be collimated with a single transmitter optical system. The combining technique is relatively insensitive to drifts in the laser-diode wavelength and provides both increased power output and laser-diode source redundancy. Combination of more than 100 laser-diodes producing an output greater than 5 watts appears feasible with this technique.

  3. [Opportunities for confocal and laser biomicroscopy of corneal nerves in diabetic polyneuropathy].

    PubMed

    Surnina, Z V

    2015-01-01

    The review concerns corneal nerves involvement in diabetes mellitus (DM), a pressing issue for ophthalmology and endocrinology. The history of research in this field along with anatomical, physiological, and biochemical features of corneal nerves is provided. Corneal nerves anatomy is described in accordance with Soviet scientific school and contemporary foreign sources. The most part of the paper is devoted to technical description of a confocal microscope and Heidelberg Retina Tomograph with corneal module as well as the feasibility of corneal nerves visualization. Diabetic neuropathy, a threatening complication of DM that can result in lower limb amputations, is discussed. A number of authors suggest confocal biomicroscopy for early diagnosis of polyneuropathy, yet few relevant publications can be found. If effective, confocal biomicroscopy can be considered as a possible screening tool able to detect early signs of diabetes complications and thus to ensure the treatment initiated in a timely manner. The latter is crucial to prevent DM progression to its terminal stage--diabetic polyneuropathy, which is dangerous of lower limb amputations. PMID:25872394

  4. Raman beam combining for laser brightness enhancement

    SciTech Connect

    Dawson, Jay W.; Allen, Graham S.; Pax, Paul H.; Heebner, John E.; Sridharan, Arun K.; Rubenchik, Alexander M.; Barty, Chrisopher B. J.

    2015-10-27

    An optical source capable of enhanced scaling of pulse energy and brightness utilizes an ensemble of single-aperture fiber lasers as pump sources, with each such fiber laser operating at acceptable pulse energy levels. Beam combining involves stimulated Raman scattering using a Stokes' shifted seed beam, the latter of which is optimized in terms of its temporal and spectral properties. Beams from fiber lasers can thus be combined to attain pulses with peak energies in excess of the fiber laser self-focusing limit of 4 MW while retaining the advantages of a fiber laser system of high average power with good beam quality.

  5. Combination free-electron and gaseous laser

    SciTech Connect

    Brau, C.A.; Rockwood, S.D.; Stein, W.E.

    1981-06-08

    A multiple laser having one or more gaseous laser stages and one or more free electron stages is described. Each of the free electron laser stages is sequentially pumped by a microwave linear accelerator. Subsequently, the electron beam is directed through a gaseous laser, in the preferred embodiment, and in an alternative embodiment, through a microwave accelerator to lower the energy level of the electron beam to pump one or more gaseous lasers. The combination laser provides high pulse repetition frequencies, on the order of 1 kHz or greater, high power capability, high efficiency, and tunability in the synchronous production of multiple beams of coherent optical radiation.

  6. Combination free electron and gaseous laser

    DOEpatents

    Brau, Charles A.; Rockwood, Stephen D.; Stein, William E.

    1980-01-01

    A multiple laser having one or more gaseous laser stages and one or more free electron stages. Each of the free electron laser stages is sequentially pumped by a microwave linear accelerator. Subsequently, the electron beam is directed through a gaseous laser, in the preferred embodiment, and in an alternative embodiment, through a microwave accelerator to lower the energy level of the electron beam to pump one or more gaseous lasers. The combination laser provides high pulse repetition frequencies, on the order of 1 kHz or greater, high power capability, high efficiency, and tunability in the synchronous production of multiple beams of coherent optical radiation.

  7. Surface emitting lasers with combined output

    NASA Technical Reports Server (NTRS)

    Carlin, Donald B. (Inventor)

    1990-01-01

    Surface emitting lasers are laterally aligned and coupled together and also have their light output signals combined. This results in greater phase and frequency coherency and narrower and reduced amplitude sidelobes. Preferably, not more than two lasers are longitudinally aligned along the same axis for still greater coherency compared with adding the light output signals of more than two longitudinally aligned lasers. The lasers can be of the DH-LOC type or of the QW type.

  8. Physiological and morphological characterization of honeybee olfactory neurons combining electrophysiology, calcium imaging and confocal microscopy.

    PubMed

    Galizia, C G; Kimmerle, B

    2004-01-01

    The insect antennal lobe is the first brain structure to process olfactory information. Like the vertebrate olfactory bulb the antennal lobe is substructured in olfactory glomeruli. In insects, glomeruli can be morphologically identified, and have characteristic olfactory response profiles. Local neurons interconnect glomeruli, and output (projection) neurons project to higher-order brain centres. The relationship between their elaborate morphology and their physiology is not understood. We recorded electrophysiologically from antennal lobe neurons, and iontophoretically injected a calcium-sensitive dye. We then measured their spatio-temporal calcium responses to a variety of odours. Finally, we confocally reconstructed the neurons, and identified the innervated glomeruli. An increase or decrease in spiking frequency corresponded to an intracellular calcium increase or decrease in the cell. While intracellular recordings generally lasted between 10 and 30 min, calcium imaging was stable for up to 2 h, allowing a more detailed physiological analysis. The responses indicate that heterogeneous local neurons get input in the glomerulus in which they branch most strongly. In many cases, the physiological response properties of the cells corresponded to the known response profile of the innervated glomerulus. In other words, the large variety of response profiles generally found when comparing antennal lobe neurons is reduced to a more predictable response profile when the innervated glomerulus is known. PMID:14639486

  9. Laser jamming technique research based on combined fiber laser

    NASA Astrophysics Data System (ADS)

    Jie, Xu; Shanghong, Zhao; Rui, Hou; Shengbao, Zhan; Lei, Shi; Jili, Wu; Shaoqiang, Fang; Yongjun, Li

    2009-06-01

    A compact and light laser jamming source is needed to increase the flexibility of laser jamming technique. A novel laser jamming source based on combined fiber lasers is proposed. Preliminary experimental results show that power levels in excess of 10 kW could be achieved. An example of laser jamming used for an air-to-air missile is given. It shows that the tracking system could complete tracking in only 4 s and came into a steady state with its new tracking target being the laser jamming source.

  10. 3D digital image processing for biofilm quantification from confocal laser scanning microscopy: Multidimensional statistical analysis of biofilm modeling

    NASA Astrophysics Data System (ADS)

    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

  11. Velocity gradients in spatially resolved laser Doppler flowmetry and dynamic light scattering with confocal and coherence gating

    NASA Astrophysics Data System (ADS)

    Uribe-Patarroyo, Néstor; Bouma, Brett E.

    2016-08-01

    Dynamic light scattering (DLS) is widely used to characterize diffusive motion to obtain precise information on colloidal suspensions by calculating the autocorrelation function of the signal from a heterodyne optical system. DLS can also be used to determine the flow velocity field in systems that exhibit mass transport by incorporating the effects of the deterministic motion of scatterers on the autocorrelation function, a technique commonly known as laser Doppler flowmetry. DLS measurements can be localized with confocal and coherence gating techniques such as confocal microscopy and optical coherence tomography, thereby enabling the determination of the spatially resolved velocity field in three dimensions. It has been thought that spatially resolved DLS can determine the axial velocity as well as the lateral speed in a single measurement. We demonstrate, however, that gradients in the axial velocity of scatterers exert a fundamental influence on the autocorrelation function even in well-behaved, nonturbulent flow. By obtaining the explicit functional relation between axial-velocity gradients and the autocorrelation function, we show that the velocity field and its derivatives are intimately related and their contributions cannot be separated. Therefore, a single DLS measurement cannot univocally determine the velocity field. Our extended theoretical model was found to be in good agreement with experimental measurements.

  12. Velocity gradients in spatially resolved laser Doppler flowmetry and dynamic light scattering with confocal and coherence gating.

    PubMed

    Uribe-Patarroyo, Néstor; Bouma, Brett E

    2016-08-01

    Dynamic light scattering (DLS) is widely used to characterize diffusive motion to obtain precise information on colloidal suspensions by calculating the autocorrelation function of the signal from a heterodyne optical system. DLS can also be used to determine the flow velocity field in systems that exhibit mass transport by incorporating the effects of the deterministic motion of scatterers on the autocorrelation function, a technique commonly known as laser Doppler flowmetry. DLS measurements can be localized with confocal and coherence gating techniques such as confocal microscopy and optical coherence tomography, thereby enabling the determination of the spatially resolved velocity field in three dimensions. It has been thought that spatially resolved DLS can determine the axial velocity as well as the lateral speed in a single measurement. We demonstrate, however, that gradients in the axial velocity of scatterers exert a fundamental influence on the autocorrelation function even in well-behaved, nonturbulent flow. By obtaining the explicit functional relation between axial-velocity gradients and the autocorrelation function, we show that the velocity field and its derivatives are intimately related and their contributions cannot be separated. Therefore, a single DLS measurement cannot univocally determine the velocity field. Our extended theoretical model was found to be in good agreement with experimental measurements. PMID:27627357

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

    NASA Astrophysics Data System (ADS)

    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.

  14. 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

    PubMed Central

    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 < 0.05 (R software version 3.1.0). Results: Comparison of microleakage showed maximum peak value of 0.45 mm for Biodentine, 0.85 mm for MTA, and 1.05 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, the difference was found to be statistically significant (P < 0.05). Conclusions: Root-end cavities prepared with Er:YAG laser and restored with Biodentine showed superior sealing ability compared to those prepared with ultrasonics. PMID:26180420

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

    PubMed

    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. PMID:24428443

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

    NASA Astrophysics Data System (ADS)

    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.

  17. In-Situ Observation of Crystallization and Growth in High-Temperature Melts Using the Confocal Laser Microscope

    NASA Astrophysics Data System (ADS)

    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.

  18. In-Situ Observation of Crystallization and Growth in High-Temperature Melts Using the Confocal Laser Microscope

    NASA Astrophysics Data System (ADS)

    Sohn, Il; Dippenaar, Rian

    2016-04-01

    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.

  19. Investigation of biological cell-protein interactions using SPR sensor through laser scanning confocal imaging-surface plasmon resonance system

    NASA Astrophysics Data System (ADS)

    Zhang, Hongyan; Yang, Liquan; Zhou, Bingjiang; Wang, Xueliang; Liu, Guiying; Liu, Weimin; Wang, Pengfei

    2014-03-01

    A new method for investigating biological cell-protein interactions was developed by using a laser scanning confocal imaging-surface plasmon resonance (LSCI-SPR) system. Mouse normal IgG was modified on the SPR chip. The suspension mouse lymphocyte cancer cells (L5178Y cells) labeled by Hoechst33342 freely flowed into the surface of the SPR sensor chip. By changing the concentration of the cells, the fluorescence images and the SPR signal were synchronously recorded in real time. The red fluorescence points in the imaging region increased with increase in the concentration of the mouse lymphocyte cancer cells and fit well with the change in the SPR signal. Different suspending cells were chosen to investigate cell-protein interactions through antigen-antibody reactions on the biological cell surfaces through binding detection. This method has potential application in cell biology and pharmacology.

  20. Biofilms on tracheoesophageal voice prostheses: a confocal laser scanning microscopy demonstration of mixed bacterial and yeast biofilms.

    PubMed

    Kania, Romain E; Lamers, Gerda E M; van de Laar, Nicole; Dijkhuizen, Marloes; Lagendijk, Ellen; Huy, Patrice Tran Ba; Herman, Philippe; Hiemstra, Pieter; Grote, Jan J; Frijns, Johan; Bloemberg, Guido V

    2010-07-01

    The aim of this study was to demonstrate the presence of yeast and bacterial biofilms on the surface of tracheoesophageal voice prostheses (TVPs) by a double-staining technique with confocal laser scanning microscopy (CLSM). Biofilms of 12 removed TVPs were visualized by scanning electron microscopy, then stained with ConA-FITC and propidium iodide for CLSM. Microbial identification was by partial 16S rRNA gene analysis and ITS-2 sequence analysis. Microbial biofilms on the TVPs consisted of bacteria and filamentous cells. Bacterial cells were attached to the filamentous and unicellular yeast cells, thus forming a network. Sequence analyses of six voice prostheses identified the presence of a variety of bacterial and yeast species. In vivo studies showed that Klebsiella oxytoca and Micrococcus luteus efficiently attached to Candida albicans. CLSM with double fluorescence staining can be used to demonstrate biofilm formations composed of a mixture of yeast and bacterial cells on the surface of TVPs. PMID:20473799

  1. A novel approach to pseudopodia proteomics: excimer laser etching, two-dimensional difference gel electrophoresis, and confocal imaging

    PubMed Central

    Mimae, Takahiro; Ito, Akihiko; Hagiyama, Man; Nakanishi, Jun; Hosokawa, Yoichiroh; Okada, Morihito; Murakami, Yoshinori; Kondo, Tadashi

    2014-01-01

    Pseudopodia are actin-rich ventral cellular protrusions shown to facilitate the migration and metastasis of tumor cells. Here, we present a novel approach to perform pseudopodia proteomics. Tumor cells growing on porous membranes extend pseudopodia into the membrane pores. In our method, cell bodies are removed by horizontal ablation at the basal cell surface with the excimer laser while pseudopodia are left in the membrane pores. For protein expression profiling, whole cell and pseudopodia proteins are extracted with a lysis buffer, labeled with highly sensitive fluorescent dyes, and separated by two-dimensional gel electrophoresis. Proteins with unique expression patterns in pseudopodia are identified by mass spectrometry. The effects of the identified proteins on pseudopodia formation are evaluated by measuring the pseudopodia length in cancer cells with genetically modified expression of target proteins using confocal imaging. This protocol allows global identification of pseudopodia proteins and evaluation of their functional significance in pseudopodia formation within one month. PMID:25309719

  2. In vivo analysis of THz wave irradiation induced acute inflammatory response in skin by laser-scanning confocal microscopy.

    PubMed

    Hwang, Yoonha; Ahn, Jinhyo; Mun, Jungho; Bae, Sangyoon; Jeong, Young Uk; Vinokurov, Nikolay A; Kim, Pilhan

    2014-05-19

    The recent development of THz sources in a wide range of THz frequencies and power levels has led to greatly increased interest in potential biomedical applications such as cancer and burn wound diagnosis. However, despite its importance in realizing THz wave based applications, our knowledge of how THz wave irradiation can affect a live tissue at the cellular level is very limited. In this study, an acute inflammatory response caused by pulsed THz wave irradiation on the skin of a live mouse was analyzed at the cellular level using intravital laser-scanning confocal microscopy. Pulsed THz wave (2.7 THz, 4 μs pulsewidth, 61.4 μJ per pulse, 3Hz repetition), generated using compact FEL, was used to irradiate an anesthetized mouse's ear skin with an average power of 260 mW/cm(2) for 30 minutes using a high-precision focused THz wave irradiation setup. In contrast to in vitro analysis using cultured cells at similar power levels of CW THz wave irradiation, no temperature change at the surface of the ear skin was observed when skin was examined with an IR camera. To monitor any potential inflammatory response, resident neutrophils in the same area of ear skin were repeatedly visualized before and after THz wave irradiation using a custom-built laser-scanning confocal microscopy system optimized for in vivo visualization. While non-irradiated control skin area showed no changes in the number of resident neutrophils, a massive recruitment of newly infiltrated neutrophils was observed in the THz wave irradiated skin area after 6 hours, which suggests an induction of acute inflammatory response by the pulsed THz wave irradiation on the skin via a non-thermal process. PMID:24921268

  3. Influence of confocal scanning laser microscopy specific acquisition parameters on the detection and matching of speeded-up robust features.

    PubMed

    Stanciu, Stefan G; Hristu, Radu; Stanciu, George A

    2011-04-01

    The robustness and distinctiveness of local features to various object or scene deformations and to modifications of the acquisition parameters play key roles in the design of many computer vision applications. In this paper we present the results of our experiments on the behavior of a recently developed technique for local feature detection and description, Speeded-Up Robust Features (SURF), regarding image modifications specific to Confocal Scanning Laser Microscopy (CSLM). We analyze the repeatability of detected SURF keypoints and the precision-recall of their matching under modifications of three important CSLM parameters: pinhole aperture, photomultiplier (PMT) gain and laser beam power. During any investigation by CSLM these three parameters have to be modified, individually or together, in order to optimize the contrast and the Signal Noise Ratio (SNR), being also inherently modified when changing the microscope objective. Our experiments show that an important amount of SURF features can be detected at the same physical locations in images collected at different values of the pinhole aperture, PMT gain and laser beam power, and further on can be successfully matched based on their descriptors. In the final part, we exemplify the potential of SURF in CSLM imaging by presenting a SURF-based computer vision application that deals with the mosaicing of images collected by this technique. PMID:21349249

  4. Optical Coherence Tomography Angiography in Mice: Comparison with Confocal Scanning Laser Microscopy and Fluorescein Angiography

    PubMed Central

    Giannakaki-Zimmermann, Helena; Kokona, Despina; Wolf, Sebastian; Ebneter, Andreas; Zinkernagel, Martin S.

    2016-01-01

    Purpose Optical coherence tomography angiography (OCT-A) allows noninvasive visualization of retinal vessels in vivo. OCT-A was used to characterize the vascular network of the mouse retina and was compared with fluorescein angiography (FA) and histology. Methods In the present study, OCT-A based on a Heidelberg Engineering Spectralis system was used to investigate the vascular network in mice. Data was compared with FA and confocal microscopy of flat-mount histology stained with isolectin IB4. For quantitative analysis the National Cancer Institute's AngioTool software was used. Vessel density, the number of vessel junctions, and endpoints were measured and compared between the imaging modalities. Results The configuration of the superficial capillary network was comparable with OCT-A and flat-mount histology in BALBc mice. However, vessel density and the number of vessel junctions per region of interest (P = 0.0161 and P = 0.0015, respectively) in the deep vascular network of BALBc mice measured by OCT-A was significantly higher than with flat-mount histology. In C3A.Cg-Pde6b+Prph2Rd2/J mice, where the deep capillary plexus is absent, analysis of the superficial network provided similar results for all three imaging modalities. Conclusion OCT-A is a helpful imaging tool for noninvasive, in vivo imaging of the vascular plexus in mice. It may offer advantages over FA and confocal microscopy especially for imaging the deep vascular plexus. Translational Relevance The present study shows that OCT-A can be employed for small animal imaging to assess the vascular network and offers advantages over flat-mount histology and FA. PMID:27570710

  5. Scan-less, line-field confocal microscopy by combination of wavelength/space conversion with dual optical comb

    NASA Astrophysics Data System (ADS)

    Yasui, Takeshi; Hase, Eiji; Miyamoto, Shuji; Hsieh, Yi-Da; Minamikawa, Takeo; Yamamoto, Hirotsugu

    2016-03-01

    Optical frequency comb (OFC) has attracted attentions for optical frequency metrology in visible and infrared regions because the mode-resolved OFC spectrum can be used as a precise frequency ruler due to both characteristics of broadband radiation and narrow-line CW radiation. Furthermore, the absolute accuracy of all frequency modes in OFC is secured by phase-locking a repetition frequency frep and a carrier-envelope-offset frequency fceo to a frequency standard. However, application fields of OFC other than optical frequency metrology are still undeveloped. One interesting aspect of OFC except for the frequency ruler is optical carrier having a huge number of discrete frequency channels because OFC is composed of a series of frequency spikes regularly separated by frep in the broad spectral range. If a certain quantity to be measured is encoded on each comb mode by dimensional conversion, a huge number of data for the measured quantity can be obtained from a single mode-resolved spectrum of OFC. In this paper, we encode the confocal microscopic line-image of a sample on the mode-resolved OFC spectrum by the dimensional conversion between wavelength and 1D-space. The resulting image-encoded OFC spectrum is acquired by an optical spectrum analyzer or dual comb spectrometer. Finally, the line image of the sample is decoded from the spectral amplitude of the mode-resolved OFC spectrum. The combination of OFC with the dimensional conversion enables to establish both confocal modality and line-field imaging under the scan-less condition.

  6. Flow assisted assembly of multilayer colloidal crystals studied using confocal laser scanning microscopy

    NASA Astrophysics Data System (ADS)

    Shereda, Laura T.

    Colloidal crystals are highly ordered particle arrays with potential applications including sensors, optical switches, and photonic materials. For production on an industrially viable scale, processes must be developed to form crystals with low defect densities, good long range order, and favorable kinetics. Application of a field to a concentrated colloidal suspension accelerates crystal formation. Ackerson et al. (Ackerson, 1991) established that systems with stress-based Peclet numbers above one resulted in crystal formation. We investigate formation of colloidal crystals by studying structural changes that occur upon shearing using confocal microscopy. Charge-stabilized poly(methylmethacrylate) particles (phi = 0.35) suspended in dioctyl phthalate were used for experiments. After application of shear, assembled structures were immobilized by UV exposure. The full sample thickness was imaged using confocal microscopy. Particle centroids were located in 3D by means of image processing and local crystallinity was quantified by application of local bond order parameter criteria (tenWolde, 1996). We present microstructural analysis of structures formed by both spin coating and uniform shear flow. Spin coating produces spatiotemporal variation in the ordering of concentrated colloidal dispersions that is a universal function of the local reduced critical stress and macroscopic strain. Samples produced at Peclet numbers greater than one and macroscopic strains above two resulted in crystal formation. A plot of the cryrstalline fraction versus Peclet number yielded a sharp order to disorder transition at Peclet number of order unity. The effect of volume fraction on the Peclet number theory was studied. Results indicated that the theory applied to volume fractions within the crystalline regime. Strain requirements for crystal formation of samples undergoing step strain deformation in a parallel plate geometry were investigated by applying stains of 1--300 to samples

  7. Blinking correlation in nanocrystal quantum dots probed with novel laser scanning confocal microscopy methods

    NASA Astrophysics Data System (ADS)

    Hefti, Ryan Alf

    Semiconductor quantum dots have a vast array of applications: as fluorescent labels in biological systems, as physical or chemical sensors, as components in photovoltaic technology, and in display devices. An attribute of nearly every quantum dot is its blinking, or fluorescence intermittency, which tends to be a disadvantage in most applications. Despite the fact that blinking has been a nearly universal phenomenon among all types of fluorescent constructs, it is more prevalent in quantum dots than in traditional fluorophores. Furthermore, no unanimously accepted model of quantum dot blinking yet exists. The work encompassed by this dissertation began with an in-depth study of molecular motor protein dynamics in a variety of environments using two specially developed techniques, both of which feature applicability to live cell systems. Parked-beam confocal microscopy was utilized to increase temporal resolution of molecular motor motion dynamics by an order of magnitude over other popular methods. The second technique, fast-scanning confocal microscopy (FSCM), was used for long range observation of motor proteins. While using FSCM on motor protein assays, we discovered an unusual phenomenon. Single quantum dots seemingly communicated with neighboring quantum dots, indicated by a distinct correlation in their blinking patterns. In order to explain this novel correlation phenomenon, the majority of blinking models developed thus far would suggest a dipole-dipole interaction or a Coulomb interaction between singly charged quantum dots. However, our results indicate that the interaction energy is higher than supported by current models, thereby prompting a renewed examination. We propose that the blinking correlation we observed is due to a Coulomb interaction on the order of 3-4 elementary charges per quantum dot and that multiple charging of individual quantum dots may be required to plunge them into a non-emissive state. As a result of charging, charge carriers are

  8. Optical biopsy of early gastroesophageal cancer by catheter-based reflectance-type laser-scanning confocal microscopy.

    PubMed

    Nakao, Madoka; Yoshida, Shigeto; Tanaka, Shinji; Takemura, Yoshito; Oka, Shiro; Yoshihara, Masaharu; Chayama, Kazuaki

    2008-01-01

    Magnified endoscopic observation of the gastrointestinal tract has become possible. However, such observation at the cellular level remains difficult. Laser-scanning confocal microscopy (LCM) is a novel, noninvasive optical imaging method that provides instant microscopic images of untreated tissue under endoscopy. We compare prototype catheter-based reflectance-type LCM images in vivo and histologic images of early gastroesophageal cancer to assess the usefulness of LCM in diagnosing such cancer. 20 sites in the esophagus and 40 sites in the stomach are examined by LCM under endoscopy prior to endoscopic or surgical resection. A prototype catheter LCM system, equipped with a semiconductor laser that oscillates at 685 nm and analyzes reflected light (Mauna Kea Technologies, Paris, France; Fujinon, Saitama, Japan) is used in vivo without fluorescent agent. In all normal esophageal mucosa and esophageal cancers, the nuclei are visualized. In nine of the ten normal esophageal mucosa, cell membranes are visualized, and in five of the ten esophageal cancers, cell membranes are visualized. In all normal gastric mucosa, nuclei and cell membranes are not visualized, but in ten of the 20 gastric cancers, nuclei are visualized. This novel method will aid in immediate diagnosis under endoscopy without the need for biopsy. PMID:19021423

  9. In-situ investigation of thermal instabilities and solid state dewetting in polycrystalline platinum thin films via confocal laser microscopy

    SciTech Connect

    Jahangir, S.; Cheng, Xuan; Huang, H. H.; Nagarajan, V.; Ihlefeld, J.

    2014-10-28

    Solid state dewetting and the subsequent morphological changes for platinum thin films grown on zinc oxide (ZnO) buffered (001) silicon substrates (Pt/ZnO/SiO{sub 2}/(001)Si system) is investigated under vacuum conditions via a custom-designed confocal laser microscope coupled with a laser heating system. Live imaging of thin film dewetting under a range of heating and quenching vacuum ambients reveals events including hillock formation, hole formation, and hole growth that lead to formation of a network of Pt ligaments, break up of Pt ligaments to individual islands and subsequent Pt islands shape reformation, in chronological fashion. These findings are corroborated by ex-situ materials characterization and quantitative electron microscopy analysis. A secondary hole formation via blistering before film rupture is revealed to be the critical stage, after which a rapid dewetting catastrophe occurs. This process is instantaneous and cannot be captured by ex-situ methods. Finally, an intermetallic phase forms at 900 °C and alters the morphology of Pt islands, suggesting a practical limit to the thermal environments that may be used for these platinized silicon wafers in vacuum conditions.

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

    PubMed Central

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

    2014-01-01

    Aim: The aim of this study is to address the cause of persistent infection of root cementum by Enterococcus faecalis. Materials and Methods: 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. Results: E. faecalis penetrated 160 μm deep into the root cementum in group III samples and only showed adhesion in group II samples. Conclusion: Penetration and survival of E. faecalis deep inside the cementum in extreme conditions could be the reason for persistent infection. PMID:24778505

  11. Automated Confocal Laser Scanning Microscopy and Semiautomated Image Processing for Analysis of Biofilms

    PubMed Central

    Kuehn, Martin; Hausner, Martina; Bungartz, Hans-Joachim; Wagner, Michael; Wilderer, Peter A.; Wuertz, Stefan

    1998-01-01

    The purpose of this study was to develop and apply a quantitative optical method suitable for routine measurements of biofilm structures under in situ conditions. A computer program was designed to perform automated investigations of biofilms by using image acquisition and image analysis techniques. To obtain a representative profile of a growing biofilm, a nondestructive procedure was created to study and quantify undisturbed microbial populations within the physical environment of a glass flow cell. Key components of the computer-controlled processing described in this paper are the on-line collection of confocal two-dimensional (2D) cross-sectional images from a preset 3D domain of interest followed by the off-line analysis of these 2D images. With the quantitative extraction of information contained in each image, a three-dimensional reconstruction of the principal biological events can be achieved. The program is convenient to handle and was generated to determine biovolumes and thus facilitate the examination of dynamic processes within biofilms. In the present study, Pseudomonas fluorescens or a green fluorescent protein-expressing Escherichia coli strain, EC12, was inoculated into glass flow cells and the respective monoculture biofilms were analyzed in three dimensions. In this paper we describe a method for the routine measurements of biofilms by using automated image acquisition and semiautomated image analysis. PMID:9797255

  12. Skeletal remodeling dynamics: New approaches with imaging instrumentation. [Laser confocal microscopy:a2

    SciTech Connect

    Parks, N.J.; Pinkerton, K.E.; Seibert, J.A.; Pool, R.R.

    1991-01-01

    This report of progress and future objectives timetable is based on an included schematic of goals and objectives and the project abstract which is included as Appendix 1. Five matters are summarized in the order of (1) novel methods of calcified bone confocal microscopy and reconstruction image analysis of decalcified beagle and human cortical bone serial sections, (2) macroscopic cross-correlation of beagle and human cortical and cancellous bone fractions with CT analysis, (3) guidance to the most radiobiologically important skeletal regions of interest with the just completed {sup 90}Sr bone tumor map from life time beagle studies, (4) deposition patterns of radioactive agents that participate in apatite crystal nucleation processes in bone and leave radiation-excited electrons trapped in bone mineral, and (5) the budget period timetable. The discovery that beta particles from {sup 166}Ho (T{sub {1/2}} =26 hr, {beta}{sub max} = 1.8 MeV) phosphonic acid bone agents leave detectable, long-lived, electron paramagnetic resonance signals in bone is included in Appendix 2 as a joint report.

  13. Reconstruction and representation of caudal vasculature of zebrafish embryo from confocal scanning laser fluorescence microscopic images.

    PubMed

    Feng, Jun; Cheng, Shuk Han; Chan, Po K; Ip, Horace H S

    2005-12-01

    Three-dimensional (3D) reconstruction from a series of sections is an important technique in medical imaging, particularly for visualization of blood vessels from angiography. Here, we present a framework for automatic segmentation and registration of different kind of blood vessels from 2-day-old zebrafish embryos. Series of optical sections were acquired from confocal microscopy with the blood vessels labeled by fluorescent microbeads (0.02 microm) injected into blood stream of 2-day-old zebrafish embryos. Blood vessels were extracted and their morphological parameters, including length and diameter, were calculated. At the same time, individual blood vessels were registered automatically. Vasculature was represented by attributed vessel represent graph (AVRG), which contained morphological data and connectivity of every blood vessel. Using AVRG to represent a vasculature made the comparison between vasculatures of different embryos more easy. Visualization, as well as quantification, of reconstructed 3D model of AVRG was presented in an interactive interface. The framework was implemented by Visual C++ as Windows-based program. PMID:16263106

  14. Noninvasive in vivo confocal laser scanning microscopy is effective in differentiating allergic from nonallergic equivocal patch test reactions.

    PubMed

    Slodownik, D; Levi, A; Lapidoth, M; Ingber, A; Horev, L; Enk, C D

    2015-04-01

    Patch testing is the gold standard for the validation of contact dermatitis. It relies on the subjective scoring by an evaluator of the inflammatory reaction induced by an allergen applied to the skin. Equivocal reactions imply faint erythema and could represent allergic, irritant, or negative reactions. They constitute approximately 1 % of the positive reactions encountered in patch test practice. Histological evaluation of the equivocal reaction has proven helpful for the correct interpretation but is however time consuming, and its invasive nature is often unacceptable to the patient. In vivo confocal laser scanning microscopy (CLSM) is a novel, noninvasive imaging technique which permits real-time visualization of skin structures and lesions at a resolution close to that obtained by conventional histology. CLSM has been successfully applied for the differentiation between clinically clear-cut allergic and irritant patch test reactions. The objective of this study is to determine the relevance of CLSM in differentiating between allergic, irritant, and negative equivocal patch test reactions. Fifteen patients who underwent patch testing in our clinic were observed as having 20 equivocal reactions. All 20 reactions were evaluated using in vivo CLSM and compared with adjacent normal skin. In vivo CLSM evaluation revealed that 8 of the 20 equivocal reactions (40 %) showed confocal patterns consistent with the patterns encountered in positive allergic reactions. Anamnestic exposure, i.e., detailed assessment of previous related contact with these allergens, confirmed high relevance rates. In vivo CLSM is useful in differentiating between allergic, irritant, and negative equivocal patch test reactions, a differentiation that cannot be made by conventional clinical patch test reading. PMID:25604734

  15. In situ detection of the Zn(2+) release process of ZnO NPs in tumour cells by confocal laser scanning fluorescence microscopy.

    PubMed

    Song, Wenshuang; Tang, Xiaoling; Li, Yong; Sun, Yang; Kong, Jilie; Qingguang, Ren

    2016-08-01

    The use of zinc oxide (ZnO) nanoparticles (NPs) for cancer is not yet clear for human clinical applications, which is primarily due to the lack of a better understanding of the action mechanisms and cellular consequences of the direct exposure of cells to these NPs. In this work, the authors have selected zinquin ethyl ester, a Zn(2+)-specific fluorescent molecular probe, to efficiently differentiate ZnO NPs and Zn(2+), and combined with confocal laser scanning microscopy (CLSM) to in situ study the Zn(2+) release process of ZnO NPs in cancer cell system through detecting the change of Zn(2+) level over time. During the experiments, the authors have designed the test group ZnO-2 in addition to assess the influence of a long-term storage on the characteristics of ZnO NPs in aqueous solution, and the Zn(2+) release process of ZnO NPs in cancer cell system. After three-month storage at room temperature, the release process became earlier and faster, which was consistent with previous results of transmission electron microscope, UV-Vis and PL spectra. It is a good detection method that combination of Zn(2+)-specific fluorescent molecular probe and CLSM, which will be helpful for ZnO NPs using in clinical research. PMID:27463786

  16. Evaluation of the extracellular polymeric substances by confocal laser scanning microscopy in conventional activated sludge and advanced membrane bioreactors treating hospital wastewater.

    PubMed

    Alrhmoun, Mousaab; Carrion, Claire; Casellas, Magali; Dagot, Christophe

    2014-01-01

    Confocal laser scanning microscopy (CLSM) combined with fluorescent viability indicators, was used in this study to investigate the impact of hospital wastewaters on floc structure and composition. In this work, three pilot-scale projects, two membrane bioreactors (MBRs) with a submerged or external membrane bioreactor and a conventional activated sludge, were installed and operated for 65 days. They were fed with an influent sampled directly from the hospital drainage system, which contained micropollutant concentrations ranging from ng/L to mg/L. Samples of flocs were observed using CLSM to characterize the extracellular polymeric substances (EPS) stained with concanavalin A-tetra methylrhodamine and fluorescein isothiocyanate solution and combined with a fluorescent viability indicator (Baclight(®) Bacterial Viability Kit, Molecular Probes), allowing visualization of isolated stained cells in the three-dimensional structure of flocs (damaged or not). The results of CLSM of the sludge composition were compared with classical biochemical analysis of EPS made through a thermal extraction method. The results showed a good relation between these analyses and the statistical treatment of microscopic pictures. PMID:24901624

  17. Classification of nanoparticle diffusion processes in vital cells by a multifeature random forests approach: application to simulated data, darkfield, and confocal laser scanning microscopy

    NASA Astrophysics Data System (ADS)

    Wagner, Thorsten; Kroll, Alexandra; Wiemann, Martin; Lipinski, Hans-Gerd

    2016-04-01

    Darkfield and confocal laser scanning microscopy both allow for a simultaneous observation of live cells and single nanoparticles. Accordingly, a characterization of nanoparticle uptake and intracellular mobility appears possible within living cells. Single particle tracking makes it possible to characterize the particle and the surrounding cell. In case of free diffusion, the mean squared displacement for each trajectory of a nanoparticle can be measured which allows computing the corresponding diffusion coefficient and, if desired, converting it into the hydrodynamic diameter using the Stokes-Einstein equation and the viscosity of the fluid. However, within the more complex system of a cell's cytoplasm unrestrained diffusion is scarce and several other types of movements may occur. Thus, confined or anomalous diffusion (e.g. diffusion in porous media), active transport, and combinations thereof were described by several authors. To distinguish between these types of particle movement we developed an appropriate classification method, and simulated three types of particle motion in a 2D plane using a Monte Carlo approach: (1) normal diffusion, using random direction and step-length, (2) subdiffusion, using confinements like a reflective boundary with defined radius or reflective objects in the closer vicinity, and (3) superdiffusion, using a directed flow added to the normal diffusion. To simulate subdiffusion we devised a new method based on tracks of different length combined with equally probable obstacle interaction. Next we estimated the fractal dimension, elongation and the ratio of long-time / short-time diffusion coefficients. These features were used to train a random forests classification algorithm. The accuracy for simulated trajectories with 180 steps was 97% (95%-CI: 0.9481-0.9884). The balanced accuracy was 94%, 99% and 98% for normal-, sub- and superdiffusion, respectively. Nanoparticle tracking analysis was used with 100 nm polystyrene particles

  18. Investigating the correlation between white matter and microvasculature changes in aging using large scale optical coherence tomography and confocal fluorescence imaging combined with tissue sectioning

    NASA Astrophysics Data System (ADS)

    Castonguay, Alexandre; Avti, Pramod K.; Moeini, Mohammad; Pouliot, Philippe; Tabatabaei, Maryam S.; Bélanger, Samuel; Lesage, Frédéric

    2015-03-01

    Here, we present a serial OCT/confocal scanner for histological study of the mouse brain. Three axis linear stages combined with a sectioning vibratome allows to cut thru the entire biological tissue and to image every section at a microscopic resolution. After acquisition, each OCT volume and confocal image is re-stitched with adjacent acquisitions to obtain a reconstructed, digital volume of the imaged tissue. This imaging platform was used to investigate correlations between white matter and microvasculature changes in aging mice. Three age groups were used in this study (4, 12, 24 months). At sacrifice, mice were transcardially perfused with a FITC containing gel. The dual imaging capability of the system allowed to reveal different contrast information: OCT imaging reveals changes in refractive indices giving contrast between white and grey matter in the mouse brain, while transcardial perfusion of a FITC shows microsvasculature in the brain with confocal imaging.

  19. Optical coherence tomography and confocal fluorescence microscopy as a combined method for studying morphological changes in lung dynamics

    NASA Astrophysics Data System (ADS)

    Gaertner, Maria; Cimalla, Peter; Knels, Lilla; Meissner, Sven; Schnabel, Christian; Kuebler, Wolfgang M.; Koch, Edmund

    2011-03-01

    Acute lung injury (ALI) is a severe pulmonary disease leading to hypoxemia accompanied by a reduced compliance and partial edema of the lung. Most of the patients have to be ventilated to compensate for the lack of oxygen. The treatment is strongly connected with ventilator induced lung injury (VILI), which is believed to introduce further stress to the lung and changes in its elastic performance. A thorough understanding of the organs micro-structure is crucial to gain more insight into the course of the disease. Due to backscattering of near-infrared light, detailed description of lung morphology can be obtained using optical coherence tomography (OCT), a non-invasive, non-contact, high resolution and fast three-dimensional imaging technique. One of its drawbacks lies in the non-specificity of light distribution in relation to defined substances, like elastic biomolecules. Using fluorescence detection, these chemical components can be visualized by introducing specifically binding fluorophores. This study presents a combined setup for studying alveolar compliance depending on volume changes and elastic fiber distributions. Simultaneously acquired OCT and confocal fluorescence images allow an entire view into morphological rearrangements during ventilation for an ex vivo mouse model using continuous pulmonary airway pressure at different values.

  20. Imaging genes, chromosomes, and nuclear structures using laser-scanning confocal microscopy

    NASA Astrophysics Data System (ADS)

    Ballard, Stephen G.

    1990-08-01

    condensed metaphase chromosomes and in interphase nuclei. The ability to image the loci of fluorescent-labelled gene probes hybridized to chromosomes and to interphase nuclei will play a major role in the mapping of the human genome. This presentation is an overview of our laboratory's efforts to use confocal imaging to address fundamental questions about the structure and organization of genes, chromosomes and cell nuclei, and to develop applications useful in clinical diagnosis of inherited diseases.

  1. Crystallization Behavior of Perovskite in the Synthesized High-Titanium-Bearing Blast Furnace Slag Using Confocal Scanning Laser Microscope

    NASA Astrophysics Data System (ADS)

    Hu, Meilong; Liu, Lu; Lv, Xuewei; Bai, Chenguang; Zhang, Shengfu

    2013-10-01

    The isothermal phase composition of high-titanium-bearing slag (23 mass pct TiO2) under an argon atmosphere during cooling process from 1723 K (1450 °C) was calculated by FactSage.6.3 (CRCT-ThermFact Inc., Montréal, Canada). Three main phases, which were perovskite, titania spinel, and clinopyroxene, could form during the cooling process and they precipitated at 1713 K, 1603 K, and 1498 K (1440 °C, 1330 °C, and 1225 °C), respectively. The nonisothermal crystallization process of perovskite in synthesized high-titanium-bearing slag was studied in situ by a confocal scanning laser microscope (CSLM) with cooling rate of 30 K/min. The results showed that the primary phase was perovskite that precipitated at 1703 K (1430 °C). The whole precipitation and growth process of perovskite was obtained, whereas other phases formed as glass under the current experimental conditions. Perovskite grew along a specific growth track and finally appeared with snowflake morphology. The growing kinetics of perovskite formation from molten slag were also mentioned.

  2. Direct observation of the asphaltene structure in paving-grade bitumen using confocal laser-scanning microscopy.

    PubMed

    Bearsley, S; Forbes, A; Haverkamp, R G

    2004-08-01

    The structure of the asphaltene phase in the bitumen is believed to have a significant effect on its rheological properties. It has traditionally been difficult to observe the asphaltene phase in unaltered samples of bitumen. The maltenes are thought to form a continuous phase in which the asphaltenes are 'dispersed'. In this study, confocal laser-scanning microscopy (CLSM) operating in fluorescence mode was used to examine the structure of paving-grade Safaniya and San Joaquin bitumen. The asphaltene fraction fluoresces in the 515-545 nm wavelength range when irradiated with light with a wavelength of 488 nm. The major advantages of CLSM are that the bitumen sample requires little pretreatment or preparation that may affect the original dispersion of asphaltenes and the bitumen is observed at ambient temperature and pressure. This reduces the possibility of producing images that are not representative of the original material. CLSM was able to show the distribution of maltene and asphaltene components in bitumen. The asphaltene aggregates in the bitumen were observed to be 2-7 micro m in size and formed a dispersed 'sol' structure in the continuous maltene matrix rather than a network 'gel' structure. Surprisingly, the structure and fluorescence of the asphaltene phase does not appear to alter radically upon oxidative ageing. The structure of the asphaltene phase of an AR4000 San Joaquin bitumen was found to be more homogeneous than that of Safaniya bitumen, illustrating the range of structures that can be observed in bitumens by this method. PMID:15315501

  3. Tracking features in retinal images of adaptive optics confocal scanning laser ophthalmoscope using KLT-SIFT algorithm.

    PubMed

    Li, Hao; Lu, Jing; Shi, Guohua; Zhang, Yudong

    2010-01-01

    With the use of adaptive optics (AO), high-resolution microscopic imaging of living human retina in the single cell level has been achieved. In an adaptive optics confocal scanning laser ophthalmoscope (AOSLO) system, with a small field size (about 1 degree, 280 μm), the motion of the eye severely affects the stabilization of the real-time video images and results in significant distortions of the retina images. In this paper, Scale-Invariant Feature Transform (SIFT) is used to abstract stable point features from the retina images. Kanade-Lucas-Tomasi(KLT) algorithm is applied to track the features. With the tracked features, the image distortion in each frame is removed by the second-order polynomial transformation, and 10 successive frames are co-added to enhance the image quality. Features of special interest in an image can also be selected manually and tracked by KLT. A point on a cone is selected manually, and the cone is tracked from frame to frame. PMID:21258443

  4. Precision Automation of Cell Type Classification and Sub-Cellular Fluorescence Quantification from Laser Scanning Confocal Images

    PubMed Central

    Hall, Hardy C.; Fakhrzadeh, Azadeh; Luengo Hendriks, Cris L.; Fischer, Urs

    2016-01-01

    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. PMID:26904081

  5. Precision Automation of Cell Type Classification and Sub-Cellular Fluorescence Quantification from Laser Scanning Confocal Images.

    PubMed

    Hall, Hardy C; Fakhrzadeh, Azadeh; Luengo Hendriks, Cris L; Fischer, Urs

    2016-01-01

    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. PMID:26904081

  6. Nano-zymography Using Laser-Scanning Confocal Microscopy Unmasks Proteolytic Activity of Cell-Derived Microparticles.

    PubMed

    Briens, Aurélien; Gauberti, Maxime; Parcq, Jérôme; Montaner, Joan; Vivien, Denis; Martinez de Lizarrondo, Sara

    2016-01-01

    Cell-derived microparticles (MPs) are nano-sized vesicles released by activated cells in the extracellular milieu. They act as vectors of biological activity by carrying membrane-anchored and cytoplasmic constituents of the parental cells. Although detection and characterization of cell-derived MPs may be of high diagnostic and prognostic values in a number of human diseases, reliable measurement of their size, number and biological activity still remains challenging using currently available methods. In the present study, we developed a protocol to directly image and functionally characterize MPs using high-resolution laser-scanning confocal microscopy. Once trapped on annexin-V coated micro-wells, we developed several assays using fluorescent reporters to measure their size, detect membrane antigens and evaluate proteolytic activity (nano-zymography). In particular, we demonstrated the applicability and specificity of this method to detect antigens and proteolytic activities of tissue-type plasminogen activator (tPA), urokinase and plasmin at the surface of engineered MPs from transfected cell-lines. Furthermore, we were able to identify a subset of tPA-bearing fibrinolytic MPs using plasma samples from a cohort of ischemic stroke patients who received thrombolytic therapy and in an experimental model of thrombin-induced ischemic stroke in mice. Overall, this method is promising for functional characterization of cell-derived MPs. PMID:27022410

  7. The effect of copper on different phototrophic microorganisms determined in vivo and at cellular level by confocal laser microscopy.

    PubMed

    Seder-Colomina, M; Burgos, A; Maldonado, J; Solé, A; Esteve, I

    2013-01-01

    Microbial mats are coastal ecosystems that consist mainly of cyanobacteria, primary producers in these habitats that play an important role in stabilising delta sediments. However, these ecosystems are subject to various kinds of pollution, including metal contamination, placing their survival at risk. Among heavy metals, copper is an essential metal at low doses and toxic at high doses. This metal is present in different pesticides used in rice production, a thriving agro-industry in the Ebro Delta (Spain). For several years, our group has been studying the Ebro Delta microbial mats and has developed a method for determining the effect that metals cause on cyanobacteria populations. This method is based on confocal laser microscopy coupled to a spectrofluorometer, which rapidly provides simultaneous three-dimensional information on photosynthetic microorganisms and their fluorescence spectra profiles. The current study determines the copper effect on different photosynthetic microorganisms from culture collection (Chroococcus sp. PCC 9106 and Spirulina sp. PCC 6313) and isolated from the environment (Microcoleus-like and the microalga DE2009). Comparing all results obtained it can be observed that the minimum dose of Cu that is capable of significantly altering chlorophyll a (chl a) fluorescence intensity were 1 × 10(-7) M in Chroococcus sp. PCC 9106; 1 × 10(-7) M in Spirulina sp. PCC 6313; 3 × 10(-7) M in Microcoleus and 5 × 10(-6) M in the microalga DE2009. Moreover, the sensitivity of the technique used was 1 × 10(-7) M. PMID:23138333

  8. Nano-zymography Using Laser-Scanning Confocal Microscopy Unmasks Proteolytic Activity of Cell-Derived Microparticles

    PubMed Central

    Briens, Aurélien; Gauberti, Maxime; Parcq, Jérôme; Montaner, Joan; Vivien, Denis; Martinez de Lizarrondo, Sara

    2016-01-01

    Cell-derived microparticles (MPs) are nano-sized vesicles released by activated cells in the extracellular milieu. They act as vectors of biological activity by carrying membrane-anchored and cytoplasmic constituents of the parental cells. Although detection and characterization of cell-derived MPs may be of high diagnostic and prognostic values in a number of human diseases, reliable measurement of their size, number and biological activity still remains challenging using currently available methods. In the present study, we developed a protocol to directly image and functionally characterize MPs using high-resolution laser-scanning confocal microscopy. Once trapped on annexin-V coated micro-wells, we developed several assays using fluorescent reporters to measure their size, detect membrane antigens and evaluate proteolytic activity (nano-zymography). In particular, we demonstrated the applicability and specificity of this method to detect antigens and proteolytic activities of tissue-type plasminogen activator (tPA), urokinase and plasmin at the surface of engineered MPs from transfected cell-lines. Furthermore, we were able to identify a subset of tPA-bearing fibrinolytic MPs using plasma samples from a cohort of ischemic stroke patients who received thrombolytic therapy and in an experimental model of thrombin-induced ischemic stroke in mice. Overall, this method is promising for functional characterization of cell-derived MPs. PMID:27022410

  9. The binding of cellulase variants to dislocations: a semi-quantitative analysis based on CLSM (confocal laser scanning microscopy) images.

    PubMed

    Hidayat, Budi J; Weisskopf, Carmen; Felby, Claus; Johansen, Katja S; Thygesen, Lisbeth G

    2015-12-01

    Binding of enzymes to the substrate is the first step in enzymatic hydrolysis of lignocellulose, a key process within biorefining. During this process elongated plant cells such as fibers and tracheids have been found to break into segments at irregular cell wall regions known as dislocations or slip planes. Here we study whether cellulases bind to dislocations to a higher extent than to the surrounding cell wall. The binding of fluorescently labelled cellobiohydrolases and endoglucanases to filter paper fibers was investigated using confocal laser scanning microscopy and a ratiometric method was developed to assess and quantify the abundance of the binding of cellulases to dislocations as compared to the surrounding cell wall. Only Humicola insolens EGV was found to have stronger binding preference to dislocations than to the surrounding cell wall, while no difference in binding affinity was seen for any of the other cellulose variants included in the study (H. insolens EGV variants, Trichoderma reesei CBHI, CBHII and EGII). This result favours the hypothesis that fibers break at dislocations during the initial phase of hydrolysis mostly due to mechanical failure rather than as a result of faster degradation at these locations. PMID:26626331

  10. Real-time in vivo confocal laser scanning microscopy of melanin-containing cells: A promising diagnostic intervention.

    PubMed

    Xiang, Wenzhong; Song, Xiuzu; Peng, Jianzhong; Xu, Aie; Bi, Zhigang

    2015-12-01

    The use of noninvasive imaging techniques to evaluate different types of skin lesions is increasing popular. In vivo confocal laser scanning microscopy (CLSM) is a new method for high resolution non-invasive imaging of intact skin in situ and in vivo. Although many studies have investigated melanin-containing cells in lesions by in vivo CLSM, few studies have systematically characterized melanin-containing cells based on their morphology, size, arrangement, density, borders, and brightness. In this study, the characteristics of melanin-containing cells were further investigated by in vivo CLSM. A total of 130 lesions, including common nevi, giant congenital pigmented nevi, vitiligo, melasma, melanoma, and chronic eczema, were imaged by in vivo CLSM. This research helps dermatologists understand the characteristics of melanin-containing cells and facilitate the clinical application of melanin-containing cells in the investigation of dermatological disease. In summary, melanin-containing cells include keratinocytes, melanocytes, macrophages, and melanocytic skin tumor cells. Our study presents the CLSM characteristics of melanin-containing cells to potentially facilitate in vivo diagnosis based on shape, size, arrangement, density, borders, and brightness. PMID:26515646

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

    NASA Astrophysics Data System (ADS)

    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.

  12. Mechanism and kinetics of protein transport in chromatographic media studied by confocal laser scanning microscopy. Part II. Impact on chromatographic separations.

    PubMed

    Hubbuch, Jürgen; Linden, Thomas; Knieps, Esther; Thömmes, Jörg; Kula, Maria-Regina

    2003-12-22

    The impact of different transport mechanism on chromatographic performance was studied by confocal laser scanning microscopy (CLSM) for solutions containing bovine serum albumin (BSA) and monoclonal IgG 2a under different solid- and fluid-phase conditions. During this investigation, a clear influence of the uptake mechanism on the affinity of the respective proteins for the different adsorbents and thus separation performance of the chromatographic process could be observed. For the system SP Sepharose Fast Flow at pH 4.5 pore diffusion could be ascribed to be the dominant transport mechanism for both proteins and the adsorption profiles resembled a pattern similar to that described by the 'shrinking core' model. Under these conditions a significantly higher affinity towards the adsorbent was found for BSA when compared to IgG 2a. With changing fluid- and solid-phase conditions, however, a change of the transport mode for IgG 2a could be detected. While the exact mechanism is still unresolved it could be concluded that both occurrence and magnitude of the now governing transport mechanism depended on protein properties and interaction with the adsorbent surface. For the system SP Sepharose XL at pH 5.0 both parameters leading to the change in IgG 2a uptake were combined resulting in a clear change of the system affinity towards the IgG 2a molecule, while BSA adsorption was restricted to the most outer shell of the sorbent. PMID:14735979

  13. Differentiation of Methanosaeta concilii and Methanocarcina barkeri in anaerobic mesophilic granular sludge by fluorescent in situ hybridization and confocal scanning laser microscopy

    SciTech Connect

    Rocheleau, S.; Greer, C.W.; Cantin, C.; Laramee, L.; Guiot, S.R.; Lawrence, J.R.

    1999-05-01

    Oligonucleotide probes, designed from genes coding for 16S rRNA, were developed to differentiate Methanosaeta concilii, Methanosarcina barkeri, and mesophilic methanogens. All M. concilii oligonucleotide probes (designated MS1, MS2, and MS5) hybridized specifically with the target DNA, but MS5 was the most specific M. concilii oligonucleotide probe. Methanosarcina barkeri oligonucleotide probes (designated MB1, MB3, and MB4) hybridized with different Methanosarcina species. The MB4 probe specifically detected Methanosarcina barkeri, and the MB3 probe detected the presence of al mesophilic Methanosarcina species. These new oligonucleotide probes facilitated the identification, localization, and quantification of the specific relative abundance of M. concilii and Methanosarcina barkeri, which play important roles in methanogenesis. The combined use of fluorescent in situ hybridization with confocal scanning laser microscopy demonstrated that anaerobic granule topography depends on granule origin and feeding. Protein-fed granules showed no layered structure with a random distribution of M. concilii. In contrast, a layered structure developed in methanol-enriched granules, where M. barkeri growth was induced in an outer layer. This outer layer was followed by a layer composed of M. concilii, with an inner core of M. concilii and other bacteria.

  14. The case of diagnostics of invasive pulmonary aspergillosis by in vivo probe-based confocal laser endomicroscopy of central and distal airways

    PubMed Central

    Danilevskaya, Olesya; Averyanov, Alexander; Klimko, Nikolay; Lesnyak, Viktor; Sorokina, Anastasia; Sazonov, Dmitry; Zabozlaev, Fedor

    2014-01-01

    We present a case of 41-year-old patient with invasive pulmonary aspergillosis (IPA) in which probe-based confocal laser endomicroscopy (pCLE) imaging of central and distal airways was first performed in vivo. pCLE imaging showed the signs of complete or partial destruction of elastin network of alveolar wall with fibrillar branching fluorescent structures in the zone with typical IPA changes on HRCT. PMID:25180153

  15. Mapping of Heavy Metal Ion Sorption to Cell-Extracellular Polymeric Substance-Mineral Aggregates by Using Metal-Selective Fluorescent Probes and Confocal Laser Scanning Microscopy

    PubMed Central

    Li, Jianli; Kappler, Andreas; Obst, Martin

    2013-01-01

    Biofilms, organic matter, iron/aluminum oxides, and clay minerals bind toxic heavy metal ions and control their fate and bioavailability in the environment. The spatial relationship of metal ions to biomacromolecules such as extracellular polymeric substances (EPS) in biofilms with microbial cells and biogenic minerals is complex and occurs at the micro- and submicrometer scale. Here, we review the application of highly selective and sensitive metal fluorescent probes for confocal laser scanning microscopy (CLSM) that were originally developed for use in life sciences and propose their suitability as a powerful tool for mapping heavy metals in environmental biofilms and cell-EPS-mineral aggregates (CEMAs). The benefit of using metal fluorescent dyes in combination with CLSM imaging over other techniques such as electron microscopy is that environmental samples can be analyzed in their natural hydrated state, avoiding artifacts such as aggregation from drying that is necessary for analytical electron microscopy. In this minireview, we present data for a group of sensitive fluorescent probes highly specific for Fe3+, Cu2+, Zn2+, and Hg2+, illustrating the potential of their application in environmental science. We evaluate their application in combination with other fluorescent probes that label constituents of CEMAs such as DNA or polysaccharides and provide selection guidelines for potential combinations of fluorescent probes. Correlation analysis of spatially resolved heavy metal distributions with EPS and biogenic minerals in their natural, hydrated state will further our understanding of the behavior of metals in environmental systems since it allows for identifying bonding sites in complex, heterogeneous systems. PMID:23974141

  16. Evaluation of the therapeutic results of actinic keratosis treated with topical 5% fluorouracil by reflectance confocal laser microscopy: preliminary study*

    PubMed Central

    Ishioka, Priscila; Maia, Marcus; Rodrigues, Sarita Bartholomei; Marta, Alessandra Cristina; Hirata, Sérgio Henrique

    2015-01-01

    Topical treatment for actinic keratosis with 5% fluorouracil has a recurrence rate of 54% in 12 months of follow-up. This study analyzed thirteen actinic keratoses on the upper limbs through confocal microscopy, at the time of clinical diagnosis and after 4 weeks of treatment with fluorouracil. After the treatment was established and evidence of clinical cure was achieved, in two of the nine actinic keratoses, confocal microscopy enabled visualization of focal areas of atypical honeycomb pattern in the epidermis indicating therapeutic failure. Preliminary data suggest the use of confocal microscopy as a tool for diagnosis and therapeutic control of actinic keratosis. PMID:26131881

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

    PubMed Central

    Latha, Jothi; Velmurugan, Natanasabapathy

    2015-01-01

    Objectives 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). Materials and Methods 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. Results 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 < 0.01 for coronal third and p < 0.001 for middle and apical thirds). On intragroup analysis, both groups showed statistically significant difference among three levels (p < 0.001). Conclusions 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. PMID:25984477

  18. Mapping microclimate pH distribution inside protein-encapsulated PLGA microspheres using confocal laser scanning microscopy

    PubMed Central

    Liu, Yajun; Schwendeman, Steven P.

    2012-01-01

    The pH in the aqueous pores of poly(lactide-co-glycolide) (PLGA) matrix, also referred to microclimate pH (μpH), is often uncontrolled ranging from highly acidic to neutral pH range. The μpH distribution inside protein-encapsulated PLGA microspheres was quantitatively evaluated using confocal laser scanning microscopy. The fluorescent response of Lysosensor yellow/blue® dextran used to map μpH in PLGA was influenced by the presence of encapsulated protein. The nonprotonated form of pyridyl group on the fluorescence probe at neutral pH was responsible for the interference, which was dependent on the type and concentration of protein. A method for correction of this interference based on estimating protein concentration inside the microspheres was established and validated. After correction of the influence, the μpH distribution kinetics inside microspheres was evaluated for different PLGA 50/50 microsphere formulations under physiological conditions for 4 weeks. Generally, the μpH acidity increased with the progression of incubation time. The co-incorporation of poorly soluble base, magnesium carbonate, in the microspheres prolonged the appearance of detectable acidity for up to 3 weeks. Co-addition of an acetate buffer was able to control the μpH over a slightly acidic range (around pH 4.7) after two weeks incubation. Microspheres prepared from a lower polymer concentration exhibited a higher μpH, likely owing to reduced diffusional resistance to acidic degradation products. The stability of protein was enhanced by addition of MgCO3, acetate buffer, or by reduced polymer concentration in the preparation, as evidenced by more soluble protein recovered after incubation. Hence, the μpH imaging technique developed can be employed in the future for optimization of formulation strategies for controlling μpH and stabilizing encapsulated proteins. PMID:22428586

  19. A first approach for digital representation and automated classification of toolmarks on locking cylinders using confocal laser microscopy

    NASA Astrophysics Data System (ADS)

    Clausing, Eric; Kraetzer, Christian; Dittmann, Jana; Vielhauer, Claus

    2012-10-01

    An important part of criminalistic forensics is the analysis of toolmarks. Such toolmarks often consist of plenty of single striations, scratches and dents which can allow for conclusions in regards to the sequence of events or used tools. To receive qualified results with an automated analysis and contactless acquisition of such toolmarks, a detailed digital representation of these and their orientation as well as placing to each other is required. For marks of firearms and tools the desired result of an analysis is a conclusion whether or not a mark has been generated by a tool under suspicion. For toolmark analysis on locking cylinders, the aim is not an identification of the used tool but rather an identification of the opening method. The challenge of such an identification is that a one-to-one comparison of two images is not sufficient - although two marked objects look completely different in regards to the specific location and shape of found marks they still can represent a sample for the identical opening method. This paper provides the first approach for modelling toolmarks on lock pins and takes into consideration the different requirements necessary to generate a detailed and interpretable digital representation of these traces. These requirements are 'detail', i.e. adequate features which allow for a suitable representation and interpretation of single marks, 'meta detail', i.e. adequate representation of the context and connection between all marks and 'distinctiveness', i.e. the possibility to reliably distinguish different sample types by the according model. The model is evaluated with a set of 15 physical samples (resulting in 675 digital scans) of lock pins from cylinders opened with different opening methods, contactlessly scanned with a confocal laser microscope. The presented results suggest a high suitability for the aspired purpose of opening method determination.

  20. Direct observation by laser scanning confocal microscopy of microstructure and phase migration of PVC gels in an applied electric field.

    PubMed

    Xia, Hong; Ueki, Takamitsu; Hirai, Toshihiro

    2011-02-01

    The fluorescent probe lucigenin was incorporated in poly(vinyl chloride) (PVC) gels, and laser scanning confocal microscopy (LSCM) was used to clarify the internal structures of the gels. From the two-dimensional and three-dimensional information by LSCM, we first observed the internal structure of the PVC gel at a wet status, where the PVC gels comprised a polymer-rich phase and a polymer-poor phase uniformly with a three-dimensional network structure. After an electric field was applied, an effect of the electric field resulted in the change of internal structure in the gels. The polymer-poor phase moved from the cathode to the anode and the polymer-rich phase formed linelike arrangement between electrodes due to the attraction force. On the other hand, the freeze-dried PVC gels with/without in-situ dc voltage casting were particularly fabricated to confirm above results by the field emission scanning electron microscopy (FE-SEM). It was found that many craters remained on the surface of the gel near the anode due to sublimation in freeze-drying. This phenomenon did not appear on the surface near the cathode. The results of in-situ dc voltage casting also suggested that a substantial amount of polymer-poor phase was moved and fixed at the anode. Thus, results of both LSCM and in-situ dc voltage casting corresponded to the effect of electric field on PVC gels and provided a convincing evidence for the interpretation of the deformation mechanism of PVC gel actuators by an applied electric field. PMID:21174424

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

    PubMed Central

    Mueller, Lukas N; de Brouwer, Jody FC; Almeida, Jonas S; Stal, Lucas J; Xavier, João 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 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 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. PMID:16412253

  2. Value of Magnifying Endoscopy With Narrow-Band Imaging and Confocal Laser Endomicroscopy in Detecting Gastric Cancerous Lesions

    PubMed Central

    Gong, Shuai; Xue, Han-Bing; Ge, Zhi-Zheng; Dai, Jun; Li, Xiao-Bo; Zhao, Yun-Jia; Zhang, Yao; Gao, Yun-Jie; Song, Yan

    2015-01-01

    Abstract Although the respective potentials of magnifying endoscopy with narrow-band imaging (ME-NBI) and confocal laser endomicroscopy (CLE) in predicting gastric cancer has been well documented, there is a lack of studies in comparing the value and diagnostic strategy of these 2 modalities. Our primary aim is to investigate whether CLE is superior to ME-NBI for differentiation between gastric cancerous and noncancerous lesions. A secondary aim is to propose an applicable clinical strategy. We conducted a diagnostic accuracy study involving patients with suspected gastric superficial cancerous lesions. White light endoscopy, ME-NBI, and CLE were performed diagnostic accuracy, sensitivity, specificity, positive predictive value, and negative predictive value between ME-NBI and CLE were assessed, as well as agreements between ME-NBI/CLE and histopathology. This study involved 86 gastric lesions in 82 consecutive patients who underwent white light endoscopy, ME-NBI, and CLE before biopsy. The accuracy, sensitivity, and specificity for ME-NBI were 93.75%, 91.67%, and 95.45%, compared with 91.86%, 90%, and 93.48%, respectively, for CLE, for discrimination cancerous/noncancerous lesion (all P > 0.05). For undifferentiated/differentiated adenocarcinoma, CLE had a numerically but not statistically significantly higher accuracy than ME-NBI (81.25% vs 73.33%, P = 0.46). Agreements between ME-NBI/CLE and histopathology were near perfect (ME-NBI, κ = 0.87; CLE, κ = 0.84). CLE is not superior to ME-NBI for discriminating gastric cancerous from noncancerous lesions. Endoscopist could make an optimal choice according to the specific indication and advantages of ME-NBI and CLE in daily practices. PMID:26554797

  3. Real-time in-vivo microscopic imaging of the cervix using confocal laser endomicroscopy: preliminary observations and feasibility study.

    PubMed

    Degueldre, Michel; Vandromme, Jean; de Wind, Alexander; Feoli, Francesco

    2016-07-01

    Confocal laser endomicroscopy (CLE) enables in-vivo, real-time, imaging of tissues with a micron-scale resolution through a fiber optic probe. CLE could be a valuable tool for the detection and characterization of suspicious (dysplastic) areas on the uterine cervix in a minimally invasive manner. This study evaluates the technical feasibility and safety of CLE on the cervix. The study also aims to create a preliminary iconography of normal and dysplastic squamous and columnar cervical epithelium. In-vivo CLE was performed on nine patients scheduled for a cervical loop electric excision procedure for high-grade superficial intraepithelial lesions. The CLE images were compared with standard hematoxylin and eosin analysis of loop electric excision procedure specimens. The histopathological diagnosis on the surgical specimen was established as per standard of care. CLE images were then reviewed by pathologists to point out specific histopathological features. pCLE of the exocervix and the transformation zone was performed successfully on seven out of nine patients. Uninterpretable images were obtained in two other cases: one using the AlveoFlex and one using the GastroFlex UHD after the application of acetic acid 2%. A total of 82.5% of the sequences recorded with the GastroFlex were suitable for interpretation. No adverse event or complications occurred. CLE enables proper in-vivo imaging of healthy and dysplastic cervical tissue. Images correlate well with the histopathological features established through traditional histology. Future blinded prospective analysis will determine the reliability of the real-time diagnosis and its potential use in the assessment and treatment of cervical lesions. PMID:26287698

  4. Characterization and quantification of wound-induced hair follicle neogenesis using in vivo confocal scanning laser microscopy

    PubMed Central

    Fan, Chengxiang; Luedtke, Michael A.; Prouty, Stephen M.; Burrows, Michelle; Kollias, Nikiforos

    2011-01-01

    Background In vivo confocal scanning laser microscopy (CSLM) is a recently-developed non-invasive technique for visualizing microscopic structures with the skin. CSLM has been used to characterize proliferative and inflammatory skin diseases, neoplastic skin lesions and pigmented lesions. Objective Here, we assessed the ability of CSLM to evaluate the formation of neogenic hair follicles after a full thickness wound in mice. Methods Full-thickness wounds were made on the dorsal skin of 3-week old mice. After scab detachment (SD), the number, width, length, space and volume of neogenic hair follicles were analyzed using CSLM. The results were compared with those from conventional methods, including staining for alkaline phosphatase (AP) and keratin 17 (K17) as well as histology. Results Quantification of neogenic hair follicles using CSLM compared favorably with results from direct measurements on isolated epidermal tissue after immunostaining for K17, a marker for the epithelial portion of new hair follicles. CSLM detected 89% of K17-stained follicles. CSLM more accurately quantitated the number of new follicles compared to AP staining, which detects the dermal portion of the new follicle. The width and length measurement from CSLM and histology were very close and correlated with each other. The minimum length of a neogenic hair follicle that could be detected by CSLM was 21 μm. The space between neogenic hair follicles was decreased in histological sections compared to CSLM. Conclusions CSLM is an accurate and valuable method for counting and measuring neogenic hair follicles non-invasively. CSLM produces images similar to histology in mice. Measurements of microstructures using CSLM more accurately reflect actual sizes since this technique avoids fixation artifact. In vivo visualization of developing follicles with CSLM permits detection of serial changes in hair follicle formation, thus conserving numbers of mice required for studies and improving detection of

  5. Confocal Comparison of Corneal Reinnervation after Small Incision Lenticule Extraction (SMILE) and Femtosecond Laser In Situ Keratomileusis (FS-LASIK)

    PubMed Central

    Qin, Bing; Zhou, Zimei; Ni, Katherine; Le, Qihua; Xiang, Jun; Wei, Anji; Ma, Weiping; Zhou, Xingtao

    2013-01-01

    Purpose To evaluate corneal reinnervation, and the corresponding corneal sensitivity and keratocyte density after small incision lenticule extraction (SMILE) and femtosecond laser in situ keratomileusis (FS-LASIK). Methods In this prospective, non-randomized observational study, 18 patients (32 eyes) received SMILE surgery, and 22 patients (42 eyes) received FS-LASIK surgery to correct myopia. The corneal subbasal nerve density and microscopic morphological changes in corneal architecture were evaluated by confocal microscopy prior to surgery and at 1 week, 1 month, 3 months, and 6 months after surgery. A correlation analysis was performed between subbasal corneal nerve density and the corresponding keratocyte density and corneal sensitivity. Results The decrease in subbasal nerve density was less severe in SMILE-treated eyes than in FS-LASIK-treated eyes at 1 week (P = 0.0147), 1 month (P = 0.0243), and 3 months (P = 0.0498), but no difference was detected at the 6-month visit (P = 0.5277). The subbasal nerve density correlated positively with central corneal sensitivity in both groups (r = 0.416, P<0.0001, and r = 0.2567, P = 0.0038 for SMILE group and FS-LASIK group, respectively). The SMILE-treated eyes have a lower risk of developing peripheral empty space with epithelial cells filling in (P = 0.0005). Conclusions The decrease in subbasal nerve fiber density was less severe in the SMILE group than the FS-LASIK group in the first 3 months following the surgeries. The subbasal nerve density was correlated with central corneal sensitivity. PMID:24349069

  6. Mapping microclimate pH distribution inside protein-encapsulated PLGA microspheres using confocal laser scanning microscopy.

    PubMed

    Liu, Yajun; Schwendeman, Steven P

    2012-05-01

    The pH in the aqueous pores of poly(lactide-co-glycolide) (PLGA) matrix, also referred to as microclimate pH (μpH), is often uncontrolled, ranging from highly acidic to neutral pH range. The μpH distribution inside protein-encapsulated PLGA microspheres was quantitatively evaluated using confocal laser scanning microscopy. The fluorescent response of Lysosensor yellow/blue dextran used to map μpH in PLGA was influenced by the presence of encapsulated protein. The nonprotonated form of pyridyl group on the fluorescence probe at neutral pH was responsible for the interference, which was dependent on the type and concentration of protein. A method for correction of this interference based on estimating protein concentration inside the microspheres was established and validated. After correction of the influence, the μpH distribution kinetics inside microspheres was evaluated for different PLGA 50/50 microsphere formulations under physiological conditions for 4 weeks. Generally, the μpH acidity increased with the progression of incubation time. The coincorporation of poorly soluble base, magnesium carbonate, in the microspheres prolonged the appearance of detectable acidity for up to 3 weeks. Co-addition of an acetate buffer was able to control the μpH over a slightly acidic range (around pH 4.7) after two week incubation. Microspheres prepared from a lower polymer concentration exhibited a higher μpH, likely owing to reduced diffusional resistance to acidic degradation products. The stability of protein was enhanced by addition of MgCO(3), acetate buffer, or by reduced polymer concentration in the preparation, as evidenced by more soluble protein recovered after incubation. Hence, the μpH imaging technique developed can be employed in the future for optimization of formulation strategies for controlling μpH and stabilizing encapsulated proteins. PMID:22428586

  7. Reversibility of gastric mucosal lesions induced by sodium phosphate tablets and characterized by probe-based confocal laser endomicroscopy

    PubMed Central

    Coron, Emmanuel; Dewitte, Marie; Aubert, Philippe; Musquer, Nicolas; Neunlist, Michel; Bruley des Varannes, Stanislas

    2015-01-01

    Background and study aims: Adequate bowel preparation is key for the optimal quality of colonoscopy. The sodium phosphate laxatives used for preparation may induce gastric injuries. However, in vivo studies monitoring the effects of sodium phosphate on the gastric mucosa are currently lacking. We aimed to characterize the effects of sodium phosphate tablets (Colokit®; Mayoly Spindler, Chatou, France) on the gastric mucosa in a large-animal model. Methods: Fourteen anesthetized pigs were used for this study. Fundic mucosal sites were analyzed at 1.5, 24, and 72 hours after the endoscopically guided application of sodium phosphate tablets (NaPT) and placebo tablets (PlaT) and were compared with unexposed sites. Different mucosal parameters were assessed with white light endoscopy, probe-based confocal laser endomicroscopy (pCLE), histology, and ex vivo permeability measurements. Results: At 90 minutes after the application of NaPT, significant increases in epithelial irregularity and crypt pit intensity were observed with pCLE. These microscopic lesions persisted at 24 hours but were resolved at 72 hours. In addition, white light endoscopy revealed local exanthema in 57 % of the animals at 1.5 hours after NaPT application. Such lesions were observed in 22 % of the pigs at 24 hours and disappeared at 72 hours after application. After 1.5 hours, PlaT induced a slight but significant increase in epithelial irregularity, as well as architectural scores that were significantly lower than the ones induced by NaPT and that disappeared after 72 hours. Conclusions: The direct and prolonged gastric application of NaPT in pigs can induce acute superficial macroscopic and microscopic injuries that are reversible within 72 hours. PMID:26134776

  8. Poor agreement between endoscopists and gastrointestinal pathologists for the interpretation of probe-based confocal laser endomicroscopy findings

    PubMed Central

    Peter, Shajan; Council, Leona; Bang, Ji Young; Neumann, Helmut; Mönkemüller, Klaus; Varadarajulu, Shyam; Wilcox, Charles Melbern

    2014-01-01

    AIM: To compare the interpretation of probe-based confocal laser endomicroscopy (pCLE) findings between endoscopists and gastrointestinal (GI)-pathologists. METHODS: All pCLE procedures were undertaken and the endoscopist rendered assessment. The same pCLE videos were then viewed offline by an expert GI pathologist. Histopathology was considered the gold standard for definitive diagnosis. The sensitivity, specificity and accuracy for diagnosis of dysplastic/ neoplastic GI lesions and interobserver agreement between endoscopists and experienced gastrointestinal pathologist for pCLE findings were analyzed. RESULTS: Of the 66 included patients, 40 (60.6%) had lesions in the esophagus, 7 (10.6%) in the stomach, 15 (22.7%) in the biliary tract, 3 (4.5%) in the ampulla and 1 (1.5%) in the colon. The overall sensitivity, specificity and accuracy for diagnosing dysplastic/neoplastic lesions using pCLE were higher for endoscopists than pathologist at 87.0% vs 69.6%, 80.0% vs 40.0% and 84.8% vs 60.6% (P = 0.0003), respectively. Area under the ROC curve (AUC) was greater for endoscopists than the pathologist (0.83 vs 0.55, P = 0.0001). Overall agreement between endoscopists and pathologist was moderate for all GI lesions (K = 0.43; 95%CI: 0.26-0.61), luminal lesions (K = 0.40; 95%CI: 0.20-0.60) and those of dysplastic/neoplastic pathology (K = 0.55; 95%CI: 0.37-0.72), the agreement was poor for benign (K = 0.13; 95%CI: -0.097-0.36) and pancreaticobiliary lesions (K = 0.19; 95%CI: -0.26-0.63). CONCLUSION: There is a wide discrepancy in the interpretation of pCLE findings between endoscopists and pathologist, particularly for benign and malignant pancreaticobiliary lesions. Further studies are needed to identify the cause of this poor agreement. PMID:25548499

  9. Rhodamine 123 phototoxicity in laser-irradiated MGH-U1 human carcinoma cells studied in vitro by electron microscopy and confocal laser scanning microscopy

    SciTech Connect

    Shea, C.R.; Sherwood, M.E.; Flotte, T.J.; Chen, N.; Scholz, M.; Hasan, T. )

    1990-07-01

    Rhodamine 123 (R123) is a permeant, cationic, fluorescent dye that localizes preferentially within mitochondria of living carcinoma cells. MGH-U1 human bladder carcinoma cells incubated in vitro with 10 microM R123 for 30 min and then irradiated at 514.5 nm with an argon ion laser underwent selective, phototoxic injury to mitochondria. Ultrastructurally, treatment with R123 plus irradiation with 10 J/cm2 caused selective, progressive mitochondrial alterations consisting of disruption of cristae, vacuolization, swelling, increasing numbers of ring-shaped and angulated mitochondria at 4 to 8 h after irradiation, and obliteration of many mitochondria at 24 to 48 h. Confocal laser scanning microscopy after treatment with R123 plus irradiation with 10 to 30 J/cm2 demonstrated altered uptake and localization of subsequently administered R123, accompanied by striking mitochondrial fragmentation. Irradiation caused a dose-dependent depletion of extractable R123, due to a photosensitized efflux that began immediately and progressed by 4 h after irradiation with 10 to 30 J/cm2; further uptake after reincubation in the presence of R123 was also quantitatively impaired in cells previously irradiated with 30 J/cm2.

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

    NASA Astrophysics Data System (ADS)

    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.

  11. Cascaded injection resonator for coherent beam combining of laser arrays

    DOEpatents

    Kireev, Vassili [Sunnyvale, CA; Liu, Yun; Protopopescu, Vladimir [Knoxville, TN; Braiman, Yehuda [Oak Ridge, TN

    2008-10-21

    The invention provides a cascaded injection resonator for coherent beam combining of laser arrays. The resonator comprises a plurality of laser emitters arranged along at least one plane and a beam sampler for reflecting at least a portion of each laser beam that impinges on the beam sampler, the portion of each laser beam from one of the laser emitters being reflected back to another one of the laser emitters to cause a beam to be generated from the other one of the laser emitters to the beam reflector. The beam sampler also transmits a portion of each laser beam to produce a laser output beam such that a plurality of laser output beams of the same frequency are produced. An injection laser beam is directed to a first laser emitter to begin a process of generating and reflecting a laser beam from one laser emitter to another laser emitter in the plurality. A method of practicing the invention is also disclosed.

  12. The Effect of Addition of an EPS Degrading Enzyme with and without Detergent to 2% Chlorhexidine on Disruption of Enterococcus faecalis Biofilm: A Confocal Laser Scanning Microscopic Study

    PubMed Central

    Nagendrababu, Venkateshbabu; John, Aby; Deivanayagam, Kandaswamy

    2015-01-01

    Background Enterococcus faecalis is one of the most commonly occurring organisms retrieved from root canal treated teeth that show refractory apical periodontitis. Though it is well known that the ability of E. faecalis to form a matrix-encased biofilm contributes to its pathogenicity, the role of extracellular dextran and DNA in biofilm formation and its effect on the susceptibility of the biofilm to chlorhexidine remains poorly understood. It was hypothesized that the addition of an Extracellular Polymeric Substance (EPS) degrading enzyme along with a detergent to chlorhexidine may increase the susceptibility of the E. faecalis biofilm. Aim To evaluate the sensitivity of Enterococcus faecalis biofilms treated with DNase enzyme and their susceptibility to 2% chlorhexidine used alone or in conjunction with a detergent in a dentin disinfection model and examine under confocal laser scanning microscopy (CLSM). Materials and Methods Semi cylindrical shaped dentin specimens were infected with E. faecalis and incubated for 24 hours. Following incubation, the infected dentin specimens were exposed for 3 minutes to the four disinfecting solutions and grouped accordingly. {Group I- Sterile saline, Group II- 2% Chlorhexidine (CHX), Group III– Dnase1 Enzyme + 2% CHX, Group IV- DNase1 Enzyme + 2% CHX & Tween 80. Bacterial viability was then assessed by staining the specimens and examining under CLSM to analyse the proportion of dead and live bacteria within the dentinal tubules. Results The Groups II, III and IV showed statistically significant (p<0.05) percentage of dead bacteria compared to the control (Group I). However there was no significant difference in the killing effectiveness within the experimental groups (II-IV) at (p<0.05). Conclusion EPS degrading enzyme (DNase I) disrupts the biofilm and increases the susceptibility of E.faecalis when exposed to 2% Chlorhexidine and the use of a surfactant with this combination significantly contributes to improving the

  13. A first approach to the detection and equalization of distorted latent fingerprints and microtraces on non-planar surfaces with confocal laser microscopy

    NASA Astrophysics Data System (ADS)

    Kirst, Stefan; Clausing, Eric; Dittmann, Jana; Vielhauer, Claus

    2012-10-01

    Fingerprints and microtraces play an important role as evidence within the field of criminalistics. Their conservative acquisition processes, are established, but are altering and impurifying the traces often. In case of microtraces even the integrity of the trace complex is affected. Using contactless methods, the acquisition process becomes non-invasiv and repeatable, but might be distorting on the other hand, when non-planar substrates are in use. Detecting and dealing with distortion in contactless aquired scans of non-planar surfaces is a novel field of research. Nowadays highly distorted fingerprints can only be used, if the substrate can be manually distorted by destroying or deforming it. In this paper we suggest methods for detection and equalization of distortion for use in combination of types of traces. Therefore we define different types of distortion in fingerprints and microtraces. A standardization of types is necessary to develop different solution for equalization. For usage within the field of forensics, each method is evaluated via proper error rates and adaptively used to acquire fingerprints and microtraces. Using our techniques, we are able to detect distortion and equalize fingerprints to support the investigators work. In case of microtraces the presented methods can even be used to equalize mircotraces themselves for better determination of their scale and topology. For all scans the confocal 3D laser microscope "Keyence VK-X110" is used to gather color-, intensity- and topography information in 22 different measurement conditions within 6 different samples consisting of a total of 880 scans. Despite our achievements in the field of distortion detection and equalization there are still challenges, like the non-isometric projection, that need to be focused on. Also, the presented equalization methods may not completely remove any kind of distortion, such as added by deformation. Therefore we suggest and discuss future work for improving the

  14. Early detection of acute graft-versus-host disease by wireless capsule endoscopy and probe-based confocal laser endomicroscopy: results of a pilot study

    PubMed Central

    Laurent, Valerie; Malard, Florent; Le Rhun, Marc; Chevallier, Patrice; Guillaume, Thierry; Mosnier, Jean-François; Galmiche, Jean-Paul; Mohty, Mohamad

    2014-01-01

    Objective Acute gastrointestinal graft-versus-host disease (GI-GVHD) is usually diagnosed using endoscopic examinations and biopsies for conventional histology. The aim of this pilot study was to determine whether mini-invasive techniques such as probe-based confocal laser endomicroscopy (pCLE) combined with wireless capsule endoscopy (WCE) could detect early lesions of GI-GVHD prior to symptoms. Design Fifteen patients undergoing allogeneic haematopoietic stem cell transplantation (allo-HSCT) were prospectively examined with a small bowel WCE, duodenal and colorectal pCLE, and standard biopsies. Per study protocol, all these examinations were scheduled between day 21 and day 28 after allo-HSCT, independently of the presence or absence of digestive symptoms. Results During follow up, eight patients developed acute GI-GVHD. Sensitivity of WCE, pCLE, and histology were 50, 87.5, and 50%, respectively. Specificity of WCE, pCLE, and histology were 80, 71.5, and 80%, respectively. We showed a positive correlation between the Glücksberg scoring system and WCE (rho = 0.543, p = 0.036) and pCLE (rho = 0.727, p = 0.002) but not with standard histology (rho = 0.481, p = 0.069). Conclusions The results from this pilot study suggest that novel methods such as pCLE and WCE could be part of a mini-invasive algorithm for early detection of GI-GVHD. PMID:25360304

  15. Morphologic evaluation of meibomian glands in chronic graft-versus-host disease using in vivo laser confocal microscopy

    PubMed Central

    Ban, Yumiko; Ibrahim, Osama M.A.; Tatematsu, Yukako; Kamoi, Mizuka; Uchino, Miki; Yaguchi, Saori; Dogru, Murat; Tsubota, Kazuo

    2011-01-01

    Purpose To evaluate the morphological changes of the meibomian glands (MGs) using in vivo laser confocal microscopy (CM) in dry eye (DE) patients with chronic graft-versus-host disease (cGVHD). Methods Seventeen eyes from 9 patients with a diagnosis of DE associated with cGVHD (DE/cGVHD group; 6 males, 3 females; median 50.5 years) and 16 eyes of 8 hematopoietic stem cell transplantation (HSCT) recipients without DE (non-DE/non-cGVHD group; 5 males, 3 females; median 47.0 years) were enrolled. CM was used to investigate the MG and MG acinar unit density (MGAUD), MG acinar longest diameter (MGALD), MG acinar shortest diameter (MGASD), and the fibrosis grade. Clinical findings of the lid margin were obtained. Tear dynamics, ocular surface vital staining, meibography, and MG expressibility were also examined. Data were compared between the 2 groups using the unpaired t and Mann–Whitney tests. Results The mean MGAUD value was significantly lower in the DE/cGVHD group than in the non-DE/non-cGVHD group (p=0.01, 57.8±38.3 glands/mm2, 88.8±26.6 glands/mm2, respectively), and the mean MGALD and MGASD were significantly shorter in the DE/cGVHD group than in the non-DE/non-cGVHD group (p=0.0018, 37.3±24.4 μm and 60.4±11.8 μm, p=0.0106, 17.7±11.8 μm and 26.6±6.03 μm, respectively). The mean fibrosis grade was significantly higher in the DE/cGVHD group than the non-DE/non-cGVHD group (p<0.0001, 1.39±0.71 grade, 0.06±0.25 grade, respectively). Clinical findings in the lid margin, tear dynamics, and ocular surface findings were significantly worse in the DE/cGVHD group than in the non-DE/non-cGVHD group. Conclusions CM clearly depicted the morphological changes of the MG in the DE/cGVHD group, and revealed the severity of the meibomian gland dysfunction. Patients with severe DE after HSCT showed atrophic MG and excessive fibrosis. PMID:22025888

  16. Development of Useful Recombinant Promoter and Its Expression Analysis in Different Plant Cells Using Confocal Laser Scanning Microscopy

    PubMed Central

    Kumar, Deepak; Sahoo, Dipak K.; Maiti, Indu B.; Dey, Nrisingha

    2011-01-01

    Background Designing functionally efficient recombinant promoters having reduced sequence homology and enhanced promoter activity will be an important step toward successful stacking or pyramiding of genes in a plant cell for developing transgenic plants expressing desired traits(s). Also basic knowledge regarding plant cell specific expression of a transgene under control of a promoter is crucial to assess the promoter's efficacy. Methodology/Principal Findings We have constructed a set of 10 recombinant promoters incorporating different up-stream activation sequences (UAS) of Mirabilis mosaic virus sub-genomic transcript (MS8, -306 to +27) and TATA containing core domains of Figwort mosaic virus sub-genomic transcript promoter (FS3, −271 to +31). Efficacies of recombinant promoters coupled to GUS and GFP reporter genes were tested in tobacco protoplasts. Among these, a 369-bp long hybrid sub-genomic transcript promoter (MSgt-FSgt) showed the highest activity in both transient and transgenic systems. In a transient system, MSgt-FSgt was 10.31, 2.86 and 2.18 times more active compared to the CaMV35S, MS8 and FS3 promoters, respectively. In transgenic tobacco (Nicotiana tabaccum, var. Samsun NN) and Arabidopsis plants, the MSgt-FSgt hybrid promoter showed 14.22 and 7.16 times stronger activity compared to CaMV35S promoter respectively. The correlation between GUS activity and uidA-mRNA levels in transgenic tobacco plants were identified by qRT-PCR. Both CaMV35S and MSgt-FSgt promoters caused gene silencing but the degree of silencing are less in the case of the MSgt-FSgt promoter compared to CaMV35S. Quantification of GUS activity in individual plant cells driven by the MSgt-FSgt and the CaMV35S promoter were estimated using confocal laser scanning microscopy and compared. Conclusion and Significance We propose strong recombinant promoter MSgt-FSgt, developed in this study, could be very useful for high-level constitutive expression of transgenes in a wide variety

  17. Spiral ganglion neuron quantification in the guinea pig cochlea using Confocal Laser Scanning Microscopy compared to embedding methods.

    PubMed

    Wrzeszcz, Antonina; Reuter, Günter; Nolte, Ingo; Lenarz, Thomas; Scheper, Verena

    2013-12-01

    Neuron counting in the cochlea is a crucial but time-consuming operation for which various methods have been developed. To improve simplicity and efficiency, we tested an imaging method of the cochlea, and based on Confocal Laser Scanning Microscopy (CLSM), we visualised Rosenthal's Canal and quantified the spiral ganglion neurons (SGN) within. Cochleae of 8 normal hearing guinea pigs and one implanted with a silicone filament were fixed in paraformaldehyde (PFA), decalcified, dehydrated and cleared in Spalteholz solution. Using the tissue's autofluorescence, CLSM was performed at 100 fold magnification generating z-series stacks of about 20 slices of the modiolus. In 5 midmodiolar slices per cochlea the perimeters of the Rosenthal's Canal were surveyed, representative neuron diameters were measured and the neurons first counted manually and then software-assisted. For comparison, 8 normal hearing guinea pig cochleae were embedded in paraffin and examined similarly. The CLSM method has the advantage that the cochleae remain intact as an organ and keep their geometrical structure. Z-stack creation is nearly fully-automatic and frequently repeatable with various objectives and step sizes and without visible bleaching. The tissue shows minimal or no shrinking artefacts and damage typical of embedding and sectioning. As a result, the cells in the cleared cochleae reach an average diameter of 21 μm and a density of about 18 cells/10,000 μm(2) with no significant difference between the manual and the automatical counts. Subsequently we compared the CLSM data with those generated using the established method of paraffin slides, where the SGN reached a mean density of 9.5 cells/10,000 μm(2) and a mean soma diameter of 13.6 μm. We were able to prove that the semi-automatic CLSM method is a simple and effective technique for auditory neuron count. It provides a high grade of tissue preservation and the automatic stack-generation as well as the counter software reduces

  18. Biofilm forming capacity of Enterococcus faecalis on Gutta-percha points treated with four disinfectants using confocal scanning laser microscope: An in vitro study

    PubMed Central

    Ravi Chandra, Polavarapu Venkata; Kumar, Vemisetty Hari; Reddy, Surakanti Jayaprada; Kiran, Dandolu Ram; Krishna, Muppala Nagendra; Kumar, Golla Vinay

    2015-01-01

    Background: The aim of this study was to evaluate and compare the in vitro biofilm forming capacity of Enterococcus faecalis on Gutta-percha points disinfected with four disinfectants. Materials and Methods: A total of 50 Gutta-percha points used in this study were divided into four test groups based on disinfectant (5.25% sodium hypochlorite, 2% chlorhexidine gluconate, 20% neem, 13% benzalkonium chloride [BAK]), and one control group. The Gutta-percha points were initially treated with corresponding disinfectants followed by anaerobic incubation in Brain Heart Infusion broth suspended with human serum and E. faecalis strain for 14 days. After incubation, these Gutta-percha points were stained with Acridine Orange (Sigma – Aldrich Co., St. Louis, MO, USA) and 0.5 mm thick cross section samples were prepared. The biofilm thickness of E. faecalis was analyzed quantitatively using a confocal scanning laser microscope. Results statistically analyzed using analysis of variance. P < 0.05 was considered to be significant. Results: Confocal scanning laser microscope showed reduced amount of E. faecalis biofilm on Gutta-percha points treated with BAK and sodium hypochlorite. Post-hoc (least square differences) test revealed that there is no statistically significant difference between BAK and sodium hypochlorite groups (P > 0.05). Conclusion: This study illustrates that the Gutta-percha points disinfected with sodium hypochlorite and BAK showed minimal biofilm growth on its surface. PMID:26288622

  19. Confocal scanning laser microscopy with complementary 3D image analysis allows quantitative studies of functional state of ionoregulatory cells in the Nile tilapia (Oreochromis niloticus) following salinity challenge.

    PubMed

    Fridman, Sophie; Rana, Krishen J; Bron, James E

    2013-04-01

    The development of a novel three-dimensional image analysis technique of stacks generated by confocal laser scanning microscopy is described allowing visualization of mitochondria-rich cells (MRCs) in the seawater-adapted Nile tilapia in relation to their spatial location. This method permits the assessment and classification of both active and nonactive MRCs based on the distance of the top of the immunopositive cell from the epithelial surface. In addition, this technique offers the potential for informative and quantitative studies, for example, densitometric and morphometric measurements based on MRC functional state. Confocal scanning laser microscopy used with triple staining whole-mount immunohistochemistry was used to detect integumental MRCs in the yolk-sac larvae tail of the Nile tilapia following transfer from freshwater to elevated salinities, that is, 12.5 and 20 ppt. Mean active MRC volume was always significantly larger and displayed a greater staining intensity (GLM; P<0.05) than nonactive MRCs. Following transfer, the percentage of active MRCs was seen to increase as did MRC volume (GLM; P<0.05). PMID:23390074

  20. Cytogenetic Characterization of the TM4 Mouse Sertoli Cell Line. II. Chromosome Microdissection, FISH, Scanning Electron Microscopy, and Confocal Laser Scanning Microscopy.

    PubMed

    Schmid, Michael; Guttenbach, Martina; Steinlein, Claus; Wanner, Gerhard; Houben, Andreas

    2015-01-01

    The chromosomes and interphase cell nuclei of the permanent mouse Sertoli cell line TM4 were examined by chromosome microdissection, FISH, scanning electron microscopy, and confocal laser scanning microscopy. The already known marker chromosomes m1-m5 were confirmed, and 2 new large marker chromosomes m6 and m7 were characterized. The minute heterochromatic marker chromosomes m4 and m5 were microdissected and their DNA amplified by DOP-PCR. FISH of this DNA probe on TM4 metaphase chromosomes demonstrated that the m4 and m5 marker chromosomes have derived from the centromeric regions of normal telocentric mouse chromosomes. Ectopic pairing of the m4 and m5 marker chromosomes with the centromeric region of any of the other chromosomes (centromeric associations) was apparent in ∼60% of the metaphases. Scanning electron microscopy revealed DNA-protein bridges connecting the centromeric regions of normal chromosomes and the associated m4 and m5 marker chromosomes. Interphase cell nuclei of TM4 Sertoli cells did not exhibit the characteristic morphology of Sertoli cells in the testes of adult mice as shown by fluorescence microscopy and confocal laser scanning microscopy. PMID:26900862

  1. High-temperature laser-scanning confocal microscopy as a tool to study the interface instability during unsteady-state solidification of low-carbon steel.

    PubMed

    Niknafs, S; Phelan, D; Dippenaar, R

    2013-01-01

    Solidification microstructure is a defining link between production techniques and the mechanical properties of metals and in particular steel. Due to the difficulty of conducting solidification studies at high temperature, knowledge of the development of solidification microstructure in steel is scarce. In this study, a laser-scanning confocal microscopy (LSCM) has been used to observe in situ and in real-time the planar to cellular to dendritic transition of the progressing solid/liquid interface in low carbon steel. Because the in situ observations in the laser-scanning confocal microscopy are restricted to the surface, the effect of sample thickness on surface observations was determined. Moreover, the effect of cooling rate and alloy composition on the planar to cellular interface transition was investigated. In the low-alloyed, low-carbon steel studied, the cooling rate does not seem to have an effect on the spacing of the cellular microstructure. However, in the presence of copper and manganese, the cell spacing decreased at higher cooling rates. Higher concentrations of copper in steel resulted on an increased cell spacing at the same cooling rates. PMID:23170969

  2. Attachment of Escherichia coli O157:H7 to the Surfaces and Internal Structures of Apples as Detected by Confocal Scanning Laser Microscopy

    PubMed Central

    Burnett, Scott L.; Chen, Jinru; Beuchat, Larry R.

    2000-01-01

    Confocal scanning laser microscopy (CSLM) was used to demonstrate the attachment of Escherichia coli O157:H7 transformed with a plasmid encoding for green fluorescent protein (GFP) to the surface and within the internal structures of nonwaxed Red Delicious cv. apples. Apples at 2 or 25°C were inoculated with an E. coli O157:H7 cell suspension at 2 or 25°C. The effect of a negative temperature differential (cold inoculum, warm apple), a positive differential (warm inoculum, cold apple), and no differential (warm inoculum, warm apple), in combination with a pressure differential (atmospheric versus 10,130 Pa), on the attachment and infiltration of cells was determined. CSLM stereo images of external surfaces of apples subjected to all combinations of test parameters showed preferential cellular attachment to discontinuities in the waxy cuticle on the surface and to damaged tissue surrounding puncture wounds, where the pathogen was observed at depths up to 70 μm below the skin surface. Attachment to lenticels was sporadic but was occasionally observed at depths of up to 40 μm. Infiltration through the floral tube and attachment to seeds, cartilaginous pericarp, and internal trichomes were observed in all apples examined, regardless of temperature differential during inoculation. The pressure differential had no effect on infiltration or attachment of E. coli O157:H7. Image analysis to count cells at various depths within tissues was used to quantitatively compare the extent of infiltration into various apple structures as well as the effects of the temperature differential. Puncture wounds harbored greater numbers of the pathogen at greater depths than did other sites examined. Attachment or infiltration of cells was greater on the intact skin and in lenticels, russet areas, and the floral tube of apples inoculated under a negative temperature differential compared to those inoculated under no temperature differential. The results suggest that E. coli O157:H7

  3. High power semiconductor laser beam combining technology and its applications

    NASA Astrophysics Data System (ADS)

    Wang, Lijun; Tong, Cunzhu; Peng, Hangyu; Zhang, Jun

    2013-05-01

    With the rapid development of laser applications, single elements of diode lasers are not able to meet the increasing requirements on power and beam quality in the material processing and defense filed, whether are used as pumping sources or directly laser sources. The coupling source with high power and high beam quality, multiplexed by many single elements, has been proven to be a promising technical solution. In this paper, the authors review the development tendency of efficiency, power, and lifetime of laser elements firstly, and then introduce the progress of laser beam combining technology. The authors also present their recent progress on the high power diode laser sources developed by beam combining technology, including the 2600W beam combining direct laser source, 1000W fiber coupled semiconductor lasers and the 1000W continuous wave (CW) semiconductor laser sources with beam quality of 12.5×14[mm. mrad]2.

  4. Confocal microscopy and exfoliative cytology

    PubMed Central

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

    2013-01-01

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

  5. Coherent beam combiner for a high power laser

    DOEpatents

    Dane, C. Brent; Hackel, Lloyd A.

    2002-01-01

    A phase conjugate laser mirror employing Brillouin-enhanced four wave mixing allows multiple independent laser apertures to be phase locked producing an array of diffraction-limited beams with no piston phase errors. The beam combiner has application in laser and optical systems requiring high average power, high pulse energy, and low beam divergence. A broad range of applications exist in laser systems for industrial processing, especially in the field of metal surface treatment and laser shot peening.

  6. Skin healing and collagen changes of rats after fractional erbium:yttrium aluminum garnet laser: observation by reflectance confocal microscopy with confirmed histological evidence.

    PubMed

    Yang, Jing; Wang, Sha; Dong, Liyun; An, Xiangjie; Li, Yan; Li, Jun; Tu, Yating; Tao, Juan

    2016-08-01

    The fractional erbium:yttrium aluminum garnet (Er:YAG) laser is widely applied. Microstructural changes after laser treatment have been observed with histopathology. Epidermal and dermal microstructures have also been analyzed using reflectance confocal microscopy (RCM). However, no studies have compared these two types of microstructural changes in the same subject at multiple time points after irradiation, and it is unclear if these two types of changes are consistent. We use RCM to observe the effect of different laser energies on skin healing and collagen changes in the skin of Sprague-Dawley rats that had been irradiated by fractional Er:YAG lasering at different energies. RCM was used to observe skin healing and detect collagen changes at different time points. Collagen changes were observed using hematoxylin and eosin (H&E) staining and quantitatively analyzed by western blot. RCM showed that, irrespective of laser energy, microscopic treatment zones (MTZs) were larger at 1 day after irradiation. The MTZs then reduced in size from 3 to 7 days after irradiation. The higher the energy, the larger the MTZ area. The amount of collagen also increased with time from 1 day to 8 weeks. However, the increase in the collagen amount on both RCM and H&E staining was not influenced by the laser energy. Western blotting confirmed that the amount of type I and type III collagens increased over time, but there were no significant differences between the different energy groups (p > 0.05). In conclusion, RCM is a reliable technique for observing and evaluating skin healing and collagen expression after laser irradiation. PMID:27272747

  7. CONFOCAL MICROSCOPY SYSTEM PERFORMANCE: SPECTROSCOPY

    EPA Science Inventory

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

  8. EVALUATION OF CONFOCAL MICROSCOPY SYSTEM PERFORMANCE

    EPA Science Inventory

    BACKGROUND. The confocal laser scanning microscope (CLSM) has enormous potential in many biological fields. Currently there is a subjective nature in the assessment of a confocal microscope's performance by primarily evaluating the system with a specific test slide provided by ea...

  9. Europium Uptake and Partitioning in Oat (Avena sativa) Roots as studied By Laser-Induced Fluorescence Spectroscopy and Confocal Microscopy Profiling Technique

    SciTech Connect

    Fellows, Robert J.; Wang, Zheming; Ainsworth, Calvin C.

    2003-11-15

    The uptake of Eu3+ by elongating oat plant roots was studied by fluorescence spectroscopy, fluorescence lifetime measurement, as well as laser excitation time-resolved confocal fluorescence profiling technique. The results of this work indicated that the initial uptake of Eu(III) by oat root was most evident within the apical meristem of the root just proximal to the root cap. Distribution of assimilated Eu(III) within the roots differentiation and elongation zone was non-uniform. Higher concentrations were observed within the vascular cylinder, specifically in the phloem and developing xylem parenchyma. Elevated levels of the metal were also observed in the root hairs of the mature root. The concentration of assimilated Eu3+ dropped sharply from the apical meristem to the differentiation and elongation zone and then gradually decreased as the distance from the root cap increased. Fluorescence spectroscopic characteristics of the assimilated Eu3+ suggested that the Eu3+ exists a s inner-sphere mononuclear complexes inside the root. This work has also demonstrated the effectiveness of a time-resolved Eu3+ fluorescence spectroscopy and confocal fluorescence profiling techniques for the in vivo, real-time study of metal[Eu3+] accumulation by a functioning intact plant root. This approach can prove valuable for basic and applied studies in plant nutrition and environmental uptake of actinide radionuclides.

  10. Time-variant analysis of organelle and vesicle movement during phagocytosis in Paramecium primaurelia by means of fluorescence confocal laser scanning microscopy.

    PubMed

    Ramoino, P; Beltrame, F; Diaspro, A; Fato, M

    1996-12-01

    Vital fluorescent dyes (FITC-albumin, Texas Red-albumin, and acridine orange) were used together with a confocal laser scanning optical microscope (CLSM) to display and analyze formation, movement, and fusion of vesicles during the phagocytosis of Paramecium primaurelia, in the x-y-z-t space. By immobilizing living cells pulsed with a food vacuole marker at successive times after chasing in unlabeled medium, the intracellular movement of food vacuoles from their formation at the cytostome to their egestion at the cytoproct was visualized, and food vacuoles were selected in a specific digestion stage. Small pinocytic vesicles are shown to evaginate from the vacuoles and move in the cytoplasm. These vesicles are transported toward the cytopharynx where they enlarge the membrane of the nascent food vacuoles or fuse with stage II food vacuoles, when the vacuoles of stage II increase their size, changing from an acidic to an alkaline status. A multimodal analysis of confocal fluorescence images and the false-color technique were used to visualize vesicle movement vs. time. Starting from three images of the same cell at succeeding time points, a composite image was generated by associating with each originally acquired image a different color corresponding to each sampling point in time. The composite image shows that vesicles move away from the food vacuole in a scattered manner exhibiting changes in direction. PMID:8989767

  11. The Superficial Stromal Scar Formation Mechanism in Keratoconus: A Study Using Laser Scanning In Vivo Confocal Microscopy

    PubMed Central

    Song, Peng; Wang, Shuting; Zhang, Peicheng; Sui, Wenjie; Zhang, Yangyang; Liu, Ting; Gao, Hua

    2016-01-01

    To investigate the mechanism of superficial stromal scarring in advanced keratoconus using confocal microscopy, the keratocyte density, distribution, micromorphology of corneal stroma, and SNP in three groups were observed. Eight corneal buttons of advanced keratoconus were examined by immunohistochemistry. The keratocyte densities in the sub-Bowman's stroma, anterior stroma, and posterior stroma and the mean SNP density were significantly different among the three groups. In the mild-to-moderate keratoconus group, activated keratocyte nuclei and comparatively highly reflective ECM were seen in the sub-Bowman's stroma, while fibrotic structures with comparatively high reflection were visible in the anterior stroma in advanced keratoconus. The alternating dark and light bands in the anterior stroma of the mild-to-moderate keratoconus group showed great variability in width and direction. The wide bands were localized mostly in the posterior stroma that corresponded to the Vogt striae in keratoconus and involved the anterior stroma only in advanced keratoconus. Histopathologically, high immunogenicity of α-SMA, vimentin, and FAP was expressed in the region of superficial stromal scarring. In vivo confocal microscopy revealed microstructural changes in the keratoconic cone. The activation of superficial keratocytes and abnormal remodeling of ECM may both play a key role in the superficial stromal scar formation in advanced keratoconus. PMID:26885515

  12. Histopathological confirmation of similar intramucosal distribution of fluorescein in both intravenous administration and local mucosal application for probe-based confocal laser endomicroscopy of the normal stomach

    PubMed Central

    Nonaka, Kouichi; Ohata, Ken; Ban, Shinichi; Ichihara, Shin; Takasugi, Rumi; Minato, Yohei; Tashima, Tomoaki; Matsuyama, Yasushi; Takita, Maiko; Matsuhashi, Nobuyuki; Neumann, Helmut

    2015-01-01

    Probe-based confocal laser endomicroscopy (pCLE) is capable of acquiring in vivo magnified cross-section images of the gastric mucosa. Intravenous injection of fluorescein sodium is used for confocal imaging. However, it is still under debate if local administration of the dye to the mucosa is also effective for confocal imaging as it is not yet clear if topical application also reveals the intramucosal distribution of fluorescein. The objective of this study was to evaluate the intramucosal distribution of fluorescein sodium after topical application and to compare the distribution to the conventional intravenous injection used for confocal imaging. pCLE of the stomach uninfected with Helicobacter pylori was performed in a healthy male employing intravenous administration and local mucosal application of fluorescein. The mucosa of the lower gastric body was biopsied 1 min and 5 min after intravenous administration or local mucosal application of fluorescein, and the distribution of fluorescein in the biopsy samples was examined histologically. Green fluorescence was already observed in the cytoplasm of fundic glandular cells in the biopsied deep mucosa 1 min after local mucosal application of fluorescein. It was also observed in the foveolar lumen and inter-foveolar lamina propria, although it was noted at only a few sites. In the tissue biopsied 5 min after the local mucosal application of fluorescein, green fluorescence was more frequently noted in the cytoplasm of fundic glandular cells than in that 1 min after the local mucosal application of fluorescein, although obvious green fluorescence was not identified in the foveolar lumen or inter-foveolar lamina propria. The distribution of intravenously administered fluorescein in the cytoplasm of fundic glandular cells was also clearly observed similarly to that after local mucosal application of fluorescein. Green fluorescence in more cells was observed in many cells 5 min after intravenous administration compared

  13. In-situ Crystallization of Highly Volatile Commercial Mold Flux Using an Isolated Observation System in the Confocal Laser Scanning Microscope

    NASA Astrophysics Data System (ADS)

    Park, Jun-Yong; Ryu, Jae Wook; Sohn, Il

    2014-08-01

    The in situ crystallization behavior of highly volatile commercial mold fluxes for medium carbon steels was investigated using the confocal laser scanning microscope (CLSM) equipped with an optimized isolated observation system. The highly volatile compounds of the mold flux were suppressed during heating allowing direct observation in the CLSM. Cooling rates of 25, 50, 100, 400, and 800 K/min were incorporated and continuous cooling transformation (CCT) diagrams of 4 different commercial mold fluxes for medium carbon steels were developed. Identification of the crystalline phase was conducted with XRD and SEM-EDS analysis. A cuspidine crystalline was observed in all samples at various cooling rates. With higher basicity, CaF2, and NaF, the crystallization of the fluxes was enhanced according to the CCT diagram. As the slag structure becomes depolymerized, the diffusion rate of the cathodic ions seems to increase.

  14. Atomic force microscopy and laser confocal scanning microscopy analysis of callose fibers developed from protoplasts of embryogenic cells of a conifer.

    PubMed

    Fukumoto, Takeshi; Hayashi, Noriko; Sasamoto, Hamako

    2005-12-01

    Efficiency of novel fiber formation was much improved in protoplast culture of embryogenic cells (ECs) of a conifer, Larix leptolepis (Sieb. et Zucc.) Gord., by pre-culturing ECs in a medium containing a high concentration of glutamine (13.7 mM). The fibrillar substructures of large and elongated fibers of protoplasts isolated from Larix ECs were investigated by laser confocal scanning microscopy (LCSM) after Aniline Blue staining and atomic force microscopy (AFM) using a micromanipulator without any pre-treatment. Fibers were composed of bundles of fibrils and subfibrils, whose diameters were defined as 0.7 and 0.17 mum, respectively, by image analysis after LCSM and AFM. These fibers were proven to be composed of callose by using specific degrading enzymes for beta-1,4-glucan and beta-1,3-glucan. PMID:16034590

  15. In vivo molecular imaging of somatostatin receptors in pancreatic islet cells and neuroendocrine tumors by miniaturized confocal laser-scanning fluorescence microscopy.

    PubMed

    Fottner, C; Mettler, E; Goetz, M; Schirrmacher, E; Anlauf, M; Strand, D; Schirrmacher, R; Klöppel, G; Delaney, P; Schreckenberger, M; Galle, P R; Neurath, M F; Kiesslich, R; Weber, M M

    2010-05-01

    The aim of the study was to evaluate real time in vivo molecular imaging of somatostatin receptors (sstrs) using a handheld miniaturized confocal laser scan microscope (CLM) in conjunction with fluorescein-labeled octreotate (OcF) in healthy mice and murine models of neuroendocrine tumors. For CLM a small rigid probe (diameter 7 mm) with an integrated single line laser (488 nm) was used (optical slice thickness 7 mum; lateral resolution 0.7 mum). OcF was synthesized via Fmoc solid-phase peptide synthesis and purified by HPLC showing high-affinity binding to the sstr2 (IC(50) 6.2 nmol). For in vitro evaluation, rat and human pancreatic cancer cells were used and characterized with respect to its sstr subtype expression and functional properties. For in vivo confocal imaging, healthy mouse pancreatic islet and renal tubular cells as well as immunoincompetent nude mice harboring sstr-expressing tumors were evaluated. Incubation of sstr-positive cells with OcF showed a specific time- and dose-dependent staining of sstr-positive cells. CLM showed rapid internalization and homogenous cytoplasmatic distribution. After systemic application to mice (n = 8), specific time-dependent internalization and cytoplasmatic distribution into pancreatic islet cells and tubular cells of the renal cortex was recorded. After injection in tumor-harboring nude mice (n = 8), sstr-positive cells selectively displayed a cell surface and cytoplasmatic staining. CLM-targeted biopsies detected sstr-positive tumor cells with a sensitivity of 87.5% and a specificity of 100% as correlated with ex vivo immunohistochemistry. CLM with OcF permits real-time molecular, functional, and morphological imaging of sstr-expressing cell structures, allowing the specific visualization of pancreatic islet cells and neuroendocrine tumors in vivo. PMID:20233796

  16. Confocal Raman microscopy of protein adsorbed in chromatographic particles.

    PubMed

    Xiao, Yuewu; Stone, Thomas; Bell, David; Gillespie, Christopher; Portoles, Marta

    2012-09-01

    Confocal Raman microscopy is a nondestructive analytical technique that combines the chemical information from vibrational spectroscopy with the spatial resolution of confocal microscopy. It was applied, for the first time, to measure conformation and distribution of protein adsorbed in wetted chromatographic particles. Monoclonal antibody was loaded into the Fractogel EMD SO(3) (M) cation exchanger at 2 mS/cm or 10 mS/cm. Amide I and III frequencies in the Raman spectrum of the adsorbed protein suggest that there are no detectable changes of the original β-sheet conformation in the chromatographic particles. Protein depth profile measurements indicate that, when the conductivity is increased from 2 mS/cm to 10 mS/cm, there is a change in mass transport mechanism for protein adsorption, from the shrinking-core model to the homogeneous-diffusion model. In this study, the use of confocal Raman microscopy to measure protein distribution in chromatographic particles fundamentally agrees with previous confocal laser scanning microscopic investigations, but confocal Raman spectroscopy enjoys additional advantages: use of unlabeled protein to eliminate fluorescent labeling, ability for characterization of protein secondary structure, and ability for spectral normalization to provide a nondestructive experimental approach to correct light attenuation effects caused by refractive index (RI) mismatching in semiopaque chromatographic particles. PMID:22803776

  17. Expression of keratin 14 in the basal cells of the lingual epithelium of mice during the morphogenesis of filiform papillae: visualization by fluorescent immunostaining and confocal laser-scanning microscopy in the transmission mode.

    PubMed

    Iwasaki, Shin-Ichi; Aoyagi, Hidekazu

    2007-07-01

    We examined the expression of keratin 14 (K14) on the lingual epithelium by immunofluorescent staining while monitoring morphological changes in the filiform papillae of mice by confocal laser-scanning microscopy in the transmission mode of the same sections to define both the histology and the morphology of cells. It is difficult to visualize histological details of the fetal lingual epithelium of the mouse on semi-ultrathin sections by light microscopy after immunohistochemical staining because the histological structures in such sections cannot be distinguished by standard counterstaining. To solve this problem and to visualize the immunoreactivity specific for K14, we analyzed the results of immunofluorescent staining of semi-ultrathin sections in combination with an examination of the corresponding images by laser-scanning microscopy in the transmission mode after staining of specimens with toluidine blue. No immunoreactivity specific for K14 was detected on the lingual epithelium of fetuses on embryonic day 15 (E15), but immunoreactivity was distinct at all postnatal stages from postnatal day 0 (P0) to P21. PMID:17660983

  18. A microscopic setup for combined, and time-coordinated electrophysiological and confocal fluorescence microscopic experiments on neurons in living brain slices

    NASA Astrophysics Data System (ADS)

    Helm, P. J.

    1996-02-01

    In this paper, a microscopic system for cell physiological research is presented. The setup which is to a large extent based on commercially available products was designed to establish a platform for time-coordinated electrophysiological and fluorescence optical compound experiments on living neurons in brain slices. Instruments for infrared differential interference contrast video microscopy (IRDICM), confocal scanning laser microscopy (CSLM), and for patch clamp studies have been assembled into one unit. Using the IRDICM equipment, a neuron can be patched somatically and dendritically. Loading the neuron with a Ca2+ indicating dye substance can be examined epifluorescence optically using the Hg lamp or Xe lamp of the microscope. A stimulus initiating the propagation of an action potential through a dendrite can be synchronized to the electronic control unit of the CSLM, and changes in the concentration of Ca2+ in the dendrite can be recorded in a time-coordinated way. The setup has been used successfully in order to study in vitro the dynamics of intracellular Ca2+ in the dendritic system of living neurons in brain slices.

  19. Assessment of a superficial chemical peel combined with a multimodal, hydroquinone-free skin brightener using in vivo reflectance confocal microscopy.

    PubMed

    Goberdhan, Lisa T; Mehta, Rahul C; Aguilar, Caroline; Makino, Elizabeth T; Colvan, Lora

    2013-03-01

    The combination of in-office procedures such as chemical peels with topical maintenance therapies has been shown to provide greater efficacy than either treatment by itself in the management of melasma. A series of 3 case studies were conducted to evaluate the efficacy and tolerability of one superficial chemical peel (containing a proprietary blend of resorcinol, lactic acid, salicylic acid, and retinol) combined with a topical multimodal, hydroquinone-free skin brightener as postpeel maintenance therapy. Patients presented with moderate to severe facial hyperpigmentation. At baseline, subjects received the superficial chemical peel treatment followed by a standard postpeel skin care regimen (cleanser, moisturizer, and SPF 30+ sunscreen). Approximately 1 week after the peel procedure, subjects initiated twice-daily application of the skin brightener. Subjects were then evaluated for Global Improvement in Hyperpigmentation by the investigator for up to 7 weeks postpeel. Standardized digital photographs of the subjects facial skin and in vivo reflectance confocal microscopy (RCM) images were taken of a target hyperpigmented lesion at baseline and at follow-up. Standardized photography and in vivo RCM images at baseline and at postpeel show the improvements observed by the investigator. Results from these case studies suggest that the combination of a superficial chemical peel with topical maintenance and the multimodal skin brightener may provide an effective treatment approach for subjects with moderate to severe facial hyperpigmentation. PMID:23545932

  20. Micromixing visualization and quantification in a microscale multi-inlet vortex nanoprecipitation reactor using confocal-based reactive micro laser-induced fluorescence

    PubMed Central

    Shi, Yanxiang; Fox, Rodney O.; Olsen, Michael G.

    2014-01-01

    A technique for visualizing and quantifying reactive mixing for laminar and turbulent flow in a microscale chemical reactor using confocal-based microscopic laser induced fluorescence (confocal μ-LIF) was demonstrated in a microscale multi-inlet vortex nanoprecipitation reactor. Unlike passive scalar μ-LIF, the reactive μ-LIF technique is able to visualize and quantify micromixing effects. The confocal imaging results indicated that the flow in the reactor was laminar and steady for inlet Reynolds numbers of 10, 53, and 93. Mixing and reaction were incomplete at each of these Reynolds numbers. The results also suggested that although mixing by diffusion was enhanced near the midplane of the reactor at Rej = 53 and 93 due to very thin bands of acidic and basic fluid forming as the fluid spiraled towards the center of the reactor, near the top, and bottom walls of the reactor, the lower velocities due to fluid friction with the walls hindered the formation of these thin bands, and, thus, resulted in large regions of unmixed and unreacted fluid. At Rej = 240, the flow was turbulent and unsteady. The mixing and reaction processes were still found to be incomplete even at this highest Reynolds number. At the reactor midplane, the flow images at Rej = 240 showed unmixed base fluid near the center of the reactor, suggesting that just as in the Rej = 53 and 93 cases, lower velocities near the top and bottom walls of the reactor hinder the mixing and rection of the acidic and basic streams. Ensemble averages of line-scan profiles for the Rej = 240 were then calculated to provide statistical quantification of the microscale mixing in the reactor. These results further demonstrate that even at this highest Reynolds number investigated, mixing and reaction are incomplete. Visualization and quantification of micromixing using this reactive μ-LIF technique can prove useful in the validation of computational fluid dynamics models of micromixing within

  1. Atmospheric propagation and combining of high-power lasers.

    PubMed

    Nelson, W; Sprangle, P; Davis, C C

    2016-03-01

    In this paper, we analyze beam combining and atmospheric propagation of high-power lasers for directed-energy (DE) applications. The large linewidths inherent in high-power fiber and slab lasers cause random phase and intensity fluctuations that occur on subnanosecond time scales. Coherently combining these high-power lasers would involve instruments capable of precise phase control and operation at rates greater than ∼10  GHz. To the best of our knowledge, this technology does not currently exist. This presents a challenging problem when attempting to phase lock high-power lasers that is not encountered when phase locking low-power lasers, for example, at milliwatt power levels. Regardless, we demonstrate that even if instruments are developed that can precisely control the phase of high-power lasers, coherent combining is problematic for DE applications. The dephasing effects of atmospheric turbulence typically encountered in DE applications will degrade the coherent properties of the beam before it reaches the target. Through simulations, we find that coherent beam combining in moderate turbulence and over multikilometer propagation distances has little advantage over incoherent combining. Additionally, in cases of strong turbulence and multikilometer propagation ranges, we find nearly indistinguishable intensity profiles and virtually no difference in the energy on the target between coherently and incoherently combined laser beams. Consequently, we find that coherent beam combining at the transmitter plane is ineffective under typical atmospheric conditions. PMID:26974640

  2. Laser-jamming effectiveness analysis of combined-fiber lasers for airborne defense systems.

    PubMed

    Jie, Xu; Shanghong, Zhao; Rui, Hou; Shengbao, Zhan; Lei, Shi; Jili, Wu; Shaoqiang, Fang; Yongjun, Li

    2008-12-20

    The laser-jamming effectiveness of combined fiber lasers for airborne defense systems is analyzed in detail. Our preliminary experimental results are proof of the concept of getting a high-power laser through a beam combination technique. Based on combined fiber lasers, the jamming effectiveness of four-quadrant guidance and imaging guidance systems are evaluated. The simulation results have proved that for a four-quadrant guidance system, the tracking system takes only two seconds to complete tracking, and the new tracking target is the jamming laser; for the imaging guidance system, increasing the power of the jamming laser or the distance between the target and the jamming laser are both efficient ways to achieve a successful laser jamming. PMID:19104536

  3. Optics clustered to output unique solutions: a multi-laser facility for combined single molecule and ensemble microscopy.

    PubMed

    Clarke, David T; Botchway, Stanley W; Coles, Benjamin C; Needham, Sarah R; Roberts, Selene K; Rolfe, Daniel J; Tynan, Christopher J; Ward, Andrew D; Webb, Stephen E D; Yadav, Rahul; Zanetti-Domingues, Laura; Martin-Fernandez, Marisa L

    2011-09-01

    Optics clustered to output unique solutions (OCTOPUS) is a microscopy platform that combines single molecule and ensemble imaging methodologies. A novel aspect of OCTOPUS is its laser excitation system, which consists of a central core of interlocked continuous wave and pulsed laser sources, launched into optical fibres and linked via laser combiners. Fibres are plugged into wall-mounted patch panels that reach microscopy end-stations in adjacent rooms. This allows multiple tailor-made combinations of laser colours and time characteristics to be shared by different end-stations minimising the need for laser duplications. This setup brings significant benefits in terms of cost effectiveness, ease of operation, and user safety. The modular nature of OCTOPUS also facilitates the addition of new techniques as required, allowing the use of existing lasers in new microscopes while retaining the ability to run the established parts of the facility. To date, techniques interlinked are multi-photon/multicolour confocal fluorescence lifetime imaging for several modalities of fluorescence resonance energy transfer (FRET) and time-resolved anisotropy, total internal reflection fluorescence, single molecule imaging of single pair FRET, single molecule fluorescence polarisation, particle tracking, and optical tweezers. Here, we use a well-studied system, the epidermal growth factor receptor network, to illustrate how OCTOPUS can aid in the investigation of complex biological phenomena. PMID:21974592

  4. Optics clustered to output unique solutions: A multi-laser facility for combined single molecule and ensemble microscopy

    NASA Astrophysics Data System (ADS)

    Clarke, David T.; Botchway, Stanley W.; Coles, Benjamin C.; Needham, Sarah R.; Roberts, Selene K.; Rolfe, Daniel J.; Tynan, Christopher J.; Ward, Andrew D.; Webb, Stephen E. D.; Yadav, Rahul; Zanetti-Domingues, Laura; Martin-Fernandez, Marisa L.

    2011-09-01

    Optics clustered to output unique solutions (OCTOPUS) is a microscopy platform that combines single molecule and ensemble imaging methodologies. A novel aspect of OCTOPUS is its laser excitation system, which consists of a central core of interlocked continuous wave and pulsed laser sources, launched into optical fibres and linked via laser combiners. Fibres are plugged into wall-mounted patch panels that reach microscopy end-stations in adjacent rooms. This allows multiple tailor-made combinations of laser colours and time characteristics to be shared by different end-stations minimising the need for laser duplications. This setup brings significant benefits in terms of cost effectiveness, ease of operation, and user safety. The modular nature of OCTOPUS also facilitates the addition of new techniques as required, allowing the use of existing lasers in new microscopes while retaining the ability to run the established parts of the facility. To date, techniques interlinked are multi-photon/multicolour confocal fluorescence lifetime imaging for several modalities of fluorescence resonance energy transfer (FRET) and time-resolved anisotropy, total internal reflection fluorescence, single molecule imaging of single pair FRET, single molecule fluorescence polarisation, particle tracking, and optical tweezers. Here, we use a well-studied system, the epidermal growth factor receptor network, to illustrate how OCTOPUS can aid in the investigation of complex biological phenomena.

  5. New Algorithm to Determine True Colocalization in Combination with Image Restoration and Time-Lapse Confocal Microscopy to Map Kinases in Mitochondria

    PubMed Central

    Iacaruso, María Florencia; Antico Arciuch, Valeria Gabriela; Poderoso, Juan José; Jares-Erijman, Elizabeth Andrea; Pietrasanta, Lía Isabel

    2011-01-01

    The subcellular localization and physiological functions of biomolecules are closely related and thus it is crucial to precisely determine the distribution of different molecules inside the intracellular structures. This is frequently accomplished by fluorescence microscopy with well-characterized markers and posterior evaluation of the signal colocalization. Rigorous study of colocalization requires statistical analysis of the data, albeit yet no single technique has been established as a standard method. Indeed, the few methods currently available are only accurate in images with particular characteristics. Here, we introduce a new algorithm to automatically obtain the true colocalization between images that is suitable for a wide variety of biological situations. To proceed, the algorithm contemplates the individual contribution of each pixel's fluorescence intensity in a pair of images to the overall Pearsońs correlation and Manders' overlap coefficients. The accuracy and reliability of the algorithm was validated on both simulated and real images that reflected the characteristics of a range of biological samples. We used this algorithm in combination with image restoration by deconvolution and time-lapse confocal microscopy to address the localization of MEK1 in the mitochondria of different cell lines. Appraising the previously described behavior of Akt1 corroborated the reliability of the combined use of these techniques. Together, the present work provides a novel statistical approach to accurately and reliably determine the colocalization in a variety of biological images. PMID:21559502

  6. Near-infrared confocal micro-Raman spectroscopy combined with PCA-LDA multivariate analysis for detection of esophageal cancer

    NASA Astrophysics Data System (ADS)

    Chen, Long; Wang, Yue; Liu, Nenrong; Lin, Duo; Weng, Cuncheng; Zhang, Jixue; Zhu, Lihuan; Chen, Weisheng; Chen, Rong; Feng, Shangyuan

    2013-06-01

    The diagnostic capability of using tissue intrinsic micro-Raman signals to obtain biochemical information from human esophageal tissue is presented in this paper. Near-infrared micro-Raman spectroscopy combined with multivariate analysis was applied for discrimination of esophageal cancer tissue from normal tissue samples. Micro-Raman spectroscopy measurements were performed on 54 esophageal cancer tissues and 55 normal tissues in the 400-1750 cm-1 range. The mean Raman spectra showed significant differences between the two groups. Tentative assignments of the Raman bands in the measured tissue spectra suggested some changes in protein structure, a decrease in the relative amount of lactose, and increases in the percentages of tryptophan, collagen and phenylalanine content in esophageal cancer tissue as compared to those of a normal subject. The diagnostic algorithms based on principal component analysis (PCA) and linear discriminate analysis (LDA) achieved a diagnostic sensitivity of 87.0% and specificity of 70.9% for separating cancer from normal esophageal tissue samples. The result demonstrated that near-infrared micro-Raman spectroscopy combined with PCA-LDA analysis could be an effective and sensitive tool for identification of esophageal cancer.

  7. Wound healing anomalies after excimer laser photorefractive keratectomy: correlation of clinical outcomes, corneal topography, and confocal microscopy.

    PubMed Central

    Steinert, R F

    1997-01-01

    PURPOSE: To further the understanding of wound healing anomalies affecting visual function after myopic photorefractive keratectomy (PRK). METHOD: Analysis of a clinical database of PRK on 133 eyes with myopia of -1.5 to -7.0 D and 43 eyes with myopia of -6.0 to -12.0 D. Visual function was analyzed by subgroups of 1) no topographic anomalies; 2) topographic central islands; and 3) topographic keyhole patterns. The natural course of healing was documented over 6 months with visual acuity measurements, clinical observation, and corneal topography. In vivo clinical-pathologic correlations were made by scanning confocal microscopy. RESULTS: Topographic anomalies were identified 1 month post-PRK in 48 eyes (40.3%) with low-moderate myopia and in 14 eyes (32.5%) with moderate-high myopia. For patients with 6 month follow-up, these rates declined to 25% and 23%, respectively. At 1 month post-PRK, topographic anomalies significantly reduced uncorrected and best-corrected visual acuity and refractive predictability. By 6 months post-PRK, the small number of eyes with persistent anomalies had visual outcomes similar to patients with normal topography. A simple approach to anti-island pre-treatment reduced islands slightly and keyhole anomalies significantly (anti-island pre-treatment vs no pretreatment: islands 25% vs 31.8%; keyholes 2.3% vs 17.6%; p = 0.021) but with decreased predictability of induced refractive change at 1 month post-PRK. Confocal microscopy in vivo demonstrated prominent deposition of subepithelial extracellular material 1 to 2 months after PRK that diminished by 6 to 8 months, but persisted in the presence of central islands. Scar formation appeared to represent an elevated plaque of new collagen with active keratocytes. CONCLUSIONS: Topographic anomalies of wound healing are common after PRK. Vision and predictability are reduced by anomalies 1 month post-PRK but anomalies often resolve by 6 months. Marked improvement of vision occurs even when

  8. Transmission Of Power Via Combined Laser Beams

    NASA Technical Reports Server (NTRS)

    Kwon, Jin H.; Lee, Ja H.

    1992-01-01

    Laser Diode Array (LDA) appears to be most efficient means of transferring power from Earth to satellites and between satellites, in terms of mass and size, of various laser configurations. To form large-scale-array amplifier (LSAA), element LDA's must generate well-defined diffraction-limited beams. Coherent matching of phases among LDA's enables system to generate good beam pattern in far field over thousands of kilometers. By passing beam from master laser through number of LDA amplifiers simultaneously, one realizes coherence among amplified output beams. LSAA used for transmission of power with efficiency of approximately 80 percent into receiver of moderate size at 5,000 km. Also transmits data at high rates by line-of-sight rather than fiber optics.

  9. Interferometer combines laser light source and digital counting system

    NASA Technical Reports Server (NTRS)

    1965-01-01

    Measurement of small linear displacements in digital readouts with extreme accuracy and sensitivity is achieved by an interferometer. The instrument combines a digital electro-optical fringe-counting system and a laser light source.

  10. Power combining of semiconductor lasers: A review

    NASA Technical Reports Server (NTRS)

    Katz, J.

    1982-01-01

    Several methods of coherent power combining are described and compared. A comparison is also made between coherent and incoherent power combining, and important operational characteristics are considered. It is found that in communication links with demanding requirements coherent power combining is necessary.

  11. Combining depth analysis with surface morphology analysis to analyse the prehistoric painted pottery from Majiayao Culture by confocal 3D-XRF

    NASA Astrophysics Data System (ADS)

    Yi, Longtao; Liu, Zhiguo; Wang, Kai; Lin, Xue; Chen, Man; Peng, Shiqi; Yang, Kui; Wang, Jinbang

    2016-04-01

    The Majiayao Culture (3300 BC-2900 BC) formed one of the three painted pottery centres of the Yellow River basin, China, in prehistoric times. Painted pottery from this period is famous for its exquisite workmanship and meticulous painting. Studying the layer structure and element distribution of the paint on the pottery is conducive to investigating its workmanship, which is important for archaeological research. However, the most common analysis methods are destructive. To investigate the layers of paint on the pottery nondestructively, a confocal three-dimensional micro-X-ray fluorescence set-up combined with two individual polycapillary lenses has been used to analyse two painted pottery fragments. Nondestructive elemental depth analyses and surface topographic analysis were performed. The elemental depth profiles of Mn, Fe and Ca obtained from these measurements were consistent with those obtained using an optical microscope. The depth profiles show that there are layer structures in two samples. The images show that the distribution of Ca is approximately homogeneous in both painted and unpainted regions. In contrast, Mn appeared only in the painted regions. Meanwhile, the distributions of Fe in the painted and unpainted regions were not the same. The surface topographic shows that the pigment of dark-brown region was coated above the brown region. These conclusions allowed the painting process to be inferred.

  12. Laser beam combiner for Thomson scattering core LIDARa)

    NASA Astrophysics Data System (ADS)

    Balboa, I.; Huang, B.; Naylor, G.; Walsh, M.; Sirinelli, A.; Parsons, P.; Fessey, J.; Townsend, M.; Beurskens, M.; Conway, N.; Flanagan, J.; Kempenaars, M.; Kirk, A.

    2010-10-01

    The light detection and ranging Thomson scattering (TS) diagnostic is advantageous since it only requires a single view port into the tokamak. This technique requires a short pulse laser at high energy, usually showing a limited repetition rate. Having multiple lasers will increase the repetition rate. This paper presents a scanning mirror as a laser beam combiner. Measurements of the position accuracy and jitter show that the pointing stability of the laser beam is within ±25 μrad for over tens of seconds. A control feedback loop is implemented to demonstrate the long term stability. Such a system could be applied for ITER and JET.

  13. Laser beam combiner for Thomson scattering core LIDAR.

    PubMed

    Balboa, I; Huang, B; Naylor, G; Walsh, M; Sirinelli, A; Parsons, P; Fessey, J; Townsend, M; Beurskens, M; Conway, N; Flanagan, J; Kempenaars, M; Kirk, A

    2010-10-01

    The light detection and ranging Thomson scattering (TS) diagnostic is advantageous since it only requires a single view port into the tokamak. This technique requires a short pulse laser at high energy, usually showing a limited repetition rate. Having multiple lasers will increase the repetition rate. This paper presents a scanning mirror as a laser beam combiner. Measurements of the position accuracy and jitter show that the pointing stability of the laser beam is within ±25 μrad for over tens of seconds. A control feedback loop is implemented to demonstrate the long term stability. Such a system could be applied for ITER and JET. PMID:21033888

  14. Laser beam combiner for Thomson scattering core LIDAR

    SciTech Connect

    Balboa, I.; Naylor, G.; Sirinelli, A.; Parsons, P.; Fessey, J.; Townsend, M.; Beurskens, M.; Conway, N.; Kempenaars, M.; Kirk, A.; Walsh, M. [Diagnostics Division, Department of CHD, ITER Organization, CS 90 046, Bulding 155 Flanagan, J.

    2010-10-15

    The light detection and ranging Thomson scattering (TS) diagnostic is advantageous since it only requires a single view port into the tokamak. This technique requires a short pulse laser at high energy, usually showing a limited repetition rate. Having multiple lasers will increase the repetition rate. This paper presents a scanning mirror as a laser beam combiner. Measurements of the position accuracy and jitter show that the pointing stability of the laser beam is within {+-}25 {mu}rad for over tens of seconds. A control feedback loop is implemented to demonstrate the long term stability. Such a system could be applied for ITER and JET.

  15. Fluorescence imaging of reactive oxygen species by confocal laser scanning microscopy for track analysis of synchrotron X-ray photoelectric nanoradiator dose: X-ray pump-optical probe.

    PubMed

    Jeon, Jae Kun; Han, Sung Mi; Kim, Jong Ki

    2016-09-01

    penetration by nanoradiators. In conclusion, the combined use of a synchrotron X-ray microbeam-irradiated three-dimensional ROS gel and confocal laser scanning fluorescence microscopy provides a simple dosimetry method for track analysis of X-ray photoelectric nanoradiator radiation, suggesting extensive cellular damage with dose-enhancement beyond a single cell containing IONs. PMID:27577774

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

    PubMed

    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. PMID:16862444

  17. Measurement of the retinal arteriolar response to a hyperoxic provocation in nonsmokers and smokers, using a high-resolution confocal scanning laser ophthalmoscope

    NASA Astrophysics Data System (ADS)

    O' Halloran, Margaret; O'Donoghue, Eamonn; Dainty, Chris

    2014-07-01

    We used a high-resolution confocal scanning laser ophthalmoscope to measure the magnitude of change in retinal arteriolar diameters in response to oxygen breathing in young, healthy nonsmokers and smokers. Image sequences were obtained before and during oxygen breathing. Image sequences were desinusoided, registered, and averaged, before vessel diameters were measured using a sliding linear regression filter. Arteriole diameters were observed to constrict during the first 5 min. of oxygen breathing, plateau, and remain stable while hyperoxia was maintained, returning to baseline at the end of the hyperoxic period. Blood flow to the temporal retina was found to be higher than to the nasal retina (p=0.008). The percentage constriction of vessels did not vary across retinal quadrants (p=0.372, analysis of variance) and did not depend on vessel size (p=0.538). Baseline diameters were unaffected by acute cigarette smoking. The magnitude of vasoconstriction was diminished in smokers compared to nonsmokers (p=0.017), while acute smoking did not influence the percentage constriction attained by the vessels (p=0.621). Using a high-resolution imaging technique allowed us to measure reactivity to a high degree of accuracy and to assess it in vessels of smaller caliber than were previously studied.

  18. Attachment of Escherichia coli O157:H7 to lettuce leaf surface and bacterial viability in response to chlorine treatment as demonstrated by using confocal scanning laser microscopy.

    PubMed

    Seo, K H; Frank, J F

    1999-01-01

    Confocal scanning laser microscopy was used to observe the location of Escherichia coli O157:H7 on and within lettuce leaves. Sections of leaves (ca. 0.5 by 0.5 cm) were inoculated by submersion in a suspension of E. coli O157:H7 (ca. 10(7) to 10(8) CFU/ml) overnight at 7 degrees C. Fluorescein isothiocyanate-labeled antibody was used to visualize the attached bacteria. E. coli O157:H7 was found attached to the surface, trichomes, stomata, and cut edges. Three-dimensional volume reconstruction of interior portions of leaves showed that E. coli O157:H7 was entrapped 20 to 100 microm below the surface in stomata and cut edges. Agar plate culturing and microscopic observation indicated that E. coli O157:H7 preferentially attached to cut edges, as opposed to the intact leaf surface. Dual staining with fluorescein isothiocyanate-labeled antibody and propidium iodide was used to determine viability of cells on artificially contaminated lettuce leaves after treatment with 20 mg/liter chlorine solution for 5 min. Many live cells were found in stomata and on cut edges following chlorine treatment. E. coli O157:H7 did not preferentially adhere to biofilm produced by Pseudomonas fluorescens on the leaf surface. In contrast to E. coli O157:H7, Pseudomonas adhered to and grew mainly on the intact leaf surface rather than on the cut edges. PMID:9921820

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

    NASA Astrophysics Data System (ADS)

    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.

  20. Novel Application of Confocal Laser Scanning Microscopy and 3D Volume Rendering toward Improving the Resolution of the Fossil Record of Charcoal

    PubMed Central

    Belcher, Claire M.; Punyasena, Surangi W.; Sivaguru, Mayandi

    2013-01-01

    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. PMID:23977267

  1. Considerations in the use of fluorescence in situ hybridization (FISH) and confocal laser scanning microscopy to characterize rumen methanogens and define their spatial distributions.

    PubMed

    Valle, Edith R; Henderson, Gemma; Janssen, Peter H; Cox, Faith; Alexander, Trevor W; McAllister, Tim A

    2015-06-01

    In this study, methanogen-specific coenzyme F420 autofluorescence and confocal laser scanning microscopy were used to identify rumen methanogens and define their spatial distribution in free-living, biofilm-, and protozoa-associated microenvironments. Fluorescence in situ hybridization (FISH) with temperature-controlled hybridization was used in an attempt to describe methanogen diversity. A heat pretreatment (65 °C, 1 h) was found to be a noninvasive method to increase probe access to methanogen RNA targets. Despite efforts to optimize FISH, 16S rRNA methanogen-specific probes, including Arch915, bound to some cells that lacked F420, possibly identifying uncharacterized Methanomassiliicoccales or reflecting nonspecific binding to other members of the rumen bacterial community. A probe targeting RNA from the methanogenesis-specific methyl coenzyme M reductase (mcr) gene was shown to detect cultured Methanosarcina cells with signal intensities comparable to those of 16S rRNA probes. However, the probe failed to hybridize with the majority of F420-emitting rumen methanogens, possibly because of differences in cell wall permeability among methanogen species. Methanogens were shown to integrate into microbial biofilms and to exist as ecto- and endosymbionts with rumen protozoa. Characterizing rumen methanogens and defining their spatial distribution may provide insight into mitigation strategies for ruminal methanogenesis. PMID:25924182

  2. Serotonin-immunoreactive neurones in the visual system of the praying mantis: an immunohistochemical, confocal laser scanning and electron microscopic study.

    PubMed

    Leitinger, G; Pabst, M A; Kral, K

    1999-03-27

    The distribution, number, and morphology of serotonin-immunoreactive (5-HTi) neurones in the optic lobe of the praying mantis Tenodera sinensis were studied using conventional microscopy and confocal laser scanning microscopy. Five or six 5-HTi neurones connect the lobula complex with the medulla, and at least 50 5-HTi neurones appear to be confined to the medulla. In addition, a few large 5-HTi processes from the protocerebrum supply the lobula complex, and two large 5-HTi processes from the protocerebrum ramify in the medulla and lamina, where they show wide field arborisations. In order to provide a basis for understanding the action of serotonin in the lamina, the ultrastructure of its 5-HTi terminals was examined by conventional and immunohistochemical electron microscopy. The 5-HTi profiles were filled with dense core vesicles and made synapses. Output synapses from 5-HTi profiles outnumbered inputs by about 3 to 1. The terminals of the 5-HTi neurones were in close contact with cells of various types, including large monopolar cells, but close apposition to photoreceptor terminals was rare, and no synapses were found between 5-HTi terminals and photoreceptor terminals. PMID:10095007

  3. Structural characterization by confocal laser scanning microscopy and electrochemical study of multi-walled carbon nanotube tyrosinase matrix for phenol detection.

    PubMed

    Guix, Maria; Pérez-López, Briza; Sahin, Melike; Roldán, Mònica; Ambrosi, Adriano; Merkoçi, Arben

    2010-08-01

    A novel visualization methodology based on the use of immunofluorescence and Confocal Laser Scanning Microscopy (CLSM) was used to quantify and visualize tyrosinase enzyme within a MWCNTs matrix immobilized onto carbon based screen-printed electrodes. CLSM was shown to be an extremely powerful technique which allowed a clear visualization of the distribution of the enzyme within both the MWCNTs and carbon based layers and provided additional and useful morphological data for a better understanding of the interaction between biomolecules and electrode materials. Transmission Electron Microscopy (TEM) and Scanning Electron Microscopy (SEM) were also employed to fully characterize the system components. The proposed MWCNT/Tyrosinase matrix was applied to the detection of phenol, as an alternative biosensor material. Electrochemical analytical performances of the biosensor were investigated in order to determine the optimal fabrication design along with the enzyme stability. The biosensor based on the developed biomaterial matrix proved promising results in terms of cost, simplicity and analytical performance. A detection limit of 1.35 microM and a sensitivity of 47.4 microA mM(-1) within a linear response range of 2.5 to 75 microM phenol were obtained. The biosensor performed well as a disposable device and could be stored in a refrigerator (-18 degrees C) without loss of activity for up to 2 months. PMID:20532304

  4. Corneal Backscatter Analysis by In Vivo Confocal Microscopy: Fellow Eye Comparison of Small Incision Lenticule Extraction and Femtosecond Laser-Assisted LASIK

    PubMed Central

    Agca, Alper; Ozgurhan, Engin Bilge; Yildirim, Yusuf; Cankaya, Kadir Ilker; Guleryuz, Nimet Burcu; Ozkaya, Abdullah; Demirok, Ahmet; Yilmaz, Omer Faruk

    2014-01-01

    Purpose. To evaluate and compare corneal backscatter from anterior stroma between small incision lenticule extraction (SMILE) and femtosecond laser-assisted LASIK (femto-LASIK). Methods. A cohort of 60 eyes of 30 patients was randomized to receive SMILE in one eye and femto-LASIK in the fellow eye. In vivo confocal microscopy was performed at 1 week and 1, 3, and 6 months after surgery. The main outcome measurements were maximum backscattered intensity and the depth from which it was measured, the backscattered light intensity 30 μm below Bowman's membrane at the flap interface and 150 μm below the superficial epithelium, and the number of refractive particles at the flap interface. Results. The mean backscattered light intensity (LI) at all measured depths and the maximum backscattered LI were higher in the SMILE group than the femto-LASIK group at all postoperative visits. LI differences at 1 week and 1- and 3-month visits were statistically significant (P < 0,05). LI differences at 6 months were not statistically significant. There was no difference in the number of refractive particles at the flap interface between the groups at any visit. Conclusions. SMILE results in increased backscattered LI in the anterior stroma when compared with femto-LASIK were evaluated. PMID:24734168

  5. Corneal backscatter analysis by in vivo confocal microscopy: fellow eye comparison of small incision lenticule extraction and femtosecond laser-assisted LASIK.

    PubMed

    Agca, Alper; Ozgurhan, Engin Bilge; Yildirim, Yusuf; Cankaya, Kadir Ilker; Guleryuz, Nimet Burcu; Alkin, Zeynep; Ozkaya, Abdullah; Demirok, Ahmet; Yilmaz, Omer Faruk

    2014-01-01

    Purpose. To evaluate and compare corneal backscatter from anterior stroma between small incision lenticule extraction (SMILE) and femtosecond laser-assisted LASIK (femto-LASIK). Methods. A cohort of 60 eyes of 30 patients was randomized to receive SMILE in one eye and femto-LASIK in the fellow eye. In vivo confocal microscopy was performed at 1 week and 1, 3, and 6 months after surgery. The main outcome measurements were maximum backscattered intensity and the depth from which it was measured, the backscattered light intensity 30  μ m below Bowman's membrane at the flap interface and 150  μ m below the superficial epithelium, and the number of refractive particles at the flap interface. Results. The mean backscattered light intensity (LI) at all measured depths and the maximum backscattered LI were higher in the SMILE group than the femto-LASIK group at all postoperative visits. LI differences at 1 week and 1- and 3-month visits were statistically significant (P < 0,05). LI differences at 6 months were not statistically significant. There was no difference in the number of refractive particles at the flap interface between the groups at any visit. Conclusions. SMILE results in increased backscattered LI in the anterior stroma when compared with femto-LASIK were evaluated. PMID:24734168

  6. Scanning electron and confocal scanning laser microscopy imaging of the ultrastructure and viability of vaginal Candida albicans and non- albicans species adhered to an intrauterine contraceptive device.

    PubMed

    Paiva, Luciene C Farias; Donatti, Lucélia; Patussi, Eliana V; Svizdinski, Terezinha I E; Lopes-Consolaro, Márcia E

    2010-10-01

    Although bacterial biofilms have been studied in detail, adhesion of Candida albicans and non-albicans species to an intrauterine contraceptive device (IUD) is not clear. The objective of this study was to evaluate aspects of imaging of the ultrastructure and viability of vaginal yeasts adhered to different parts of an IUD, through scanning electron microscopy (SEM) and confocal scanning laser microscopy (CSLM). We studied yeasts isolated from different patients with vulvovaginal candidiasis: C. albicans, C. glabrata, C. guillermondii, C. parapsilosis, C. tropicalis, and Saccharomyces cerevisiae. A suspension of the each yeast was prepared and incubated with IUD parts (tail, without copper, and copper-covered). SEM and CSLM showed that all the vaginal yeasts adhered to all the parts of the IUD and demonstrated viability, including 30 days after contact for C. albicans. Possibly irregularities of IUD surface contribute to the adherence process. Although all of the IUD parts contribute to retention of yeasts in the genital tract, high concentration of yeast cells on the tail may indicate the importance of this segment in maintaining the colonization by yeast cells because the tail forms a bridge between the external environment, the vagina that is colonized by yeast cells, and the upper genital tract where there is no colonization. PMID:20804637

  7. Use of probe-based confocal laser endomicroscopy (pCLE) in gastrointestinal applications. A consensus report based on clinical evidence

    PubMed Central

    Wang, Kenneth K; Carr-Locke, David L; Singh, Satish K; Neumann, Helmut; Bertani, Helga; Arsenescu, Razvan I; Caillol, Fabrice; Chang, Kenneth J; Chaussade, Stanislas; Coron, Emmanuel; Costamagna, Guido; Dlugosz, Aldona; Ian Gan, S; Giovannini, Marc; Gress, Frank G; Haluszka, Oleh; Ho, Khek Y; Kahaleh, Michel; Konda, Vani J; Prat, Frederic; Shah, Raj J; Sharma, Prateek; Slivka, Adam; Wolfsen, Herbert C; Zfass, Alvin

    2015-01-01

    Background Probe-based confocal laser endomicroscopy (pCLE) provides microscopic imaging during an endoscopic procedure. Its introduction as a standard modality in gastroenterology has brought significant progress in management strategies, affecting many aspects of clinical care and requiring standardisation of practice and training. Objective This study aimed to provide guidance on the standardisation of its practice and training in Barrett’s oesophagus, biliary strictures, colorectal lesions and inflammatory bowel diseases. Methods Initial statements were developed by five group leaders, based on the available clinical evidence. These statements were then voted and edited by the 26 participants, using a modified Delphi approach. After two rounds of votes, statements were validated if the threshold of agreement was higher than 75%. Results Twenty-six experts participated and, among a total of 77 statements, 61 were adopted (79%) and 16 were rejected (21%). The adoption of each statement was justified by the grade of evidence. Conclusion pCLE should be used to enhance the diagnostic arsenal in the evaluation of these indications, by providing microscopic information which improves the diagnostic performance of the physician. In order actually to implement this technology in the clinical routine, and to ensure good practice, standardised initial and continuing institutional training programmes should be established. PMID:26137298

  8. Outcome of probe-based confocal laser endomicroscopy (pCLE) during endoscopic retrograde cholangiopancreatography: A single-center prospective study in 45 patients

    PubMed Central

    Lönnebro, Ragnar; Stigliano, Serena; Haas, Stephan L; Swahn, Fredrik; Enochsson, Lars; Noel, Rozh; Segersvärd, Ralf; Chiaro, Marco Del; Verbeke, Caroline S; Arnelo, Urban

    2015-01-01

    Background Diagnosis of pre-malignant and malignant lesions in the bile duct and the pancreas is sometimes cumbersome. This applies in particular to intraductal papillary mucinous neoplasia (IPMN) and bile duct strictures in primary sclerosing cholangitis (PSC). Aims To evaluate in a prospective cohort study the sensitivity and specificity of probe-based confocal laser microscopy (pCLE) during endoscopic retrograde cholangiopancreatography (ERCP). Methods We performed pCLE together with mother-baby endoscopy (SpyGlass) during 50 ERCP sessions in 45 patients. The Miami and Paris criteria were applied. Clinical diagnosis via imaging was compared to pCLE and the final pathological diagnosis from surgically-resected, biopsy, or cytology specimens. Patients were followed up for at least 1 year. Results We were able to perform pCLE in all patients. Prior to endoscopy, the diagnosis was benign in 23 patients and undetermined (suspicious) in 16 patients, while six patients had an unequivocal diagnosis of malignancy. Sensitivity was 91% and specificity 52%. The positive (PPV) and negative predictive value (NPV) was 82% and 100%, respectively. Apart from mild post-ERCP pancreatitis in two patients, no complications occurred. Conclusions Our study showed that pCLE is a safe, expert endoscopic method with high technical feasibility, high sensitivity and high NPV. It provided diagnostic information that can be helpful for decisions on patient management, especially in the case of IPMN and unclear pancreatic lesions, in individuals whom are at increased risk for pancreatic cancer. PMID:26668748

  9. Confocal fluorescence microendoscopy of bronchial epithelium

    NASA Astrophysics Data System (ADS)

    Lane, Pierre M.; Lam, Stephen; McWilliams, Annette; Leriche, Jean C.; Anderson, Marshall W.; Macaulay, Calum E.

    2009-03-01

    Confocal microendoscopy permits the acquisition of high-resolution real-time confocal images of bronchial mucosa via the instrument channel of an endoscope. We report here on the construction and validation of a confocal fluorescence microendoscope and its use to acquire images of bronchial epithelium in vivo. Our objective is to develop an imaging method that can distinguish preneoplastic lesions from normal epithelium to enable us to study the natural history of these lesions and the efficacy of chemopreventive agents without biopsy removal of the lesion that can introduce a spontaneous regression bias. The instrument employs a laser-scanning engine and bronchoscope-compatible confocal probe consisting of a fiber-optic image guide and a graded-index objective lens. We assessed the potential of topical application of physiological pH cresyl violet (CV) as a fluorescence contrast-enhancing agent for the visualization of tissue morphology. Images acquired ex vivo with the confocal microendoscope were first compared with a bench-top confocal fluorescence microscope and conventional histology. Confocal images from five sites topically stained with CV were then acquired in vivo from high-risk smokers and compared to hematoxylin and eosin stained sections of biopsies taken from the same site. Sufficient contrast in the confocal imagery was obtained to identify cells in the bronchial epithelium. However, further improvements in the miniature objective lens are required to provide sufficient axial resolution for accurate classification of preneoplastic lesions.

  10. Cascaded combiners for a high power CW fiber laser

    NASA Astrophysics Data System (ADS)

    Tan, Qirui; Ge, Tingwu; Zhang, Xuexia; Wang, Zhiyong

    2016-02-01

    We report cascaded combiners for a high power continuous wave (CW) fiber laser in this paper. The cascaded combiners are fabricated with an improved lateral splicing process. During the fusing process, there is no stress or tension between the pump fiber and the double-cladding fiber. Thus, the parameters of the combiner are better than those that have been reported. The coupling efficiency is 98.5%, and the signal insertion loss is 1%. The coupling efficiency of the cascaded combiners is 97.5%. The pump lights are individually coupled into the double-cladding fiber via five combiners. The thermal effects cannot cause damage to the combiners and the cascaded combiners can operate stably in high power CW fiber lasers. We also develop a high power CW fiber laser that generates a maximum 780 W of CW signal power at 1080 nm with 71% optical-to-optical conversion efficiency. The fiber laser is pumped via five intra-cavity cascaded combiners and five extra-cavity cascaded combiners with a maximum pump power of 1096 W and a pump wavelength of 975 nm.

  11. Influence of erbium, chromium-doped: Yttrium scandium-gallium-garnet laser etching and traditional etching systems on depth of resin penetration in enamel: A confocal laser scanning electron microscope study

    PubMed Central

    Vijayan, Vishal; Rajasigamani, K.; Karthik, K.; Maroli, Sasidharan; Chakkarayan, Jitesh; Haris, Mohamed

    2015-01-01

    Objective: This study was performed to assess the resin tag length penetration in enamel surface after bonding of brackets to identify which system was most efficient. Methodology: Our study was based on a more robust confocal microscopy for visualizing the resin tags in enamel. Totally, 100 extracted human first and second premolars have been selected for this study and were randomly divided into ten groups of 10 teeth each. In Group 1, the buccal enamel surface was etched with 37% phosphoric acid (3M ESPE), Group 2 with 37% phosphoric (Ultradent). In Groups 5, 6, and 7, erbium, chromium-doped: Yttrium scandium-gallium-garnet (Er, Cr: YSGG) laser (Biolase) was used for etching the using following specifications: Group 5 (1.5 W/20 Hz, 15 s), Group 6 (2 W/10 Hz, 15 s), and Group 7 (2 W/20 Hz, 15 s). In Groups 8, 9, and 10, Er, Cr: YSGG laser (Biolase) using same specifications and additional to this step, conventional etching on the buccal enamel surface was etched with 37% (3M ESPE) after laser etching. In Groups 1, 5, 6, 7, 8, 9, and 10 3M Unitek Transbond XT primer was mixed with Rhodamine B dye (Sigma-Aldrich, Germany) to etched surface and then cured for 20 s. In Group 2, Ultradents bonding agent was mixed with Rhodamine B. In Group 3, 3M Unitek Transbond PLUS, Monrovia, USA, which was mixed with Rhodamine B dye (Sigma-Aldrich, Germany). Group 4, with self-etching primer (Ultradent-Peak SE, USA) was mixed with Rhodamine B dye (Sigma-Aldrich, Germany). Later (3M Unitek, Transbond XT, Monrovia USA) [Figure 1] was used to bond the modified Begg brackets (T. P. Orthodontics) in Groups 1, 3, 5, 6, 7, 8, 9, and 10. In Groups 2, 4 Ultradent-Peak LC Bond was used to bond the modified brackets. After curing brackets were debonded, and enamel depth penetration was assessed using confocal laser scanning microscope. Results: Group J had a mean maximum depth of penetration of 100.876 μm, and Group D was the least having a maximum value of 44.254 μm. Conclusions: Laser

  12. Ultrarelativistic laser systems based on coherent beam combining

    NASA Astrophysics Data System (ADS)

    Bagayev, S. N.; Trunov, V. I.; Pestryakov, E. V.; Frolov, S. A.; Leschenko, V. E.; Kirpichnikov, A. V.; Kokh, A. E.; Petrov, V. V.; Vasiliev, V. A.

    2012-07-01

    Conceptual design for femtosecond laser system of exawatt class, based on multi-channel amplifier and coherent field combining of petawatt amplifier channels with phase-frequency controlled radiation by optical clock are discussed. The scheme of start petawatt level few-cycle laser system with stable phase-frequency parameters determinated by the accuracy of the optical standard based on parametric amplification in big-size LBO crystals pumped by picosecond pulses is analyzed.

  13. Mid-IR laser source using hollow waveguide beam combining

    NASA Astrophysics Data System (ADS)

    Elder, Ian F.; Thorne, Daniel H.; Lamb, Robert A.; Jenkins, R. M.

    2016-03-01

    Hollow waveguide technology is a route to efficient beam combining of multiple laser sources in a compact footprint. It is a technology appropriate for combining free-space or fibre-coupled beams generated by semiconductor, fibre or solidstate laser sources. This paper will present results of a breadboard mid-IR system comprising four laser sources combined using a hollow waveguide optical circuit. In this approach the individual dichroic beam combiner components are held in precision alignment slots in the hollow waveguide circuit and the different input wavelengths are guided between the components to a common output port. The hollow waveguide circuit is formed in the surface of a Macor (machinable glass-ceramic) substrate using precision CNC machining techniques. The hollow waveguides have fundamentally different propagation characteristics to solid core waveguides leading to transmission characteristics close to those of the atmosphere while still providing useful light guidance properties. The transmission efficiency and power handling of the hollow waveguide circuit can be designed to be very high across a broad waveband range. Three of the sources are quantum cascade lasers (QCLs), a semiconductor laser technology providing direct generation of midwave IR output. The combined beams provide 4.2 W of near diffraction-limited output co-boresighted to better than 20 µrad. High coupling efficiency into the waveguides is demonstrated, with negligible waveguide transmission losses. The overall transmission of the hollow waveguide beam combining optical circuit, weighted by the laser power at each wavelength, is 93%. This loss is dominated by the performance of the dichroic optics used to combine the beams.

  14. A novel technique for differentiation of proteins in the development of acid gel structure from control and heat treated milk using confocal scanning laser microscopy.

    PubMed

    Dubert-Ferrandon, Alix; Niranjan, Keshaven; Grandison, Alistair S

    2006-11-01

    The incorporation of caseins and whey proteins into acid gels produced from unheated and heat treated skimmed milk was studied by confocal scanning laser microscopy (CSLM) using fluorescent labelled proteins. Bovine casein micelles were labelled using Alexa Fluor 594, while whey proteins were labelled using Alexa Fluor 488. Samples of the labelled protein solutions were introduced into aliquots of pasteurised skim milk, and skim milk heated to 90 degrees C for 2 min and 95 degrees C for 8 min. The milk was acidified at 40 degrees C to a final pH of 4.4 using 20 g glucono-delta-lactone/l (GDL). The formation of gels was observed with CSLM at two wavelengths (488 nm and 594 nm), and also by visual and rheological methods. In the control milk, as pH decreased distinct casein aggregates appeared, and as further pH reduction occurred, the whey proteins could be seen to coat the casein aggregates. With the heated milks, the gel structure was formed of continuous strands consisting of both casein and whey protein. The formation of the gel network was correlated with an increase in the elastic modulus for all three treatments, in relation to the severity of heat treatment. This model system allows the separate observation of the caseins and whey proteins, and the study of the interactions between the two protein fractions during the formation of the acid gel structure, on a real-time basis. The system could therefore be a valuable tool in the study of structure formation in yoghurt and other dairy protein systems. PMID:16834815

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

    PubMed Central

    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

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

    PubMed

    Li, Qingqing; Chen, Baoliang

    2014-05-01

    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. PMID:24678956

  17. Prospective evaluation of the utility of intraoperative confocal laser endomicroscopy in patients with brain neoplasms using fluorescein sodium: experience with 74 cases.

    PubMed

    Martirosyan, Nikolay L; Eschbacher, Jennifer M; Kalani, M Yashar S; Turner, Jay D; Belykh, Evgenii; Spetzler, Robert F; Nakaji, Peter; Preul, Mark C

    2016-03-01

    OBJECTIVE This study evaluated the utility, specificity, and sensitivity of intraoperative confocal laser endomicroscopy (CLE) to provide diagnostic information during resection of human brain tumors. METHODS CLE imaging was used in the resection of intracranial neoplasms in 74 consecutive patients (31 male; mean age 47.5 years; sequential 10-month study period). Intraoperative in vivo and ex vivo CLE was performed after intravenous injection of fluorescein sodium (FNa). Tissue samples from CLE imaging-matched areas were acquired for comparison with routine histological analysis (frozen and permanent sections). CLE images were classified as diagnostic or nondiagnostic. The specificities and sensitivities of CLE and frozen sections for gliomas and meningiomas were calculated using permanent histological sections as the standard. RESULTS CLE images were obtained for each patient. The mean duration of intraoperative CLE system use was 15.7 minutes (range 3-73 minutes). A total of 20,734 CLE images were correlated with 267 biopsy specimens (mean number of images/biopsy location, in vivo 84, ex vivo 70). CLE images were diagnostic for 45.98% in vivo and 52.97% ex vivo specimens. After initiation of CLE, an average of 14 in vivo images and 7 ex vivo images were acquired before identification of a first diagnostic image. CLE specificity and sensitivity were, respectively, 94% and 91% for gliomas and 93% and 97% for meningiomas. CONCLUSIONS CLE with FNa provided intraoperative histological information during brain tumor removal. Specificities and sensitivities of CLE for gliomas and meningiomas were comparable to those for frozen sections. These data suggest that CLE could allow the interactive identification of tumor areas, substantially improving intraoperative decisions during the resection of brain tumors. PMID:26926051

  18. Fully Automatic Determination of Soil Bacterium Numbers, Cell Volumes, and Frequencies of Dividing Cells by Confocal Laser Scanning Microscopy and Image Analysis

    PubMed Central

    Bloem, J.; Veninga, M.; Shepherd, J.

    1995-01-01

    We describe a fully automatic image analysis system capable of measuring cell numbers, volumes, lengths, and widths of bacteria in soil smears. The system also determines the number of cells in agglomerates and thus provides the frequency of dividing cells (FDC). Images are acquired from a confocal laser scanning microscope. The grey images are smoothed by convolution and by morphological erosion and dilation to remove noise. The background is equalized by flooding holes in the image and is then subtracted by two top hat transforms. Finally, the grey image is sharpened by delineation, and all particles above a fixed threshold are detected. The number of cells in each detected particle is determined by counting the number of local grey-level maxima in the particle. Thus, up to 1,500 cells in 10 fields of view in a soil smear are analyzed in 30 min without human intervention. Automatic counts of cell numbers and FDC were similar to visual counts in field samples. In microcosms, automatic measurements showed significant increases in cell numbers, FDC, mean cell volume, and length-to-width ratio after amendment of the soil. Volumes of fluorescent microspheres were measured with good approximation, but the absolute values obtained were strongly affected by the settings of the detector sensitivity. Independent measurements of bacterial cell numbers and volumes by image analysis and of cell carbon by a total organic carbon analyzer yielded an average specific carbon content of 200 fg of C (mu)m(sup-3), which indicates that our volume estimates are reasonable. PMID:16534976

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

    PubMed Central

    de ANDRADE, Flaviana Bombarda; ARIAS, Marcela Paola Castro; MALIZA, Amanda Garcia Alves; DUARTE, Marco Antonio Hungaro; GRAEFF, Márcia Sirlene Zardin; AMOROSO-SILVA, Pablo Andrés; MIDENA, Raquel Zanin; de MORAES, Ivaldo Gomes

    2015-01-01

    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 (p<0.05). No differences were observed between group A and B (p>0.05). 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. PMID:26200524

  20. Polarization/Spatial Combining of Laser-Diode Pump Beams

    NASA Technical Reports Server (NTRS)

    Gelsinger, Paul; Liu, Duncan

    2008-01-01

    A breadboard version of an optical beam combiner is depicted which make it possible to use the outputs of any or all of four multimode laser diodes to pump a non-planar ring oscillator (NPRO) laser. The output of each laser diode has a single-mode profile in the meridional plane containing an axis denoted the 'fast' axis and a narrower multimode profile in the orthogonal meridional plane, which contains an axis denoted the 'slow' axis and a narrower multimode profile in the orthogonal meridional plane, which contains an axis denoted the 'slow' axis. One of the purposes served by the beam-combining optics is to reduce the fast-axis numerical aperture (NA) of the laser-diode output to match the NA of the optical fiber. Along the slow axis, the unmodified laser-diode NA is already well matched to the fiber optic NA, so no further slow-axis beam shaping is needed. In this beam combiner, the laser-diode outputs are collimated by aspherical lenses, then half-wave plates and polarizing beam splitters are used to combine the four collimated beams into two beams. Spatial combination of the two beams and coupling into the optical fiber is effected by use of anamorphic prisms, mirrors, and a focusing lens. The anamorphic prisms are critical elements in the NA-matching scheme, in that they reduce the fast-axis beam width to 1/6 of its original values. Inasmuch as no slow-axis beam shaping is needed, the collimating and focusing lenses are matched for 1:1 iumaging. Because these lenses are well corrected for infinite conjugates the combiner offers diffraction-limited performance along both the fast and slow axes.

  1. Virtual pinhole confocal microscope

    SciTech Connect

    George, J.S.; Rector, D.M.; Ranken, D.M.; Peterson, B.; Kesteron, J.

    1999-06-01

    Scanned confocal microscopes enhance imaging capabilities, providing improved contrast and image resolution in 3-D, but existing systems have significant technical shortcomings and are expensive. Researchers at Los Alamos National Laboratory have developed a novel approach--virtual pinhole confocal microscopy--that uses state of the art illumination, detection, and data processing technologies to produce an imager with a number of advantages: reduced cost, faster imaging, improved efficiency and sensitivity, improved reliability and much greater flexibility. Work at Los Alamos demonstrated proof of principle; prototype hardware and software have been used to demonstrate technical feasibility of several implementation strategies. The system uses high performance illumination, patterned in time and space. The authors have built functional confocal imagers using video display technologies (LCD or DLP) and novel scanner based on a micro-lens array. They have developed a prototype system for high performance data acquisition and processing, designed to support realtime confocal imaging. They have developed algorithms to reconstruct confocal images from a time series of spatially sub-sampled images; software development remains an area of active development. These advances allow the collection of high quality confocal images (in fluorescence, reflectance and transmission modes) with equipment that can inexpensively retrofit to existing microscopes. Planned future extensions to these technologies will significantly enhance capabilities for microscopic imaging in a variety of applications, including confocal endoscopy, and confocal spectral imaging.

  2. Combination of erbium and holmium laser radiation for tissue ablation

    NASA Astrophysics Data System (ADS)

    Pratisto, Hans S.; Frenz, Martin; Koenz, Flurin; Altermatt, Hans J.; Weber, Heinz P.

    1996-05-01

    Erbium lasers emitting at 2.94 micrometers and holmium lasers emitting at 2.1 micrometers are interesting tools for cutting, drilling, smoothing and welding of water containing tissues. The high absorption coefficient of water at these wavelengths leads to their good ablation efficiency with controlled thermally altered zones around the ablation sites. Combination of pulses with both wavelengths transmitted through one fiber were used to perform incisions in soft tissue and impacts in bone disks. Histological results and scanning electron microscope evaluations reveal the strong influence of the absorption coefficient on tissue effects, especially on the ablation efficiency and the zone of thermally damaged tissue. It is demonstrated that the combination of high ablation rates and deep coagulation zones can be achieved. The results indicate that this laser system can be considered as a first step towards a multi-functional medical instrument.

  3. 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

    PubMed Central

    Girish, C Sabari; Ponnappa, KC; Girish, TN; Ponappa, MC

    2013-01-01

    Background: 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. Aim: (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: 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. Results: 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. Conclusion: 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. PMID:23956530

  4. Lossless beam combiners for nearly equal laser frequencies

    NASA Astrophysics Data System (ADS)

    Haubrich, D.; Dornseifer, M.; Wynands, R.

    2000-02-01

    We discuss three ways to combine two laser beams with equal linear polarizations and very closely spaced frequencies into a single output beam containing up to 100% of the input power of each beam. One setup, a modified Mach-Zehnder interferometer, is examined in detail; it allows to adjust the combined output power electronically with the help of a simple servo loop. With off-the-shelf optical components we obtained a 98% efficiency.

  5. Confocal microscopy in microgravity research

    NASA Astrophysics Data System (ADS)

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

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

  6. Neurosurgical confocal endomicroscopy: A review of contrast agents, confocal systems, and future imaging modalities

    PubMed Central

    Zehri, Aqib H.; Ramey, Wyatt; Georges, Joseph F.; Mooney, Michael A.; Martirosyan, Nikolay L.; Preul, Mark C.; Nakaji, Peter

    2014-01-01

    Background: The clinical application of fluorescent contrast agents (fluorescein, indocyanine green, and aminolevulinic acid) with intraoperative microscopy has led to advances in intraoperative brain tumor imaging. Their properties, mechanism of action, history of use, and safety are analyzed in this report along with a review of current laser scanning confocal endomicroscopy systems. Additional imaging modalities with potential neurosurgical utility are also analyzed. Methods: A comprehensive literature search was performed utilizing PubMed and key words: In vivo confocal microscopy, confocal endomicroscopy, fluorescence imaging, in vivo diagnostics/neoplasm, in vivo molecular imaging, and optical imaging. Articles were reviewed that discussed clinically available fluorophores in neurosurgery, confocal endomicroscopy instrumentation, confocal microscopy systems, and intraoperative cancer diagnostics. Results: Current clinically available fluorescent contrast agents have specific properties that provide microscopic delineation of tumors when imaged with laser scanning confocal endomicroscopes. Other imaging modalities such as coherent anti-Stokes Raman scattering (CARS) microscopy, confocal reflectance microscopy, fluorescent lifetime imaging (FLIM), two-photon microscopy, and second harmonic generation may also have potential in neurosurgical applications. Conclusion: In addition to guiding tumor resection, intraoperative fluorescence and microscopy have the potential to facilitate tumor identification and complement frozen section analysis during surgery by providing real-time histological assessment. Further research, including clinical trials, is necessary to test the efficacy of fluorescent contrast agents and optical imaging instrumentation in order to establish their role in neurosurgery. PMID:24872922

  7. Fiber-matrix interface studies on bioabsorbable composite materials for internal fixation of bone fractures. II. A new method using laser scanning confocal microscopy.

    PubMed

    Slivka, M A; Chu, C C

    1997-12-01

    In this study, a new visual characterization method was developed using laser scanning confocal microscopy (LSCM) to study morphologic properties, particularly at the fiber-matrix interface, by optical sectioning of bioabsorbable single-fiber composites. The interface gap width (IGW) between the fiber and matrix, and the changes in IGW after in vitro hydrolysis, named the gap rate (Rg), were measured from images obtained using the LSCM. Higher values for IGW and Rg showed faster degradation of the fiber-matrix interface. These parameters were used to investigate the effects of strain, wicking, different reinforcing fibers, and gamma-irradiation on the fiber-matrix interface morphology. The component materials used were nonbioabsorbable AS4 carbon (C) fibers, bioabsorbable calcium phosphate (CaP), poly(glycolic acid) (PGA), and chitin fibers, and bioabsorbable poly(L-lactic acid) (PLLA) matrix. The application of strain on CaP/PLLA composites increased the IGW up to about 15%, after which there was no change up to 25%. The Rg for CaP/PLLA composites with the fiber ends exposed in vitro (permitting wicking) was greater than for CaP/PLLA with the fiber ends embedded completely within the matrix (preventing wicking). Open-end C/PLLA composites had the slowest rate of interface degradation in vitro, followed by chitin/PLLA, PGA/PLLA, and CaP/PLLA. The exposure of closed-end CaP/PLLA composites to 4 Mrad of gamma-irradiation, in air at room temperature or in vaccuum at 77K, accelerated the rate of interface degradation in vitro. In conclusion, an effective new visual characterization method was developed using LSCM, and it was used to show that (a) moderate strain could accelerate the degradation of the interface, (b) fiber-matrix interface wicking could accelerate the rate of degradation of the interface, (c) the rate of interface degradation depends on the type of fiber used, and (d) gamma-irradiation could accelerate the rate of interface degradation. Furthermore, the

  8. Changes in F-actin organization induced by hard metal particle exposure in rat pulmonary epithelial cells using laser scanning confocal microscopy.

    PubMed

    Antonini, J M; Starks, K; Roberts, J R; Millecchia, L; Yang, H M; Rao, K M

    2000-01-01

    Chronic inhalation of hard metal (WC-Co) particles causes alveolitis and the eventual development of pulmonary fibrosis. The initial inflammatory response includes a change in the alveolar epithelial cell-capillary barrier, which has been shown to be regulated by the state of assembly and organization of the actin cytoskeletal network. The objective of this study was to evaluate the effect WC-Co particles have on F-actin organization of lung epithelial cells in an in vitro culture system. Rat lung epithelial (L2) cells were exposed to 5, 25, and 100 microg/mL of WC-Co particles, as well as the individual components (Co and WC) of the hard metal mixture particles for 24 h. The effect on F-actin organization was visualized by laser scanning confocal microscopy (LSCM) following Bodipy-Phallacidin staining. Minimal changes in the F-actin microfilaments of L2 cells were observed by LSCM after exposure to WC and WC-Co at 5 and 25 microg/mL, while at 100 microg/mL, there was a noticeable disruption in the uniform distribution of L2 cell F-actin microfilaments. After exposure to Co, a dose-dependent change in the F-actin organization of the L2 cells was observed. Little change in F-actin assembly was observed after treatment with 5 microg/mL of Co (the concentration equivalent to the 5% amount of Co commonly present in 100 microg/mL of the WC-Co sample mixture). However, at 100 microg/mL of Co, the microfilaments aggregated into homogeneous masses within the cells, and a significant loss in the organization of L2 F-actin was observed. These dramatic alterations in F-actin organization seen after exposure to the higher doses of Co were attributed to an increase in L2 cell injury as measured by lactate dehydrogenase and trypan blue exclusion. We conclude the pulmonary response evoked in the lung by inhalation of high levels of WC-Co particles is unlikely due to alterations in the F-actin microfilaments of lung-epithelial cells. PMID:10900403

  9. Advances in fiber combined pump modules for fiber lasers

    NASA Astrophysics Data System (ADS)

    Crum, Trevor; Romero, Oscar; Li, Hanxuan; Jin, Xu; Towe, Terry; Chyr, Irving; Truchan, Tom; Liu, Daming; Cutillas, Serge; Johnson, Kelly; Park, Sang-Ki; Wolak, Ed; Miller, Robert; Bullock, Robert; Mott, Jeff; Fidric, Bernard; Harrison, James

    2009-02-01

    Fiber combining multiple pump sources for fiber lasers has enabled the thermal and reliability advantages of distributed architectures. Recently, mini-bar based modules have been demonstrated which combine the advantages of independent emitter failures previously shown in single-stripe pumps with improved brightness retention yielding over 2 MW/cm2Sr in compact economic modules. In this work multiple fiber-coupled mini-bars are fiber combined to yield an output of over 400 W with a brightness exceeding 1 MW/cm2Sr in an economic, low loss architecture.

  10. Combination visible and infrared lasers for skin rejuvenation.

    PubMed

    Lee, Min-Wei Christine

    2002-12-01

    Noninvasive techniques for skin rejuvenation are quickly being established as a new standard in the treatment of mild rhytides and overall skin toning. Multiple laser wavelengths and modalities have been tried for this procedure with varying degrees of success. These lasers include 532 nm, 585 nm, 1064 nm, 1320 nm, 1450 nm, and 1540 nm wavelengths. This study evaluates a combination technique by using a long-pulsed 532 nm potassium titanyl phosphate (KTP) laser and a long-pulsed 1064 nm Neodynium:yttrium aluminum garnet (Nd:YAG) laser, both separately and combined, for noninvasive photorejuvenation and skin toning/collagen enhancement, and establishes efficacy and degree of success. A total of 150 patients were treated with the long-pulsed KTP 532 nm (Aura; Laserscope, San Jose, CA) and long-pulsed Nd:YAG 1064 nm (Lyra; Laserscope) lasers both separately and combined. Patients included skin types I through V. The fluences varied between 7 and 15 J/cm2 at 7 to 20 ms pulse duration with a 2-mm handpiece, and 6 to 15 J/cm2 and 30 to 50 ms with a 4-mm handpiece for KTP. The Nd:YAG fluences were set at 24 to 30 J/cm2 for a 10-mm handpiece and 30 J/cm2 for a SmartScan Plus scanner (Laserscope, San Jose, CA). These energies were delivered at 30 to 65 ms pulse durations. All patients were treated at least 3 times and at most 6 times at monthly intervals, and were observed for up to 18 months after the last treatment. All 150 patients exhibited a mild to moderate degree of improvement in the appearance of rhytides, moderate degree of improvement in skin toning and texture, and great improvement in the reduction of redness and pigmentation. The KTP used alone was superior to the Nd:YAG laser in terms of results. The KTP and Nd:YAG laser combination was superior to either laser used alone. PMID:12512652

  11. Parallel excitation-emission multiplexed fluorescence lifetime confocal microscopy for live cell imaging

    PubMed Central

    Zhao, Ming; Li, Yu; Peng, Leilei

    2014-01-01

    We present a novel excitation-emission multiplexed fluorescence lifetime microscopy (FLIM) method that surpasses current FLIM techniques in multiplexing capability. The method employs Fourier multiplexing to simultaneously acquire confocal fluorescence lifetime images of multiple excitation wavelength and emission color combinations at 44,000 pixels/sec. The system is built with low-cost CW laser sources and standard PMTs with versatile spectral configuration, which can be implemented as an add-on to commercial confocal microscopes. The Fourier lifetime confocal method allows fast multiplexed FLIM imaging, which makes it possible to monitor multiple biological processes in live cells. The low cost and compatibility with commercial systems could also make multiplexed FLIM more accessible to biological research community. PMID:24921725

  12. Confocal microscopy imaging of solid tissue

    EPA Science Inventory

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

  13. Comparative Evaluation Between Ranibizumab Combined with Laser and Bevacizumab Combined with Laser Versus Laser Alone for Macular Oedema Secondary to Branch Retinal Vein Occlusion

    PubMed Central

    Azad, Shorya Vardhan; Salman, Amjad; Mahajan, Deepankur; Sain, Siddharth; Azad, Rajvardhan

    2014-01-01

    Purpose: To evaluate the anatomical and functional efficacy of combination therapy of intravitreal ranibizumab with laser or intravitreal bevacizumab with laser treatment compared to only laser treatment for macular edema due to branch retinal vein occlusion (BRVO). Materials and Methods: Thirty eyes of 30 patients with BRVO of at least 6 weeks duration were randomized into three groups: Group 1 received a single dose of intravitreal Ranibizumab followed by grid laser treatment, Group 2 received a single dose of intravitreal Bevacizumab followed by grid laser treatment, and Group 3 received grid laser alone. Outcomes at 6 months follow-up were reported. Data were collected on best corrected visual acuity (BCVA), central foveal thickness (CFT), and gain in lines of Snellen acuity. Results: At 6 month follow-up, the difference in the mean BCVA and CFT between the three treatment groups was not statistically significant (P > 0.05, all comparisons). Six eyes (60%) in Group 1, four eyes (40%) in Group 2 and two eyes (20%) in Group 3 had a statistically significant gain of ≥3 lines of Snellen acuity (P < 0.05). Conclusion: Both ranibizumab and bevacizumab combined with laser photocoagulation, resulted in better outcomes than grid laser treatment. PMID:25371633

  14. Combined fringe and Fabry-Perot laser anemometer for 3 component velocity measurements in turbine stator cascade facility

    NASA Technical Reports Server (NTRS)

    Seasholtz, Richard G.; Goldman, Louis J.

    1986-01-01

    A laser anemometer is described that was developed for use in a 508 mm diameter annular turbine stator cascade facility. All three velocity components are measured through a single restricted optical port, both within the stator vane row and downstream of the vanes. The measurements are made through a cylindrical window in the casing that matches the tip radius of the cascade. The stator tested has a contoured hub endwall that results in a large radial flow near the hub. The anemometer uses a standard fringe configuration (LFA) with a fluorescent aerosol seed to measure the axial and circumferential velocity components. The radial component is measured with a confocal Fabry-Perot interferometer. The two configurations are combined in a single optical system and can operate simultaneously. Data are presented to illustrate the capabilities of the system.

  15. Combined fringe and Fabry-Perot laser anemometer for three component velocity measurements in turbine stator cascade facility

    NASA Technical Reports Server (NTRS)

    Seasholtz, R. G.; Goldman, L. J.

    1986-01-01

    A laser anemometer is described that was developed for use in a 508 mm diameter annular turbine stator cascade facility. All three velocity components are measured through a single restricted optical port, both within the stator vane row and downstream of the vanes. The measurements are made through a cylindrical window in the casing that matches the tip radius of the cascade. The stator tested has a contoured hub endwall that results in a large radial flow near the hub. The anemometer uses a standard fringe configuration (LFA) with a fluorescent aerosol seed to measure the axial and circumferential velocity components. The radial component is measured with a confocal Fabry-Perot interferometer. The two configurations are combined in a single optical system and can operate simultaneously. Data are presented to illustrate the capabilities of the system.

  16. Confocal simultaneous phase-shifting interferometry

    SciTech Connect

    Zhao Chenguang; Tan Jiubin; Tang Jianbo; Liu Tao; Liu Jian

    2011-02-10

    In order to implement the ultraprecise measurement with large range and long working distance in confocal microscopy, confocal simultaneous phase-shifting interferometry (C-SPSI) has been presented. Four channel interference signals, with {pi}/2 phase shift between each other, are detected simultaneously in C-SPSI. The actual surface height is then calculated by combining the optical sectioning with the phase unwrapping in the main cycle of the interference phase response, and the main cycle is determined using the bipolar property of differential confocal microscopy. Experimental results showed that 1 nm of axial depth resolution was achieved for either low- or high-NA objective lenses. The reflectivity disturbance resistibility of C-SPSI was demonstrated by imaging a typical microcircuit specimen. C-SPSI breaks through the restriction of low NA on the axial depth resolution of confocal microscopy effectively.

  17. High-speed multispectral confocal biomedical imaging

    PubMed Central

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

    2014-01-01

    Abstract. 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. PMID:24658777

  18. Fiber laser microjoining for novel dissimilar material combinations

    NASA Astrophysics Data System (ADS)

    Patwa, R.; Herfurth, H.; Heinemann, S.; Ehrenmann, S.; Newaz, G.; Baird, R. J.

    2009-02-01

    Today's complexity in packaging of MEMS and BioMEMS requires advanced joining techniques that take the specific package integration for each device into account. Current focus on reducing investment and operating costs for device packaging require a flexible and reliable joining approach for similar and dissimilar materials such as metals, polymers, glass and silicon to manage increasing system complexity. Depending on the application, packaged devices must fulfill tough requirements regarding strength, thermal stress, fatigue and hermeticity and long-term stability. This research is focused on laser microjoining of polyimide and PEEK polymers to metals such as nitinol, chromium and titanium using fiber laser. Our earlier investigations have demonstrated the potential of this unique joining technique, which successfully addresses the existing microjoining challenges including high precision, localized processing capability and biocompatibility. Our current study further defines the key processing parameters for joining novel dissimilar material combinations based on the characterization of such laser joints by means of mechanical failure tests and the bond area analysis using optical microscope, scanning electron microscopy (SEM) and X-ray photoelectron spectroscopy (XPS). The results compare operating windows for generating quality bonds for different material joining configurations. They also provide an initial approach to characterize laser-fabricated microjoints that can be potentially used for the optimization of the design process of devices utilizing these materials. Potential packaging applications include microsystems used for chemical or biological assays (lab-on-a-chip), implantable devices used for pressure or temperature sensing, neural stimulation and drug delivery.

  19. Optomechanical design of the grating laser beam combiner (GLBC) laser diode header

    NASA Technical Reports Server (NTRS)

    Rall, Jonathan A. R.; Spadin, Paul L.

    1989-01-01

    A laser diode header has been fabricated for a grating laser beam combiner (GLBC). The laser diode header provides the thermal control, the drive electronics, and the optical system necessary for proper operation of the beam combiner. The diode header is required to provide diffraction limited optical performance while providing correction for worst case defocus aberration, 0.6 mrad excess divergence, and worst case decenter aberration, 1.0 mrad pointing error. The design of the header considered the mechanical design and the optical design together resulting in a small, self-contained header with 0.7 mrad range for focus correction and +/- 2.5 mrad of beam steering. The complete diode header is currently undergoing optical and mechanical performance testing.

  20. Spatial distribution of neutral oxygen vacancies on ZnO nanowire surfaces: An investigation combining confocal microscopy and first principles calculations

    NASA Astrophysics Data System (ADS)

    Mun Wong, Kin; Alay-e-Abbas, S. M.; Fang, Yaoguo; Shaukat, A.; Lei, Yong

    2013-07-01

    A qualitative approach using room-temperature confocal microscopy is employed to investigate the spatial distribution of shallow and deep oxygen vacancy (VO) concentrations on the polar (0001) and non-polar (101¯0) surfaces of zinc oxide (ZnO) nanowires (NWs). Using the spectral intensity variation of the confocal photoluminescence of the green emission at different spatial locations on the surface, the VO concentrations of an individual ZnO NW can be obtained. The green emission at different spatial locations on the ZnO NW polar (0001) and non-polar (101¯0) surfaces is found to have maximum intensity near the NW edges, decreasing to a minimum near the NW center. First-principles calculations using simple supercell-slab (SS) models are employed to approximate/model the defects on the ZnO NW (101¯0) and (0001) surfaces. These calculations give increased insight into the physical mechanism behind the green emission spectral intensity and the characteristics of an individual ZnO NW. The highly accurate density functional theory (DFT)-based full-potential linearized augmented plane-wave plus local orbitals (FP-LAPW + lo) method is used to compute the defect formation energy (DFE) of the SSs. Previously, using these SS models, it was demonstrated through the FP-LAPW + lo method that in the presence of oxygen vacancies at the (0001) surface, the phase transformation of the SSs in the graphite-like structure to the wurtzite lattice structure will occur even if the thickness of the graphite-like SSs are equal to or less than 4 atomic graphite-like layers [Wong et al., J. Appl. Phys. 113, 014304 (2013)]. The spatial profile of the neutral VO DFEs from the DFT calculations along the ZnO [0001] and [101¯0] directions is found to reasonably explain the spatial profile of the measured confocal luminescence intensity on these surfaces, leading to the conclusion that the green emission spectra of the NWs likely originate from neutral oxygen vacancies. Another significant

  1. Multidepth imaging by chromatic dispersion confocal microscopy

    NASA Astrophysics Data System (ADS)

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

    2012-03-01

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

  2. Double-label confocal laser-scanning microscopy, image restoration, and real-time three-dimensional reconstruction to study axons in the central nervous system and their contacts with target neurons.

    PubMed

    Wouterlood, Floris G; van Haeften, Theo; Blijleven, Nico; Pérez-Templado, Pepa; Pérez-Templado, Helena

    2002-03-01

    The current double tracing-double confocal laser-scanning method was developed to reconstruct identified nerve fibers and their contacts with identified target neurons in the rat brain in three dimensions. It intends to fill the gap between conventional light microscopic and electron microscopic neuroanatomic tracing. The steps involved are as follows: (1) injection of two neuroanatomic tracers--Phaseolus vulgaris leucoagglutinin (PHA-L) to label fibers innervating a particular brain area and Neurobiotin to label prospective target neurons in that area; (2) immunofluorescence detection of the labeled fibers (fluorophore Cy5, infrared emission), together with fluorochromated avidin detection of the taken-up Neurobiotin (Cy2 or Alexa 488; green emission); (3) acquisition of Z-series of confocal images at high magnification with a laser-scanning microscope using the laser lines 488 nm and 647 nm; and (4) computer-processing and three-dimensional reconstruction of the labeled fibers and the presumed target dendrites. Rotation on the computer of the three-dimensional reconstructed fibers and dendrites along all three spatial axes enabled the authors to determine whether "true" or "false" contacts occur. In a true contact no space was present between the apposing structures, whereas a false contact consisted of two differently stained structures close to each other but separated by a narrow, optically empty space. One important phenomenon in the three-dimensional reconstruction of double-stained structures that needed correction was "twin image mismatch"--i.e., the observation that a three-dimensional reconstruction of a small test object (double-stained on purpose) produced two slightly shifted objects, each associated with its particular fluorochrome. To measure the actual twin image mismatch of the confocal instrument and to obtain accurate correction factors the authors took in each session in which they obtained image series of the real experiments, with both laser

  3. Manufacturing of Medical Implants by Combination of Selective Laser Melting and Laser Ablation

    NASA Astrophysics Data System (ADS)

    Hallmann, S.; Glockner, P.; Daniel, C.; Seyda, V.; Emmelmann, C.

    2015-09-01

    The perfect fit of hip stem prostheses is supposed to have positive effects on their lifetime performance. Moreover, the ingrowth of tissue into the surface of the implant has to be assured to create a firm and load bearing contact. For the manufacturing of customized hip stem prostheses, the technology of Selective Laser Melting has shown promising results. Poor surface quality, however, makes it necessary to finish up the part by e.g., sand blasting or polishing. With the use of laser ablation for post-processing, reproducible and functionalized surface morphologies might be achievable. Hence, with the motive to produce customized hip stem prostheses, a combined process chain for both mentioned laser technologies is developed. It is examined what type of surface should be produced at which part of the process chain. The produced implants should contain the demanded final surface characteristics without any conventional post-processing. Slight advantages for the Selective Laser Melting regarding the accuracy for different geometrical structures of 400 μm depth were observed. However, an overall improvement of surface quality after the laser ablation process in terms of osseointegration could be achieved. A complete laser based production of customized hip stem implants is found to be with good prospects.

  4. Combination of Er:YAG laser and CO2 laser treatment on skin tissue.

    PubMed

    Anayb Baleg, Sana Mohammed; Bidin, Noriah; Suan, Lau Pik; Sidi Ahmad, Muhammad Fakarruddin; Krishnan, Ganesan; Johari, Abd Rahman; Hamid, Asmah

    2015-01-01

    Skin is the most important organ in our body, as it protects us from external environmental effects. Study the ability of the skin to stretch and the histological examinations of irradiated tissues have significant values in scientific and medical applications. Only a few studies have been done to study the correlation between epidermis ablation and the changes that occur at dermal levels when using dual lasers in ablative resurfacing mode. The aim of this work is to determine this correlation and to estimate the effects of multiple pulses on induced collagen remodeling and the strength of skin exposed with dual lasers in an in vivo rat model. All laser exposures led to mark improvement in the skin's strength compared to their own controls. The histological investigation indicated that there was a thickness loss in the epidermis layer with the induction of deep collagen coagulation in the dermis layer as the dual laser pulses increased. Additionally, more collagen fibers were remolded in the treated samples by dual wavelengths. We conclude that by combining dual lasers with multiple pulses targeted at not only the epidermis layer of the skin, it could also induce some heat diffusion in the dermis layer which causes more coagulation of collagen fibers. The tensile results confirmed by our histological data demonstrate that the strength of irradiated skin with dual wavelengths increased more than using both lasers separately on the skin tissue since more collagen is induced. PMID:25327511

  5. Confocal endomicroscopy of the larynx

    NASA Astrophysics Data System (ADS)

    Just, T.; Wiechmann, T.; Stachs, O.; Stave, J.; Guthoff, R.; Hüttmann, G.; Pau, H. W.

    2012-02-01

    Beside the good image quality with the confocal laser scanning microscope (HRTII) and the Rostock Cornea Module (RCM), this technology can not be used to investigate the human larynx in vivo. To accomplish this, a rigid custom-made endoscope (KARL STORZ GmbH & Co. KG; Tuttlingen Germany) was developed. A connector was developed to connect the scanner head of the HRTII to the rigid endoscope. With the connector, the starting plane can be set manually. To achieve optical sectioning of the laryngeal tissue (80 μm per volume scan), the scanning mechanism of the HRTII needs to be activated using a foot switch. The devices consisting of the endoscope, HRTII, and the connector supply images of 400 x 400 μm and reach average penetration depths of 100-300 μm (λ/4 plate of the scanner head of the HRTII was removed). The lateral and axial resolutions are about 1-2 μm and 2 μm, respectively. In vivo rigid confocal endoscopy is demonstrated with an acquisition time for a volume scan of 6 s. The aim of this study was to differentiate pre-malignant laryngeal lesions from micro-invasive carcinoma of the larynx. 22 patients with suspicious lesions of the true vocal cords were included. This pilot study clearly demonstrates the possibility to detect dysplastic cells close to the basal cell layer and within the subepithelial space in lesions with small leukoplakia (thin keratin layer). These findings may have an impact on microlaryngoscopy to improve the precision for biopsy and on microlaryngoscopic laser surgery of the larynx to identify the margins of the pre-malignant lesion.

  6. Confocal Imaging of porous media

    NASA Astrophysics Data System (ADS)

    Shah, S.; Crawshaw, D.; Boek, D.

    2012-12-01

    Carbonate rocks, which hold approximately 50% of the world's oil and gas reserves, have a very complicated and heterogeneous structure in comparison with sandstone reservoir rock. We present advances with different techniques to image, reconstruct, and characterize statistically the micro-geometry of carbonate pores. The main goal here is to develop a technique to obtain two dimensional and three dimensional images using Confocal Laser Scanning Microscopy. CLSM is used in epi-fluorescent imaging mode, allowing for the very high optical resolution of features well below 1μm size. Images of pore structures were captured using CLSM imaging where spaces in the carbonate samples were impregnated with a fluorescent, dyed epoxy-resin, and scanned in the x-y plane by a laser probe. We discuss the sample preparation in detail for Confocal Imaging to obtain sub-micron resolution images of heterogeneous carbonate rocks. We also discuss the technical and practical aspects of this imaging technique, including its advantages and limitation. We present several examples of this application, including studying pore geometry in carbonates, characterizing sub-resolution porosity in two dimensional images. We then describe approaches to extract statistical information about porosity using image processing and spatial correlation function. We have managed to obtain very low depth information in z -axis (~ 50μm) to develop three dimensional images of carbonate rocks with the current capabilities and limitation of CLSM technique. Hence, we have planned a novel technique to obtain higher depth information to obtain high three dimensional images with sub-micron resolution possible in the lateral and axial planes.

  7. Combined Endoscopic Optical Coherence Tomography and Laser Induced Fluorescence

    NASA Astrophysics Data System (ADS)

    Barton, Jennifer K.; Tumlinson, Alexandre R.; Utzinger, Urs

    Optical coherence tomography (OCT) and laser-induced fluorescence (LIF) are promising modalities for tissue characterization in human patients and animal models. OCT detects coherently backscattered light, whereas LIF detects fluorescence emission of endogenous biochemicals, such as reduced nicotinamide adenine dinucleotide (NADH), flavin adenine dinucleotide (FAD), collagen, and fluorescent proteins, or exogenous substances such as cyanine dyes. Given the complementary mechanisms of contrast for OCT and LIF, the combination of the two modalities could potentially provide more sensitive and specific detection of disease than either modality alone. Sample probes for both OCT and LIF can be implemented using small diameter optical fibers, suggesting a particular synergy for endoscopic applications. In this chapter, the mechanisms of contrast and diagnostic capability for both OCT and LIF are briefly examined. Evidence of complementary capability is described. Example published combined OCT-LIF systems are reviewed, one successful commercial instrument is discussed, and example applications are provided.

  8. Laser beam joining of material combinations for automotive applications

    NASA Astrophysics Data System (ADS)

    Schubert, Emil; Zerner, Ingo; Sepold, Gerd

    1997-08-01

    An ideal material for automotive applications would combine the following properties: high corrosion resistance, high strength, high stiffness and not at least a low material price. Today a single material is not able to meet all these requirements. Therefore, in the future different materials will be placed where they meet the requirements best. The result of this consideration is a car body with many different alloys and metals, which have to be joined to one another. BIAS is working on the development of laser based joining technologies for different material combinations, especially for thin sheets used in automotive applications. One result of the research is a joining technology for an aluminum-steel-joint. Using a Nd:YAG laser the problem of brittle intermetallic phases between these materials was overcome. Using suitable temperature-time cycles, elected by a FEM-simulation, the thickness of intermetallic phases was kept below 10 micrometers . This technology was also applied to coated steels, which were joined with different aluminum alloys. Further it is demonstrated that titanium alloys, e.g. used for racing cars, can also be joined with aluminum alloys.

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

    PubMed Central

    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

  10. Analysis of doxorubicin distribution in MCF-7 cells treated with drug-loaded nanoparticles by combination of two fluorescence-based techniques, confocal spectral imaging and capillary electrophoresis.

    PubMed

    Gautier, Juliette; Munnier, Emilie; Soucé, Martin; Chourpa, Igor; Douziech Eyrolles, Laurence

    2015-05-01

    The intracellular distribution of the antiancer drug doxorubicin (DOX) was followed qualitatively by fluorescence confocal spectral imaging (FCSI) and quantitatively by capillary electrophoresis (CE). FCSI permits the localization of the major fluorescent species in cell compartments, with spectral shifts indicating the polarity of the respective environment. However, distinction between drug and metabolites by FCSI is difficult due to their similar fluorochromes, and direct quantification of their fluorescence is complicated by quantum yield variation between different subcellular environments. On the other hand, capillary electrophoresis with fluorescence detection (CE-LIF) is a quantitative method capable of separating doxorubicin and its metabolites. In this paper, we propose a method for determining drug and metabolite concentration in enriched nuclear and cytosolic fractions of cancer cells by CE-LIF, and we compare these data with those of FCSI. Significant differences in the subcellular distribution of DOX are observed between the drug administered as a molecular solution or as a suspension of drug-loaded iron oxide nanoparticles coated with polyethylene glycol. Comparative analysis of the CE-LIF vs FCSI data may lead to a tentative calibration of this latter method in terms of DOX fluorescence quantum yields in the nucleus and more or less polar regions of the cytosol. PMID:25749791

  11. Combining femtosecond laser ablation and diode laser welding in lamellar and endothelial corneal transplants

    NASA Astrophysics Data System (ADS)

    Pini, Roberto; Rossi, Francesca; Matteini, Paolo; Ratto, Fulvio; Menabuoni, Luca; Lenzetti, Ivo; Yoo, Sonia H.; Parel, Jean-Marie

    2008-02-01

    Based on our previous clinical experiences in minimally invasive diode laser-induced welding of corneal tissue in penetrating keratoplasty (PK), i.e. full-thickness transplant of the cornea, we combined this technique with the use of a femtosecond laser for applications in lamellar (LK) and endothelial (EK) keratoplasty. In LK, the femtosecond laser was used to prepare donor button and recipient corneal bed; the wound edges were stained with a water solution of Indocyanine Green (ICG) and then irradiated with a diode laser emitting in CW mode to induce stromal welding. Intraoperatory observations and follow-up results up to 6 months indicated the formation of a smooth stromal interface, total absence of edema as well as inflammation, and reduction of post-operative astigmatism, as compared with conventional suturing procedures. In EK the femtosecond laser was used for the preparation of a 100 μm thick, 8.5mm diameter donor corneal endothelium flap. The flap stromal side was stained with ICG. After stripping the recipient Descemet's membrane and endothelium, the donor flap was positioned in the anterior chamber on the inner face of the cornea by an air bubble and secured to the recipient cornea by diode laser pulses delivered by means of a fiberoptic contact probe introduced in the anterior chamber, which produced welding spots of 200 μm diameter. Femtosecond laser sculpturing of the donor cornea provided lamellar and endothelial flaps of preset and constant thickness. Diode laserinduced welding showed a unique potential to permanently secure the donor flap in place, avoiding postoperative displacement and inflammation reaction.

  12. Synchrotron radiation as a light source in confocal microscopy of biological processes

    NASA Astrophysics Data System (ADS)

    Gerritsen, Hans C.; van der Oord, C. J. R.; Levine, Yehudi K.; Munro, Ian H.; Myring, Wendy J.; Shaw, D. A.; Rommerts, Fokko F.

    1992-04-01

    A novel confocal microscope is presented using the Daresbury Synchrotron Radiation source as its light source. The broad spectrum of synchrotron radiation in combination with the UV compatible microscope allows the extension of confocal microscopy from the visible to the UV region down to about 200 nm. It is envisaged that structures separated by about 70 nm can be resolved at a wavelength of 200 nm. In addition, the tunability of synchrotron radiation affords the selective excitation of any specific fluorescent molecule at the maximum of the absorption band. This avoids the restriction of working at fixed laser lines. A further advantage of using synchrotron radiation is the realization of multiwavelength excitation. Test results using laser systems in the visible and in the UV are presented. Fluorescence images of test targets using UV excitation reveal the superior resolution of the microscope. Furthermore, images of Leydig cells incubated with a fluorescent cholesterol derivative whose maximum of absorption is at 325 nm are shown. These images cannot be produced by conventional confocal laser microscopes. Finally, promising preliminary results obtained with synchrotron radiation are presented.

  13. Confocal multiview light-sheet microscopy

    PubMed Central

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

    2015-01-01

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

  14. Combined laser and photodynamic treatment in extensive purulent wounds

    NASA Astrophysics Data System (ADS)

    Solovieva, A. B.; Tolstih, P. I.; Melik-Nubarov, N. S.; Zhientaev, T. M.; Kuleshov, I. G.; Glagolev, N. N.; Ivanov, A. V.; Karahanov, G. I.; Tolstih, M. P.; Timashev, P. S.

    2010-05-01

    Recently, photodynamic therapy (PDT) has been used for the treatment of festering wounds and trophic ulcers. An important advantage of PDT is its ability to affect bacterial cultures that are resistant to antibiotics. However the use of PDT alone does not usually guarantee a stable antiseptic effect and cannot prevent an external infection of wounds and burns. In this work attention is focused on the healing of the extensive soft tissues wounds with combined laser therapy (LT) and PDT treatment. At the first stage of this process festering tissues (for example spacious purulent wounds with area more than 100 cm2) were illuminated with high-energy laser beam (with power 20 W) in continues routine. The second stage involves “softer” PDT affect, which along with the completion stages of destruction pathological cells, stimulating the process of wound granulation and epithelization. Also, according to our previous results, photosensitizer (photoditazin) is introduced inside the wound with different amphiphilic polymers for increasing the PDT efficacy.

  15. Image inpainting for the differential confocal microscope

    NASA Astrophysics Data System (ADS)

    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.

  16. CONFOCAL MICROSCOPY SYSTEM PERFORMANCE: SPECTROSCOPY AND FOUNDATIONS FOR QUANTITATION

    EPA Science Inventory

    The confocal laser-scanning microscope (CLSM) has enormous potential in many biological fields. The reliability of the CLSM to obtain specific measurements and quantify fluorescence data is dependent on using a correctly aligned machine that contains a stable laser power. For man...

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

    PubMed Central

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

    2014-01-01

    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. PMID:24847258

  18. Scanning computed confocal imager

    DOEpatents

    George, John S.

    2000-03-14

    There is provided a confocal imager comprising a light source emitting a light, with a light modulator in optical communication with the light source for varying the spatial and temporal pattern of the light. A beam splitter receives the scanned light and direct the scanned light onto a target and pass light reflected from the target to a video capturing device for receiving the reflected light and transferring a digital image of the reflected light to a computer for creating a virtual aperture and outputting the digital image. In a transmissive mode of operation the invention omits the beam splitter means and captures light passed through the target.

  19. Confocal imaging of ionised calcium in living plant cells.

    PubMed

    Williams, D A; Cody, S H; Gehring, C A; Parish, R W; Harris, P J

    1990-04-01

    Laser-scanning confocal microscopy has been used in conjunction with Fluo-3, a highly fluorescent visible wavelength probe for Ca2+, to visualize Ca2(+)-dynamics in the function of living plant cells. This combination has overcome many of the problems that have limited the use of fluorescence imaging techniques in the study of the role of cations (Ca2+ and H+) in plant cell physiology and enables these processes to be studied in single cells within intact plant tissue preparations. Maize coleoptiles respond to application of ionophores and plant growth hormones with elevations in cytosolic Ca2+ that can be resolved with a high degree of spatial resolution and can be interpreted quantitatively. PMID:2113832

  20. In vivo Imaging of Tumor Angiogenesis using Fluorescence Confocal Videomicroscopy

    PubMed Central

    Fitoussi, Victor; Faye, Nathalie; Chamming's, Foucauld; Clement, Olivier; Cuenod, Charles-Andre; Fournier, Laure S.

    2013-01-01

    Fibered confocal fluorescence in vivo imaging with a fiber optic bundle uses the same principle as fluorescent confocal microscopy. It can excite fluorescent in situ elements through the optical fibers, and then record some of the emitted photons, via the same optical fibers. The light source is a laser that sends the exciting light through an element within the fiber bundle and as it scans over the sample, recreates an image pixel by pixel. As this scan is very fast, by combining it with dedicated image processing software, images in real time with a frequency of 12 frames/sec can be obtained. We developed a technique to quantitatively characterize capillary morphology and function, using a confocal fluorescence videomicroscopy device. The first step in our experiment was to record 5 sec movies in the four quadrants of the tumor to visualize the capillary network. All movies were processed using software (ImageCell, Mauna Kea Technology, Paris France) that performs an automated segmentation of vessels around a chosen diameter (10 μm in our case). Thus, we could quantify the 'functional capillary density', which is the ratio between the total vessel area and the total area of the image. This parameter was a surrogate marker for microvascular density, usually measured using pathology tools. The second step was to record movies of the tumor over 20 min to quantify leakage of the macromolecular contrast agent through the capillary wall into the interstitium. By measuring the ratio of signal intensity in the interstitium over that in the vessels, an 'index leakage' was obtained, acting as a surrogate marker for capillary permeability. PMID:24056503

  1. In vivo imaging of tumor angiogenesis using fluorescence confocal videomicroscopy.

    PubMed

    Fitoussi, Victor; Faye, Nathalie; Chamming's, Foucauld; Clement, Olivier; Cuenod, Charles-Andre; Fournier, Laure S

    2013-01-01

    Fibered confocal fluorescence in vivo imaging with a fiber optic bundle uses the same principle as fluorescent confocal microscopy. It can excite fluorescent in situ elements through the optical fibers, and then record some of the emitted photons, via the same optical fibers. The light source is a laser that sends the exciting light through an element within the fiber bundle and as it scans over the sample, recreates an image pixel by pixel. As this scan is very fast, by combining it with dedicated image processing software, images in real time with a frequency of 12 frames/sec can be obtained. We developed a technique to quantitatively characterize capillary morphology and function, using a confocal fluorescence videomicroscopy device. The first step in our experiment was to record 5 sec movies in the four quadrants of the tumor to visualize the capillary network. All movies were processed using software (ImageCell, Mauna Kea Technology, Paris France) that performs an automated segmentation of vessels around a chosen diameter (10 μm in our case). Thus, we could quantify the 'functional capillary density', which is the ratio between the total vessel area and the total area of the image. This parameter was a surrogate marker for microvascular density, usually measured using pathology tools. The second step was to record movies of the tumor over 20 min to quantify leakage of the macromolecular contrast agent through the capillary wall into the interstitium. By measuring the ratio of signal intensity in the interstitium over that in the vessels, an 'index leakage' was obtained, acting as a surrogate marker for capillary permeability. PMID:24056503

  2. Quantitative phase-contrast confocal microscope

    PubMed Central

    Liu, Changgeng; Marchesini, Stefano; Kim, Myung K.

    2014-01-01

    We present a quantitative phase-contrast confocal microscope (QPCCM) by combining a line-scanning confocal system with digital holography (DH). This combination can merge the merits of these two different imaging modalities. High-contrast intensity images with low coherent noise, and the optical sectioning capability are made available due to the confocality. Phase profiles of the samples become accessible thanks to DH. QPCCM is able to quantitatively measure the phase variations of optical sections of the opaque samples and has the potential to take high-quality intensity and phase images of non-opaque samples such as many biological samples. Because each line scan is recorded by a hologram that may contain the optical aberrations of the system, it opens avenues for a variety of numerical aberration compensation methods and development of full digital adaptive optics confocal system to emulate current hardware-based adaptive optics system for biomedical imaging, especially ophthalmic imaging. Preliminary experiments with a microscope objective of NA 0.65 and 40 × on opaque samples are presented to demonstrate this idea. The measured lateral and axial resolutions of the intensity images from the current system are ~0.64μm and ~2.70μm respectively. The noise level of the phase profile by QPCCM is ~2.4nm which is better than the result by DH. PMID:25089404

  3. An active alignment method for post launch co-alignment of laser beam combiner systems

    NASA Technical Reports Server (NTRS)

    Yu, A. W.; Green, J. W.; Maynard, W. L.; Minott, P. O.; Krainak, M. A.

    1992-01-01

    A laser transmitter for high bandwidth geosynchronous satellite communications is described. High optical power is achieved by combining semiconductor laser diodes. An active alignment scheme is proposed for achieving the +/- 20 microrad post launch multiple laser angular co-alignment requirement.

  4. Invasive leg vein treatment with 1064/1319 Nd:YAG laser: combination with dye laser treatment

    NASA Astrophysics Data System (ADS)

    Smucler, Roman; Horak, Ladislav; Mazanek, Jiri

    1999-06-01

    More than 2 500 leg veins patients were treated with dye laser / ScleroPlus, Candela, USA / successfully in our clinic and we use this therapy as the basic cosmetics treatment. But especially diameter of leg vein is limiting factor. Very often we have to treat some cases that are not ideal for classical surgical or for dye laser method. We decided to make invasive perivenous laser coagulation. We adapted original Czech 1064/1319 nm Nd:YAG laser / US patent pending /, which is new combine tool, for invasive application. Principe: After we have penetrated the cutis with laser fiber we coagulate leg veins during slowly perivenous motion. Perfect preoperative examination is a condition of success. After 15 months we have very interesting results. Some patients / 15%/ were perfect treated only with this possibility but excellent results are acquired from combination with dye laser.

  5. Enhanced confocal microscopy and ophthalmoscopy with polarization imaging

    NASA Astrophysics Data System (ADS)

    Campbell, Melanie C. W.; Bueno, Juan M.; Cookson, Christopher J.; Liang, Qingyuan; Kisilak, Marsha L.; Hunter, Jennifer J.

    2005-09-01

    We previously developed a Mueller matrix formalism to improve confocal imaging in microscopes and ophthalmoscopes. Here we describe a procedure simplified by firstly introducing a generator of polarization states in the illumination pathway of a confocal scanning laser microscope and secondly computing just four elements of the Mueller matrix of any sample and instrument combination. Using a subset of Mueller matrix elements, the best images are reconstructed. The method was tested for samples with differing properties (specular, diffuse and partially depolarizing). Images were also studied of features at the rear of the eye. The best images obtained with this technique were compared to the original images and those obtained from frame averaging. Images corresponding to non-polarized incident light were also computed. For all cases, the best reconstructed images were of better quality than both the original and frame-averaged images. The best reconstructed images also showed an improvement compared with the images corresponding to non polarized light. This methodology will have broad application in biomedical imaging.

  6. Exploring the diversity of Listeria monocytogenes biofilm architecture by high-throughput confocal laser scanning microscopy and the predominance of the honeycomb-like morphotype.

    PubMed

    Guilbaud, Morgan; Piveteau, Pascal; Desvaux, Mickaël; Brisse, Sylvain; Briandet, Romain

    2015-03-01

    Listeria monocytogenes is involved in food-borne illness with a high mortality rate. The persistence of the pathogen along the food chain can be associated with its ability to form biofilms on inert surfaces. While most of the phenotypes associated with biofilms are related to their spatial organization, most published data comparing biofilm formation by L. monocytogenes isolates are based on the quantitative crystal violet assay, which does not give access to structural information. Using a high-throughput confocal-imaging approach, the aim of this work was to decipher the structural diversity of biofilms formed by 96 L. monocytogenes strains isolated from various environments. Prior to large-scale analysis, an experimental design was created to improve L. monocytogenes biofilm formation in microscopic-grade microplates, with special emphasis on the growth medium composition. Microscopic analysis of biofilms formed under the selected conditions by the 96 isolates revealed only weak correlation between the genetic lineages of the isolates and the structural properties of the biofilms. However, a gradient in their geometric descriptors (biovolume, mean thickness, and roughness), ranging from flat multilayers to complex honeycomb-like structures, was shown. The dominant honeycomb-like morphotype was characterized by hollow voids hosting free-swimming cells and localized pockets containing mixtures of dead cells and extracellular DNA (eDNA). PMID:25548046

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

    NASA Astrophysics Data System (ADS)

    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.

  8. Richardson-Lucy algorithm with total variation regularization for 3D confocal microscope deconvolution.

    PubMed

    Dey, Nicolas; Blanc-Feraud, Laure; Zimmer, Christophe; Roux, Pascal; Kam, Zvi; Olivo-Marin, Jean-Christophe; Zerubia, Josiane

    2006-04-01

    Confocal laser scanning microscopy is a powerful and popular technique for 3D imaging of biological specimens. Although confocal microscopy images are much sharper than standard epifluorescence ones, they are still degraded by residual out-of-focus light and by Poisson noise due to photon-limited detection. Several deconvolution methods have been proposed to reduce these degradations, including the Richardson-Lucy iterative algorithm, which computes maximum likelihood estimation adapted to Poisson statistics. As this algorithm tends to amplify noise, regularization constraints based on some prior knowledge on the data have to be applied to stabilize the solution. Here, we propose to combine the Richardson-Lucy algorithm with a regularization constraint based on Total Variation, which suppresses unstable oscillations while preserving object edges. We show on simulated and real images that this constraint improves the deconvolution results as compared with the unregularized Richardson-Lucy algorithm, both visually and quantitatively. PMID:16586486

  9. Three-dimensional reconstruction of topological deformation in chiral nematic microspheres using fluorescence confocal polarizing microscopy.

    PubMed

    Guo, Jin-Kun; Song, Jang-Kun

    2016-04-01

    Chiral nematic droplets exhibit abundant topological defect structures, which have been intensively studied, both theoretically and experimentally. However, to observe and reconstruct the exact shape of three-dimensional (3D) defect structures has been a challenging task. In this study, we successfully reconstruct the 3D defect structures within a CLC microsphere with long helical pitches by combining polarized optical microscopy (POM) and laser scanning type fluorescence confocal polarizing microscopy (FCPM). The obtained confocal stack images provide us with the vertical location of disclination defects, to allow reconstruction of the full 3D structures. The reconstructed 3D structures can be viewed from different directions, providing a better understanding of the topological structure. Moreover, the defect lines are identified to be + 1 defects, different from the previous prediction. Thus, FCPM provides an excellent tool to study the complex topological configuration in microspheres, and fosters its potential applicability in new devices based on topologically structured soft media. PMID:27137028

  10. Optical sectioning using a digital Fresnel incoherent-holography-based confocal imaging system

    PubMed Central

    Kelner, Roy; Katz, Barak; Rosen, Joseph

    2015-01-01

    We propose a new type of confocal microscope using Fresnel incoherent correlation holography (FINCH). Presented here is a confocal configuration of FINCH using a phase pinhole and point illumination that is able to suppress out-of-focus information from the recorded hologram and hence combine the super-resolution capabilities of FINCH with the sectioning capabilities of confocal microscopy. PMID:26413560

  11. Laser-induced breakdown spectroscopy combined with spatial heterodyne spectroscopy.

    PubMed

    Gornushkin, Igor B; Smith, Ben W; Panne, Ulrich; Omenetto, Nicoló

    2014-01-01

    A spatial heterodyne spectrometer (SHS) is tested for the first time in combination with laser-induced breakdown spectroscopy (LIBS). The spectrometer is a modified version of the Michelson interferometer in which mirrors are replaced by diffraction gratings. The SHS contains no moving parts and the gratings are fixed at equal distances from the beam splitter. The main advantage is high throughput, about 200 times higher than that of dispersive spectrometers used in LIBS. This makes LIBS-SHS a promising technique for low-light standoff applications. The output signal of the SHS is an interferogram that is Fourier-transformed to retrieve the original plasma spectrum. In this proof-of-principle study, we investigate the potential of LIBS-SHS for material classification and quantitative analysis. Brass standards with broadly varying concentrations of Cu and Zn were tested. Classification via principal component analysis (PCA) shows distinct groupings of materials according to their origin. The quantification via partial least squares regression (PLS) shows good precision (relative standard deviation < 10%) and accuracy (within ± 5% of nominal concentrations). It is possible that LIBS-SHS can be developed into a portable, inexpensive, rugged instrument for field applications. PMID:25226262

  12. Withania somnifera prevents morphine withdrawal-induced decrease in spine density in nucleus accumbens shell of rats: a confocal laser scanning microscopy study.

    PubMed

    Kasture, Sanjay; Vinci, Stefania; Ibba, Federico; Puddu, Alessandro; Marongiu, Mara; Murali, Balasubramanian; Pisanu, Augusta; Lecca, Daniele; Zernig, Gerald; Acquas, Elio

    2009-11-01

    Opiate withdrawal is associated with morphological changes of dopamine neurons in the ventral tegmental area and with reduction of spine density of second-order dendrites of medium size spiny neurons in the nucleus accumbens shell but not core. Withania somnifera has long been used in the Middle East, Africa, and India as a remedy for different conditions and diseases and a growing body of evidence points to its beneficial effects on a number of experimental models of neurological disorders. Recently, many studies focused on the potential neuritic regeneration and synaptic reconstruction properties of its methanolic extract and its constituents (withanolides). This study investigates whether morphine withdrawal-induced spine reduction in the nucleus accumbens is affected by the administration of a Withania somnifera extract. To this end, rats were chronically treated with Withania somnifera extract along with morphine or saline and, upon spontaneous (1 and 3 days) or pharmacologically precipitated withdrawal, their brains were fixed in Golgi-Cox stain for confocal microscopic examination. In a separate group of animals, Withania somnifera extract was administered during three days of spontaneous withdrawal. Withania somnifera extract treatment reduced the severity of the withdrawal syndrome when given during chronic morphine but not during withdrawal. In addition, treatment with Withania somnifera extract during chronic morphine, but not during withdrawal, fully prevented the reduction of spine density in the nucleus accumbens shell in spontaneous and pharmacologically precipitated morphine withdrawal. These results indicate that pretreatment with Withania somnifera extract protects from the structural changes induced by morphine withdrawal potentially providing beneficial effects on the consequences related to this condition. PMID:19551457

  13. Diagnostic values of dual focus narrow band imaging and probe-based confocal laser endomicroscopy in FAP-related duodenal adenoma

    PubMed Central

    Pittayanon, Rapat; Rerknimitr, Rungsun; Imraporn, Boonlert; Wisedopas, Naruemon; Kullavanijaya, Pinit

    2015-01-01

    Background and study aims: Familial adenomatous polyposis (FAP) is associated with an increased risk of development of periampullary and nonampullary adenoma. Either routine biopsy or endoscopic removal of the lesion is generally required to identify the presence of adenoma. Because the risk of tissue sampling from the ampulla is high and nonampullary polyps are sometimes numerous, resection of all the lesions is time-consuming. This study aimed to evaluate the diagnostic values of duodenal adenoma by dual focus NBI (dNBI) and probe-based confocal endomicroscopy (pCLE) in FAP patients. Patients and methods: The authors conducted a diagnostic study in a single tertiary-care referral center. Surveillance esophagogastroduodenoscopy with dNBI and pCLE was performed on 26 patients with FAP for real-time adenoma diagnosis by two different endoscopists; one used dNBI and the other pCLE. Histology from the matched lesion was used as the gold standard. Results: A total of 55 matched biopsies (25 ampullas, 30 nonampullas) were performed. The sensitivity, specificity, post predictive value (PPV), negative predictive value (NPV), and accuracy of dNBI vs. pCLE from all duodenal lesions were 96.9 % vs. 93.8 %, 78.3 % vs. 81 %, 86.1 % vs. 88.2 %, 94.7 vs. 89.5 %, and 92.4 % vs. 88.6 %, respectively. Conclusions: For surveillance of periampullary and nonampullary adenoma in patients with FAP, the real-time readings provided a high degree of diagnostic value when histology was used as the gold standard. (Clinical trial registration number: NCT02162173). PMID:26528500

  14. Multiply scattered light tomography and confocal imaging: detecting neovascularization in age-related macular degeneration

    NASA Astrophysics Data System (ADS)

    Elsner, Ann E.; Miura, Masahiro; Burns, Stephen Allan; Beausencourt, E.; Kunze, C.; Kelley, L. M.; Walker, J. P.; Wing, G. L.; Raskauskas, P. A.; Fletcher, D. C.; Zhou, Qienyuan; Dreher, Andreas W.

    2000-07-01

    A novel technique, Multiply Scattered Light Tomography (MSLT), and confocal Infrared Imaging are used to provide diagnostic information using a comfortable, rapid, and noninvasive method. We investigated these techniques in detecting neovascularization in age-related macular degeneration. The MSLT used a Vertical Cavity Surface Emitting Laser (VCSEL) at 850 nm, while the confocal imaging technique used either the VCSEL or a 790 nm laser diode. Both were implemented into the topographical scanning system (TopSS, Laser Diagnostic Technologies, Inc.) Confocal imaging with both lasers provided different information about neovascularization as a function of focal plane, and different also from MSLT.

  15. A survey of beam-combining technologies for laser space power transmission

    NASA Technical Reports Server (NTRS)

    Kwon, J. H.; Williams, M. D.; Lee, J. H.

    1988-01-01

    The combination of laser beams holds much promise for obtaining powerful beams. Methods are surveyed for beam combination (coherent and incoherent) and two of them are identified as the most effective means for achieving high power transmission in space. The two methods as applied to laser diode arrays are analyzed, and potentially productive work areas for the advancement of technology are delineated.

  16. Hyperspectral confocal microscope

    NASA Astrophysics Data System (ADS)

    Sinclair, Michael B.; Haaland, David M.; Timlin, Jerilyn A.; Jones, Howland D. T.

    2006-08-01

    We have developed a new, high performance, hyperspectral microscope for biological and other applications. For each voxel within a three-dimensional specimen, the microscope simultaneously records the emission spectrum from 500 nm to 800 nm, with better than 3 nm spectral resolution. The microscope features a fully confocal design to ensure high spatial resolution and high quality optical sectioning. Optical throughput and detection efficiency are maximized through the use of a custom prism spectrometer and a backside thinned electron multiplying charge coupled device (EMCCD) array. A custom readout mode and synchronization scheme enable 512-point spectra to be recorded at a rate of 8300 spectra per second. In addition, the EMCCD readout mode eliminates curvature and keystone artifacts that often plague spectral imaging systems. The architecture of the new microscope is described in detail, and hyperspectral images from several specimens are presented.

  17. Hyperspectral confocal microscope.

    PubMed

    Sinclair, Michael B; Haaland, David M; Timlin, Jerilyn A; Jones, Howland D T

    2006-08-20

    We have developed a new, high performance, hyperspectral microscope for biological and other applications. For each voxel within a three-dimensional specimen, the microscope simultaneously records the emission spectrum from 500 nm to 800 nm, with better than 3 nm spectral resolution. The microscope features a fully confocal design to ensure high spatial resolution and high quality optical sectioning. Optical throughput and detection efficiency are maximized through the use of a custom prism spectrometer and a backside thinned electron multiplying charge coupled device (EMCCD) array. A custom readout mode and synchronization scheme enable 512-point spectra to be recorded at a rate of 8300 spectra per second. In addition, the EMCCD readout mode eliminates curvature and keystone artifacts that often plague spectral imaging systems. The architecture of the new microscope is described in detail, and hyperspectral images from several specimens are presented. PMID:16892134

  18. Confocal coded aperture imaging

    DOEpatents

    Tobin, Jr., Kenneth William; Thomas, Jr., Clarence E.

    2001-01-01

    A method for imaging a target volume comprises the steps of: radiating a small bandwidth of energy toward the target volume; focusing the small bandwidth of energy into a beam; moving the target volume through a plurality of positions within the focused beam; collecting a beam of energy scattered from the target volume with a non-diffractive confocal coded aperture; generating a shadow image of said aperture from every point source of radiation in the target volume; and, reconstructing the shadow image into a 3-dimensional image of the every point source by mathematically correlating the shadow image with a digital or analog version of the coded aperture. The method can comprise the step of collecting the beam of energy scattered from the target volume with a Fresnel zone plate.

  19. A clearer view of the insect brain—combining bleaching with standard whole-mount immunocytochemistry allows confocal imaging of pigment-covered brain areas for 3D reconstruction

    PubMed Central

    Stöckl, Anna L.; Heinze, Stanley

    2015-01-01

    In the study of insect neuroanatomy, three-dimensional (3D) reconstructions of neurons and neuropils have become a standard technique. As images have to be obtained from whole-mount brain preparations, pigmentation on the brain surface poses a serious challenge to imaging. In insects, this is a major problematic in the first visual neuropil of the optic lobe, the lamina, which is obstructed by the pigment of the retina as well as by the pigmented fenestration layer. This has prevented inclusion of this major processing center of the insect visual system into most neuroanatomical brain atlases and hinders imaging of neurons within the lamina by confocal microscopy. It has recently been shown that hydrogen peroxide bleaching is compatible with immunohistochemical labeling in insect brains, and we therefore developed a simple technique for removal of pigments on the surface of insect brains by chemical bleaching. We show that our technique enables imaging of the pigment-obstructed regions of insect brains when combined with standard protocols for both anti-synapsin-labeled as well as neurobiotin-injected samples. This method can be combined with different fixation procedures, as well as different fluorophore excitation wavelengths without negative effects on staining quality. It can therefore serve as an effective addition to most standard histology protocols used in insect neuroanatomy. PMID:26441552

  20. A clearer view of the insect brain-combining bleaching with standard whole-mount immunocytochemistry allows confocal imaging of pigment-covered brain areas for 3D reconstruction.

    PubMed

    Stöckl, Anna L; Heinze, Stanley

    2015-01-01

    In the study of insect neuroanatomy, three-dimensional (3D) reconstructions of neurons and neuropils have become a standard technique. As images have to be obtained from whole-mount brain preparations, pigmentation on the brain surface poses a serious challenge to imaging. In insects, this is a major problematic in the first visual neuropil of the optic lobe, the lamina, which is obstructed by the pigment of the retina as well as by the pigmented fenestration layer. This has prevented inclusion of this major processing center of the insect visual system into most neuroanatomical brain atlases and hinders imaging of neurons within the lamina by confocal microscopy. It has recently been shown that hydrogen peroxide bleaching is compatible with immunohistochemical labeling in insect brains, and we therefore developed a simple technique for removal of pigments on the surface of insect brains by chemical bleaching. We show that our technique enables imaging of the pigment-obstructed regions of insect brains when combined with standard protocols for both anti-synapsin-labeled as well as neurobiotin-injected samples. This method can be combined with different fixation procedures, as well as different fluorophore excitation wavelengths without negative effects on staining quality. It can therefore serve as an effective addition to most standard histology protocols used in insect neuroanatomy. PMID:26441552

  1. Applications of combination wavelength (1060-nm and 530-nm) and pulsed Nd:YAG laser for contact laser surgery.

    PubMed

    Liu, K R; Peyman, G A; Myers, J D; Hamlin, S A; Katoh, N

    1989-01-01

    Two pulsed neodimium yittrium aluminum garnet (Nd:YAG) laser systems were evaluated for contact surgery through a fiberoptic system with a sapphire tip. Pulsed Nd:YAG laser at 1060 nm was as effective as continuous-wave Nd:YAG laser in producing tissue incisions. A combination of 1060-nm and 530-nm wavelengths achieved smooth cutting at lower energy levels. Corneal endothelial cell damage occurred at the high power level (7 watts) required for smooth underwater incisions with both continuous wave and pulsed lasers. PMID:2733255

  2. Maximum permissible exposure of the retina in the human eye in optical coherence tomography systems using a confocal scanning laser ophthalmoscopy platform

    NASA Astrophysics Data System (ADS)

    Rees, Sian; Dobre, George

    2014-01-01

    When using scanning laser ophthalmoscopy to produce images of the eye fundus, maximum permissible exposure (MPE) limits must be considered. These limits are set out in international standards such as the National Standards Institute ANSI Z136.1 Safe Use of Lasers (USA) and BS EN 60825-1: 1994 (UK) and corresponding Euro norms but these documents do not explicitly consider the case of scanned beams. Our study aims to show how MPE values can be calculated for the specific case of retinal scanning by taking into account an array of parameters, such as wavelength, exposure duration, type of scanning, line rate and field size, and how each set of initial parameters results in MPE values that correspond to thermal or photochemical damage to the retina.

  3. Laser Discs, Barcodes, and Books--a Great Combination.

    ERIC Educational Resources Information Center

    Peto, Erica

    1996-01-01

    Describes the use of barcodes to link laser discs with books in school libraries. Highlights include use of a barcode reader as a remote control device as well as a scanner, guidelines for making laser disc books, and a sidebar that explains how to make barcodes and describes software. (LRW)

  4. Any Way You Slice It—A Comparison of Confocal Microscopy Techniques

    PubMed Central

    Jonkman, James

    2015-01-01

    The confocal fluorescence microscope has become a popular tool for life sciences researchers, primarily because of its ability to remove blur from outside of the focal plane of the image. Several different kinds of confocal microscopes have been developed, each with advantages and disadvantages. This article will cover the grid confocal, classic confocal laser-scanning microscope (CLSM), the resonant scanning-CLSM, and the spinning-disk confocal microscope. The way each microscope technique works, the best applications the technique is suited for, the limitations of the technique, and new developments for each technology will be presented. Researchers who have access to a range of different confocal microscopes (e.g., through a local core facility) should find this paper helpful for choosing the best confocal technology for specific imaging applications. Others with funding to purchase an instrument should find the article helpful in deciding which technology is ideal for their area of research. PMID:25802490

  5. High-power beam combining: a step to a future laser weapon system

    NASA Astrophysics Data System (ADS)

    Protz, Rudolf; Zoz, Jürgen; Geidek, Franz; Dietrich, Stephan; Fall, Michael

    2012-11-01

    Due to the enormous progress in the field of high-power fiber lasers during the last years commercial industrial fiber lasers are now available, which deliver a near-diffraction limited beam with power levels up to10kW. For the realization of a future laser weapon system, which can be used for Counter-RAM or similar air defence applications, a laser source with a beam power at the level of 100kW or more is required. At MBDA Germany the concept for a high-energy laser weapon system is investigated, which is based on such existing industrial laser sources as mentioned before. A number of individual high-power fiber laser beams are combined together, using one common beam director telescope. By this "geometric" beam coupling scheme, sufficient laser beam power for an operational laser weapon system can be achieved. The individual beams from the different lasers are steered by servo-loops, using fast tip-tilt mirrors. This principle enables the concentration of the total laser beam power at the common focal point on a distant target, also allowing fine tracking of target movements and first order compensation of turbulence effects on laser beam propagation. The proposed beam combination concept was demonstrated using several experimental set-ups. Different experiments were performed, to investigate laser beam target interaction and target fine tracking also at large distances. Content and results of these investigations are reported. An example for the lay-out of an Air Defence High Energy Laser Weapon (ADHELW ) is given. It can be concluded, that geometric high-power beam combining is an important step for the realization of a laser weapon system in the near future.

  6. Active and passive coherent beam combining of thulium-doped fiber lasers

    NASA Astrophysics Data System (ADS)

    Zhou, Pu; Wang, Xiaolin; Ma, Yanxing; Ma, Haotong; Han, Kai; Xu, Xiaojun; Liu, Zejin

    2010-11-01

    Thulium-doped fiber laser (TFL), which emitted near 2 μm laser beam, has become the latest revolution in highpower fiber laser technology. Further increasing the output power will face great challenges induced by nonlinear effects; coherent beam combining of TFL can increase laser output power while simultaneously maintaining beam quality. In this manuscript, we will present our detailed investigation on coherent beam combining of TFLs. Three different approaches, i.e., interferometric array, mutual injection locking and active phasing based on multi-dithering technique, are employed. In the interferometric array scheme, coherent combining is realized by using an intracavity fiber coupler in an all-fiber laser array configuration. Efficient coherent combining can be achieved by providing sufficient loss discrimination. High combining efficiency of 85% for two fiber laser has been obtained. In mutual injection locking scheme, mutual coherence between the two fiber lasers is established by means of mutual coupling through two 3dB couplers. High combining efficiency of 99% for two fiber laser has been obtained, and the fringe contrast of the intensity pattern at the receiving plane is as high as 93%. In active phasing scheme, when the phase control system is in the closed loop, the fringe contrast of far-field intensity pattern is improved by more than 75 % from 10 % in open loop, and the residual phase error is less than λ/20.

  7. Observations of the incidence of metastasis following laser hyperthermia in combination with chemotherapy, PDT, and excision

    NASA Astrophysics Data System (ADS)

    Wang, Mianjing; Gao, Menglin; Gao, Jin; Xue, Kexun; Xu, Zuyan; Zhang, Jingyuan; Li, Qongru; Geng, Zifan; Gong, Zhuo; Ye, Qing; Gu, Pei; Xao, Jing-Lian

    1993-03-01

    Our early observations have confirmed that laser hyperthermia or PDT alone does not promote the tumor metastasis. In order to evaluate the combined effect of local tumor laser hyperthermia on the distant metastasis, transplantable forestomach carcinoma (Fc) in 615 line mice was treated by Nd:YAG laser hyperthermia (45 degree(s)C/20 min) combined with PDT (HpD 5 mg/kg, 480 J/cm2, 20 min), chemotherapy (Cyclophosphamide 28.8 mg/kg) and excision, respectively. The results show that (1) the tumor growth inhibition by various treatment was significant compared with a control group; (2) no statistics different in metastasis rate were observed in laser hyperthermia combined with PDT, chemotherapy, or scalpel excision separately. It is suggested that laser hyperthermia combined with PDT, chemotherapy, or excision does not increase the incidence of the tumor metastasis.

  8. Quantitative tissue cytometry (Tissomics): multimodal slide-based cytometry, confocal imaging, and volume rendering is the key

    NASA Astrophysics Data System (ADS)

    Tarnok, Attila; Mittag, Anja; Kuska, Jens-Peer; Braumann, Ulf-Dietrich; Mosch, Birgit; Arendt, Thomas

    2007-02-01

    Multiplexed high-content cytometric analysis of cells is a prerequisite for Cytomics and Systems Biology. Slide Based Cytometry (SBC) analysis yields quantitative cell related data on various cell constituents. It allows to measure and identify in high-throughput hundred-thousands of objects and obtain cytometric data on light absorption, scatter and fluorescence signals. Selected cells of interest can be rescanned and morphologically evaluated. To be cytometric SBC measurement needs high focal depth in order to acquire the fluorescence of the whole cell. For tissue analysis section thickness of >30μm is needed to reduce cell sectioning leading in multiple labelled specimens to an overestimation of multiple stained cells due to stereology, mimicking co-expression or elevated expression that is in fact due to coincidences in the z-axis direction. By confocal sectioning and 3D-reconstruction these overlays could be eliminated but confocal 3D imaging is slow and the resulting data are not cytometric. To overcome this obstacle, we combined SBC analysis with confocal imaging using a Laser Scanning Cytometer (iCys, Compucyte Corp., MA). Single to triple labelled 30-120μm thick human brain sections were scanned cytometrically (up to three laser 405nm, 488nm, 633nm) and double and triple labeled cells were identified. In the second step these objects were relocated, scanned confocally and 3D-reconstructed (Mathematica®, MathGL3d). This combination of high-throughput SBC and high-resolution confocal imaging enables for unequivocal identification of multiple labelled objects and is a prerequisite for Cytomic tissue analysis, Tissomics. (Support: HBFG 036/379-1)

  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.

    PubMed

    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. PMID:20047806

  10. A combined double-tweezers and wavelength-tunable laser nanosurgery microscope

    NASA Astrophysics Data System (ADS)

    Zhu, Qingyuan; Parsa, Shahab; Shi, Linda Z.; Harsono, Marcellinus; Wakida, Nicole M.; Berns, Michael W.

    2009-08-01

    In two previous studies we have conducted combined laser subcellular microsurgery and optical trapping on chromosomes in living cells1, 2. In the latter study we used two separate microscopes, one for the trap and one for the laser scissors, thus requiring that we move the cell specimen between microscopes and relocate the irradiated cells. In the former paper we combined the 1064 nm laser trap and the 532 nm laser scissors into one microscope. However, in neither study did we have multiple traps allowing for more flexibility in application of the trapping force. In the present paper we describe a combined laser scissors and tweezers microscope that (1) has two trapping beams (both moveable via rapid scanning mirrors (FSM- 300, Newport Corp.), (2) uses a short pulsed tunable 200 fs 710-990 nm Ti:Sapphire laser for laser microsurgery, and (3) also has the option to use a 337 nm 4 ns UV laser for subcellular surgery. The two laser tweezers and either of the laser ablation beams can be used in a cell surgery experiment. The system is integrated into the robotic-controlled RoboLase system3. Experiments on mitotic chromosomes of rat kangaroo PTK2 cells are described.

  11. Combined Experimental and Numerical Investigations into Laser Propulsion Engineering Physics

    NASA Astrophysics Data System (ADS)

    Kenoyer, David Adam

    The RPI pulsed Laser Propulsion (LP) research effort focuses on the future application of launching nano- and micro-satellites (1-10 kg payloads) into Low Earth Orbit (LEO), using a remote Ground Based Laser (GBL) power station to supply the required energy for flight. This research program includes both experimental and numerical studies investigating the propulsive performance of several engine geometries (constituting a lightcraft family). Using the Lumonics twin K-922m TEA pulsed laser system, axial and lateral thrust, C m, Isp, and η measurements were made for these engine geometries, examining the effects of several critical factors including: engine orientation (e.g. lateral and angular offset), laser pulse energy, pulse repetition frequency, pulse duration, propellant type, and engine size-scaling effects. Investigation into the origins of lateral "beam riding" forces was of particular interest. Lateral impulse measurements and high speed Schlieren photography were utilized to provide an understanding of laser beam-riding/propulsive physics. The acquired lightcraft database was used to further develop an existing 7-Degree Of Freedom (DOF) flight dynamics model extensively calibrated against 16 actual trajectories of small scale model lightcraft flown at White Sands Missile Range, NM on a 10 kW pulsed CO2 laser called PLVTS. The full system 7-DOF model is comprised of updated individual aerodynamics, engine, laser beam propagation, variable vehicle inertia, reaction controls system, and dynamics models, integrated to represent all major phenomena in a consistent framework. This flight dynamics model and associated 7-DOF code provide a physics-based predictive tool for basic research investigations into laser launched lightcraft for suborbital and orbital missions. Simulations were performed to demonstrate the flight capabilities of each engine geometry using the updated lightcraft propulsion database, the results of which further demonstrate that autonomous

  12. Programmable illumination and high-speed, multi-wavelength, confocal microscopy using a digital micromirror.

    PubMed

    Martial, Franck P; Hartell, Nicholas A

    2012-01-01

    Confocal microscopy is routinely used for high-resolution fluorescence imaging of biological specimens. Most standard confocal systems scan a laser across a specimen and collect emitted light passing through a single pinhole to produce an optical section of the sample. Sequential scanning on a point-by-point basis limits the speed of image acquisition and even the fastest commercial instruments struggle to resolve the temporal dynamics of rapid cellular events such as calcium signals. Various approaches have been introduced that increase the speed of confocal imaging. Nipkov disk microscopes, for example, use arrays of pinholes or slits on a spinning disk to achieve parallel scanning which significantly increases the speed of acquisition. Here we report the development of a microscope module that utilises a digital micromirror device as a spatial light modulator to provide programmable confocal optical sectioning with a single camera, at high spatial and axial resolution at speeds limited by the frame rate of the camera. The digital micromirror acts as a solid state Nipkov disk but with the added ability to change the pinholes size and separation and to control the light intensity on a mirror-by-mirror basis. The use of an arrangement of concave and convex mirrors in the emission pathway instead of lenses overcomes the astigmatism inherent with DMD devices, increases light collection efficiency and ensures image collection is achromatic so that images are perfectly aligned at different wavelengths. Combined with non-laser light sources, this allows low cost, high-speed, multi-wavelength image acquisition without the need for complex wavelength-dependent image alignment. The micromirror can also be used for programmable illumination allowing spatially defined photoactivation of fluorescent proteins. We demonstrate the use of this system for high-speed calcium imaging using both a single wavelength calcium indicator and a genetically encoded, ratiometric, calcium

  13. Interference Confocal Microscope Integrated with Spatial Phase Shifter.

    PubMed

    Wang, Weibo; Gu, Kang; You, Xiaoyu; Tan, Jiubin; Liu, Jian

    2016-01-01

    We present an interference confocal microscope (ICM) with a new single-body four-step simultaneous phase-shifter device designed to obtain high immunity to vibration. The proposed ICM combines the respective advantages of simultaneous phase shifting interferometry and bipolar differential confocal microscopy to obtain high axis resolution, large dynamic range, and reduce the sensitivity to vibration and reflectance disturbance seamlessly. A compact single body spatial phase shifter is added to capture four phase-shifted interference signals simultaneously without time delay and construct a stable and space-saving simplified interference confocal microscope system. The test result can be obtained by combining the interference phase response and the bipolar property of differential confocal microscopy without phase unwrapping. Experiments prove that the proposed microscope is capable of providing stable measurements with 1 nm of axial depth resolution for either low- or high-numerical aperture objective lenses. PMID:27563909

  14. Separate phase-locking and coherent combining of two laser diodes in a Michelson cavity

    NASA Astrophysics Data System (ADS)

    Schimmel, G.; Doyen, I.; Janicot, S.; Ramirez, L. P.; Hanna, M.; Georges, P.; Lucas-Leclin, G.; Vilokkinen, V.; Melanen, P.; Uusimaa, P.; Decker, J.; Crump, P.; Erbert, G.; Bull, S.; Kaunga-Nyirenda, S.; Larkins, E. C.

    2015-03-01

    We describe a new coherent beam combining architecture based on passive phase-locking of two laser diodes in a Michelson external cavity on their rear facet, and their coherent combination on the front facet. As a proof-of-principle, two ridge lasers have been coherently combined with >90 % efficiency. The phase-locking range, and the resistance of the external cavity to perturbations have been thoroughly investigated. The combined power has been stabilized over more than 15 min with an optical feedback as well as with an automatic adjustment of the driving currents. Furthermore, two high-brightness high-power tapered laser diodes have been coherently combined in a similar arrangement; the combining efficiency is 70% and results in an output power of 4 W. We believe that this new configuration combines the simplicity of passive self-organizing architectures with the optical efficiency of master-oscillator power-amplifier ones.

  15. Digital confocal microscopy through a multimode fiber.

    PubMed

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

    2015-09-01

    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). PMID:26368478

  16. Rapid-flow combined-action industrial CO2 laser

    NASA Astrophysics Data System (ADS)

    Generalov, N. A.; Zimakov, V. P.; Kosynkin, V. D.; Raizer, Iu. P.; Solovev, N. G.

    1982-08-01

    A general-purpose industrial CO2 laser intended for both CW and pulse-periodic operation and emitting pulses of a duration (up to 100 microsec) suitable for industrial applications is described. The operation derives from the method of creating a highly homogeneous glow discharge of large volume in a closed-cycle system. It involves setting up a high-power longitudinal (along the direction of the gas flow) on non-self-sustained discharge whose sole function is to provide laser pumping. The relatively low-power repetitive high-voltage pulses that ionize the gas are applied capacitatively to the discharge plasma without electrodes. The laser generates an average power of 1 kW at a pulse repetition frequency of 200 Hz, or 1.5 kW CW. The maximum output powers are, respectively, 1.5 and 2 kW.

  17. Optical properties measurement of the laser-ablated tissues for the combined laser ablation with photodynamic therapy

    NASA Astrophysics Data System (ADS)

    Honda, Norihiro; Ishii, Katsunori; Awazu, Kunio

    2012-03-01

    Laser ablation therapy combined with photodynamic therapy (PDT) is studied for treatment of advanced cancers. The clinical outcome of PDT may be improved by the accurate knowledge about the light distribution within tissue. Optical properties [absorption coefficient (μa), scattering coefficient (μs), anisotropy factor (g), refractive index, etc.] of tissues help us realizing a light propagation through the tissue. It is important to understand of the effect of laser coagulation formed by laser ablation to PDT. The aim of this study is to estimate of influence of coagulated region to PDT for effective PDT combined laser ablation therapy. We evaluated the optical property of mouse tumor tissue in native and coagulated state using a double integrating sphere system and an inverse Monte Carlo method in the wavelength range from 350 to 1000 nm. After laser ablation, the μa and reduced scattering coefficient spectra of coagulated tissues were increased in the wavelength range from 350 to 1000 nm. The optical penetration depth of coagulated tissues is 1.2-2.9 times lower than the native state in the wavelength range from 350 to 1000 nm. The intensity of the light energy inside the coagulated tissue falls to about 60% of its original value at the end of coagulated layer. The evaluation of light energy distribution by the determination of the tissues optical properties could be useful for optimization of the treatment procedure in combined laser ablation with PDT.

  18. CONFOCAL MICROSCOPY SYSTEM PERFORMANCE: FOUNDATIONS FOR MEASUREMENTS, QUANTITATION AND SPECTROSCOPY

    EPA Science Inventory

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

  19. CONFOCAL MICROSCOPY SYSTEM PERFORMANCE: FOUNDATIONS FOR CALIBRATION, QUANTITATION AND SPECTROSCOPY

    EPA Science Inventory

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

  20. Narrow linewidth operation of a spectral beam combined diode laser bar.

    PubMed

    Zhu, Zhanda; Jiang, Menghua; Cheng, Siqi; Hui, Yongling; Lei, Hong; Li, Qiang

    2016-04-20

    Our experiment is expected to provide an approach for realizing ultranarrow linewidth for a spectral beam combined diode laser bar. The beams of a diode laser bar are combined in a fast axis after a beam transformation system. With the help of relay optics and a transform lens with a long focal length of 1.5 m, the whole wavelength of a spectral combined laser bar can be narrowed down to 0.48 nm from more than 10 nm. We have achieved 56.7 W cw from a 19-element single bar with an M2 of 1.4  (in horizontal direction)×11.6  (in vertical direction). These parameters are good evidence that all the beams from the diode laser bar are combined together to increase the brightness. PMID:27140101

  1. High-power pump combiners for Tm-doped fibre lasers

    NASA Astrophysics Data System (ADS)

    Stachowiak, D.; Kaczmarek, P.; Abramski, K. M.

    2015-12-01

    In this paper our results of investigation on a pump power combiner in a configuration of 7×1 are presented. The performed combiner, with pump power of 80-85% transmission level, was successfully applied in a thulium doped fibre laser. The performed all-fibre laser setup reached a total CW output power of 6.42 W, achieving the efficiency on a 32.1% level.

  2. Combined proton acceleration from foil targets by ultraintense short laser pulses

    NASA Astrophysics Data System (ADS)

    Fang, Yuan; Yu, Tongpu; Ge, Xulei; Yang, Su; Wei, Wenqing; Yuan, Tao; Liu, Feng; Chen, Min; Liu, Jingquan; Li, Yutong; Yuan, Xiaohui; Sheng, Zhengming; Zhang, Jie

    2016-04-01

    Proton emission from solid foil targets irradiated by relativistically intense femtosecond laser pulses is studied experimentally. Broad plateaus in energy spectra are measured from micron-thick targets when the incident laser pulses have relatively low intensity contrasts. It is proposed that such proton spectra can be attributed to the combined processes of laser-driven collisionless shock acceleration and target normal sheath acceleration. Simple analytic estimation and two-dimensional particle-in-cell simulations are performed, which support our interpretation. The obtained plateau-shape spectrum may also serve as an effective tool to diagnose the plasma state and verify the ion acceleration mechanisms in laser-solid interactions.

  3. Combined single-pulse holography and time-resolved laser schlieren for flow visualization

    NASA Technical Reports Server (NTRS)

    Burner, A. W.; Goad, W. K.

    1981-01-01

    A pulsed ruby laser and continuous-wave argon ion laser were used in a combined setup at the Langley Expansion Tube for single pulse holography and time resolved laser schlieren with a common optical axis. The systems can be operated simultaneously for a single run. For a single frame, the pulsed holographic setup offers the options of shadowgraph, Schlieren, and interferometry from the reconstructed hologram as well as the advantage of post-run sensitivity adjustments. For flow establishment studies the time resolved laser Schlieren provides visualization of the flow field every 12.5 microns for up to 80 frames with an exposure time per frame of 5.4 microns.

  4. Clinical applications of a real-time scanning-slit confocal microscope designed for real-time observations of the in-vivo human cornea

    NASA Astrophysics Data System (ADS)

    Masters, Barry R.

    1995-05-01

    We describe a new, real-time, flying slit confocal microscope, that has unique features and imaging characteristics for in vivo human ocular imaging. In vivo real-time confocal microscopy is currently used to investigate the tear film, renewal of the ocular surface, the role of epithelial innervation in epithelial cell proliferation, wound healing, kinetics of drug penetration, the effects of laser refractive surgery on the keratocyte activation and distribution in the stroma, and the nature of endothelial defects. The following clinical examples will be presented and discussed: confocal microscopy of normal human basal and wing cells in the epithelium, confocal microscopy of lamellar and penetrating corneal grafts, confocal microscopy of corneal ulcer, confocal microscopy of scar formation after herpes keratitis, and confocal microscopy of corneal innervation. The use of scanning slit confocal microscopes has unique advantages over other instrumental systems based on pinhole-containing Nipkow disks (tandem-scanning confocal microscopes) for clinical in vivo confocal microscopy.

  5. Mechanical scanner-less multi-beam confocal microscope with wavefront modulation

    NASA Astrophysics Data System (ADS)

    Takiguchi, Yu; Seo, Min-Woong; Kagawa, Keiichiro; Takamoto, Hisayoshi; Inoue, Takashi; Kawahito, Shoji; Terakawa, Susumu

    2016-04-01

    We propose a novel full-electronically controlled laser confocal microscope in which a liquid-crystal-on-silicon spatial light modulator and a custom CMOS imaging sensor are synchronized for performing multi-beam confocal imaging. Adaptive wavefront modulation for functional multi-beam excitation can be achieved by displaying appropriate computer generated holograms on the spatial light modulator, in consideration of the numerical aperture of the focusing objective. We also adopted a custom CMOS imaging sensor to realize multi-beam confocal microscopy without any physical pinhole. The confocality of this microscope was verified by improvements in transverse and axial resolutions of fluorescent micro-beads.

  6. Combination of vascular endothelial growth factor inhibitors and laser therapy for diabetic macular oedema: a review.

    PubMed

    Mehta, Hemal; Gillies, Mark C; Fraser-Bell, Samantha

    2016-05-01

    This review provides a perspective on published and ongoing clinical trials of vascular endothelial growth factor inhibitors (anti-VEGF agents) combined with laser therapy for diabetic macular oedema (DMO). Although there was little short-term benefit in combining prompt macular laser with anti-VEGF therapy for centre-involving DMO in the Diabetic Retinopathy Clinical Research Network (DRCRnet) Protocol I study, deferred macular laser was still required in over 40% of study eyes in DRCRnet Protocol T. Macular laser was applied in more than 30% of eyes with centre-involving DMO receiving ranibizumab in the RISE and RIDE studies. For non centre-involving DMO the evidence-base still supports use of focal macular laser alone, although clinicians should be cautious about applying laser too close to the foveal avascular zone with the availability of pharmacotherapy. Ongoing clinical trials are assessing whether selectively targeting areas of peripheral retinal ischaemia with laser reduces the number of anti-VEGF injections to stabilise DMO and whether combining macular micropulse laser with anti-VEGF therapy is beneficial in DMO. PMID:27061760

  7. Fusion welding studies using laser on Ti-SS dissimilar combination

    NASA Astrophysics Data System (ADS)

    Shanmugarajan, B.; Padmanabham, G.

    2012-11-01

    Laser welding investigations were carried out on dissimilar Ti-SS combination. The study is aimed to improve the weld strength and ductility by minimizing harmful intermetallics and taking advantage of high cooling rates in laser welding. Results of continuous wave 3.5 kW CO2 laser welding of totally dissimilar combination of Titanium and stainless steel (304) have been discussed. Bead on plate welding experiments were conducted to identify the laser welding parameters using depth of penetration as criteria. The welding of dissimilar combination has been attempted both autogenously and with interlayers such as Vanadium (V) and Tantalum (Ta) in the form of laser cladding as well as strip. Autogenous welds were carried out by varying the laser power, welding speed and position of the laser beam with respect to the joint centre. The resultant welds are characterized by macrostructure analysis, SEM/EDAX and XRD and as welded tensile test in UTM. The autogenous welds have exhibited extensive cracking even when welded at high speeds or by manipulating the beam position with respect to the joint. Similarly Vandaium as interlayer could not achieve crack free joint. A joint with 40 MPa strength could be made with Ta as interlayer. Results and analysis of these variants of laser welded joints are reported and discussed.

  8. Development of high damage threshold multilayer thin film beam combiner for laser application

    NASA Astrophysics Data System (ADS)

    Nand, Mangla; Babita, Jena, S.; Tokas, R. B.; Rajput, P.; Mukharjee, C.; Thakur, S.; Jha, S. N.; Sahoo, N. K.

    2016-05-01

    A polarized wavelength multiplexer with high laser induced damage threshold has been developed to combine two laser beam of high peak power in the visible region. The present wavelength multiplexer is a multilayer thin film device deposited by reactive electron beam evaporation. The developed device is capable of combining two p-polarized laser beams of peak power density of 1.7 GW/cm2 at an angle of incidence of 45°. High transmission (T> 90%) in high pass region and high reflection (R> 99%) in stop band region have been achieved.

  9. High-power spectral beam combining of linearly polarized Tm:fiber lasers.

    PubMed

    Shah, Lawrence; Sims, R Andrew; Kadwani, Pankaj; Willis, Christina C C; Bradford, Joshua B; Sincore, Alex; Richardson, Martin

    2015-02-01

    To date, high-power scaling of Tm:fiber lasers has been accomplished by maximizing the power from a single fiber aperture. In this work, we investigate power scaling by spectral beam combination of three linearly polarized Tm:fiber MOPA lasers using dielectric mirrors with a steep transition from highly reflective to highly transmissive that enable a minimum wavelength separation of 6 nm between individual laser channels within the wavelength range from 2030 to 2050 nm. Maximum output power is 253 W with M(2)<2, ultimately limited by thermal lensing in the beam combining elements. PMID:25967785

  10. Confocal stereology: an efficient tool for measurement of microscopic structures.

    PubMed

    Kubínová, Lucie; Janáček, Jiří

    2015-04-01

    Quantitative measurements of geometric forms or counting of objects in microscopic specimens is an essential tool in studies of microstructure. Confocal stereology represents a contemporary approach to the evaluation of microscopic structures by using a combination of stereological methods and confocal microscopy. 3-D images acquired by confocal microscopy can be used for the estimation of geometrical characteristics of microscopic structures by stereological methods, based on the evaluation of optical sections within a thick slice and using computer-generated virtual test probes. Such methods can be used for estimating volume, number, surface area and length using relevant spatial probes, which are generated by specific software. The interactions of the probes with the structure under study are interactively evaluated. An overview of the methods of confocal stereology developed during the past 30 years is presented. Their advantages and pitfalls in comparison with other methods for measurement of geometrical characteristics of microscopic structures are discussed. PMID:25743691

  11. A near-infrared confocal scanner

    NASA Astrophysics Data System (ADS)

    Lee, Seungwoo; Yoo, Hongki

    2014-06-01

    In the semiconductor industry, manufacturing of three-dimensional (3D) packages or 3D integrated circuits is a high-performance technique that requires combining several functions in a small volume. Through-silicon vias, which are vertical electrical connections extending through a wafer, can be used to direct signals between stacked chips, thus increasing areal density by stacking and connecting multiple patterned chips. While defect detection is essential in the semiconductor manufacturing process, it is difficult to identify defects within a wafer or to monitor the bonding results between bonded surfaces because silicon and many other semiconductor materials are opaque to visible wavelengths. In this context, near-infrared (NIR) imaging is a promising non-destructive method to detect defects within silicon chips, to inspect bonding between chips and to monitor the chip alignment since NIR transmits through silicon. In addition, a confocal scanner provides high-contrast, optically-sectioned images of the specimen due to its ability to reject out-of-focus noise. In this study, we report an NIR confocal scanner that rapidly acquires high-resolution images with a large field of view through silicon. Two orthogonal line-scanning images can be acquired without rotating the system or the specimen by utilizing two orthogonally configured resonant scanning mirrors. This NIR confocal scanner can be efficiently used as an in-line inspection system when manufacturing semiconductor devices by rapidly detecting defects on and beneath the surface.

  12. Evaluation of dermal extracellular matrix and epidermal-dermal junction modifications using matrix-assisted laser desorption/ionization mass spectrometric imaging, in vivo reflectance confocal microscopy, echography, and histology: effect of age and peptide applications.

    PubMed

    Mondon, Philippe; Hillion, Mélanie; Peschard, Olivier; Andre, Nada; Marchand, Thibault; Doridot, Emmanuel; Feuilloley, Marc Gj; Pionneau, Cédric; Chardonnet, Solenne

    2015-06-01

    This study was conducted to establish a new methodology for evaluating elements of dermal extracellular matrix (ECM), of epidermal-dermal junction (EDJ), and effects of molecules which can modulate their synthesis. This methodology is based on matrix-assisted laser desorption/ionization mass spectrometric imaging (MALDI-MSI). In vivo reflectance confocal microscopy (in vivo RCM) and echography were also used. Using immunohistochemistry methods on explants, age-related modification data were obtained for selected dermal ECM and EDJ proteins (collagen I, collagen IV, collagen VII, collagen XVII, nidogen I, decorin/decorunt) and used as reference for MALDI-MSI studies. A methodology was developed with MALDI-MSI to map epidermis and dermis proteins. Then MALDI-MSI was used to study age modifications. In vivo RCM and high-frequency ultrasounds were used to evaluate ECM and EDJ undulation modifications caused by aging. Anti-aging molecule evaluations were performed with a blend of palmitoyl oligopeptide and palmitoyl tetrapeptide-7. Immunohistochemistry studies demonstrated that the selected proteins were found to be less abundant in aged group explants vs. young group except for decorin. MALDI-MSI studies correlated the results obtained for decorin. In vivo RCM measurements indicated a decrease of EDJ undulation depth with age and ECM modifications in the upper part of dermis. Echography demonstrated that the peptide blend reduced subepidermal low-echogenic band thickness and improved its density. In vivo RCM studies indicated that the peptides improved the ECM structure vs. placebo. This preliminary MALDI-MSI study raised some technical difficulties that were overcome. Further studies will be conducted to identify more proteins and to demonstrate the interest of this method for cosmetic evaluations. PMID:25817264

  13. Removal of metals and ceramics by combined effects of micro liquid jet and laser pulse

    NASA Astrophysics Data System (ADS)

    Ahn, Daehwan; Seo, Changho; Kim, Dongsik

    2012-12-01

    In this work, we analyze a hybrid laser/liquid jet micromachining process for several metals and ceramics based on the optical breakdown of a microdroplet. In the process, materials are removed by the combined effects of a laser pulse and a high-speed pulsed microjet ejected from the microdroplet. The opto-hydrodynamic phenomena occurring during this process and the interaction of the laser/liquid jet with various materials, including copper, aluminum, stainless steel, alumina, and boron nitride, are investigated experimentally. The results show that the laser/liquid jet can remove the materials with substantially increased removal rates and reduced thermal side effects compared with the conventional pulsed laser ablation process. Visualization of the process reveals that the materials are partially ablated and melted by the laser pulse during the early stage of the process and that the molten material is subsequently eliminated by the hydrodynamic impact of the liquid jet.

  14. Interstitial laser irradiation of metastatic mammary tumors in combination with intratumoral injection of immunoadjuvant

    NASA Astrophysics Data System (ADS)

    Joshi, Chet; Jose, Jessnie; Figueroa, Daniel; Goddard, Jessica; Li, Xiaosong; Liu, Hong; Nordquist, Robert E.; Hode, Tomas; Chen, Wei R.

    2012-03-01

    Laser immunotherapy (LIT) was developed to treat metastatic cancers using a combination of laser irradiation and immunological stimulation. The original design of LIT employs a non-invasive, selective laser photothermal interaction, using an in situ light-absorbing dye. However, this non-invasive treatment mode faces challenges in treating deep, large tumors. Furthermore, it has difficulties in the cases of highly pigmented skin overlying target tumors. To overcome these limitations, interstitial laser immunotherapy (ILIT) was proposed. In ILIT, a cylindrical, side-fire fiber diffuser is placed inside the target tumor to induce thermal damage. To enhance the interstitial irradiation induced photothermal interaction, an immunological modifier, glycated chitosan (GC), is injected into the tumor after the laser treatment. In this study, a cylindrical diffuser with an active length of 1 cm was used to treat tumors of 1 to 1.5 cm in size. Different laser powers (1 to 3 watts) and different irradiation durations (10 to 30 minutes) were used to test the thermal effects of ILIT. Different doses of the GC (1.0%, 0.1 to 0.6 ml per rat) were used to determine the immunological effects of ILIT. Our results show that the animal survival depends on both laser dose and GC dose. A dose of 0.2 ml per tumor appeared to result in the highest survival rate under interstitial laser irradiation with 2.5 watts and 20 minutes. While the results in this study are not conclusive, they indicate that interstitial laser irradiation can be combined with immunotherapy to treat metastatic cancers. Furthermore, our results suggest that an optimal combination of laser dose and GC dose could be obtained for future clinical protocols using interstitial laser immunotherapy.

  15. Diffractive Combiner of Single-Mode Pump Laser-Diode Beams

    NASA Technical Reports Server (NTRS)

    Liu, Duncan; Wilson, Daniel; Qiu, Yueming; Forouhar, Siamak

    2007-01-01

    An optical beam combiner now under development would make it possible to use the outputs of multiple single-mode laser diodes to pump a neodymium: yttrium aluminum garnet (Nd:YAG) nonplanar ring oscillator (NPRO) laser while ensuring that the laser operates at only a single desired frequency. Heretofore, an Nd:YAG NPRO like the present one has been pumped by a single multimode laser-diode beam delivered via an optical fiber. It would be desirable to use multiple pump laser diodes to increase reliability beyond that obtainable from a single pump laser diode. However, as explained below, simplistically coupling multiple multimode laser-diode beams through a fiber-optic combiner would entail a significant reduction in coupling efficiency, and lasing would occur at one or more other frequencies in addition to the single desired frequency. Figure 1 schematically illustrates the principle of operation of a laser-diode-pumped Nd:YAG NPRO. The laser beam path is confined in a Nd:YAG crystal by means of total internal reflections on the three back facets and a partial-reflection coating on the front facet. The wavelength of the pump beam - 808 nm - is the wavelength most strongly absorbed by the Nd:YAG crystal. The crystal can lase at a wavelength of either 1,064 nm or 1,319 nm - which one depending on the optical coating on the front facet. A thermal lens effect induced by the pump beam enables stable lasing in the lowest-order transverse electromagnetic mode (the TEM00 mode). The frequency of this laser is very stable because of the mechanical stability of the laser crystal and the unidirectional nature of the lasing. The unidirectionality is a result of the combined effects of (1) a Faraday rotation induced by an externally applied magnetic field and (2) polarization associated with non-normal incidence and reflection on the front facet.

  16. Laser surgery for Zenker's diverticulum: European combined study.

    PubMed

    Papaspyrou, Giorgos; Schick, Bernhard; Papaspyrou, Spyros; Wiegand, Susanne; Al Kadah, Basel

    2016-01-01

    Surgical intervention is the gold standard of treatment for Zenker's diverticulum. The aim of this study was to examine the role of laser surgery in a large number of patients with this pathological entity. The data of 91 consecutive patients treated due to Zenker's diverticulum with the aid of CO2 laser in three institutions (Homburg/Saar and Marburg, Germany/Athens, Greece) during the last 10 years were retrospectively analyzed. Parameters examined were sex, age, preoperative symptoms, length of operation and complications, revision surgery necessity and degree of patient satisfaction. All patients had a minimum follow-up of one year. Dysphagia was the most common preoperative symptom (78 %). The most common minor complication was dental injury (6.6 %), but a serious complication in form of emphysema was observed in only two patients (2.2 %). A surgical revision was necessary in 8 (8.8 %) of the treated patients. The majority of treated patients was free of symptoms (86.8 %), or presented mild symptoms (9.9 %) one year after intervention, and only three patients (3.3 %) were dissatisfied. Our study shows that laser treatment of Zenker's diverticulum is an efficient operative technique associated with low complications rates and significant improvement of patients' symptoms in most of the examined cases. PMID:25567345

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

    PubMed Central

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

    2014-01-01

    We present an interferometric confocal microscope using an array of 1200 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. PMID:25078199

  18. Spectral confocal reflection microscopy using a white light source

    NASA Astrophysics Data System (ADS)

    Booth, M.; Juškaitis, R.; Wilson, T.

    2008-08-01

    We present a reflection confocal microscope incorporating a white light supercontinuum source and spectral detection. The microscope provides images resolved spatially in three-dimensions, in addition to spectral resolution covering the wavelength range 450-650nm. Images and reflection spectra of artificial and natural specimens are presented, showing features that are not normally revealed in conventional microscopes or confocal microscopes using discrete line lasers. The specimens include thin film structures on semiconductor chips, iridescent structures in Papilio blumei butterfly scales, nacre from abalone shells and opal gemstones. Quantitative size and refractive index measurements of transparent beads are derived from spectral interference bands.

  19. Combination of fiber-guided pulsed erbium and holmium laser radiation for tissue ablation under water

    NASA Astrophysics Data System (ADS)

    Pratisto, Hans; Frenz, Martin; Ith, Michael; Altermatt, Hans J.; Jansen, E. Duco; Weber, Heinz P.

    1996-07-01

    Because of the high absorption of near-infrared laser radiation in biological tissue, erbium lasers and holmium lasers emitting at 3 and 2 mu m, respectively, have been proven to have optimal qualities for cutting or welding and coagulating tissue. To combine the advantages of both wavelengths, we realized a multiwavelength laser system by simultaneously guiding erbium and holmium laser radiation by means of a single zirconium fluoride (ZrF4) fiber. Laser-induced channel formation in water and poly(acrylamide) gel was investigated by the use of a time-resolved flash-photography setup, while pressure transients were recorded simultaneously with a needle hydrophone. The shapes and depths of vapor channels produced in water and in a submerged gel after single erbium and after combination erbium-holmium radiation delivered by means of a 400- mu m ZrF4 fiber were measured. Transmission measurements were performed to determine the amount of pulse energy available for tissue ablation. The effects of laser wavelength and the delay time between pulses of different wavelengths on the photomechanical and photothermal responses of meniscal tissue were evaluated in vitro by the use of histology. It was observed that the use of a short (200- mu s, 100-mJ) holmium laser pulse as a prepulse to generate a vapor bubble through which the ablating erbium laser pulse can be transmitted (delay time, 100 mu s) increases the cutting depth in meniscus from 450 to 1120 mu m as compared with the depth following a single erbium pulse. The results indicate that a combination of erbium and holmium laser radiation precisely and efficiently cuts tissue under water with 20-50- mu m collateral tissue damage. wave, cavitation, channel formation, infrared-fiber-delivery system, tissue damage, cartilage.

  20. Spatial Combining of Laser-Diode Beams for Pumping an NPRO

    NASA Technical Reports Server (NTRS)

    Gelsinger, Paul; Liu, Duncan; Mulder, Jerry; Aguayo, Francisco

    2008-01-01

    A free-space optical beam combiner now undergoing development makes it possible to use the outputs of multiple multimode laser diodes to pump a neodymium-doped yttrium aluminum garnet (Nd:YAG) non-planar ring oscillator (NPRO) laser while ensuring that the laser operates at only a single desired frequency. Heretofore, a Nd:YAG NPRO like the present one has been pumped by a single multimode laser-diode beam delivered via an optical fiber. It would be desirable to use multiple pump laser diodes to increase reliability beyond that obtainable from a single pump laser diode. However, as explained in this article, simplistically coupling multiple multimode laser-diode beams through a fiber-optic combiner would entail a significant reduction in coupling efficiency, and lasing would occur at one or more other frequencies in addition to the single desired frequency. To minimize coupling loss, one must ensure that the NA (approximately equal to 0.3) of the combined laser-diode beams is less than the NA of the fiber. The A(Omega) of the laser-diode beam in the slow-axis plane is 1/1.3 as large as that of the fiber. This A(Omega) is small enough to enable efficient coupling of light into the optical fiber, but too large for combining of beams in the slow-axis plane. Therefore, a pair of cylindrical lenses is used to cancel the slow-axis plane magnification introduced by the on-cylindrical lenses used to effect magnification in the fast-axis plane.

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

    PubMed

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

    2013-12-01

    The combination of genetically encoded fluorescent proteins and three-dimensional imaging enables cell-type-specific studies of embryogenesis. Light sheet microscopy, in which fluorescence excitation is provided by a plane of laser light, is an appealing approach to live imaging due to its high speed and efficient use of photons. While the advantages of rapid imaging are apparent from recent work, the importance of low light levels to studies of development is not well established. We examine the zebrafish opercle, a craniofacial bone that exhibits pronounced shape changes at early developmental stages, using both spinning disk confocal and light sheet microscopies of fluorescent osteoblast cells. We find normal and aberrant opercle morphologies for specimens imaged with short time intervals using light sheet and spinning disk confocal microscopies, respectively, under equivalent exposure conditions over developmentally-relevant time scales. Quantification of shapes reveals that the differently imaged specimens travel along distinct trajectories in morphological space. PMID:23242824

  2. To see the unseeable: confocal miniprobes for routine microscopic imaging during endoscopy

    NASA Astrophysics Data System (ADS)

    Osdoit, A.; Lacombe, F.; Cavé, C.; Loiseau, S.; Peltier, E.

    2007-02-01

    Confocal fluorescence high resolution imaging during standard endoscopic procedures has been presented as a very promising tool to enhance patient care and physician practice by providing supplementary diagnostic information in real-time. The purpose of this paper is to show not only potential, but convincing results of endoscopic microscopy using a catheter-based approach. Mauna Kea Technologies' core technology, Cellvizio, delivers dynamic imaging at 12 frames/second using confocal miniprobes inserted through the operating channel of regular endoscopes. Cellvizio is composed of 3 parts including (a) a Laser Scanning Unit, (b) Confocal Miniprobe TM with the following characteristics: 5-15 μm axial resolution, 2-5 μm lateral resolution, 15-100 μm depth of penetration, field of view of 600x500 μm and (c) a software package with onthe- fly processing capabilities. With several tens of patients examined during routine GI endoscopy procedures, the most relevant clinical parameters could be assessed in a doubled-blinded fashion between the endoscopist and a pathologist and results showing very high accuracy in the differentiation of neoplasia from normal and hyperplastic tissue were obtained. In the field of pulmonology, the micro-autofluorescence properties of tissues could be assessed and structures never before accessed in vivo were observed. Cellvizio® may be useful to study bronchial remodeling in asthma and chronic obstructive pulmonary diseases. Using appropriate topical fluorescent dye, the Confocal Miniprobes may also make it possible to perform optical biopsy of precancerous and superficial bronchial cancers. Cellvizio® is as a new tool towards "targeted biopsies", leading to earlier, more reliable and cost effective diagnostic procedures. Other applications, specifically in molecular imaging are also made possible by the miniaturization of the probe (combination with biopsy needle for solid organs use or lymph node detection) and by the

  3. Noncontact microsurgery and micromanipulation of living cells with combined system femtosecond laser scalpel-optical tweezers

    NASA Astrophysics Data System (ADS)

    Il'ina, Inna V.; Sitnikov, Dmitry S.; Ovchinnikov, Andrey V.; Agranat, Mikhail B.; Khramova, Yulia V.; Semenova, Maria L.

    2012-06-01

    We report on the results of using self-developed combined laser system consisting of a femtosecond laser scalpel (Cr:Forsterite seed oscillator and a regenerative amplifier, 620 nm, 100 fs, 10 Hz) and optical tweezers (cw laser, 1064 nm) for performing noncontact laser-mediated polar body (PB) and trophectoderm (TE) biopsy of early mammalian embryos. To perform PB biopsy the femtosecond laser scalpel was initially used to drill an opening in the zona pellucida, and then the PB was extracted out of the zygote with the optical tweezers. Unlike PB biopsy, TE biopsy allows diagnosing maternally-derived as well as paternally-derived defects. Moreover, as multiple TE cells can be taken from the embryo, more reliable diagnosis can be done. TE biopsy was performed by applying laser pulses to dissect the desired amount of TE cells that had just left the zona pellucida during the hatching. Optical tweezers were then used to trap and move the dissected TE cells in a prescribed way. Laser power in optical tweezers and energy of femtosecond laser pulses were thoroughly optimized to prevent cell damage and obtain high viability rates. In conclusion, the proposed techniques of laser-based embryo biopsy enable accurate, contamination-free, simple and quick microprocessing of living cells.

  4. Detection limits of confocal surface plasmon microscopy

    PubMed Central

    Pechprasarn, Suejit; Somekh, Michael G.

    2014-01-01

    This paper applies rigorous diffraction theory to evaluate the minimum mass sensitivity of a confocal optical microscope designed to excite and detect surface plasmons operating on a planar metallic substrate. The diffraction model is compared with an intuitive ray picture which gives remarkably similar predictions. The combination of focusing the surface plasmons and accurate phase measurement mean that under favorable but achievable conditions detection of small numbers of molecules is possible, however, we argue that reliable detection of single molecules will benefit from the use of structured surfaces. System configurations needed to optimize performance are discussed. PMID:24940537

  5. Hybrid detectors improved time-lapse confocal microscopy of PML and 53BP1 nuclear body colocalization in DNA lesions.

    PubMed

    Foltánková, Veronika; Matula, Pavel; Sorokin, Dmitry; Kozubek, Stanislav; Bártová, Eva

    2013-04-01

    We used hybrid detectors (HyDs) to monitor the trajectories and interactions of promyelocytic leukemia (GFP-PML) nuclear bodies (NBs) and mCherry-53BP1-positive DNA lesions. 53BP1 protein accumulates in NBs that occur spontaneously in the genome or in γ-irradiation-induced foci. When we induced local DNA damage by ultraviolet irradiation, we also observed accumulation of 53BP1 proteins into discrete bodies, instead of the expected dispersed pattern. In comparison with photomultiplier tubes, which are used for standard analysis by confocal laser scanning microscopy, HyDs significantly eliminated photobleaching of GFP and mCherry fluorochromes during image acquisition. The low laser intensities used for HyD-based confocal analysis enabled us to observe NBs for the longer time periods, necessary for studies of the trajectories and interactions of PML and 53BP1 NBs. To further characterize protein interactions, we used resonance scanning and a novel bioinformatics approach to register and analyze the movements of individual PML and 53BP1 NBs. The combination of improved HyD-based confocal microscopy with a tailored bioinformatics approach enabled us to reveal damage-specific properties of PML and 53BP1 NBs. PMID:23410959

  6. Video-rate Scanning Confocal Microscopy and Microendoscopy

    PubMed Central

    Nichols, Alexander J.; Evans, Conor L.

    2011-01-01

    Confocal microscopy has become an invaluable tool in biology and the biomedical sciences, enabling rapid, high-sensitivity, and high-resolution optical sectioning of complex systems. Confocal microscopy is routinely used, for example, to study specific cellular targets1, monitor dynamics in living cells2-4, and visualize the three dimensional evolution of entire organisms5,6. Extensions of confocal imaging systems, such as confocal microendoscopes, allow for high-resolution imaging in vivo7 and are currently being applied to disease imaging and diagnosis in clinical settings8,9. Confocal microscopy provides three-dimensional resolution by creating so-called "optical sections" using straightforward geometrical optics. In a standard wide-field microscope, fluorescence generated from a sample is collected by an objective lens and relayed directly to a detector. While acceptable for imaging thin samples, thick samples become blurred by fluorescence generated above and below the objective focal plane. In contrast, confocal microscopy enables virtual, optical sectioning of samples, rejecting out-of-focus light to build high resolution three-dimensional representations of samples. Confocal microscopes achieve this feat by using a confocal aperture in the detection beam path. The fluorescence collected from a sample by the objective is relayed back through the scanning mirrors and through the primary dichroic mirror, a mirror carefully selected to reflect shorter wavelengths such as the laser excitation beam while passing the longer, Stokes-shifted fluorescence emission. This long-wavelength fluorescence signal is then passed to a pair of lenses on either side of a pinhole that is positioned at a plane exactly conjugate with the focal plane of the objective lens. Photons collected from the focal volume of the object are collimated by the objective lens and are focused by the confocal lenses through the pinhole. Fluorescence generated above or below the focal plane will

  7. Video-rate scanning confocal microscopy and microendoscopy.

    PubMed

    Nichols, Alexander J; Evans, Conor L

    2011-01-01

    Confocal microscopy has become an invaluable tool in biology and the biomedical sciences, enabling rapid, high-sensitivity, and high-resolution optical sectioning of complex systems. Confocal microscopy is routinely used, for example, to study specific cellular targets, monitor dynamics in living cells, and visualize the three dimensional evolution of entire organisms. Extensions of confocal imaging systems, such as confocal microendoscopes, allow for high-resolution imaging in vivo and are currently being applied to disease imaging and diagnosis in clinical settings. Confocal microscopy provides three-dimensional resolution by creating so-called "optical sections" using straightforward geometrical optics. In a standard wide-field microscope, fluorescence generated from a sample is collected by an objective lens and relayed directly to a detector. While acceptable for imaging thin samples, thick samples become blurred by fluorescence generated above and below the objective focal plane. In contrast, confocal microscopy enables virtual, optical sectioning of samples, rejecting out-of-focus light to build high resolution three-dimensional representations of samples. Confocal microscopes achieve this feat by using a confocal aperture in the detection beam path. The fluorescence collected from a sample by the objective is relayed back through the scanning mirrors and through the primary dichroic mirror, a mirror carefully selected to reflect shorter wavelengths such as the laser excitation beam while passing the longer, Stokes-shifted fluorescence emission. This long-wavelength fluorescence signal is then passed to a pair of lenses on either side of a pinhole that is positioned at a plane exactly conjugate with the focal plane of the objective lens. Photons collected from the focal volume of the object are collimated by the objective lens and are focused by the confocal lenses through the pinhole. Fluorescence generated above or below the focal plane will therefore not

  8. High Resolution Surface Geometry and Albedo by Combining Laser Altimetry and Visible Images

    NASA Technical Reports Server (NTRS)

    Morris, Robin D.; vonToussaint, Udo; Cheeseman, Peter C.; Clancy, Daniel (Technical Monitor)

    2001-01-01

    The need for accurate geometric and radiometric information over large areas has become increasingly important. Laser altimetry is one of the key technologies for obtaining this geometric information. However, there are important application areas where the observing platform has its orbit constrained by the other instruments it is carrying, and so the spatial resolution that can be recorded by the laser altimeter is limited. In this paper we show how information recorded by one of the other instruments commonly carried, a high-resolution imaging camera, can be combined with the laser altimeter measurements to give a high resolution estimate both of the surface geometry and its reflectance properties. This estimate has an accuracy unavailable from other interpolation methods. We present the results from combining synthetic laser altimeter measurements on a coarse grid with images generated from a surface model to re-create the surface model.

  9. Designing and optimizing highly efficient grating for high-brightness laser based on spectral beam combining

    SciTech Connect

    Yang, Ying-Ying E-mail: yangyy@semi.ac.cn; Zhao, Ya-Ping; Wang, Li-Rong; Zhang, Ling; Lin, Xue-Chun E-mail: yangyy@semi.ac.cn

    2015-03-14

    A highly efficient nano-periodical grating is theoretically investigated for spectral beam combining (SBC) and is experimentally implemented for attaining high-brightness laser from a diode laser array. The rigorous coupled-wave analysis with the S matrix method is employed to optimize the parameters of the grating. According the optimized parameters, the grating is fabricated and plays a key role in SBC cavity. The diffraction efficiency of this grating is optimized to 95% for the output laser which is emitted from the diode laser array. The beam parameter product of 3.8 mm mrad of the diode laser array after SBC is achieved at the output power of 46.3 W. The optical-to-optical efficiency of SBC cavity is measured to be 93.5% at the maximum operating current in the experiment.

  10. Non-collinear spectral coherent combination of ultrashort laser pulses.

    PubMed

    Ionel, Laura; Ursescu, Daniel

    2016-04-01

    Non-collinear spectral coherent combining (NCSCC) of ultrashort pulses is analyzed. 2D modeling of the electromagnetic field is performed in case of NCSCC using two or three pulses with different wavelengths. In the case of two pulses, a potentially unwanted spatio-temporal structure of the field appears, corresponding to spatial and temporal modulation of the pulse. By using NCSCC of three 62 fs long pulses with different spectral composition, such spatial-temporal coupling is eliminated and the combined pulse duration in the focal region drops to less than half. The method is scalable to a large number of ultrashort pulses. PMID:27136998

  11. Combined Laser Ultrasonics, Laser Heating and Raman Scattering in Diamond Anvil Cell System

    NASA Astrophysics Data System (ADS)

    Zinin, Pavel; Prakapenka, Vitali; Odake, Shoko; Burgess, Katherine

    2013-06-01

    We developed a unique and multifunctional in-situ measurement system under high pressure equipped with laser ultrasonics system, Raman device, and laser heating system (LH-LU-DAC) at the University of Hawaii. The system consists of four components: (1) LU-DAC system (probe and pump lasers, photodetector, and oscilloscope); (2) a fiber laser (1064 nm), which is designed to allow precise control of the total power in the range from 2 to 100 W by changing the diode current, for heating samples; (3) a spectrometer for measuring the temperature of the sample (using Black body radiation), fluorescence spectrum (spectrum of the ruby for pressure measurement), and Raman scattering measurements inside DAC under high pressure and high temperature (HPHT) conditions; and (4) an optical system for focusing laser beams (pump, probe, and 100 W CW lasers) on the sample in DAC and for imaging a sample inside the DAC. The system allows us to: (a) measure acoustical properties of materials under HPHT; (b) synthesize new phases under HPHT; and (c) measure Raman scattering under HPHT conditions for detection of phase transition. This work was supported by the U.S. DOE Grant, NO. DE-FG02-07ER46408, and NSF Grant, NO. EAR-1215796.

  12. Laryngocele resection by combined external and endoscopic laser approach.

    PubMed

    Ettema, Sandra L; Carothers, Daniel G; Hoffman, Henry T

    2003-04-01

    Options in the management of laryngoceles include observation, endoscopic resection, and resection via an external approach. We introduce a combined endoscopic and external approach that we have employed on several occasions to ensure complete removal of the laryngocele and the saccule from which it originated. A case is presented to help define the technique. PMID:12731632

  13. Use alone or in Combination of Red and Infrared Laser in Skin Wounds

    PubMed Central

    de Lima, Fernando José Camello; Barbosa, Fabiano Timbó; de Sousa-Rodrigues, Célio Fernando

    2014-01-01

    A systematic review was conducted covering the action of red laser, infrared and combination of both, with emphasis on cutaneous wound therapy, showing the different settings on parameters such as fluency, power, energy density, time of application, frequency mode and even the type of low-power lasers and their wavelengths. It was observed that in general, the lasers brings good clinical and histological results mainly, but there is not a protocol that defines a dosage of use that has predictability of therapeutic success in repairing these wounds. PMID:25653799

  14. Optical bistability in erbium-doped yttrium aluminum garnet crystal combined with a laser diode.

    PubMed

    Maeda, Y

    1994-01-10

    Optical bistability was observed in a simple structure of an injection laser diode combined with an erbium-doped yttrium aluminum garnet crystal. Since a hysteresis characteristic exists in the relationship between the wavelength and the injection current of a laser diode, an optical memory function capable of holding the output status is confirmed. In addition, an optical signal inversion was caused by the decrease of transmission of the erbium-doped yttrium aluminum garnet crystal against the red shift (principally mode hopping) of the laser diode. It is suggested that the switching time of this phenomenon is the time necessary for a mode hopping by current injection. PMID:20862016

  15. Coherent combining of four slab laser amplifiers with high beam quality

    NASA Astrophysics Data System (ADS)

    Yan, Hong; Ye, Yidong; Tian, Fei; Li, Guohui; Pan, Xundong; Zhang, Wei; Gao, Qingsong; Zhou, Tangjian; Liao, Yuan; Chen, Li; Lu, Fei; Luo, Jia

    2014-10-01

    We report a coherent combining of four slab laser amplifiers with high beam quality. The long strip laser beam is reshaped into a square beam using adjustable beam expander which removes the enormous astigmatism aberration. A filling ratio of 90% is achieved by two-dimensional splicing. A compact optical system with high sampling frequency is designed to detect the pointing direction of lasers. Fast steering mirror (FSM) driven by piezoelectric ceramics is applied in laser stabilizing. Thanks to the closed loop pointing control, the root mean square error of the optical axis is significantly reduced to be less than 2 microradians. The piston phases of the lasers are locked by an active phase control system based on Field Programmable Gate Array (FPGA) using stochastic parallel gradient descent (SPGD) algorithm. When the total output power of four lasers is 400W, the in-phase peak intensity of the far field spot is increased by a factor of 3.8, reaching 95% of the ideal case. The beam quality of the combined beam is improved by CBC from 1.52x diffraction limit (DL) to 1.26x DL. When the output power is increased to 805W, the phase locking and pointing control still work stably. The results suggest that CBC of solid-state lasers with higher energy could be achieved by using the techniques presented here.

  16. Determination of cobalt in low-alloy steels using laser-induced breakdown spectroscopy combined with laser-induced fluorescence.

    PubMed

    Li, Jiaming; Guo, Lianbo; Zhao, Nan; Yang, Xinyan; Yi, Rongxing; Li, Kuohu; Zeng, Qingdong; Li, Xiangyou; Zeng, Xiaoyan; Lu, Yongfeng

    2016-05-01

    Cobalt element plays an important role for the properties of magnetism and thermology in steels. In this work, laser-induced breakdown spectroscopy combined with laser-induced fluorescence (LIBS-LIF) was studied to selectively enhance the intensities of Co lines. Two states of Co atoms were resonantly excited by a wavelength-tunable laser. LIBS-LIF with ground-state atom excitation (LIBS-LIFG) and LIBS-LIF with excited-state atom excitation (LIBS-LIFE) were compared. The results show that LIBS-LIFG has analytical performance with LoD of 0.82μg/g, R(2) of 0.982, RMSECV of 86μg/g, and RE of 9.27%, which are much better than conventional LIBS and LIBS-LIFE. This work provided LIBS-LIFG as a capable approach for determining trace Co element in the steel industry. PMID:26946032

  17. 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

    NASA Astrophysics Data System (ADS)

    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.

  18. Value of probe-based confocal laser endomicroscopy (pCLE) and dual focus narrow-band imaging (dNBI) in diagnosing early squamous cell neoplasms in esophageal Lugol’s voiding lesions

    PubMed Central

    Prueksapanich, Piyapan; Pittayanon, Rapat; Rerknimitr, Rungsun; Wisedopas, Naruemon; Kullavanijaya, Pinit

    2015-01-01

    Background and study aims: Lugol’s chromoendoscopy provides excellent sensitivity for the detection of early esophageal squamous cell neoplasms (ESCN), but its specificity is suboptimal. An endoscopy technique for real-time histology is required to decrease the number of unnecessary biopsies. This study aimed to compare the ESCN diagnostic capability of probed-based confocal laser endomicroscopy (pCLE) and dual focus narrow-band imaging (dNBI) in Lugol’s voiding lesions. Patients and methods: Patients with a history of head and neck cancer without dysphagia were recruited. Lugol’s voiding lesions larger than 5 mm were sequentially characterized by dNBI and pCLE by two independent operators. Finally, all lesions larger than 5 mm were biopsied followed by histological analysis, which is considered to be the gold standard in cancer diagnosis. The primary outcomes were the sensitivity, specificity, positive predictive value (PPV), negative predictive value (NPV), and the accuracy of the two techniques. Results: In total, 44 patients were enrolled with a mean age of 60 years; 80 % were male. Twenty-one Lugol’s voiding lesions larger than 5 mm were detected in 12 patients. Seven lesions (33 %) from four patients were histologically diagnosed as ESCNs (four with high grade dysplasia and three with low grade dysplasia). The other 14 lesions were histologically confirmed as non-neoplastic: active esophagitis, glycogenation with inflammation, acute ulcer, inlet patch, and unremarkable changes. The sensitivity, specificity, PPV, NPV, and accuracy of pCLE vs. dNBI were 83 % vs. 85 %, 92 % vs. 62 %, 83 % vs. 54 %, 92 % vs. 89 %, and 89 % vs. 70 %, respectively (NS). Conclusions: Asymptomatic patients with a history of head and neck cancer underwent Lugol’s chromoendoscopy based ESCN surveillance. Further characterization of the Lugol’s voiding lesions by advanced imaging showed that both pCLE and dNBI provided good sensitivity in

  19. [Combined helium-neon laser therapy in patients with ischemic heart disease].

    PubMed

    Korochkin, I M; Kartelishev, A V; Babushkina, G V; Kapustina, G M

    1990-03-01

    The paper describes the combined helium-neon-laser (HNL) therapy (intravenous and topical) developed by the authors to treat patients with coronary heart disease. A high efficacy of this therapy mode was demonstrated in patients over 70 years of age with Functional Classes III-IV angina refractory to antianginal agents. The mechanisms responsible for therapeutic efficiency of laser irradiation were studied at the membraneous and cellular levels. There is evidence that the combined HNL-therapy had advantages over topical HNL exposure in terms of higher clinical efficiency and patterns of abnormal chemical changes. PMID:2381119

  20. Diversity combining in laser Doppler vibrometry for improved signal reliability

    SciTech Connect

    Dräbenstedt, Alexander

    2014-05-27

    Because of the speckle nature of the light reflected from rough surfaces the signal quality of a vibrometer suffers from varying signal power. Deep signal outages manifest themselves as noise bursts and spikes in the demodulated velocity signal. Here we show that the signal quality of a single point vibrometer can be substantially improved by diversity reception. This concept is widely used in RF communication and can be transferred into optical interferometry. When two statistically independent measurement channels are available which measure the same motion on the same spot, the probability for both channels to see a signal drop-out at the same time is very low. We built a prototype instrument that uses polarization diversity to constitute two independent reception channels that are separately demodulated into velocity signals. Send and receive beams go through different parts of the aperture so that the beams can be spatially separated. The two velocity channels are mixed into one more reliable signal by a PC program in real time with the help of the signal power information. An algorithm has been developed that ensures a mixing of two or more channels with minimum resulting variance. The combination algorithm delivers also an equivalent signal power for the combined signal. The combined signal lacks the vast majority of spikes that are present in the raw signals and it extracts the true vibration information present in both channels. A statistical analysis shows that the probability for deep signal outages is largely decreased. A 60 fold improvement can be shown. The reduction of spikes and noise bursts reduces the noise in the spectral analysis of vibrations too. Over certain frequency bands a reduction of the noise density by a factor above 10 can be shown.

  1. Wavelength beam combining of a 980-nm tapered diode laser bar in an external cavity

    NASA Astrophysics Data System (ADS)

    Vijayakumar, Deepak; Jensen, Ole Bjarlin; Thestrup, Birgitte

    2010-04-01

    High power diode lasers are used in a large number of applications. A limiting factor for more widespread use of broad area lasers is the poor beam quality. Gain guided tapered diode lasers are ideal candidates for industrial applications that demands watt level output power with good beam quality. By adapting a bar geometry, the output power could be scaled even up to several tens of watts. Unfortunately, the high divergence which is a characteristic feature of the bar geometry could lead to a degradation of the overall beam quality of the laser bar. However, spectral beam combining is an effective solution for preserving the beam quality of the bar in the range of that of a single emitter and at the same time, enabling the power scaling. We report spectral beam combining applied to a 12 emitter tapered laser bar at 980 nm. The external cavity has been designed for a wavelength separation of 4.0 nm between the emitters. An output power of 9 W has been achieved at an operating current of 30 A. The combined beam had an M2 value (1/e2) of 5.3 along the slow axis which is comparable to that of a single tapered emitter on the laser bar. The overall beam combining efficiency was measured to be 63%. The output spectrum of the individual emitters was narrowed considerably. In the free running mode, the individual emitters displayed a broad spectrum of the order of 0.5-1.0 nm while the spectral width has been reduced to 30-100 pm in the spectral beam combining mode.

  2. Effect of Diode Laser Irradiation Combined with Topical Fluoride on Enamel Microhardness of Primary Teeth

    PubMed Central

    Bahrololoomi, Zahra; Lotfian, Malihe

    2015-01-01

    Objectives: Laser irradiation has been suggested as an adjunct to traditional caries prevention methods. But little is known about the cariostatic effect of diode laser and most studies available are on permanent teeth.The purpose of the present study was to investigate the effect of diode laser irradiation combined with topical fluoride on enamel surface microhardness. Materials and Methods: Forty-five primary teeth were used in this in vitro study. The teeth were sectioned to produce 90 slabs. The baseline Vickers microhardness number of each enamel surface was determined. The samples were randomly divided into 3 groups. Group 1: 5% NaF varnish, group 2: NaF varnish+ diode laser at 5 W power and group 3: NaF varnish+ diode laser at 7 W power. Then, the final microhardness number of each surface was again determined. The data were statistically analyzed by repeated measures ANOVA at 0.05 level of significance. Results: In all 3 groups, microhardness number increased significantly after surface treatment (P<0.05). However, Microhardness change after treatment was not significantly different among groups (P >0.05). Conclusion: The combined application of diode laser and topical fluoride varnish on enamel surface did not show any significant additional effect on enamel resistance to caries. PMID:26056517

  3. Interstitial laser and chemotherapy combined for treatment of human squamous cell carcinoma

    NASA Astrophysics Data System (ADS)

    Saxton, Romaine E.; Graeber, Ines P.; Suh, Michael J.; Paek, Woo H.; Paiva, Marcos B.; Castro, Dan J.

    1997-05-01

    We have tested a combined treatment for squamous cell carcinoma based on laser activation of anti-cancer drugs in human solid tumors. Cisplatinum and the new anthrapyrazole CI- 941 are reported to interact with photothermal energy. Combined intratumor drug and interstitial laser therapy were tested in nude mice bearing human squamous cell carcinomas grown as subcutaneous tumors. Cisplatinum injection (1.2 mg/500 mg tumor) 4 hours before KTP laser fiberoptic treatment (532 nm, 0.8 W, 10 sec/site, 300 J) resulted in complete tumor regression in 6/8 animals, while intratumor drug alone led to partial regression and tumor regrowth in 10/10 mice during 12 weeks followup. Laser treatment alone resulted in ablation followed by recurrence in 7/8 cases. Similar laser treatment 4 hours after injection of the light sensitive anthrapyrazole CI-941 led to complete tumor regression in 15/22 cases. CI-941 alone at drug levels up to 1.2 mg/gm tumor in 30 mice induced stasis followed by progression in all cases. Finally, tumor retention of 14C-CI-941 in mice 4 hrs after intralesional injection was 200-fold higher than via the systemic route and by 24 hrs remained 40-fold higher, but drug levels in normal tissues were reduced 10 - 100 fold. The data suggest laser chemotherapy may be a useful new treatment for human cancer.

  4. Fluorescence performance standards for confocal microscopy

    NASA Astrophysics Data System (ADS)

    Rüttinger, Steffen; Kapusta, Peter; Völlkopf, Volker; Koberling, Felix; Erdmann, Rainer; Macdonald, Rainer

    2010-02-01

    State of the art confocal microscopes offer diffraction limited (or even better) spatial resolution, highest (single molecule) sensitivity and ps-fluorescence lifetime measurement accuracy. For developers, manufacturers, as well as users of confocal microscopes it is mandatory to assign values to these qualities. In particular for users, it is often not easy to ascertain that the instrument is properly aligned as a large number of factors influence resolution or sensitivity. Therefore, we aspire to design a set of performance standards to be deployed on a day-to-day fashion in order to check the instruments characteristics. The main quantities such performance standard must address are: • Spatial resolution • Sensitivity • Fluorescence lifetime To facilitate the deployment and thus promote wide range adoption in day-to-day performance testing the corresponding standards have to be ready made, easy to handle and to store. The measurement procedures necessary should be available on as many different setups as possible and the procedures involved in their deployment should be as easy as possible. To this end, we developed two performance standards to accomplish the mentioned goals: • Resolution reference • Combined molecular brightness and fluorescence lifetime reference The first one is based on sub-resolution sized Tetra-SpeckTM fluorescent beads or alternatively on single molecules on a glass surface to image and to determine quantitatively the confocal volume, while the latter is a liquid sample containing fluorescent dyes of different concentrations and spectral properties. Both samples are sealed in order to ease their use and prolong their storage life. Currently long-term tests are performed to ascertain durability and road capabilities.

  5. Combined pulsed dye and CO2 lasers in the treatment of angiolymphoid hyperplasia with eosinophilia.

    PubMed

    Sagi, Lior; Halachmi, Shlomit; Levi, Assi; Amitai, Dan Ben; Enk, Claes D; Lapidoth, Moshe

    2016-08-01

    Angiolymphoid hyperplasia with eosinophilia (ALHE) is an uncommon dermatosis of unknown etiology that manifests as characteristic red nodules and papules with a predilection for the scalp and periauricular region. Treatment is required for both esthetic and functional reasons, as lesions may ulcerate and bleed. Many treatment approaches have been reported, including excision, systemic medical approaches, topical or intralesional therapies, and non-invasive modalities including cryotherapy, electrosurgery, and laser. Treatments have exhibited variable efficacy, and the recurrence rate is 100 %. We report the combination of pulsed dye laser and CO2 laser in the treatment of ALHE in 14 patients. All patients exhibited clinical response after a mean of 2.4 ± 0.4 treatment sessions. The clinical efficacy of the combined treatment, together with its well-tolerated nature, render the use of pulsed dye laser in combination with CO2 laser, a viable treatment for debulking ALHE lesions. Ongoing maintenance treatments are needed to due to the high degree of relapse. PMID:27184154

  6. Combining harmonic generation and laser chirping to achieve high spectral density in Compton sources

    NASA Astrophysics Data System (ADS)

    Terzić, Balša; Reeves, Cody; Krafft, Geoffrey A.

    2016-04-01

    Recently various laser-chirping schemes have been investigated with the goal of reducing or eliminating ponderomotive line broadening in Compton or Thomson scattering occurring at high laser intensities. As a next level of detail in the spectrum calculations, we have calculated the line smoothing and broadening expected due to incident beam energy spread within a one-dimensional plane wave model for the incident laser pulse, both for compensated (chirped) and unchirped cases. The scattered compensated distributions are treatable analytically within three models for the envelope of the incident laser pulses: Gaussian, Lorentzian, or hyperbolic secant. We use the new results to demonstrate that the laser chirping in Compton sources at high laser intensities: (i) enables the use of higher order harmonics, thereby reducing the required electron beam energies; and (ii) increases the photon yield in a small frequency band beyond that possible with the fundamental without chirping. This combination of chirping and higher harmonics can lead to substantial savings in the design, construction and operational costs of the new Compton sources. This is of particular importance to the widely popular laser-plasma accelerator based Compton sources, as the improvement in their beam quality enters the regime where chirping is most effective.

  7. Combination of a Laser and Stem Cells in Posterior Eye Ophthalmology

    NASA Astrophysics Data System (ADS)

    Lukashev, Alexei; Baranov, Eugene; Gavrilova, Natalia; Saburina, Irina; Revischin, Alexander; Tornambe, Paul

    2010-05-01

    Investigation of combined application of different type of cells, delivery methods with laser irradiation of retina on an animal model in vivo(rabbit eye) was the purpose of the study. An argon at 514nm and a dye laser at 577nm with were used to provide a controlled damage on the rabbit retina. Two type of human progenitor stem cells(hPSC) were tested: Mesenchymal and Neural. Four cell delivery methods were compared: Retrobulbar, Introvitreous, Subconjuctival and Suprachoroidal injections. Electroretinography(ERG) was used as a diagnostics of retina functionality. Selective immunohystochemical analysis was performed to assess cells migration and viability. Controlled laser damage on retina provides strong attracting signal for stem cells. Application of laser light enhances results of stem cells injection in posterior eye and may have benefits for treatment of different types of retinopathy and macular degeneration.

  8. Clinical efficacy of the Nd:YAG laser for combination periodontitis therapy.

    PubMed

    Neill, M E; Mellonig, J T

    1997-08-01

    Recent results of a limited clinical trial suggest that mechanical root scaling and root planing therapy alone may not be the most effective mode of treatment for patients affected by moderate to severe adult periodontitis. However, scaling and planing combined with laser therapy utilizing a low-powered pulsed Nd:YAG laser have been shown to be successful in the elimination of the bacteria commonly associated with the development of this oral condition. The double-blind, split mouth design study involved 10 human subjects randomly assigned to one of three treatments: 1) scaling and root planing alone, 2) dental laser plus scaling and root planing, and 3) control only. This article presents the clinical results of the trial, which suggest that laser therapy is a viable adjunct to local, nonsurgical therapy such as scaling and planing. PMID:9573831

  9. Numerical analysis of the beam quality and spectrum of wavelength-beam-combined laser diode arrays

    NASA Astrophysics Data System (ADS)

    Tang, Xuan; Wang, Xiao-Jun; Ke, Wei-Wei

    2015-02-01

    In this paper, a numerical model is presented to simulation the performance of the wavelength-beam-combined laser diode arrays (LDA) system. The eigen mode expansion method is used to describe the two-dimensional optical amplification and the strength of field feedback of external cavity. To describe the mode competition in laser diodes, the gain saturation effect is considered. The two-dimension distributions of the carrier concentration, recombination rates, and optical gain are calculated for solving the laser dynamic equation. The Fresnel integration, grating equation and mode overlap integration are used to obtain the feedback coefficient of extent cavity diffraction. Quantum noise is considered to evaluate the spectral linewidth of semiconductor laser. Based on the numerical model, the impact of the mutual optical feedback on the beam quality and spectrum of the LDA is present and analysis.

  10. Plasma formation on a metal surface under combined action of laser and microwave radiation

    SciTech Connect

    Gavrilyuk, A P; Shaparev, N Ya

    2013-10-31

    By means of numerical modelling of the combined effect of laser (1.06 mm) and microwave (10{sup 10} – 10{sup 13} s{sup -1}) radiation on the aluminium surface in vacuum it is shown that the additional action of microwave radiation with the frequency 10{sup 12} s{sup -1} provides complete ionisation of the metal vapour (for the values of laser radiation duration and intensity used in the calculations), while in the absence of microwave radiation the vapour remains weakly ionised. The mathematical model used accounts for the processes, occurring in the condensed phase (heat conduction, melting), the evaporation and the kinetic processes in the resulting vapour. (interaction of laser radiation with matter. laser plasma)

  11. Combined Laser and Electron Cooling of Bunched C3+ Ion Beams at the Storage Ring ESR

    SciTech Connect

    Schramm, U.; Bussmann, M.; Habs, D.; Kuehl, T.; Beller, P.; Franzke, B.; Nolden, F.; Steck, M.; Saathoff, G.; Reinhardt, S.; Karpuk, S.

    2006-03-20

    We report on first laser cooling studies of bunched beams of triply charged carbon ions stored at an energy of 1.46 GeV at the ESR (GSI). Despite for the high beam energy and charge state laser cooling provided a reduction of the momentum spread of one order of magnitude in space-charge dominated bunches as compared to electron cooling. For ion currents exceeding 10 {mu}A intra-beam-scattering losses could not be compensated by the narrow band laser system presently in use. Yet, no unexpected problems occurred encouraging the envisaged extension of the laser cooling to highly relativistic beams. At ESR, especially the combination with modest electron cooling provided three-dimensionally cold beams in the plasma parameter range of unity, where ordering effects can be expected and a still unexplained signal reduction of the Schottky signal is observed.

  12. Super-intense femtosecond multichannel laser system with coherent beam combining

    NASA Astrophysics Data System (ADS)

    Bagayev, S. N.; Trunov, V. I.; Pestryakov, E. V.; Frolov, S. A.; Leshchenko, V. E.; Kokh, A. E.; Vasiliev, V. A.

    2014-07-01

    The conceptual design of ultra-high intensity multichannel laser system with coherent beam combining is presented. Design of 1 PW and 10 PW laser channels with pulse repetition rate of 10 Hz based on optical parametric amplification in LBO crystals is considered. Requirements of the most critical pulse parameters for high efficiency coherent beam combining and their dependence on the number of channels is analyzed. Experimentally coherent beam combining of parametrically amplified compressed femtosecond pulses is demonstrated for the first time. Original two-loop relative timing jitter active stabilization scheme is proposed and experimentally investigated. 97% coherent beam combining efficiency is achieved with 110 as relative timing jitter.

  13. Frequency Division Multiplexed Multichannel High-Speed Fluorescence Confocal Microscope

    PubMed Central

    Wu, Fei; Zhang, Xueqian; Cheung, Joseph Y.; Shi, Kebin; Liu, Zhiwen; Luo, Claire; Yin, Stuart; Ruffin, Paul

    2006-01-01

    In this article, we report a new type of fluorescence confocal microscope: frequency division multiplexed multichannel fluorescence confocal microscope, in which we encode the spatial location information into the frequency domain. In this microscope, the exciting laser beam is first split into multiple beams and each beam is modulated at a different frequency. These multiple beams are focused at different locations of the target to form multiple focal points, which further generate multiple fluorescent emission spots. The fluorescent emissions from different focal points are also modulated at different frequencies, because the exciting beams are modulated at different frequencies (or difference carrier frequency). Then, all the fluorescent emissions (modulated at different frequencies) are collected together and detected by a highly sensitive, large-dynamic-range photomultiplier tube. By demodulating the detected signal (i.e., via the Fourier transform), we can distinguish the fluorescent light emitted from the different locations by the corresponding carrier frequencies. The major advantage of this unique fluorescence confocal microscope is that it not only has a high sensitivity because of the use of photomultiplier tube but also can get multiple-point data simultaneously, which is crucial to study the dynamic behavior of many biological process. As an initial step, to verify the feasibility of the proposed multichannel confocal microscope, we have developed a two-channel confocal fluorescence microscope and applied it to study the dynamic behavior of the changes of the calcium ion concentration during the single cardiac myocyte contraction. Our preliminary experimental results demonstrated that we could indeed realize multichannel confocal fluorescence microscopy by utilizing the frequency division multiplexed microscope, which could become an effective tool to study the dynamic behavior of many biological processes. PMID:16815894

  14. Comparison possibilities of ultrasound and its combination with laser in surgery and therapy

    NASA Astrophysics Data System (ADS)

    Zharov, Vladimir P.; Menyaev, Yulian A.; Kabisov, Ruslan K.; Alkov, Sergey V.; Nesterov, A. V.; Loshchilov, Vladimir I.; Suen, James Y.

    2000-05-01

    This article presents the further developments of combined laser-ultrasound medical technologies with paying attention the possibility ultrasound in surgery and therapy. The analyses of main effects at the low frequency ultrasonic treatment of biotissues including cavitation, acoustic streams, acoustic pressure, mechanical influence etc are analyzed. The main promising areas of application of low frequency ultrasound are considered including bactericidal treatment of infections wounds, spray treatment of wounds in head and neck surgery, tumor treatment etc. In particular the clinical result of using ultrasonic devices based on imposing ultrasonic oscillations in a range of 22-66 kHz on a cutting instrument with a special form, radiation intensity up to 10 W/cm2 and oscillation amplitude up to 40-60 micrometers with respect to oncology for halt bleeding from a tumor, liquidating pain, acoustic denervation are presented. Some limitation of medical application of ultrasound are discussed and perspective combination with laser for increasing efficiency of new combined technologies are found. Among them: combination photodynamic therapy and ultrasonic treatment of tumors, laser-ultrasonic treatment of infections wounds including using spray, laser-ultrasonic drug delivery. The preliminary result of experimental study of some of above-mentioned technologies are presented.

  15. One shot confocal microscopy based on wavelength/space conversion by use of multichannel spectrometer

    NASA Astrophysics Data System (ADS)

    Miyamoto, Shuji; Hase, Eiji; Ichikawa, Ryuji; Mnamikawa, Takeo; Yasui, Takeshi; Yamamoto, Hirotugu

    2016-03-01

    Confocal laser microscope (CLM) has been widely used in the fields of the non-contact surface topography, biomedical imaging, and other applications, because of two-dimensional (2D) or three-dimensional (3D) imaging capability with the confocal effect and the stray light elimination. Although the conventional CLM has acquired the 2D image by mechanical scanning of the focused beam spot, further reduction of image acquisition time and the robustness to various disturbances are strongly required. To this end, it is essential to omit mechanical scanning for the image acquisition. In this article, we developed the scan-less, full-field CLM by combination of the line-focused CLM with the wavelength/1D-space conversion. This combination enables us to form the 2D focal array of a 2D rainbow beam on a sample and to encode the 2D image information of a sample on the 2D rainbow beam. The image-encoded 2D rainbow beam was decoded as a spectral line image by a multi-channel spectrometer equipped with a CMOS camera without the need for the mechanical scanning. The confocal full-field image was acquired during 0.23 ms with the lateral resolution of 26.3μm and 4.9μm for the horizontal and vertical directions, respectively, and the depth resolution of 34.9μm. We further applied this scan-less, full-field CLM for biomedical imaging of a sliced specimen and non-contact surface topography of an industry products. These demonstrations highlight a high potential of the proposed scan-less, full-field CLM.

  16. Confocal endomicroscopy: Is it time to move on?

    PubMed Central

    Robles-Medranda, Carlos

    2016-01-01

    Confocal laser endomicroscopy permits in-vivo microscopy evaluation during endoscopy procedures. It can be used in all the parts of the gastrointestinal tract and includes: Esophagus, stomach, small bowel, colon, biliary tract through and endoscopic retrograde cholangiopancreatography and pancreas through needles during endoscopic ultrasound procedures. Many researches demonstrated a high correlation of results between confocal laser endomicroscopy and histopathology in the diagnosis of gastrointestinal lesions; with accuracy in about 86% to 96%. Moreover, in spite that histopathology remains the gold-standard technique for final diagnosis of any diseases; a considerable number of misdiagnosis rate could be present due to many factors such as interpretation mistakes, biopsy site inaccuracy, or number of biopsies. Theoretically; with the diagnostic accuracy rates of confocal laser endomicroscopy could help in a daily practice to improve diagnosis and treatment management of the patients. However, it is still not routinely used in the clinical practice due to many factors such as cost of the procedure, lack of codification and reimbursement in some countries, absence of standard of care indications, availability, physician image-interpretation training, medico-legal problems, and the role of the pathologist. These limitations are relative, and solutions could be found based on new researches focused to solve these barriers. PMID:26788257

  17. Determination of phosphorus in steel by the combined technique of laser induced breakdown spectrometry with laser induced fluorescence spectrometry

    NASA Astrophysics Data System (ADS)

    Kondo, Hiroyuki; Hamada, Naoya; Wagatsuma, Kazuaki

    2009-09-01

    Laser induced breakdown spectrometry (LIBS) combined with laser induced fluorescence spectrometry (LIFS) has been applied for detection of trace-level phosphorus in steel. The plasma induced by irradiation of Nd:YAG laser pulse for ablation was illuminated by the 3rd harmonic of Ti:Sapphire laser tuned to one of the resonant lines for phosphorus in the wavelength region of 253-256 nm. An excitation line for phosphorus was selected to give the highest signal-to-noise ratio. Fluorescence signals, P213.62 and P214.91 nm, were observed with high selectivity at the contents as low as several tens µg g - 1 . Fluorescence intensities were in a good linear correlation with the contents. Fluorescence intensity ratio of a collisionally assisted line (213.62 nm) to a direct transition line (214.91 nm) was discussed in terms of the analytical conditions and experimental results were compared with a calculation based on rate equations. Since the fluorescence signal light in the wavelength range longer than 200 nm can be transmitted relatively easily, even through fiber optics of moderate length, LIBS/LIFS would be a versatile technique in on-site applications for the monitoring of phosphorus contents in steel.

  18. Detection of trace phosphorus in steel using laser-induced breakdown spectroscopy combined with laser-induced fluorescence.

    PubMed

    Shen, X K; Wang, H; Xie, Z Q; Gao, Y; Ling, H; Lu, Y F

    2009-05-01

    Monitoring of light-element concentration in steel is very important for quality assurance in the steel industry. In this work, detection in open air of trace phosphorus (P) in steel using laser-induced breakdown spectroscopy (LIBS) combined with laser-induced fluorescence (LIF) has been investigated. An optical parametric oscillator wavelength-tunable laser was used to resonantly excite the P atoms within plasma plumes generated by a Q-switched Nd:YAG laser. A set of steel samples with P concentrations from 3.9 to 720 parts in 10(6) (ppm) were analyzed using LIBS-LIF at wavelengths of 253.40 and 253.56 nm for resonant excitation of P atoms and fluorescence lines at wavelengths of 213.55 and 213.62 nm. The calibration curves were measured to determine the limit of detection for P in steel, which is estimated to be around 0.7 ppm. The results demonstrate the potential of LIBS-LIF to meet the requirements for on-line analyses in open air in the steel industry. PMID:19412215

  19. Confocal Raman Imaging of Polymeric Materials

    NASA Astrophysics Data System (ADS)

    Schmidt, Ute; Müller, Jörg; Koenen, Joachim

    Polymers play an essential role in modern materials science. Due to the wide variety of mechanical and chemical properties of polymers, they are used in almost every field of application and are still a dynamic area in the development of new materials with demanding requirements. Raman spectroscopy is one of the standard characterization techniques used to uniquely determine the chemical composition of a polymer. Modern materials, however, are generally heterogeneous, in which various chemical components or polymorphs of the same chemical species can be present in a very small sample volume. For the analysis of such heterogeneous materials, the combination of Raman spectroscopy with confocal microscopy delivers information about the spatial distribution of the various chemical species with a resolution down to 200 nm. The aim of this contribution is to demonstrate the power of confocal Raman imaging for the characterization of heterogeneous polymeric materials. The first section will deal with polymorphs of polypropylene in polymer films, followed by the nondestructive analysis of polymer blends. A later section will deal with multi-layer polymer coatings and paints and finally various additives to polymer matrices will be discussed.

  20. High-resolution confocal Raman microscopy using pixel reassignment.

    PubMed

    Roider, Clemens; Ritsch-Marte, Monika; Jesacher, Alexander

    2016-08-15

    We present a practical modification of fiber-coupled confocal Raman scanning microscopes that is able to provide high confocal resolution in conjunction with high light collection efficiency. For this purpose, the single detection fiber is replaced by a hexagonal lenslet array in combination with a hexagonally packed round-to-linear multimode fiber bundle. A multiline detector is used to collect individual Raman spectra for each fiber. Data post-processing based on pixel reassignment allows one to improve the lateral resolution by up to 41% compared to a single fiber of equal light collection efficiency. We present results from an experimental implementation featuring seven collection fibers, yielding a resolution improvement of about 30%. We believe that our implementation represents an attractive upgrade for existing confocal Raman microscopes that employ multi-line detectors. PMID:27519099

  1. Combination Of Narrow Bandwidth Excimer Laser And Monochromatic Reduction Projection Lens

    NASA Astrophysics Data System (ADS)

    Kajiyama, K.; Saito, K.; Moro, N.; Maeda, Y.; Natsuaki, H.

    1988-01-01

    This paper will discuss the problems associated with excimer laser photo-lithography -the combination of a KrF narrow band width excimer laser (non-injection locked type) with a large field fused silica monochromatic reduction lens. An excimer laser with a KrF narrow bandwidth, in combination with a large field monochromatic lens which is appropriate for use with such laser, have been developed and tested. The system's resolution capability has been confirmed at 0.4 um L/S with MP2400 resist. The laser has been designed so as to be installed and maintained in a clean room environment as well as to have a very narrow spectrum line. A very narrow band-width beam, down to 0.003nm, has been attained through a stable resonator with more than 20mJ pulse energy. The ultra-compact laser head (300mm x 545mm x 1100mm) contains a small laser discharge unit (182mm x 156mm x 584mm), and no amplifier because the oscillator is highly efficient in spite of the narrow line emission. Maintenance is much easier in the clean room environment. Users can replace the discharge unit as easily as they would change Hg-lamp, only taking twenty minutes, and while they clean the window and check the electrodes of the removed unit, the laser can be operated with the easily installed replacement -already passivated discharge unit. The laser head unit is separated from a gas circulating unit and trigger pulse circuit - vibration, heat, EMI noise and particle generation. Therefore, it can be installed even in the thermal clean chamber of a stepper. The N.A. (numerical aperture) of the monochromatic lens is 0.36 and the field size is 15mm x 15mm. In fact, three kinds of lenses with N.A.s of 0.4, 0.35 and 0.3 respectively, were designed and individually evaluated for their OTF's and defocus's dependence on the light source's spectral width, and also their co-relationship. In parallel, simulations on the relationship between each lens' chromatic aberration and laser spectral width were completed and

  2. An Automatic Procedure for Combining Digital Images and Laser Scanner Data

    NASA Astrophysics Data System (ADS)

    Moussa, W.; Abdel-Wahab, M.; Fritsch, D.

    2012-07-01

    Besides improving both the geometry and the visual quality of the model, the integration of close-range photogrammetry and terrestrial laser scanning techniques directs at filling gaps in laser scanner point clouds to avoid modeling errors, reconstructing more details in higher resolution and recovering simple structures with less geometric details. Thus, within this paper a flexible approach for the automatic combination of digital images and laser scanner data is presented. Our approach comprises two methods for data fusion. The first method starts by a marker-free registration of digital images based on a point-based environment model (PEM) of a scene which stores the 3D laser scanner point clouds associated with intensity and RGB values. The PEM allows the extraction of accurate control information for the direct computation of absolute camera orientations with redundant information by means of accurate space resection methods. In order to use the computed relations between the digital images and the laser scanner data, an extended Helmert (seven-parameter) transformation is introduced and its parameters are estimated. Precedent to that, in the second method, the local relative orientation parameters of the camera images are calculated by means of an optimized Structure and Motion (SaM) reconstruction method. Then, using the determined transformation parameters results in having absolute oriented images in relation to the laser scanner data. With the resulting absolute orientations we have employed robust dense image reconstruction algorithms to create oriented dense image point clouds, which are automatically combined with the laser scanner data to form a complete detailed representation of a scene. Examples of different data sets are shown and experimental results demonstrate the effectiveness of the presented procedures.

  3. Influence of surface laser cleaning combined with substrate preheating on the splat morphology

    NASA Astrophysics Data System (ADS)

    Costil, S.; Liao, H.; Gammoudi, A.; Coddet, C.

    2005-03-01

    The morphology of sprayed splats influences the coating adhesion and properties, which are determined by the spraying parameters. Many studies in this field show that the substrate surface temperature is a very relevant factor for the splat shape: the hypotheses of substrate surface wettability and contamination or adsorption layer on the surfaces are supported by the fact that the near-disk-shaped splat can be obtained by increasing the substrate temperature. In this work, a short-duration pulsed laser was used to ablate the substrate just before powder spraying. This ablation was powerful enough to eliminate the contaminants on the substrate surface and to improve the adhesion. In this study the analyses of NiAl splat morphology on the polished TA6V (Ti-6Al-4V) substrate were carried out using laser ablation with different substrate temperatures and different heating modes: the flame and another laser. Results show that the temperature at which the disk-shaped splat can be obtained decreased dramatically by laser ablation. Moreover, laser ablation combined with another laser increased the adhesion strength of the coatings.

  4. Chromatic confocal microscopy using staircase diffractive surface.

    PubMed

    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. PMID:25320920

  5. Deep stroma investigation by confocal microscopy

    NASA Astrophysics Data System (ADS)

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

    2015-03-01

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

  6. The technique of high power laser beam combination using liquid crystal spatial light modulator

    NASA Astrophysics Data System (ADS)

    Duan, Ying-ying; Wang, Xiang-ru; Huang, Zi-qiang

    2013-09-01

    Based on the phase modulation characteristics of optically addressed liquid crystal spatial light modulator (OA-LC-SLM) which is realized by controlling the power of addressing light, a physical model of coherent beam combination fiber laser using a bunch of fibers and a device of OA-LC-SLM is established on the theory of diffraction optics and liquid crystal birefringence effect. On the basis of this model, the properties of given scheme of coherent beam combination fiber laser are investigated including main lobe distribution and ability of phase modulation. Meanwhile, the plot functions of phase modulation versus the optical power of addressing light are obtained on different given driving voltage conditions and fiber alignment parameters such as core diameter and filling factor. After the numerical simulation, it shows that, this coherent beam combination fiber laser using OA-LC-SLM demonstrates an ability of coherent beam combination on the far field. With the increase of core diameter, the combination efficiency is improved better, and the divergence angle decreases narrower.

  7. Two-photon fluorescence imaging of embryo with much less damage than confocal imaging

    NASA Astrophysics Data System (ADS)

    Liu, Bian; Xu, Hui; Jin, Lei; Ma, Hui; Chen, Die Yan

    2002-09-01

    Two-photon Laser Scanning Microscopy (TPLSM) is a novel technique based on the two-photon excitation of fluorophore. In this paper, TPLSM and traditional confocal microscopy are introduced. And the influence of femtosecond near-infrared (NIR) illumination on mouse embryos is investigated for the first time. The result shows that NIR laser has much less damage to embryos than blue laser and proves that TPLSM is superior to conventional confocal microscopy in keeping sample alive. TPLSM enables us to make a continuous observation for a longer time on embryogenesis.

  8. Enhanced laser tissue soldering using indocyanine green chromophore and gold nanoshells combination

    NASA Astrophysics Data System (ADS)

    Khosroshahi, Mohammad E.; Nourbakhsh, Mohammad S.

    2011-08-01

    Gold nanoshells (GNs) are new materials that have an optical response dictated by the plasmon resonance. The wavelength at which the resonance occurs depends on the core and shell sizes. The purposes of this study were to use the combination of indocyanine green (ICG) and different concentration of gold nanoshells for skin tissue soldering and also to examine the effect of laser soldering parameters on the properties of repaired skin. Two mixtures of albumin solder and different combinations of ICG and gold nanoshells were prepared. A full thickness incision of 2 × 20 mm2 was made on the surface and after addition of mixtures it was irradiated by an 810 nm diode laser at different power densities. The changes of tensile strength (σt) due to temperature rise, number of scan (Ns), and scan velocity (Vs) were investigated. The results showed at constant laser power density (I), σt of repaired incisions increases by increasing the concentration of gold nanoshells in solder, Ns, and decreasing Vs. It was demonstrated that laser soldering using combination of ICG + GNs could be practical provided the optothermal properties of the tissue are carefully optimized. Also, the tensile strength of soldered skin is higher than skins that soldered with only ICG or GNs. In our case, this corresponds to σt = 1800 g cm-2 at I ~ 47 Wcm-2, T ~ 85 ºC, Ns = 10, and Vs = 0.3 mms-1.

  9. Enhanced laser tissue soldering using indocyanine green chromophore and gold nanoshells combination.

    PubMed

    Khosroshahi, Mohammad E; Nourbakhsh, Mohammad S

    2011-08-01

    Gold nanoshells (GNs) are new materials that have an optical response dictated by the plasmon resonance. The wavelength at which the resonance occurs depends on the core and shell sizes. The purposes of this study were to use the combination of indocyanine green (ICG) and different concentration of gold nanoshells for skin tissue soldering and also to examine the effect of laser soldering parameters on the properties of repaired skin. Two mixtures of albumin solder and different combinations of ICG and gold nanoshells were prepared. A full thickness incision of 2 × 20 mm(2) was made on the surface and after addition of mixtures it was irradiated by an 810 nm diode laser at different power densities. The changes of tensile strength (σ(t)) due to temperature rise, number of scan (Ns), and scan velocity (Vs) were investigated. The results showed at constant laser power density (I), σ(t) of repaired incisions increases by increasing the concentration of gold nanoshells in solder, Ns, and decreasing Vs. It was demonstrated that laser soldering using combination of ICG + GNs could be practical provided the optothermal properties of the tissue are carefully optimized. Also, the tensile strength of soldered skin is higher than skins that soldered with only ICG or GNs. In our case, this corresponds to σ(t) = 1800 g cm(-2) at I ∼ 47 Wcm(-2), T ∼ 85 [ordinal indicator, masculine]C, Ns = 10, and Vs = 0.3 mms(-1). PMID:21895342

  10. Treatment of angiokeratoma of Mibelli alone or in combination with pulsed dye laser and long-pulsed Nd: YAG laser.

    PubMed

    Zeng, Ying; Li, Xi-Qing; Lin, Qiong-Zhu; Zhan, Kui

    2014-01-01

    Treatment of angiokeratoma of Mibelli is usually challenging because of the location, the pathogenetic condition and the cosmetic requirements. We present our characteristic treatment with the application of pulsed dye laser PDL and lpNd:YAG laser. All of these lesions were treated by topical anesthesia with Emla. Combined dual PDL-lpNd:YAG (PDL: 595 nm, 5 mm/7 mm, 0.5 ms, 8-10 J/cm(2) ; lpNd:YAG: 3 mm/5 mm, 15 ms, 90-120 J/cm(2) ) treatment was used to treat lesions which with moderate to severe hyperkeratosis and hyperplasia. To the maculopapule ones, the energy density of lpNd:YAG might upgrade to 150 J/cm(2) . Singular PDL (595 nm, 5 mm/7 mm, 0.5 ms, 9-12 J/cm(2) ) treatment was used to treat lesions which with slight hyperkeratosis and hyperplasia. Continuous airflow cooling was always applied during the laser treatment. The treatment interval was 6-12 weeks. Of the 5 patients, 3 of them were cured and 2 of them were improved. All of them were satisfied with the cosmetic results. We recommended the combined dual PDL-lpNd:YAG laser in treating severe hyperkeratotic and hyperplastic angiokeratoma of Mibelli. It can aid in achieving a desirable outcome whilst also reducing the required treatment sessions. However, most patients felt painful during the operation and experienced a severe long term recovery time after operation. PMID:24911941