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3D reconstruction of the material surface shape from stereo images of electron micrographs

3D reconstruction of the material surface shape from stereo ...

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conference on Three-dimensional microscopy - image acquisition and processing III

作者： Pang, LY STANFORD UNIV DEPT MECH ENGNSTANFORDCA 94305

ISBN: (纸本)0819420298

Shape reconstruction from stereo images of fracture surfaces, which are obtained by scanning electron microscopy, is applied to research in micromechanics and fracture mechanics. The shape is reconstructed by matching the corresponding points on a pair of stereo images. To deal with the problem of false-target matching and to reduce computation time, the `interesting points', which usually are pixel locations of important features in each pair of images, are determined first, and the matching is only performed on these `interesting points'. Two algorithms are used for finding the `interesting points'. The first one selects the pixels that have the maximum value of local entropy. The second one selects pixels having the local maximum gray scale value after the image has been processed by a Laplacian of Gaussian filter. In order to reduce the effects of shadows, the logarithm of intensity is calculated before the image is filtered. The algorithm of matching corresponding points in the stereo images is based on the theory of cross correlation. To reduce the false-matching, a coarse and fine matching algorithm is developed. Finally, the height of the surface is calculated by applying stereo triangulation theory.

关键词： shape reconstruction cross correlation image processing experimental mechanics

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Tradeoffs in regularized maximum-likelihood image restoration

Tradeoffs in regularized maximum-likelihood image restoratio...

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conference on Three-dimensional microscopy - image acquisition and processing IV

作者： Markham, J Conchello, JA Washington Univ. (United States)

ISBN: (纸本)0819423955

All algorithms for three-dimensional deconvolution of fluorescence microscopical images have as a common goal the estimation of a specimen function (SF) that is consistent with the recorded image and the process for image formation and recording. To check for consistency, the image of the estimated SF predicted by the imaging operator is compared to the recorded image, and the similarity between them is used as a figure of merit (FOM) in the algorithm to improve the specimen function estimate. Commonly used FOM's include squared differences, maximum entropy, and. maximum likelihood (Mi,). The imaging operator is usually characterized by the point-spread function (PSF), the image of a point source of light, or its Fourier transform, the optical transfer function (OTF). Because the OTF is non-zero only over a small region of the spatial-frequency domain, the inversion of the image formation operator is non-unique and the estimated SF is potentially artifactual. Adding a term to the FOM that penalizes some unwanted behavior of the estimated SF effectively ameliorates potential artifacts, but at the same time biases the estimation process. For example, an intensity penalty avoids overly large pixel values but biases the SF to small pixel values. A roughness penalty avoids rapid pixel to pixel variations bur biases the SF to be smooth. In this article we assess the effects of the roughness and intensity penalties on maximum likelihood image estimation.

关键词： 3D microscopy maximum penalized likelihood deconvolution evaluation

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Restoration of three-dimensional quasi-binary images from confocal microscopy and its application to dendritic trees

Restoration of three-dimensional quasi-binary images from co...

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conference on Three-dimensional microscopy - image acquisition and processing IV

作者： Herzog, A Krell, G Michaelis, B Wang, JZ Zuschratter, W Braun, K Otto-von-Guericke Univ. Magdeburg (FRG) and Federal Institute of Neurobiology (Germany)

ISBN: (纸本)0819423955

For the analysis of learning processes and the underlying changes of the shape of excitatory synapses (spines), 3-D volume samples of selected dendritic segments are scanned by a confocal laser scanning microscope. For a :more detailed analysis, such as the classification of spine types, binary images of higher resolution are required. Simple threshold methods have disadvantages for small structures because the microscope point spread function (PSF) causes a darkening and a spread. The direction-dependent PSF leads to shape errors. To reconstruct structures and edge positions with a resolution smaller that one voxel a parametric model for the dendrite and the spines is created. In our application we use the known tree-like structure of the nerve cell as a-priori information. To create the model, simple geometrical elements (cylinders with hemispheres at the ends) are connected. The model can be adapted for size and position in sub-pixel domain. To estimate the quadratic error between the microscope image and the model, the model is sampled with the same resolution as the microscope image and convolved by the microscope PSF. During an iterative process the parameters of the model are optimised. In contrast to other pixel-based methods the number of variable parameters is much slower. The influence of small deviations in the microscope image (caused by the inhomogeneous biological material) is reduced.

关键词： deconvolution binarisation restoration confocal microscopy

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3D reconstruction methods using line-scanning microscopy with a linear sensor

3D reconstruction methods using line-scanning microscopy wit...

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conference on Advanced microscopy Techniques III

作者： Macedo, Milton P. Correia, C. M. B. A. Univ Coimbra Dept Phys Ctr Instrumentat Coimbra Portugal

ISBN: (纸本)9780819496461

Line-scanning microscopy is a technique with ability to deliver images with an higher acquisition rate than confocal microscopy. But it is accomplished at expense of the degradation of resolution for details parallel to sensor if slit detectors are used. With a linear image sensor it is possible to attenuate or even cancel this effect through the use of information stored in each pixel / light distribution across line pixels of the sensor. In spite of its great potential the use of linear image sensors and in particular the development of three-dimensional (3D) reconstruction methods that take into account its specificity is scarce. This led to our motivation to build a laboratory prototype of a bench stage-scanning microscope using a linear image sensor. We aim at improving lateral resolution isotropy but also image visualization and 3D mesh reconstruction using different optical setups particularly illumination modes, e. g., widefield and line-illumination. The versatility of the laboratory prototype namely its software for image acquisition, processing and visualization is important to attain this goal in the sense that it provides excellent means to develop and test algorithms. Several algorithms for 3D reconstruction were developed and are presented and discussed in this paper. Results of the application of these 3D reconstruction methods show the improvements on lateral resolution isotropy and depth discrimination achieved using algorithms integrating sensor geometry or spatial sampling rate. Also it is evidenced the impact of an insufficient spatial sampling rate from 3D mesh reconstructions.

关键词： Three-dimensional microscopy slit-scanning microscopy image reconstruction techniques linear image sensor lateral resolution depth discrimination

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Tunable thin-film optical filters for hyperspectral microscopy

Tunable thin-film optical filters for hyperspectral microsco...

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conference on Three-dimensional and multidimensional microscopy - image acquisition and processing XX

作者： Favreau, Peter F. Rich, Thomas C. Prabhat, Prashant Leavesley, Silas J. Univ S Alabama Biomol Chem engn Mobile AL 36688 USA Univ S Alabama Pharmacol Mobile AL 36688 USA Univ S Alabama Ctr Lung Biol Mobile AL 36688 USA Semrock Inc A Unit Index Mobile AL 36688 USA

ISBN: (纸本)9780819493583

Hyperspectral imaging was originally developed for use in remote sensing applications. More recently, it has been applied to biological imaging systems, such as fluorescence microscopes. The ability to distinguish molecules based on spectral differences has been especially advantageous for identifying fluorophores in highly autofluorescent tissues. A key component of hyperspectral imaging systems is wavelength filtering. Each filtering technology used for hyperspectral imaging has corresponding advantages and disadvantages. Recently, a new optical filtering technology has been developed that uses multi-layered thin-film optical filters that can be rotated, with respect to incident light, to control the center wavelength of the pass-band. Compared to the majority of tunable filter technologies, these filters have superior optical performance including greater than 90% transmission, steep spectral edges and high out-of-band blocking. Hence, tunable thin-film optical filters present optical characteristics that may make them well-suited for many biological spectral imaging applications. An array of tunable thin-film filters was implemented on an inverted fluorescence microscope (TE 2000, Nikon Instruments) to cover the full visible wavelength range. images of a previously published model, GFP-expressing endothelial cells in the lung, were acquired using a charge-coupled device camera (Rolera EM-C2, Q-Imaging). This model sample presents fluorescently-labeled cells in a highly autofluorescent environment. Linear unmixing of hyperspectral images indicates that thin-film tunable filters provide equivalent spectral discrimination to our previous acousto-optic tunable filter-based approach, with increased signal-to-noise characteristics. Hence, tunable multi-layered thin film optical filters may provide greatly improved spectral filtering characteristics and therefore enable wider acceptance of hyperspectral widefield microscopy.

关键词： Hyperspectral imaging spectral imaging microscopy AOTF acousto-optic tunable filter TFTF thin-film tunable filter VersaChrome

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The Gray Institute Open Microscopes applied to radiobiology and protein interaction studiesZ

The Gray Institute Open Microscopes applied to radiobiology ...

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conference on Three-dimensional and multidimensional microscopy - image acquisition and processing XXI

作者： Barber, P. R. Tullis, I. D. C. Rowley, M. I. Martins, C. D. Weitsman, G. Lawler, K. Coffey, M. Woodman, N. Gillett, C. E. Ng, T. Vojnovic, B. Univ Oxford Gray Inst Radiat Oncol & Biol Dept Oncol Oxford OX1 2JD England Inst Math & Molecular Biomed Kings Coll London London England Kings Coll London Div Cancer Studies London England Kings Coll London Randall Div London England Kings Coll London Breakthrough Breast Cancer Res Unit London England Kings Coll London Guys & St Thomas Breast Tissue & Data Bank London England

ISBN: (纸本)9780819498625

We describe an 'open' design methodology for wide-field fluorescence, confocal and fluorescence lifetime imaging microscopy (FLIM), and how the resulting microscopes are being applied to radiation biology and protein activity studies in cells and human tissue biopsies. The design approach allows easy expansion as it moves away from the use of a monolithic microscope body to small, commercial off-the-shelf and custom made modular components. Details have been made available under an open license for non-commercial use at http://***/similar to atdgroup. Two radiobiology 'end-stations' have been constructed which enable fast radiation targeting and imaging of biological material opening up completely novel studies, where the consequences of ionising radiation (signaling and protein recruitment) can be studied in situ, at short times following irradiation. One is located at Surrey University, UK, where radiation is a highly focused in beam (e.g. protons, helium or higher mass ions). The second is installed at the Gray Institute linear accelerator facility, Oxford University, which uses sub-microsecond pulses of 6 MeV electrons. FLIM capabilities have enhanced the study of protein-protein interactions in cells and tissues via Frster Resonance Energy Transfer (FRET). Extracting FRET signals from breast cancer tissue is challenging because of endogenous and fixation fluorescence;we are investigating novel techniques to measure this robustly. Information on specific protein interactions from large numbers of patient tumors will reveal prognostic and diagnostic information.

关键词： Tissues Fluorescence resonance energy transfer Microscopes image filtering Atrial fibrillation Fluorescence lifetime imaging Optical filters Proteins Luminescence Linear filtering

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Hyperspectral imaging fluorescence excitation scanning (HIFEX) microscopy for live cell imaging 26

Hyperspectral imaging fluorescence excitation scanning (HIFE...

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conference on Three-dimensional and multidimensional microscopy - image acquisition and processing XXVI

作者： Leavesley, Silas J. Griswold, John Robert Deal, Joshua McAlister, Kathleen Mayes, Sam Browning, Craig Parker, Marina Mayes, Samantha Gunn Rich, Thomas C. Univ S Alabama Dept Chem & Biomol Engn Mobile AL 36688 USA Univ S Alabama Dept Pharmacol Mobile AL 36688 USA Univ S Alabama Ctr Lung Biol Mobile AL 36688 USA Univ S Alabama Dept Biomed Sci Mobile AL 36688 USA Univ S Alabama Syst Engn Mobile AL 36688 USA

ISBN: (纸本)9781510624092

In the past two decades, spectral imaging technologies have expanded the capacity of fluorescence microscopy for accurate detection of multiple labels, separation of labels from cellular and tissue autofluorescence, and analysis of autofluorescence signatures. These technologies have been implemented using a range of optical techniques, such as tunable filters, diffraction gratings, prisms, interferometry, and custom Bayer filters. Each of these techniques has associated strengths and weaknesses with regard to spectral resolution, spatial resolution, temporal resolution, and signal-to-noise characteristics. We have previously shown that spectral scanning of the fluorescence excitation spectrum can provide greatly increased signal strength compared to traditional emission-scanning approaches. Here, we present results from utilizing a Hyperspectral Imaging Fluorescence Excitation Scanning (HIFEX) microscope system for live cell imaging. Live cell signaling studies were performed using HEK 293 and rat pulmonary microvascular endothelial cells (PMVECs), transfected with either a cAMP FRET reporter or a Ca2+ reporter. Cells were further labeled to visualize subcellular structures (nuclei, membrane, mitochondria, etc.). Spectral images were acquired using a custom inverted microscope (TE2000, Nikon Instruments) equipped with a 300W Xe arc lamp and tunable excitation filter (VF-5, Sutter Instrument Co., equipped with VersaChrome filters, Semrock), and run through MicroManager. Timelapse spectral images were acquired from 350-550 nm, in 5 nm increments. Spectral image data were linearly unmixed using custom MATLAB scripts. Results indicate that the HIFEX microscope system can acquire live cell image data at acquisition speeds of 8 ms/wavelength band with minimal photobleaching, sufficient for studying moderate speed cAMP and Ca2+ events.

关键词： Spectral Spectroscopy Signature Fingerprint Optical Biopsy Classification EEM

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New techniques in confocal microscopy

New techniques in confocal microscopy

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Laser Science, LS 2006

作者： Mertz, Jerome Boston University Biomedical Engineering Department 44 Cummington St. Boston MA 02215 United States

Two new optical techniques are presented that lead to out-of-focus background rejection. The first is autoconfocal microscopy, which reveals phase-gradients. The second is dynamic speckle illumination microscopy, whic... 详细信息

ISBN: (纸本)1557528187

关键词： image processing Imaging techniques Laser speckle Speckle patterns Structured illumination microscopy Three dimensional imaging Imaging systems (110.0110) Three-dimensional image acquisition (110.6880) microscopy (180.0180) Confocal microscopy (180.1790)

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QUANTIFYING THE COMPLEXITY OF SMALL SCALE 3D LASER RANGE DATA

QUANTIFYING THE COMPLEXITY OF SMALL SCALE 3D LASER RANGE DAT...

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3-dimensional microscopy - image acquisition and processing II conference

作者： DUBUC, B TAWASHI, R MCGILL UNIV CTR INTELLIGENT MACHINESMONTREALPQ H3A 2T5CANADA

ISBN: (纸本)0819417599

Proper characterization of surface topography is critical for understanding a wide range of chemical and biological processes at the interface. This paper describes a method for characterizing the complexity of a surface from its depth map acquired with a laser vision sensor. The technique used to calculate fractal dimension was based on the variation method. Instead of trying to estimate the dimension around 0 however, this was done at specific scales over a range of values. We will show the impact of using local information to estimate the complexity of the surface and emphasize the problems that can be encountered when blindly trying to estimate the fractal dimension of 3D data. We conclude that linear local approximation of the object should be used to quantify its complexity, and to determine the scale at which analysis should be done. The method was applied to two samples of plasma coated titanium plates generated under differenty spraying conditions. The results show that our technique can provide a quantification scheme for standardization of the coating process and can improve the validation of manufacturing technologies.

关键词： Fractal analysis Cameras Titanium Data acquisition Manufacturing Natural surfaces Plasma Sensors Standards development Interfaces

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Subpixel resolution in maximum likelihood image restoration

Subpixel resolution in maximum likelihood image restoration

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conference on Three-dimensional microscopy - image acquisition and processing IV

作者： Conchello, JA McNally, JG Washington Univ. (United States)

ISBN: (纸本)0819423955

A number of algorithms have been developed for three-dimensional (3D) deconvolution of fluorescence microscopical images. These algorithms use a mathematical-physics model for the process of image formation and try estimate the specimen function, i.e. the distribution of fluorescent dye in the specimen. To keep the algorithms tractable and computational load practical, the algorithms rely on simplifying assumptions, and the extent to which these assumptions approximate the actual process of image formation and recording has a strong effect on the capabilities of the algorithms. The process of image formation is a continuous-space process, but the algorithms must be implemented using a discrete-space approximation to this process and render a sampled specimen function. A commonly-used assumption is that there is one pixel in the specimen for each pixel in the recorded image and that the pixel size in the recorded image is small compared to the size of the diffraction limited spot or Airy disk, a condition necessary to satisfy Nyquist sampling criterion. Modern CCD cameras, however, have large wells that integrate into a single pixel an area of the image that is significantly larger that the Airy disk. We derived a maximum-likelihood-based algorithm to accommodate for these large CCD pixel sizes. In this algorithm we assume that each pixel in the recorded image integrates several pixels that satisfy Nyquist criterion. The algorithm then attempts to estimate the specimen function at a resolution better than that allowed by the CCD camera. Preliminary results of this sub-pixel resolution algorithm are encouraging.

关键词： sub-pixel resolution missing information maximum-likelihood expectation-maximization super-resolution

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