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Conference on three-dimensional and multidimensional microscopy - image acquisition and processing XXIII

作者： Sheppard, Colin J. R. Castello, Marco Vicidomini, Giuseppe Duocastella, Marti Diaspro, Alberto Ist Italiano Tecnol Nanoscopy Nanophys Via Morego 30 I-16163 Genoa Italy Univ Genoa DIBRIS Via Opera Pia 13 I-16145 Genoa Italy

ISBN: (纸本)9781628419474

There are basically two types of microscope, which we call conventional and scanning. The former type is a full-field imaging system. In the latter type, the object is illuminated with a probe beam, and a signal detected. We can generalize the probe to a patterned illumination. Similarly we can generalize the detection to a patterned detection. Combining these we get a range of different modalities: confocal microscopy, structured illumination (with full-field imaging), spinning disk (with multiple illumination points), and so on. The combination allows the spatial frequency bandwidth of the system to be doubled. In general we can record a four dimensional (4D) image of a 2D object (or a 6D image from a 3D object, using an acoustic tuneable lens). The optimum way to directly reconstruct the resulting image is by image scanning microscopy (ISM). But the 4D image is highly redundant, so deconvolution-based approaches are also relevant. ISM can be performed in fluorescence, bright field or interference microscopy. Several different implementations have been described, with associated advantages and disadvantages. In two-photon microscopy, the illumination and detection point spread functions are very different. This is also the case when using pupil filters or when there is a large Stokes shift.

关键词： Confocal microscopy scanning optical microscopy fluorescence microscopy image scanning microscopy pixel reassignment

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Low-cost light-sheet microscopy for whole brain imaging 25

Low-cost light-sheet microscopy for whole brain imaging

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Conference on three-dimensional and multidimensional microscopy: image acquisition and processing XXV

作者： Kumar, Manish Nasenbeny, Jordan Kozorovitskiy, Yevgenia Northwestern Univ Dept Neurobiol Evanston IL 60208 USA

ISBN: (数字)9781510614840

ISBN: (纸本)9781510614840;9781510614833

Light-sheet microscopy has evolved as an indispensable tool in imaging biological samples. It can image 3D samples at fast speed, with high-resolution optical sectioning, and with reduced photobleaching effects. These properties make light-sheet microscopy ideal for imaging fluorophores in a variety of biological samples and organisms, e.g. zebrafish, drosophila, cleared mouse brains, etc. While most commercial turnkey light-sheet systems are expensive, the existing lower cost implementations, e.g. OpenSPIM, are focused on achieving high-resolution imaging of small samples or organisms like zebrafish. In this work, we substantially reduce the cost of light-sheet microscope system while targeting to image much larger samples, i.e. cleared mouse brains, at single-cell resolution. The expensive components of a light sheet system excitation laser, water-immersion objectives, and translation stage are replaced with an incoherent laser diode, dry objectives, and a custom-built Arduino-controlled translation stage. A low-cost CUBIC protocol is used to clear fixed mouse brain samples. The open-source platforms of Manager and Fiji support image acquisition, processing, and visualization. Our system can easily be extended to multi-color light-sheet microscopy.

关键词： Light-sheet microscopy Brain Fluorescence imaging Low-cost microscopy

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Modified K-factor image decomposition for three-dimensional super resolution microscopy 13

Modified K-factor image decomposition for three-dimensional ...

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Conference on three-dimensional and multidimensional microscopy - image acquisition and processing XXIII

作者： Ilovitsh, Tali Weiss, Aryeh Meiri, Amihai Ebeling, Carl G. Amiel, Aliza Katz, Hila Mannasse-Green, Batya Zalevsky, Zeev Bar Ilan Univ Fac Engn IL-52900 Ramat Gan Israel Univ Utah Dept Elect & Comp Engn Salt Lake City UT USA Univ Utah Dept Phys & Astron Salt Lake City UT USA Meir Med Ctr Genet Inst Kefar Sava Israel Bar Ilan Univ Fac Life Sci Ramat Gan Israel

ISBN: (纸本)9781628419474

The ability to track single fluorescent particles within a three dimensional (3D) cellular environment can provide valuable insights into cellular processes. In this paper, we present a modified nonlinear image decomposition technique called K-factor that reshapes the 3D point spread function (PSF) of an XYZ image stack into a narrow Gaussian profile. The method increases localization accuracy by similar to 60% with compare to regular Gaussian fitting, and improves minimal resolvable distance between overlapping PSFs by similar to 50%. The algorithm was tested both on simulated data and experimentally. This work presets a novel use of the nonlinear image decomposition technique called K-factor that reshapes the three dimensional (3D) point spread function (PSF) of an XYZ image stack into a narrow Gaussian profile. The experimentally obtained PSF of a Z-stack raw data that is acquired by a widefield microscope has a more elaborate shape that is given by the Gibson and Lanni model. This shape increases the computational complexity associated with the localization routine, when used in localization microscopy techniques. Furthermore, due to its nature, this PSF spreads over a larger volume, making the problem of overlapping emitters detection more pronounced. The ability to use Gaussian fitting with high accuracy on 3D data can facilitate the computational complexity, hence reduce the processing time required for the generation of the 3D superresolved image. In addition it allows the detection of overlapping PSFs and reduces the effects of the penetration of out of focus PSFs into in focused PSFs, therefore enables the increase in the activated fluorophore density by similar to 50%. The algorithm was tested both on simulated data and experimentally, where it yielded an increase in the localization accuracy by similar to 60% with compare to regular Gaussian fitting, and improved the minimal resolvable distance between overlapping PSFs by similar to 50%, making it extrem

关键词： three-dimensional microscopy image processing Fluorescence Superresolution

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Extension of spectral domain phase microscopy to three-dimensional nanoscale displacement mapping in cardiomyocytes

Extension of spectral domain phase Microscopy to three-dimen...

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Conference on three-dimensional and multidimensional microscopy - image acquisition and processing XV

作者： Ellerbee, Audrey K. Hendargo, Hansford C. Motomura, Amy R. Izatt, Joseph A. Duke Univ Dept Biomed Engn Durham NC 27708 USA

ISBN: (纸本)9780819470362

Spectral domain phase microscopy (SDPM) is a functional extension of optical coherence tomography (OCT) whose common-path interferometric design. enables phase-referenced imaging of dynamic samples. Like OCT, axial resolution in SDPM is determined by the source coherence length, while lateral resolution is limited by diffraction in the microscope optics. Nonetheless, the quantitative phase information SDPM generates is sensitive to sub-Angstrom displacements of scattering structures. Integrative quantitative phase imaging techniques, such as Fourier phase microscopy, Hilbert phase microscopy, and Digital holographic microscopy, have achieved sub-micron motion detection in live cells. In contrast with the techniques, SDPM can achieve full depth discrimination, allowing for resolution of the motion of independent, sub-cellular structures at various cross-sectional planes within the sample. The ability of SDPM to measure Doppler flow in single-celled organisms, time-resolved cellular motions, and rheological information of the cytoskeleton has been previously demonstrated. The objective of this study is to extend the use of SDPM to produce three-dimensional reconstructions of the internal and surface motions of beating cardiomyocytes. Phase information is used to the motion of quantify cellular structures in the axial dimension. Our gated acquisition process involves synchronization of the SDPM detection system with and applied electrical field used to stimulate beating in isolated cardiomyocytes. For a given pacing protocol, we obtain repeat motion measurements in two-dimensions during cellular contraction, building a volume image by repeating the process at multiple discrete slices through the cell. This experiment serves as a proof-of-principle for volumetric imaging of beating cardiomyocytes.

关键词： interferometry optical coherence tomography quantitative phase imaging microscopy

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Improved quantitative phase contrast in self-interference digital holographic microscopy and sensing dynamic refractive index changes of the cytoplasm using internalized microspheres as probes

Improved quantitative phase contrast in self-interference di...

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Conference on three-dimensional and multidimensional microscopy - image acquisition and processing XX

作者： Kemper, Bjoern Schubert, Robin Dartmann, Sebastian Vollmer, Angelika Ketelhut, Steffi von Bally, Gert Univ Munster Ctr Biomed Opt & Photon D-48149 Munster Germany

ISBN: (纸本)9780819493583

In self-interference digital holographic microscopy (DHM), scattering patterns that are induced by coherent laser light affect the resolution for the detection of optical path length changes. We present a simple and efficient approach for the reduction of coherent disturbances in quantitative DHM phase images by amplitude and phase modulation of the sample illumination. The performance of the method in quantitative phase imaging of living cells is illustrated. Moreover, the application of self-interference DHM for sensing of dynamic refractive index changes of adherent cells is demonstrated. Therefore, silica microspheres are used in living cells as optical probes to determine the refractive index of the cytoplasm from single quantitative phase images.

关键词： Digital holographic microscopy quantitative phase imaging cellular refractive index label-free minimally invasive live cell imaging

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Improvement based on FDK reconstruction algorithm of conebeam CT

Improvement based on FDK reconstruction algorithm of conebea...

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Conference on three-dimensional and multidimensional microscopy - image acquisition and processing XVII

作者： Miao, Hui Wang, Tingting Zhao, Huijuan Gao, Feng Tianjin Univ Coll Precis Instruments & Optoelect Engn Tianjin 300072 Peoples R China

ISBN: (纸本)9780819479662

As a typical 3D image algorithm, Feldkamp-David-Kress reconstruction algorithm is a filtered backprojection very similar to the 2D algorithm. Recent years, it has been widely used for its easy implementation and acceptable reconstruction precision for small cone-beam angle. For big cone-beam angles, however, the reconstruction exactness of horizontal planes will dramatically decrease with the distance from the central plane increases. Therefore, for improving the reconstruction precision in vertical direction, a similar reciprocal Gaussian function as the weighted factor was introduced into FDK algorithm in the article. The validity of the improved FDK algorithm was verified and evaluated through both the computer numerical simulation and the phantom model experiment. The quantitative analysis for the reconstruction results demonstrated that the reconstruction image using the improved FDK algorithm could primely revise the original reconstruction images and restore well closely to the tomography image of the prototype object.

关键词： FDK reconstruction cone-beam CT image reconstruction reconstruction algorithm

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Managing multiple image stacks from confocal laser scanning microscopy

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Proceedings of SPIE - The International Society for Optical Engineering 1999年 3605卷 228-235页

作者： Zerbe, Joerg Goetze, Christian Zuschratter, Werner Fraunhofer Inst for Computer Graphics Rostock Germany

ISBN: (纸本)0819430757;9780819430755

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.

关键词： Optical microscopy

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Enhanced spectral lightfield fusion microscopy via deep computational optics for whole-slide pathology 26

Enhanced spectral lightfield fusion microscopy via deep comp...

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Conference on three-dimensional and multidimensional microscopy - image acquisition and processing XXVI

作者： Kazemzadeh, Farnoud Li-Chang, Hector Wong, Alexander Univ Waterloo Waterloo ON Canada Waterloo Artificial Intelligence Inst Waterloo ON Canada Royal Victoria Reg Hlth Ctr Barrie ON Canada

ISBN: (纸本)9781510624092

Lensfree on-chip microscopy, which harnesses holography principles to capture interferometric light-field encodings without the need of lenses, is an emerging microscopy modality with widespread interest given the large field-of-view (FOV) compared to lens-based microscopy systems. In particular, there is a growing interest on the development of high-quality lensfree on-chip color microscopy. In this study, we propose a multi-laser spectral lightfield fusion microscopy using deep computational optics for achieving lensfree on-chip color microscopy. We will demonstrate that leveraging deep computational optics can enable imaging resolution beyond the diffraction limit without the use of any complex hardware-based super-resolution techniques, such as aperture scanning. The capabilities of the microscope are examined for whole-slide pathology. The superior imaging resolution of the instrument is demonstrated by imaging of a series of biological specimens demonstrating the true color imaging capability of the instrument while showcasing the large FOV of the instrument.

关键词： on-chip microscopy deep computational optics light-field fusion

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Dual-mode digital holographic and fluorescence microscopy for the study of morphological changes in cells under simulated microgravity

Dual-mode digital holographic and fluorescence microscopy fo...

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Conference on three-dimensional and multidimensional microscopy - image acquisition and processing XVII

作者： Toy, M. Fatih Pache, Christophe Parent, Jerome Kuehn, Jonas Egli, Marcel Depeursinge, Christian Ecole Polytech Fed Lausanne Adv Photon Lab CH-1015 Lausanne Switzerland ETH Space Biol Grp CH-8005 Zurich Switzerland

ISBN: (纸本)9780819479662

A dual mode microscope is developed to study morphological evolution of mouse myoblast cells under simulated microgravity in real time. Microscope operates in Digital Holographic microscopy (DHM) and widefield epifluorescence microscopy modes in a time sequential basis. DHM provides information on real time cellular morphology. EGFP transfected actin filaments in mouse myoblast cells function as the reporter for the fluorescence microscopy mode. Experimental setup is fixed in the RPM to observe microgravity induced dynamic changes in live cells. Initial results revealed two different modifications. Disorganized structures become visible in the formed lamellipodias, and proteins accumulate in the perinuclear region.

关键词： microscopy holography digital holographic microscopy fluorescence microscopy microgravity cell morphology cytoskeleton

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Wide-Field microscopy using Microcamera Arrays

Wide-Field Microscopy using Microcamera Arrays

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Conference on three-dimensional and multidimensional microscopy - image acquisition and processing XX

作者： Marks, Daniel L. Youn, Seo Ho Son, Hui S. Kim, Jungsang Brady, David J. Duke Univ Dept Elect & Comp Engn Durham NC 27708 USA

ISBN: (纸本)9780819493583

A microcamera is a relay lens paired with image sensors. Microcameras are grouped into arrays to relay overlapping views of a single large surface to the sensors to form a continuous synthetic image. The imaged surface may be curved or irregular as each camera may independently be dynamically focused to a different depth. Microcamera arrays are akin to microprocessors in supercomputers in that both join individual processors by an optoelectronic routing fabric to increase capacity and performance. A microcamera may image ten or more megapixels and grouped into an array of several hundred, as has already been demonstrated by the DARPA AWARE Wide-Field program with multiscale gigapixel photography. We adapt gigapixel microcamera array architectures to wide-field microscopy of irregularly shaped surfaces to greatly increase area imaging over 1000 square millimeters at resolutions of 3 microns or better in a single snapshot. The system includes a novel relay design, a sensor electronics package, and a FPGA-based networking fabric. Biomedical applications of this include screening for skin lesions, wide-field and resolution-agile microsurgical imaging, and microscopic cytometry of millions of cells performed in situ.

关键词： Wide-field microscopy camera array image processing agile resolution

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