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检索条件"任意字段=Three-Dimensional and Multidimensional Microscopy: Image Acquisition and Processing XIX"

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Compressive Time-resolved Multispectral Fluorescence microscopy with Single-Pixel Camera 30

Compressive Time-resolved Multispectral Fluorescence Microsc...

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three-dimensional and multidimensional microscopy: image acquisition and processing XXX 2023

作者： Ghezzi, Alberto Farina, Andrea Lenz, Armin J.M. Soldevila, Fernando Tajahuerce, Enrique Bassi, Andrea Valentini, Gianluca D’Andrea, Cosimo Politecnico di Milano Dipartimento di Fisica Piazza L. da Vinci 32 Milano20133 Italy Consiglio Nazionale delle Ricerche IFN Piazza L. da Vinci 32 Milan20133 Italy Universitat Jaume I Avda. Sos Baynat s/n Castelló12071 Spain UMR 8552 Collège de France 24 rue Lhomond Paris75005 France Center for NanoScience and Technology @PoliMi Istituto Italiano di Tecnologia via Pascoli 70/3 via Pascoli 70/3 Milan20133 Italy

ISBN: (纸本)9781510658752

Time-resolved multispectral fluorescence microscopy provides a 4D hypercube dataset with high specificity for cellular examination. However, this is generally obtained by significantly increasing the measurement time, which is quite limiting for in vivo measurement or with photosensitive sample. It is possible to reduce the measurement effort with a novel microscopy framework exploiting Compressive Sensing based on Single-Pixel Camera. In this work, we present a compressive sensing system and validate it with a cellular sample. Data fusion with a high-resolution camera image allows us to tackle the well-known problem of low-resolution in single-pixel imaging. © 2023 SPIE. All rights reserved.

关键词： Compressed sensing

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Modal wavefront sensor for adaptive confocal microscopy

Modal wavefront sensor for adaptive confocal microscopy

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three-dimensional and multidimensional microscopy: image acquisition processing VII

作者： Booth, Martin J. Neil, Mark A.A. Wilson, Tony Univ of Oxford Oxford United Kingdom

A confocal microscope permits 3-D imaging of volume objects by the inclusion of a pinhole in the detector path which eliminates out of focus light. This configuration is however very sensitive to aberrations induced by the specimen or the optical system and would therefore benefit from an adaptive optics approach. We present a wavefront sensor capable of measuring directly the Zernike components of an aberrated wavefront and show that it is particularly applicable to the confocal microscope since only those wavefronts originating in the focal region contribute to the measured aberration.

关键词： Medical imaging

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Evaluation of different illumination designs for tunable 3D structured illumination microscopy through model-based restoration 28

Evaluation of different illumination designs for tunable 3D ...

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three-dimensional and multidimensional microscopy: image acquisition and processing XXVIII 2021

作者： Mohammed, Jules Van, Cong T. S. Preza, Chrysanthe Computational Imaging Research Laboratory Department of Electrical and Computer Engineering University of Memphis MemphisTN38152 United States

ISBN: (数字)9781510641341

ISBN: (纸本)9781510641334

In a tunable 3D structured illumination microscopy (3D-SIM) based on an illumination system comprised by a multi-slit array and a Fresnel biprism, the 3D structured illumination (SI) pattern depends on the design of the slit array. Previous work studied the impact of the illumination pattern design on the achieved extension of the compact support of the tunable 3D-SIM optical transfer function. In this contribution, we use simulations with different illumination designs that utilize a different number of slits, to evaluate system performance by investigating the lateral and axial resolution achieved in the restored images computed with a regularized iterative 3D model-based algorithm, which minimizes the mean square error between the model and data. three illumination designs are investigated in which the number of slits is N = 3, 9 and 11 while the corresponding distance between the slits is x0 = 488 μm, 122 μm and 100 μm, respectively. A 3D star-like object, commonly used in resolution analysis of imaging systems, is used in the simulations. Results are quantified using the mean square error and the structured similarity index as well as intensity profiles that show the achieved resolution and robustness to noise in each case. The lateral and axial 3D-SIM theoretical resolution limits are achieved in the presence of 20-dB Poisson noise while small differences are evident in the results from 15-dB simulated data obtained using the 3 investigated designs. © 2021 Copyright SPIE.

关键词： Fluorescence microscopy

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Developments and applications of confocal theta microscopy

Developments and applications of confocal theta microscopy

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Proceedings of the 1999 three-dimensional and multidimensional microscopy: image acquisition and processing VI

作者： Haar, Frank-Martin Swoger, Jim Stelzer, Ernst H.K. Heidelberg Germany

ISBN: (纸本)0819430757

Confocal theta microscopy improves the resolution of confocal laser scanning microscopes by instrumentally solving the problem of the inferior axial resolution. A technical variation of this microscopy technique, the Single-Lens Theta microscopy (SLTM), is designed to be easily adapted to any common confocal laser scanning microscope. With SLTM, different kinds of other microscopical techniques are possible.

关键词： Optical microscopy

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MAGNIFY: molecule anchorable gel-enabled nanoscale in-situ fluorescence microscopy for nanoscale imaging of biomolecules 30

MAGNIFY: molecule anchorable gel-enabled nanoscale in-situ f...

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three-dimensional and multidimensional microscopy: image acquisition and processing XXX 2023

作者： Klimas, Aleksandra Gallagher, Brendan R. DiBernardo, Emma Cheng, Zhangyu Zhao, Yongxin Department of Biological Sciences Carnegie Mellon University PittsburghPA United States

ISBN: (纸本)9781510658752

Expansion microscopy (ExM) is a powerful imaging strategy that offers a low-cost solution for interrogating biological systems at the nanoscale using conventional optical microscopes. It achieves this by physically and isotropically magnifying preserved biological specimens embedded in a cross-linked water-swellable hydrogel. However, most reported techniques are unable to preserve endogenous epitopes due to strong protease digestion used to expand samples. In addition, these protocols rely on mechanically fragile hydrogels that only expand by at most 4.5× linearly. We present a new ExM framework, Molecule Anchorable Gel-enabled Nanoscale In-situ Fluorescence microscopy (MAGNIFY), that exhibits a broad retention of nucleic acids, proteins, and lipids without the need for a separate anchoring step. By using a mechanically sturdy hydrogel, MAGNIFY is capable of expanding biological specimens up to 11×. This facilitates nanoscale imaging (~25-nm effective resolution) using an ∼280-nm diffraction-limited objective lens on a conventional optical microscope and can be furthered to ~15 nm effective resolution if combined with computational methods such as Super-resolution Optical Fluctuation Imaging (SOFI). Here, we demonstrate that MAGNIFY provides a generalized solution for imaging nanoscale subcellular features of a broad range of biological specimens. We also show that MAGNIFY provides a novel, accessible tool for improving the precision, utility, and generality of nanoscopy. © 2023 SPIE. All rights reserved.

关键词： Fluorescence microscopy

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Design and fabrication of universal resolution targets for microsphere-assisted microscopy 31

Design and fabrication of universal resolution targets for m...

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three-dimensional and multidimensional microscopy: image acquisition and processing XXXI 2024

作者： Abbasian, Vahid Awan, Kashif Masud Darafsheh, Arash Department of Radiation Oncology Washington University School of Medicine in St. Louis St. LouisMO63110 United States Imaging Science Program McKelvey School of Engineering Washington University in St. Louis St. LouisMO63130 United States Institute of Materials Science and Engineering Washington University St. LouisMO63130 United States Department of Mechanical Engineering and Materials Science Washington University in St. Louis St. LouisMO63130 United States

ISBN: (纸本)9781510669550

Determining the achieved spatial resolution in microsphere-assisted microscopy (MAM) has remained an important and challenging task. While fluorescent-based microscopies use fluorescent nano-beads to determine the point spread function (PSF), such a method is not practical in label-free super-resolution techniques. In some previous studies, objects with certain features have been imaged and those feature sizes have been reported as the resolution;however, determining the resolution in that way is not consistent with the definition of resolution based on point sources. Furthermore, the imaging characteristics in MAM may depend on specimen-specific properties, such as materials and substrate. In this study, we propose a robust and objective technique to address these issues. We fabricate arrays of nano-pillars varying in diameter and periodicity, ranging from 50 to 1000 nm. These pillars consist of a 20 nm Ag layer with a 2 nm Cr adhesion layer, deposited on a silicon substrate. The role of specimen-specific properties can be investigated by considering different materials. Fabrication involves the bilayer PMMA resist lift-off process, ensuring accurate deposition of nanometer-scale features to serve as sub-diffraction-limited targets in optical super-resolution microscopy. A deconvolution method that provides a robust platform for resolution measurement was used. By deconvolving the object profile, obtained from scanning electron microscopy, from the image profile, the PSF of the system is determined. Our proposed technique offers a reliable and standardized approach for quantifying resolution, magnification, and field-of-view in MAM or any other label-free microscopy technique, enabling accurate and objective comparison across different novel microscopy techniques. © COPYRIGHT SPIE. Downloading of the abstract is permitted for personal use only.

关键词： Microspheres

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System for fluorescence lifetime imaging microscopy

System for fluorescence lifetime imaging microscopy

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Proceedings of the 1999 three-dimensional and multidimensional microscopy: image acquisition and processing VI

作者： van Geest, Lambertus K. Boddeke, Frank R. van Dijk, Pieter W. Kamp, Arjen F. van der Oord, Cornelis J.R. Stoop, Karel W.J. Lambert Instruments Netherlands

ISBN: (纸本)0819430757

An affordable, robust and easy-to-use fluorescence lifetime imaging microscopy (FLIM) workstation that is completely automated and does not need any difficult calibration procedure is introduced. The workstation consists of a standard fluorescence microscope, a modulated excitation light source, a camera, a modulation signal generator and acquisition/processing software. An evaluation of the system and its critical components is presented.

关键词： Optical microscopy

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Correlative FRET: New method improves rigor and reproducibility in determining distances within synaptic nanoscale architecture 25

Correlative FRET: New method improves rigor and reproducibil...

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three-dimensional and multidimensional microscopy: image acquisition and processing XXV 2018

作者： Shinogle-Decker, Heather Martinez-Rivera, Noraida O'Brien, John Powell, Richard D. Joshi, Vishwas N. Connell, Samuel Rosa-Molinar, Eduardo Microscopy and Analytical Imaging Research Resource Core Laboratory University of Kansas LawrenceKS66045 United States Biological Imaging Group University of Kansas LawrenceKS66045 United States Department of Ophthalmology and Visual Science University of Texas Health Science Center at Houston TX77030 United States Nanoprobes Inc. YaphankNY11980 United States Anvesha Laboratories SetauketNY11733 United States Intelligent Imaging Innovations DenverCO80216 United States

ISBN: (数字)9781510614840

ISBN: (纸本)9781510614833

A new correlative Förster Resonance Energy Transfer (FRET) microscopy method using FluoroNanogold™, a fluorescent immunoprobe with a covalently attached Nanogold® particle (1.4nm Au), overcomes resolution limitations in determining distances within synaptic nanoscale architecture. FRET by acceptor photobleaching has long been used as a method to increase fluorescence resolution. The transfer of energy from a donor to an acceptor generally occurs between 10-100Å, which is the relative distance between the donor molecule and the acceptor molecule. For the correlative FRET microscopy method using FluoroNanogold™, we immuno-labeled GFP-tagged-HeLa-expressing Connexin 35 (Cx35) with anti-GFP and with anti-Cx35/36 antibodies, and then photo-bleached the Cx before processing the sample for electron microscopic imaging. Preliminary studies reveal the use of Alexa Fluor® 594 FluoroNanogold™ slightly increases FRET distance to 70Å, in contrast to the 62.5Å using AlexaFluor 594®. Preliminary studies also show that using a FluoroNanogold™ probe inhibits photobleaching. After one photobleaching session, Alexa Fluor 594® fluorescence dropped to 19% of its original fluorescence;in contrast, after one photobleaching session, Alexa Fluor 594® FluoroNanogold™ fluorescence dropped to 53% of its original intensity. This result confirms that Alexa Fluor 594® FluoroNanogold™ is a much better donor probe than is Alexa Fluor 594®. The new method (a) creates a double confirmation method in determining structure and orientation of synaptic architecture, (b) allows development of a two-dimensional in vitro model to be used for precise testing of multiple parameters, and (c) increases throughput. Future work will include development of FluoroNanogold™ probes with different sizes of gold for additional correlative microscopy studies. © COPYRIGHT SPIE. Downloading of the abstract is permitted for personal use only.

关键词： Forster resonance energy transfer

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Auto-correlation for multi-view deconvolved reconstruction in light sheet microscopy 28

Auto-correlation for multi-view deconvolved reconstruction i...

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three-dimensional and multidimensional microscopy: image acquisition and processing XXVIII 2021

作者： Ancora, Daniele Valentini, Gianluca Pifferi, Antonio Giovanni Bassi, Andrea Dipartimento di Fisica Politecnico di Milano Piazza Leonardo da Vinci 32 Milano20133 Italy Istituto di Fotonica e Nanotecnologie Consiglio Nazionale delle ricerche Piazza Leonardo da Vinci 32 Milano20133 Italy

ISBN: (数字)9781510641341

ISBN: (纸本)9781510641334

Tomographic inspection of fluorescent labels distributed within a specimen is an important aspect in biology. Light sheet fluorescent microscopy (LSFM) offers a powerful and simple tool to selectively slice the sample and let us directly obtain a tomographic view of the specimen. However, due to non-isotropic resolution of the technique along the axial scanning, one may want to combine different views of the object and add deconvolution to the process in order to achieve higher resolution. Typically, multi-view Bayesian methods based on Richardson-Lucy deconvolution are used for this task once the datasets are exactly registered against each other. In this work, instead, we begin to investigate how to avoid the alignment procedure and use a direct algorithm to form a multi-view tomographic reconstruction. To do this, we developed a new framework based on auto-correlation analysis that let us achieve deconvolved reconstructions starting from blurred auto-correlations. Since the latter are insensitive to shifts, we can combine the auto-correlations coming from multi-view acquisitions without taking care of the registration procedure. © 2021 Copyright SPIE.

关键词： Inverse problems

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Doubling the lateral resolution of wide-field fluorescence microscopy using structured illumination

Doubling the lateral resolution of wide-field fluorescence m...

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three-dimensional and multidimensional microscopy: image acquisition processing VII

作者： Gustafsson, Mats G.L. Agard, David A. Sedat, John W. Univ of California San Francisco San Francisco United States

Spatial information that exceeds the classical resolution limit by a factor of two can be made visible in the widefield fluorescence microscope by illuminating the sample with spatially structured (patterned) excitation light. By computationally restoring this information to its proper location in reciprocal space, an image with twice the normal lateral resolution can be produced. The method can be applied in three dimensions, and yields an axial sectioning power equal to that of confocal microscopes. Unlike the case in confocal microscopy, however, both the lateral and axial resolution enhancements are achieved without any loss of emission light, resulting in uncompromised sensitivity. The method has been experimentally verified on both test objects and complex biological structures and performs in complete agreement with theoretical predictions. The resulting images possess a visual clarity that dramatically exceeds that of both conventional and confocal microscopes.

关键词： Medical imaging

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