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

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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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Digital aberration correction of fluorescent images with coherent holographic image reconstruction by phase transfer (CHIRPT) 13

Digital aberration correction of fluorescent images with coh...

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

作者： Field, Jeffrey J. Bartels, Randy A. Colorado State Univ Microscopy Core Facil Ft Collins CO 80523 USA Colorado State Univ Dept Elect & Comp Engn Ft Collins CO 80523 USA Colorado State Univ Dept Biochem & Mol Biol Ft Collins CO 80523 USA Colorado State Univ Inst Genome Architecture & Funct Ft Collins CO 80523 USA Colorado State Univ Sch Biomed Engn Ft Collins CO 80523 USA

ISBN: (纸本)9781628419474

Coherent holographic image reconstruction by phase transfer (CHIRPT) is an imaging method that permits digital holographic propagation of fluorescent light. The image formation process in CHIRPT is based on illuminating the specimen with a precisely controlled spatio-temporally varying intensity pattern. This pattern is formed by focusing a spatially coherent illumination beam to a line focus on a spinning modulation mask, and image relaying the mask plane to the focal plane of an objective lens. Deviations from the designed spatio-temporal illumination pattern due to imperfect mounting of the circular modulation mask onto the rotation motor induce aberrations in the recovered image. Here we show that these aberrations can be measured and removed non-iteratively by measuring the disk aberration phase externally. We also demonstrate measurement and correction of systematic optical aberrations in the CHIRPT microscope.

关键词： fluorescence holography aberration correction microscopy

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Design of adaptive objective lens for ultrabroad near infrared imaging 13

Design of adaptive objective lens for ultrabroad near infrar...

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

作者： Lan, Gongpu Li, Guoqiang Ohio State Univ Dept Ophthalmol & Visual Sci Columbus OH 43212 USA Ohio State Univ Dept Elect & Comp Engn 1330 Kinnear Road Columbus OH 43212 USA

ISBN: (纸本)9781628419474

We present a compound adaptive objective lens in which a water-filled membrane lens is inserted into a front group (one lens) and a back group (two lenses). This adaptive objective lens works in the ultrabroad near infrared waveband (760nm similar to 920nm) with the volume scan of > 1mm(3) and the resolution of 2.8 mu m (calculated at the wavelength of 840 nm). The focal range is 19.5mm similar to 20.5mm and the numerical number is 0.196. The size of the adaptive lens is 10mm (diameter) x 17mm (length). This kind of lens can be widely used in three-dimensional (3D) volume biomedical imaging instruments, such as confocal microscope, optical coherence tomography (OCT), two photon microscope, etc.

关键词： Active optics adaptive objective lens extended depth of field three-dimensional microscopy confocal microscopy optical coherence tomography two-photon microscopy optical design

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Fabrication of two-color annular hybrid wave plate for three-dimensional super-resolution microscopy 13

Fabrication of two-color annular hybrid wave plate for three...

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

作者： Kumagai, Hiroshi Iketaki, Yoshinori Jahn, Kornel Bokor, Nador Kitasato Univ Allied Hlth Sci Phys Minami Ku 1-15-1Kitasato Sagamihara Kanagawa 2520373 Japan Olympus Corp 2-3 Kuboyama Cho Hachioji Tokyo 1928512 Japan Budapest Univ Technol & Econ Dept Phys Budafoki U 8 H-1111 Budapest Hungary

ISBN: (纸本)9781628419474

In super-resolution microscopy, we use fluorescence depletion, where an erase beam quenches a molecule in the S1 state generated by a pump beam, and then prevents fluorescence from the S1 state. When a tight doughnut shaped erase beam with is focused on the dyed sample together with a Gaussian pump beam, the remaining fluorescence spot in the focal plane becomes smaller than the diffraction-limited size. Applying destructive interference to the erase beam, erase beam has a minute three-dimensional dark spot surrounded by the light near the focal region. Since this spot introduces fluorescence depletion along the optical axis as in the focal plane, we can achieve three-dimensional super-resolution microscopy. However, to overcome the diffraction limit, an extremely precise optical alignment is required for projecting the focused pump beam into the dark spot of the erase beam. To resolve this technical issue, we fabricated a two-color annular hybrid wave plate (TAHWP) by combining two multi-order wave quartz plates. Although the pump and erase beams co-axially pass through the plate;the pump beam retains its original Gaussian shape, while the erase beam undergoes destructive interference. Inserting the TAHWP into a commercial scanning laser microscope, a three-dimensional spherical fluorescence spot with a volume of (similar to 100 nm)(3) can be created. Beside eliminating alignment problems and yielding a compact setup, the TAHWP makes our proposed method very suitable for commercial microscope systems. In this study, we report about detailed fabrication procedure and three-dimensional image properties given by the TAHWP.

关键词： Super-resolution microscopy fluorescence depletion three-dimensional destructive interference laser scanning microscope diffraction limit micro beads

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Dual-detection confocal microscopy: high-speed surface profiling without depth scanning 13

Dual-detection confocal microscopy: high-speed surface profi...

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

作者： Lee, Dong-Ryoung Gweon, Dae-Gab Yoo, Hongki Korea Adv Inst Sci & Technol KAIST Dept Mech Engn Nano Optomechatron Lab 291 Daehak Ro Daejeon 34141 South Korea Hanyang Univ Dept Biomed Engn Biomed Opt & Photomed Lab 222 Wangsimni Ro Seoul 04763 South Korea

ISBN: (纸本)9781628419474

We propose a new method for three-dimensional (3-D) imaging without depth scanning that we refer to as the dual-detection confocal microscopy (DDCM). Compared to conventional confocal microscopy, DDCM utilizes two pinholes of different sizes. DDCM generates two axial response curves which have different stiffness according to the pinhole diameters. The two axial response curves can draw the characteristics curve of the system which shows the relationship between the axial position of the sample and the intensity ratio. Utilizing the characteristic curve, the DDCM reconstructs a 3-D surface profile with a single 2-D scanning. The height of each pixel is calculated by the intensity ratio of the pixel and the intensity ratio curve. Since the height information can be obtained directly from the characteristic curve without depth scanning, a major advantage of DDCM over the conventional confocal microscopy is a speed. The 3-D surface profiling time is dramatically reduced. Furthermore, DDCM can measure 3-D images without the influence of the sample condition since the intensity ratio is independent of the quantum yield and reflectance. We present two types of DDCM, such as a fluorescence microscopy and a reflectance microscopy. In addition, we extend the measurement range axially by varying the pupil function. Here, we demonstrate the working principle of DDCM and the feasibility of the proposed methods.

关键词： Confocal microscopy three-dimensional microscopy high speed imaging

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Hologram Encoding strategies for non-Bayesian noise suppression in Digital Holography reconstructions and optical display 13

Hologram Encoding strategies for non-Bayesian noise suppress...

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

作者： Bianco, V. Memmolo, P. Finizio, A. Paturzo, M. Ferraro, P. Italian Natl Res Council ISASI CNR Inst Appl Sci & Intelligent Syst Via Campi Flegrei 34 I-80078 Pozzuoli NA Italy

ISBN: (纸本)9781628419474

Here we first propose a fast, one-shot, non-Bayesian method which performs a numerical synthesis of a moving aperture in order to reduce the noise in Digital Holography without prior information on its statistics. Starting from one single hologram capture, multiple uncorrelated reconstructions are provided by random sparse resampling masks, which can be incoherently averaged. Thus, the problem of the setup complexity introduced by multiple recordings gets solved. Besides, at the scope of performing DH display using a SLM, it is highly required to operate directly on the hologram, in order to obtain its denoised version without losing the coherence between amplitude and phase information. We then move a step forward, showing a novel encoding formula allowing us to directly synthesize denoised holograms to be optically displayed by SLMs.

关键词： (090.1995) Digital holography (090.4220) Multiplex holography (090.2870) Holographic display (100.3010) image reconstruction techniques

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Dynamic photothermal interferometric phase microscopy 13

Dynamic photothermal interferometric phase microscopy

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

作者： Turko, Nir A. Blum, Omry Shaked, Natan T. Tel Aviv Univ Fac Engn Dept Biomed Engn IL-69978 Ramat Aviv Israel

ISBN: (纸本)9781628419474

We present our latest advances in highly dynamic photothermal interferometric phase microscopy for quantitative, selective contrast imaging. Gold nanoparticles can be bio-functionalized to bind specific cells. When stimulating gold nanoparticles at their plasmon-peak wavelength, local increase of temperature occurs due to plasmon resonance. This causes a rapid change of optical phase of the light beam interacting with the sample. These phase changes can be recorded by interferometric phase microscopy and analyzed to form a photothermal image of the binding sites of the nanoparticles in the cells. Furthermore, by increasing the excitation laser light, one can deplete certain cells at will. Usually, the analysis of the photothermal signals utilizes a Fourier transform, which is computational time consuming. This makes photothermal imaging not suitable for applications requiring dynamic imaging or real-time quantitative analysis, such as for analyzing and sorting cells during their fast flow. For this goal, we have developed new algorithms, based on discrete Fourier transform variants, enabling fast analysis of photothermal signals from nanoparticles in live and highly dynamic cells. For the first time, video-rate photothermal signals are obtained, which forms the basis for real-time interferometric phase microscopy with molecular specificity. This technique holds great potential for using photothermal imaging in flow cytometry.

关键词： Photothermal imaging gold nanoparticles interferometric phase microscopy digital holographic microscopy

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Compressive Sensing in Reflectance Confocal microscopy of Skin images: A Preliminary Comparative Study 13

Compressive Sensing in Reflectance Confocal Microscopy of Sk...

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

作者： Arias, Fernando X. Sierra, Heidy Rajadhyaksha, Milind Arzuaga, Emmanuel Univ Puerto Rico Dept Elect & Comp Engn Lab Appl Remote Sensing & Image Proc Mayaguez PR USA Mem Sloan Kettering Canc Ctr Dermatol Serv 1275 York Ave New York NY 10021 USA

ISBN: (纸本)9781628419474

Compressive Sensing (CS)-based technologies have shown potential to improve the efficiency of acquisition, manipulation, analysis and storage processes on signals and imagery with slight discernible loss in data performance. The CS framework relies on the reconstruction of signals that are presumed sparse in some domain, from a significantly small data collection of linear projections of the signal of interest. As a result, a solution to the underdetermined linear system resulting from this paradigm makes it possible to estimate the original signal with high accuracy. One common approach to solve the linear system is based on methods that minimize the L1-norm. Several fast algorithms have been developed for this purpose. This paper presents a study on the use of CS in high-resolution reflectance confocal microscopy (RCM) images of the skin. RCM offers a cell resolution level similar to that used in histology to identify cellular patterns for diagnosis of skin diseases. However, imaging of large areas (required for effective clinical evaluation) at such high-resolution can turn image capturing, processing and storage processes into a time consuming procedure, which may pose a limitation for use in clinical settings. We present an analysis on the compression ratio that may allow for a simpler capturing approach while reconstructing the required cellular resolution for clinical use. We provide a comparative study in compressive sensing and estimate its effectiveness in terms of compression ratio vs. image reconstruction accuracy. Preliminary results show that by using as little as 25% of the original number of samples, cellular resolution may be reconstructed with high accuracy.

关键词： compressive sensing reflectance confocal microscopy dermatology optimization optical imaging non-invasive imaging

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three dimensional measurement of cAMP gradients using hyperspectral confocal microscopy 13

Three dimensional measurement of cAMP gradients using hypers...

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

作者： Rich, Thomas C. Annamdevula, Naga Britain, Andrea L. Mayes, Samuel Favreau, Peter F. Leavelsey, Silas J. Univ S Alabama Pharmacol Mobile AL 36688 USA Univ S Alabama Ctr Lung Biol Mobile AL 36688 USA Univ S Alabama Chem & Biomol Engn Mobile AL 36688 USA

ISBN: (纸本)9781628419474

Cyclic AMP (cAMP) is a ubiquitous second messenger known to differentially regulate many cellular functions over a wide range of timescales. Several lines of evidence have suggested that the distribution of cAMP within cells is not uniform, and that cAMP compartmentalization is largely responsible for signaling specificity within the cAMP signaling pathway. However, to date, no studies have experimentally measured three dimensional (3D) cAMP distributions within cells. Here we use both 2D and 3D hyperspectral microscopy to visualize cAMP gradients in endothelial cells from the pulmonary microvasculature (PMVECs). cAMP levels were measured using a FRET-based cAMP sensor comprised of a cAMP binding domain from EPAC sandwiched between FRET donors and acceptors - Turquoise and Venus fluorescent proteins. Data were acquired using either a Nikon A1R spectral confocal microscope or custom spectral microscopy system. Analysis of hyperspectral image stacks from a single confocal slice or from summed images of all slices (2D analysis) indicated little or no cAMP gradients were formed within PMVECs under basal conditions or following agonist treatment. However, analysis of hyperspectral image stacks from 3D cellular geometries (z stacks) demonstrate marked cAMP gradients from the apical to basolateral membrane of PMVECs. These results strongly suggest that 2D imaging studies of cAMP compartmentalization whether epifluorescence or confocal microscopy - may lead to erroneous conclusions about the existence of cAMP gradients, and that 3D studies are required to assess mechanisms of signaling specificity.

关键词： Hyperspectral imaging spectral imaging microscopy FRET Forster resonance energy transfer cAMP second messenger signaling pulmonary endothelium

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Point spread function engineering with multiphoton SPIFI 13

Point spread function engineering with multiphoton SPIFI

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

作者： Wernsing, Keith A. Field, Jeffrey J. Domingue, Scott R. Allende-Motz, Alyssa M. DeLuca, Keith F. Levi, Dean H. DeLuca, Jennifer G. Young, Michael D. Squier, Jeff A. Bartels, Randy A. Colorado State Univ Dept Elect & Comp Engn Ft Collins CO 80523 USA Colorado State Univ Dept Biochem & Mol Biol Ft Collins CO 80523 USA Colorado State Univ Microscopy Core Facil Ft Collins CO 80523 USA Colorado State Univ Inst Genome Architecture & Funct Ft Collins CO 80523 USA Colorado Sch Mines Dept Phys Golden CO 80401 USA Natl Renewable Energy Lab Golden CO USA Colorado State Univ Sch Biomed Engn Ft Collins CO 80523 USA

ISBN: (纸本)9781628419474

MultiPhoton SPatIal Frequency modulated Imaging (MP-SPIFI) has recently demonstrated the ability to simultaneously obtain super-resolved images in both coherent and incoherent scattering processes - namely, second harmonic generation and two-photon fluorescence, respectively. 1 In our previous analysis, we considered image formation produced by the zero and first diffracted orders from the SPIFI modulator. However, the modulator is a binary amplitude mask, and therefore produces multiple diffracted orders. In this work, we extend our analysis to image formation in the presence of higher diffracted orders. We find that tuning the mask duty cycle offers a measure of control over the shape of super-resolved point spread functions in an MP-SPIFI microscope.

关键词： multiphoton microscopy super-resolution SPIFI

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