The Experts below are selected from a list of 17970 Experts worldwide ranked by ideXlab platform

Jagesh V Shah - One of the best experts on this subject based on the ideXlab platform.

  • microfluidics integrated Time Lapse Imaging for analysis of cellular dynamics
    Integrative Biology, 2010
    Co-Authors: Dirk R Albrecht, Gregory H Underhill, Joshua Resnikoff, Avital Mendelson, Sangeeta N Bhatia, Jagesh V Shah
    Abstract:

    An understanding of the mechanisms regulating cellular responses has recently been augmented by innovations enabling the observation of phenotypes at high spatio-temporal resolution. Technologies such as microfluidics have sought to expand the throughput of these methods, although assimilation with advanced Imaging strategies has been limited. Here, we describe the pairing of high resolution Time-Lapse Imaging with microfluidic multiplexing for the analysis of cellular dynamics, utilizing a design selected for facile fabrication and operation, and integration with microscopy instrumentation. This modular, medium-throughput platform enables the long-term Imaging of living cells at high numerical aperture (via oil immersion) by using a conserved 96-well, ∼6 × 5 mm2 Imaging area with a variable input/output channel design chosen for the number of cell types and microenvironments under investigation. In the validation of this system, we examined fundamental features of cell cycle progression, including mitotic kinetics and spindle orientation dynamics, through the high-resolution parallel analysis of model cell lines subjected to anti-mitotic agents. We additionally explored the self-renewal kinetics of mouse embryonic stem cells, and demonstrate the ability to dynamically assess and manipulate stem cell proliferation, detect rare cell events, and measure extended Time-scale correlations. We achieved an experimental throughput of >900 cells/experiment, each observed at >40× magnification for up to 120 h. Overall, these studies illustrate the capacity to probe cellular functions and yield dynamic information in Time and space through the integration of a simple, modular, microfluidics-based Imaging platform.

Scott E Fraser - One of the best experts on this subject based on the ideXlab platform.

  • Time-Lapse Imaging of the early avian embryo.
    Methods in Cell Biology, 2020
    Co-Authors: Max Ezin, Scott E Fraser
    Abstract:

    Publisher Summary This chapter focuses on the Time-Lapse Imaging of the early avian embryo. Time-Lapse analysis records a series of still images of a developing embryo over Time. When aligned and assembled into an animated sequence, Time-Lapse Imaging reveals surprisingly dynamic events, which could be captured with no other methods. Filming the development of the early chick embryo has a long history, starting with the filming of gastrulation in ovo. By combining live Imaging with molecular experiments and microdissection, it is possible to experimentally address the role of given molecules or cell types in morphogenesis. The early avian embryo (blastula and gastrula) is well suited for Time-Lapse Imaging because it is relatively transparent, flat as it develops quickly outside the mother. Quail embryos lend themselves to Time-Lapse Imaging, as they look very similar to chick embryos at all stages, although they develop slightly faster than chick embryos. All of the labeling approaches that work in chick work well in quail, including infection with virus particles modified to drive the expression of various fluorescent markers, lipophilic dye labeling, and electroporation. Capturing the development of the live embryo has benefited from key technical advances in three domains—namely, image-recording hardware and software, culturing techniques, and tissue labeling methods. The current technology in those three domains has made Time-Lapse Imaging of the avian embryo more accessible and effective.

  • Hyperspectral phasor analysis enables multiplexed 5D in vivo Imaging
    Nature Methods, 2017
    Co-Authors: Francesco Cutrale, Vikas Trivedi, Le A Trinh, Chi-li Chiu, John M Choi, Marcela S Artiga, Scott E Fraser
    Abstract:

    Time-Lapse Imaging of multiple labels is challenging for biological Imaging as noise, photobleaching and phototoxicity compromise signal quality, while throughput can be limited by processing Time. Here, we report software called Hyper-Spectral Phasors (HySP) for denoising and unmixing multiple spectrally overlapping fluorophores in a low signal-to-noise regime with fast analysis. We show that HySP enables unmixing of seven signals in Time-Lapse Imaging of living zebrafish embryos. Hyper-Spectral Phasors allow unmixing of multiple signals even under conditions with low signal-to-noise ratios, and they enable highly multiplexed 5D Imaging of live zebrafish embryos labeled with conventional fluorophores.

  • hyperspectral phasor analysis enables multiplexed 5d in vivo Imaging
    Nature Methods, 2017
    Co-Authors: Francesco Cutrale, Vikas Trivedi, Le A Trinh, Chi-li Chiu, John M Choi, Marcela S Artiga, Scott E Fraser
    Abstract:

    Time-Lapse Imaging of multiple labels is challenging for biological Imaging as noise, photobleaching and phototoxicity compromise signal quality, while throughput can be limited by processing Time. Here, we report software called Hyper-Spectral Phasors (HySP) for denoising and unmixing multiple spectrally overlapping fluorophores in a low signal-to-noise regime with fast analysis. We show that HySP enables unmixing of seven signals in Time-Lapse Imaging of living zebrafish embryos.

Francesco Cutrale - One of the best experts on this subject based on the ideXlab platform.

  • Hyperspectral phasor analysis enables multiplexed 5D in vivo Imaging
    Nature Methods, 2017
    Co-Authors: Francesco Cutrale, Vikas Trivedi, Le A Trinh, Chi-li Chiu, John M Choi, Marcela S Artiga, Scott E Fraser
    Abstract:

    Time-Lapse Imaging of multiple labels is challenging for biological Imaging as noise, photobleaching and phototoxicity compromise signal quality, while throughput can be limited by processing Time. Here, we report software called Hyper-Spectral Phasors (HySP) for denoising and unmixing multiple spectrally overlapping fluorophores in a low signal-to-noise regime with fast analysis. We show that HySP enables unmixing of seven signals in Time-Lapse Imaging of living zebrafish embryos. Hyper-Spectral Phasors allow unmixing of multiple signals even under conditions with low signal-to-noise ratios, and they enable highly multiplexed 5D Imaging of live zebrafish embryos labeled with conventional fluorophores.

  • hyperspectral phasor analysis enables multiplexed 5d in vivo Imaging
    Nature Methods, 2017
    Co-Authors: Francesco Cutrale, Vikas Trivedi, Le A Trinh, Chi-li Chiu, John M Choi, Marcela S Artiga, Scott E Fraser
    Abstract:

    Time-Lapse Imaging of multiple labels is challenging for biological Imaging as noise, photobleaching and phototoxicity compromise signal quality, while throughput can be limited by processing Time. Here, we report software called Hyper-Spectral Phasors (HySP) for denoising and unmixing multiple spectrally overlapping fluorophores in a low signal-to-noise regime with fast analysis. We show that HySP enables unmixing of seven signals in Time-Lapse Imaging of living zebrafish embryos.

Martin A Denvir - One of the best experts on this subject based on the ideXlab platform.

  • adaptive prospective optical gating enables day long 3d Time Lapse Imaging of the beating embryonic zebrafish heart
    Nature Communications, 2019
    Co-Authors: Jonathan M Taylor, Carl J Nelson, Finnius A Bruton, Aryan K Baghbadrani, Charlotte Buckley, Carl Tucker, Adriano G Rossi, John J Mullins, Martin A Denvir
    Abstract:

    Three-dimensional fluorescence Time-Lapse Imaging of the beating heart is extremely challenging, due to the heart’s constant motion and a need to avoid pharmacological or phototoxic damage. Although real-Time triggered Imaging can computationally “freeze” the heart for 3D Imaging, no previous algorithm has been able to maintain phase-lock across developmental Timescales. We report a new algorithm capable of maintaining day-long phase-lock, permitting routine acquisition of synchronised 3D + Time video Time-Lapse datasets of the beating zebrafish heart. This approach has enabled us for the first Time to directly observe detailed developmental and cellular processes in the beating heart, revealing the dynamics of the immune response to injury and witnessing intriguing proliferative events that challenge the established literature on cardiac trabeculation. Our approach opens up exciting new opportunities for direct Time-Lapse Imaging studies over a 24-hour Time course, to understand the cellular mechanisms underlying cardiac development, repair and regeneration. Imaging heart development is challenging due to constant tissue movement and changing physical landmarks. Here the authors present an algorithm capable of maintaining phase-locked Imaging throughout a 24 hour Timespan, enabling long term TimeLapse Imaging studies of zebrafish heart development, repair and regeneration.

  • Hybrid optical gating for long-term 3D Time-Lapse Imaging of the beating embryonic zebrafish heart
    bioRxiv, 2019
    Co-Authors: Jonathan M Taylor, Carl J Nelson, Finnius A Bruton, Aryan K Baghbadrani, Charlotte Buckley, John J Mullins, Carl S. Tucker, Martin A Denvir
    Abstract:

    Three-dimensional fluorescence Time-Lapse Imaging of structural, cellular and subcellular processes in the beating heart is an increasingly achievable goal using the latest Imaging and computational techniques. However, previous approaches have had significant limitations. Temporarily arresting the heart using drugs disrupts the heart9s physiological state, and the use of ultra-high frame-rates for fluorescence image acquisition causes phototoxic cell damage. Real-Time triggered Imaging, synchronized to a specific phase in the cardiac-cycle, can computationally "freeze" the heart to acquire the minimal number of fluorescence images required for 3D Time-Lapse Imaging. However, until now no solution has been able to maintain phase-lock to the same point in the cardiac cycle for more than about one hour. Our new hybrid optical gating system maintains phase-lock for up to 24 h, acquiring synchronized 3D+Time video stacks of the unperturbed heart in vivo. This approach has enabled us to observe detailed developmental, structural, cellular and subcellular processes, including live cell division and cell fate tracking, in the embryonic zebrafish heart using transgenic fish lines expressing cell-specific fluorophores. We show that our approach not only provides high spatial and temporal resolution 3D-Imaging, but also avoids phototoxic injury, where alternative approaches induce measurable harm. This provides superb cellular and subcellular Imaging of the heart while it is beating in its normal physiological state, and opens up new and exciting opportunities for further study in the heart and other moving cellular and subcellular structures in vivo.

Marcela S Artiga - One of the best experts on this subject based on the ideXlab platform.

  • Hyperspectral phasor analysis enables multiplexed 5D in vivo Imaging
    Nature Methods, 2017
    Co-Authors: Francesco Cutrale, Vikas Trivedi, Le A Trinh, Chi-li Chiu, John M Choi, Marcela S Artiga, Scott E Fraser
    Abstract:

    Time-Lapse Imaging of multiple labels is challenging for biological Imaging as noise, photobleaching and phototoxicity compromise signal quality, while throughput can be limited by processing Time. Here, we report software called Hyper-Spectral Phasors (HySP) for denoising and unmixing multiple spectrally overlapping fluorophores in a low signal-to-noise regime with fast analysis. We show that HySP enables unmixing of seven signals in Time-Lapse Imaging of living zebrafish embryos. Hyper-Spectral Phasors allow unmixing of multiple signals even under conditions with low signal-to-noise ratios, and they enable highly multiplexed 5D Imaging of live zebrafish embryos labeled with conventional fluorophores.

  • hyperspectral phasor analysis enables multiplexed 5d in vivo Imaging
    Nature Methods, 2017
    Co-Authors: Francesco Cutrale, Vikas Trivedi, Le A Trinh, Chi-li Chiu, John M Choi, Marcela S Artiga, Scott E Fraser
    Abstract:

    Time-Lapse Imaging of multiple labels is challenging for biological Imaging as noise, photobleaching and phototoxicity compromise signal quality, while throughput can be limited by processing Time. Here, we report software called Hyper-Spectral Phasors (HySP) for denoising and unmixing multiple spectrally overlapping fluorophores in a low signal-to-noise regime with fast analysis. We show that HySP enables unmixing of seven signals in Time-Lapse Imaging of living zebrafish embryos.