Time-lapse X-ray phase-contrast microtomography for in vivo imaging and analysis of morphogenesis

Julian Moosmann, Alexey Ershov, Venera Weinhardt, Tilo Baumbach, Maneeshi S. Prasad, Carole Labonne, Xianghui Xiao, Jubin Kashef, Ralf Hofmann

    Research output: Contribution to journalArticle

    16 Citations (Scopus)

    Abstract

    X-ray phase-contrast microtomography (XPCμT) is a label-free, high-resolution imaging modality for analyzing early development of vertebrate embryos in vivo by using time-lapse sequences of 3D volumes. Here we provide a detailed protocol for applying this technique to study gastrulation in Xenopus laevis (African clawed frog) embryos. In contrast to μMRI, XPCμT images optically opaque embryos with subminute temporal and micrometer-range spatial resolution. We describe sample preparation, culture and suspension of embryos, tomographic imaging with a typical duration of 2 h (gastrulation and neurulation stages), intricacies of image pre-processing, phase retrieval, tomographic reconstruction, segmentation and motion analysis. Moreover, we briefly discuss our present understanding of X-ray dose effects (heat load and radiolysis), and we outline how to optimize the experimental configuration with respect to X-ray energy, photon flux density, sample-detector distance, exposure time per tomographic projection, numbers of projections and time-lapse intervals. The protocol requires an interdisciplinary effort of developmental biologists for sample preparation and data interpretation, X-ray physicists for planning and performing the experiment and applied mathematicians/computer scientists/physicists for data processing and analysis. Sample preparation requires 9-48 h, depending on the stage of development to be studied. Data acquisition takes 2-3 h per tomographic time-lapse sequence. Data processing and analysis requires a further 2 weeks, depending on the availability of computing power and the amount of detail required to address a given scientific problem.

    Original languageEnglish
    Pages (from-to)294-304
    Number of pages11
    JournalNature Protocols
    Volume9
    Issue number2
    DOIs
    Publication statusPublished - 1 Feb 2014

    Fingerprint

    Morphogenesis
    X-Rays
    Imaging techniques
    X rays
    Gastrulation
    Embryonic Structures
    Neurulation
    Radiolysis
    Xenopus laevis
    Thermal load
    Photons
    Anura
    Magnetic resonance imaging
    Dosimetry
    Embryonic Development
    Vertebrates
    Labels
    Data acquisition
    Suspensions
    Hot Temperature

    ASJC Scopus subject areas

    • Biochemistry, Genetics and Molecular Biology(all)

    Cite this

    Moosmann, J., Ershov, A., Weinhardt, V., Baumbach, T., Prasad, M. S., Labonne, C., ... Hofmann, R. (2014). Time-lapse X-ray phase-contrast microtomography for in vivo imaging and analysis of morphogenesis. Nature Protocols, 9(2), 294-304. https://doi.org/10.1038/nprot.2014.033

    Time-lapse X-ray phase-contrast microtomography for in vivo imaging and analysis of morphogenesis. / Moosmann, Julian; Ershov, Alexey; Weinhardt, Venera; Baumbach, Tilo; Prasad, Maneeshi S.; Labonne, Carole; Xiao, Xianghui; Kashef, Jubin; Hofmann, Ralf.

    In: Nature Protocols, Vol. 9, No. 2, 01.02.2014, p. 294-304.

    Research output: Contribution to journalArticle

    Moosmann, J, Ershov, A, Weinhardt, V, Baumbach, T, Prasad, MS, Labonne, C, Xiao, X, Kashef, J & Hofmann, R 2014, 'Time-lapse X-ray phase-contrast microtomography for in vivo imaging and analysis of morphogenesis', Nature Protocols, vol. 9, no. 2, pp. 294-304. https://doi.org/10.1038/nprot.2014.033
    Moosmann J, Ershov A, Weinhardt V, Baumbach T, Prasad MS, Labonne C et al. Time-lapse X-ray phase-contrast microtomography for in vivo imaging and analysis of morphogenesis. Nature Protocols. 2014 Feb 1;9(2):294-304. https://doi.org/10.1038/nprot.2014.033
    Moosmann, Julian ; Ershov, Alexey ; Weinhardt, Venera ; Baumbach, Tilo ; Prasad, Maneeshi S. ; Labonne, Carole ; Xiao, Xianghui ; Kashef, Jubin ; Hofmann, Ralf. / Time-lapse X-ray phase-contrast microtomography for in vivo imaging and analysis of morphogenesis. In: Nature Protocols. 2014 ; Vol. 9, No. 2. pp. 294-304.
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