title: Quantitative imaging of cell dynamics in mouse embryos using light-sheet microscopy.
abstract: Single/selective-plane illumination, or light-sheet, systems offer several advantages over other fluorescence microscopy methods for live, 3D microscopy. These systems are valuable for studying embryonic development in several animal systems, such as Drosophila, C. elegans and zebrafish. The geometry of the light path in this form of microscopy requires the sample to be accessible from multiple sides and fixed in place so that it can be rotated around a single axis. Popular methods for mounting include hanging the specimen from a pin or embedding it in 1-2% agarose. These methods can be particularly problematic for certain samples, such as post-implantation mouse embryos, that expand significantly in size and are very delicate and sensitive to mounting. To overcome the current limitations and to establish a robust strategy for long-term (24 h) time-lapse imaging of E6.5-8.5 mouse embryos with light-sheet microscopy, we developed and tested a method using hollow agarose cylinders designed to accommodate for embryonic growth, yet provide boundaries to minimize tissue drift and enable imaging in multiple orientations. Here, we report the first 24-h time-lapse sequences of post-implantation mouse embryo development with light-sheet microscopy. We demonstrate that light-sheet imaging can provide both quantitative data for tracking changes in morphogenesis and reveal new insights into mouse embryogenesis. Although we have used this approach for imaging mouse embryos, it can be extended to imaging other types of embryos as well as tissue explants.
authors.affiliation: Department of Molecular Physiology and Biophysics, Baylor College of Medicine, Houston, TX 77030, USA.
authors.forename: Ryan S
authors.initials: RS
authors.lastname: Udan
authors.affiliation: Department of Molecular Physiology and Biophysics, Baylor College of Medicine, Houston, TX 77030, USA.
authors.forename: Victor G
authors.initials: VG
authors.lastname: Piazza
authors.affiliation: Department of Molecular Physiology and Biophysics, Baylor College of Medicine, Houston, TX 77030, USA.
authors.forename: Chih-Wei
authors.initials: CW
authors.lastname: Hsu
authors.affiliation: Developmental Biology Program, Memorial Sloan-Kettering Cancer Center, New York, NY 10065, USA.
authors.forename: Anna-Katerina
authors.initials: AK
authors.lastname: Hadjantonakis
authors.affiliation: Department of Molecular Physiology and Biophysics, Baylor College of Medicine, Houston, TX 77030, USA mdickins@bcm.tmc.edu.
authors.forename: Mary E
authors.initials: ME
authors.lastname: Dickinson
mesh_terms: D000818:Animals; D004622:Embryo, Mammalian / Q000166:cytology / Q000196:embryology*; D047108:Embryonic Development / Q000502:physiology*; D021621:Imaging, Three-Dimensional / Q000379:methods*; D051379:Mice; D008856:Microscopy, Fluorescence / Q000379:methods*; D012685:Sepharose; D059008:Time-Lapse Imaging / Q000379:methods*; D016610:Tissue Embedding / Q000379:methods
chemical_list: D012685:Sepharose
keywords: Cell dynamics; Light-sheet; Mouse embryo culture; Postimplantation; Quantitative