title: 3D imaging of the early embryonic chicken heart with focused ion beam scanning electron microscopy.
abstract: Early embryonic heart development is a period of dynamic growth and remodeling, with rapid changes occurring at the tissue, cell, and subcellular levels. A detailed understanding of the events that establish the components of the heart wall has been hampered by a lack of methodologies for three-dimensional (3D), high-resolution imaging. Focused ion beam scanning electron microscopy (FIB-SEM) is a novel technology for imaging 3D tissue volumes at the subcellular level. FIB-SEM alternates between imaging the block face with a scanning electron beam and milling away thin sections of tissue with a FIB, allowing for collection and analysis of 3D data. FIB-SEM was used to image the three layers of the day 4 chicken embryo heart: myocardium, cardiac jelly, and endocardium. Individual images obtained with FIB-SEM were comparable in quality and resolution to those obtained with transmission electron microscopy. Up to 1,100 serial images were obtained in 4 nm increments at 4.88 nm resolution, and image stacks were aligned to create volumes 800-1,500 μm3 in size. Segmentation of organelles revealed their organization and distinct volume fractions between cardiac wall layers. We conclude that FIB-SEM is a powerful modality for 3D subcellular imaging of the embryonic heart wall.
authors.affiliation: 1Center for Developmental Health,Knight Cardiovascular Institute,Oregon Health & Science University,Portland,OR,USA.
authors.forename: Monique Y
authors.initials: MY
authors.lastname: Rennie
authors.affiliation: 2Oregon State University,Corvallis,OR,USA.
authors.forename: Curran G
authors.initials: CG
authors.lastname: Gahan
authors.affiliation: 3Department of Molecular Microbiology and Immunology,Oregon Health & Science University,Portland,OR,USA.
authors.forename: Claudia S
authors.initials: CS
authors.lastname: López
authors.affiliation: 1Center for Developmental Health,Knight Cardiovascular Institute,Oregon Health & Science University,Portland,OR,USA.
authors.forename: Kent L
authors.initials: KL
authors.lastname: Thornburg
authors.affiliation: 1Center for Developmental Health,Knight Cardiovascular Institute,Oregon Health & Science University,Portland,OR,USA.
authors.forename: Sandra
authors.initials: S
authors.lastname: Rugonyi
mesh_terms: D000818:Animals; D002642:Chick Embryo; D006321:Heart / Q000196:embryology*; D021621:Imaging, Three-Dimensional / Q000379:methods*; D008855:Microscopy, Electron, Scanning / Q000379:methods*
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