title: Reconstructing adhesion structures in tissues by cryo-electron tomography of vitrified frozen sections.
abstract: Cryo-electron tomography enables three-dimensional insights into the macromolecular architecture of cells in a close-to-life state. However, it is limited to thin specimens, <1.0 μm in thickness, typically restricted to the peripheral areas of intact eukaryotic cells. Analysis of tissue ultrastructure, on the other hand, requires physical sectioning approaches, preferably cryo-sectioning, following which electron tomography (ET) may be performed. Nevertheless, cryo-electron microscopy of vitrified sections is a demanding technique and typically cannot be used to examine thick sections, >80-100 nm, due to surface crevasses. Here, we explore the potential use of cryo-ET of vitrified frozen sections (VFSs) for imaging cell adhesions in chicken smooth muscle and mouse epithelial tissues. By investigating 300-400 nm thick sections, which are collected on the EM grid and re-vitrified, we resolved fine 3D structural details of the membrane-associated dense plaques and flanking caveoli in smooth muscle tissue, and desmosomal adhesions in stratified epithelium. Technically, this method offers a simple approach for reconstructing thick volumes of hydrated frozen sections.
authors.affiliation: Department of Life Sciences, Ben-Gurion University of the Negev, BeerSheva 84105, Israel.
authors.forename: Melanie
authors.initials: M
authors.lastname: Bokstad
authors.forename: Helena
authors.initials: H
authors.lastname: Sabanay
authors.forename: Idit
authors.initials: I
authors.lastname: Dahan
authors.forename: Benjamin
authors.initials: B
authors.lastname: Geiger
authors.forename: Ohad
authors.initials: O
authors.lastname: Medalia
mesh_terms: D000818:Animals; D002448:Cell Adhesion / Q000502:physiology*; D002645:Chickens; D020285:Cryoelectron Microscopy / Q000379:methods*; D016616:Cryoultramicrotomy / Q000379:methods; D055032:Electron Microscope Tomography / Q000379:methods*; D005629:Frozen Sections; D009130:Muscle, Smooth / Q000648:ultrastructure*; D058989:Vitrification