title: Quantitative Assessment of Cerebral Basement Membranes Using Electron Microscopy.
abstract: In this chapter we describe in detail the tissue processing techniques we employ for the study of cerebral tissue by transmission electron microscopy (TEM). In particular, we explain a technique that enables quantification of changes in cerebral basement membranes at the ultrastructural level. This is significant, as age related pathological conditions affecting the brain are often accompanied by ultrastructural changes in the cerebral vasculature.Briefly, experimental mice are fixed by perfusion and their brains removed. Brains are then vibratomed into 100 μm slices with regions of interest microdissected and processed for TEM following a protocol optimized for the preservation of cerebral tissue. Changes in the thickness of cerebral basement membranes are then quantified using novel software. Some prior knowledge of general TEM specimen preparation and sectioning will be useful when performing this protocol.
authors.affiliation: Clinical Neurosciences, Faculty of Medicine, University of Southampton, Southampton, UK. M.T.Sharp@soton.ac.uk.
authors.forename: Matthew MacGregor
authors.initials: MM
authors.lastname: Sharp
authors.affiliation: Biomedical Imaging Unit, University of Southampton, Southampton, UK.
authors.forename: Anton
authors.initials: A
authors.lastname: Page
authors.affiliation: Clinical Neurosciences, Faculty of Medicine, University of Southampton, Southampton, UK.
authors.forename: Alan
authors.initials: A
authors.lastname: Morris
authors.affiliation: Clinical Neurosciences, Faculty of Medicine, University of Southampton, Southampton, UK.
authors.forename: Roy O
authors.initials: RO
authors.lastname: Weller
authors.affiliation: Clinical Neurosciences, Faculty of Medicine, University of Southampton, Southampton, UK.
authors.forename: Roxana O
authors.initials: RO
authors.lastname: Carare
mesh_terms: D000818:Animals; D001485:Basement Membrane / Q000098:blood supply / Q000648:ultrastructure*; D002540:Cerebral Cortex / Q000098:blood supply / Q000648:ultrastructure*; D003890:Desiccation / Q000379:methods; D005557:Formaldehyde / Q000737:chemistry; D005976:Glutaral / Q000737:chemistry; D051379:Mice; D046529:Microscopy, Electron, Transmission / Q000379:methods*; D008867:Microtomy / Q000295:instrumentation / Q000379:methods; D016610:Tissue Embedding / Q000379:methods; D016707:Tissue Fixation / Q000379:methods
chemical_list: D005557:Formaldehyde; D005976:Glutaral
keywords: Basement membrane; Dehydration; Electron micrograph; Fixation; Microtomy; Resin embedding; Tissue processing; Transmission electron microscopy; Ultra-thin sectioning