Targeted ultramicrotomy: a valuable tool for correlated light and electron microscopy of small model organisms. 2012

Irina Kolotuev, and Daniel J Bumbarger, and Michel Labouesse, and Yannick Schwab
Institut de Génétique et Développement de Rennes, UMR 6290 CNRS, Université Rennes 1, Faculté de Medecine/1, 35043 Rennes Cedex, France.

Correlative light and electron microscopy (CLEM) is used when one needs to combine both imaging modalities on the same sample. When working on living small model organisms, such as Caenorhabditis elegans, specific CLEM protocols are required to acquire high-resolution light microscopic images of a region of interest and thereafter to relocate and study the same object at the ultrastructural level using a transmission electron microscope. In this chapter, we describe how to process living specimens from the confocal microscope to the transmission electron microscopy (TEM), focusing on an improved ultramicrotomy technique that allows a precise and reliable targeting of the object of interest. This improvement significantly reduces the time consuming and frequently frustrating search for the region of interest. Our targeted ultramicrotomy protocol is versatile enough to be applied on a variety of bulk specimens, such as fly and fish embryos, or mouse tissues.

UI MeSH Term Description Entries
D000818 Animals Unicellular or multicellular, heterotrophic organisms, that have sensation and the power of voluntary movement. Under the older five kingdom paradigm, Animalia was one of the kingdoms. Under the modern three domain model, Animalia represents one of the many groups in the domain EUKARYOTA. Animal,Metazoa,Animalia
D012685 Sepharose Agarose,Sepharose 4B,Sepharose C1 4B,4B, Sepharose C1,C1 4B, Sepharose
D015925 Cryopreservation Preservation of cells, tissues, organs, or embryos by freezing. In histological preparations, cryopreservation or cryofixation is used to maintain the existing form, structure, and chemical composition of all the constituent elements of the specimens. Cryofixation,Cryonic Suspension,Cryonic Suspensions,Suspension, Cryonic
D016613 Plastic Embedding The infiltrating of histological specimens with plastics, including acrylic resins, epoxy resins and polyethylene glycol, for support of the tissues in preparation for sectioning with a microtome. Embedding, Plastic
D016616 Cryoultramicrotomy The technique of using a cryostat or freezing microtome, in which the temperature is regulated to -20 degrees Celsius, to cut ultrathin frozen sections for microscopic (usually, electron microscopic) examination. Cryosectioning,Freeze Sectioning,Sectioning, Freeze
D017173 Caenorhabditis elegans A species of nematode that is widely used in biological, biochemical, and genetic studies. Caenorhabditis elegan,elegan, Caenorhabditis
D046529 Microscopy, Electron, Transmission Electron microscopy in which the ELECTRONS or their reaction products that pass down through the specimen are imaged below the plane of the specimen. Electron Diffraction Microscopy,Electron Microscopy, Transmission,Microscopy, Electron Diffraction,Transmission Electron Microscopy,Diffraction Microscopy, Electron,Microscopy, Transmission Electron
D018613 Microscopy, Confocal A light microscopic technique in which only a small spot is illuminated and observed at a time. An image is constructed through point-by-point scanning of the field in this manner. Light sources may be conventional or laser, and fluorescence or transmitted observations are possible. Confocal Microscopy,Confocal Microscopy, Scanning Laser,Laser Microscopy,Laser Scanning Confocal Microscopy,Laser Scanning Microscopy,Microscopy, Confocal, Laser Scanning,Confocal Laser Scanning Microscopy,Confocal Microscopies,Laser Microscopies,Laser Scanning Microscopies,Microscopies, Confocal,Microscopies, Laser,Microscopies, Laser Scanning,Microscopy, Laser,Microscopy, Laser Scanning,Scanning Microscopies, Laser,Scanning Microscopy, Laser
D020285 Cryoelectron Microscopy Electron microscopy involving rapid freezing of the samples. The imaging of frozen-hydrated molecules and organelles permits the best possible resolution closest to the living state, free of chemical fixatives or stains. Electron Cryomicroscopy,Cryo-electron Microscopy,Cryo electron Microscopy,Cryo-electron Microscopies,Cryoelectron Microscopies,Cryomicroscopies, Electron,Cryomicroscopy, Electron,Electron Cryomicroscopies,Microscopies, Cryo-electron,Microscopies, Cryoelectron,Microscopy, Cryo-electron,Microscopy, Cryoelectron

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