title: Correlative light and electron microscopy analysis of the centrosome: A step-by-step protocol.
abstract: Correlative light and electron microscopy harnesses the best from each of the two modalities of microscopy it utilizes; while light microscopy provides information about the dynamic properties of the cellular structure or fluorescently labeled protein, electron microscopy provides ultrastructural information in an unsurpassed resolution. However, tracing a particular cell and its rare and small structures such as centrosomes throughout numerous steps of the experiment is not a trivial task. In this chapter, we present the experimental workflow for combining live-cell fluorescence microscopy analysis with classical transmission electron microscopy, adapted for the studies of the centrosomes and basal bodies. We describe, in a step-by-step manner, an approach that can be affordably and successfully employed in any typical cell biology laboratory. The article details all key phases of the analysis starting from cell culture, live-cell microscopy, and sample fixation, through the steps of sample preparation for electron microscopy, to the identification of the target cell on the electron microscope.
authors.affiliation: Laboratory of Protein Dynamics and Signaling, NIH/NCI/CCR-Frederick, Frederick, MD, USA.
authors.forename: Dong
authors.initials: D
authors.lastname: Kong
authors.affiliation: Laboratory of Protein Dynamics and Signaling, NIH/NCI/CCR-Frederick, Frederick, MD, USA.
authors.forename: Jadranka
authors.initials: J
authors.lastname: Loncarek
mesh_terms: D018385:Centrosome / Q000648:ultrastructure*; D006367:HeLa Cells; D006801:Humans; D046529:Microscopy, Electron, Transmission / Q000379:methods; D008856:Microscopy, Fluorescence / Q000379:methods; D008867:Microtomy; D016613:Plastic Embedding; D013194:Staining and Labeling
keywords: Centriole; Centrosome; Correlative light and electron microscopy (CLEM); Live-cell microscopy; Serial sectioning