title: A method of correlative light and electron microscopy for yeast cells.
abstract: Correlative light and electron microscopy (CLEM) is a method of imaging in which the same specimen is observed by both light microscopy and electron microscopy. Specifically, CLEM compares images obtained by light and electron microscopy and makes a correlation between them. After the advent of fluorescent proteins, CLEM was extended by combining electron microscopy with fluorescence microscopy to enable molecular-specific imaging of subcellular structures with a resolution at the nanometer level. This method is a powerful tool that is used to determine the localization of specific molecules of interest in the context of subcellular structures. Knowledge of the localization of target proteins coupled with the functions of the structures to which they are localized yields valuable information about the molecular functions of these proteins. However, this method has been mostly applied to adherent cells due to technical difficulties in immobilizing non-adherent target cells, such as yeasts, during sample preparation. We have developed a method of CLEM applicable to yeast cells. In this report, we detail this method and present its extension to Live CLEM. The Live CLEM method enabled us to link the dynamic properties of molecules of interest to cellular ultrastructures in the yeast cell. Since yeasts are premier organisms in molecular genetics, combining CLEM with yeast genetics promises to provide important new findings for understanding the molecular basis of the function of cellular structures.
authors.affiliation: Graduate School of Frontier Biosciences, Osaka University, Suita 565-0871, Japan.
authors.forename: Haruhiko
authors.initials: H
authors.lastname: Asakawa
authors.affiliation: Graduate School of Frontier Biosciences, Osaka University, Suita 565-0871, Japan; Advanced ICT Research Institute Kobe, National Institute of Information and Communications Technology, 588-2 Iwaoka, Iwaoka-cho, Nishi-ku, Kobe 651-2492, Japan; Graduate School of Science, Osaka University, Toyonaka 560-0043, Japan.
authors.forename: Yasushi
authors.initials: Y
authors.lastname: Hiraoka
authors.affiliation: Graduate School of Frontier Biosciences, Osaka University, Suita 565-0871, Japan; Advanced ICT Research Institute Kobe, National Institute of Information and Communications Technology, 588-2 Iwaoka, Iwaoka-cho, Nishi-ku, Kobe 651-2492, Japan; Graduate School of Science, Osaka University, Toyonaka 560-0043, Japan. Electronic address: tokuko@nict.go.jp.
authors.forename: Tokuko
authors.initials: T
authors.lastname: Haraguchi
mesh_terms: D007091:Image Processing, Computer-Assisted; D008854:Microscopy, Electron / Q000379:methods*; D008856:Microscopy, Fluorescence / Q000379:methods*; D012441:Saccharomyces cerevisiae / Q000378:metabolism / Q000648:ultrastructure*; D029701:Saccharomyces cerevisiae Proteins / Q000648:ultrastructure; D012568:Schizosaccharomyces / Q000378:metabolism / Q000648:ultrastructure*; D029702:Schizosaccharomyces pombe Proteins / Q000648:ultrastructure; D012984:Software
chemical_list: D029701:Saccharomyces cerevisiae Proteins; D029702:Schizosaccharomyces pombe Proteins
keywords: CLEM; Correlative light and electron microscopy; Electron microscopy; Fluorescence microscopy; Live CLEM, Non-adherent cells