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  • 标题:In situ fiducial markers for 3D correlative cryo-fluorescence and FIB-SEM imaging
  • 本地全文:下载
  • 作者:Nadav Scher ; Katya Rechav ; Perrine Paul-Gilloteaux
  • 期刊名称:iScience
  • 印刷版ISSN:2589-0042
  • 出版年度:2021
  • 卷号:24
  • 期号:7
  • 页码:1-13
  • DOI:10.1016/j.isci.2021.102714
  • 语种:English
  • 出版社:Elsevier
  • 摘要:SummaryImaging of cells and tissues has improved significantly over the last decade. Dual-beam instruments with a focused ion beam mounted on a scanning electron microscope (FIB-SEM), offering high-resolution 3D imaging of large volumes and fields-of-view are becoming widely used in the life sciences. FIB-SEM has most recently been implemented on fully hydrated, cryo-immobilized, biological samples. Correlative light and electron microscopy workflows combining fluorescence microscopy (FM) with FIB-SEM imaging exist, whereas workflows combining cryo-FM and cryo-FIB-SEM imaging are not yet commonly available. Here, we demonstrate that fluorescently labeled lipid droplets can serve asin situfiducial markers for correlating cryo-FM and FIB-SEM datasets and that this approach can be used to target the acquisition of large FIB-SEM stacks spanning tens of microns under cryogenic conditions. We also show that cryo-FIB-SEM imaging is particularly informative for questions related to organelle structure and inter-organellar contacts, nuclear organization, and mineral deposits in cells.Graphical abstractDisplay OmittedHighlights•3D CLEM of cryo-FM and FIB-SEM datasets using fluorescently labeled lipid droplets•Cryo-FIB-SEM imaging of organelle-organelle interactions and nuclear organizationCell biology; Technical aspects of cell biology; Structural biology
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