Precise targeting for 3D cryo-correlative light and electron microscopy volume imaging of tissues using a FinderTOP.
SourceCommunications Biology, 6, 1, (2023), pp. 510, article 510
Article / Letter to editor
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Cell Biology (UMC)
Animal Ecology & Physiology
SubjectAll institutes and research themes of the Radboud University Medical Center; Radboudumc 10: Reconstructive and regenerative medicine Biochemistry (UMC); Radboudumc 10: Reconstructive and regenerative medicine Cell Biology (UMC)
Cryo-correlative light and electron microscopy (cryoCLEM) is a powerful strategy to high resolution imaging in the unperturbed hydrated state. In this approach fluorescence microscopy aids localizing the area of interest, and cryogenic focused ion beam/scanning electron microscopy (cryoFIB/SEM) allows preparation of thin cryo-lamellae for cryoET. However, the current method cannot be accurately applied on bulky (3D) samples such as tissues and organoids. 3D cryo-correlative imaging of large volumes is needed to close the resolution gap between cryo-light microscopy and cryoET, placing sub-nanometer observations in a larger biological context. Currently technological hurdles render 3D cryoCLEM an unexplored approach. Here we demonstrate a cryoCLEM workflow for tissues, correlating cryo-Airyscan confocal microscopy with 3D cryoFIB/SEM volume imaging. Accurate correlation is achieved by imprinting a FinderTOP pattern in the sample surface during high pressure freezing, and allows precise targeting for cryoFIB/SEM volume imaging.
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