Automated sub-5 nm image registration in integrated correlative fluorescence and electron microscopy using cathodoluminescence pointers

Martijn Haring, Nalan Liv Hamarat, Christiaan Zonnevylle, Angela Narvaez Gonzalez, Lennard Voortman, Pieter Kruit, Jacob Hoogenboom

    Research output: Contribution to journalArticleScientificpeer-review

    11 Citations (Scopus)
    31 Downloads (Pure)

    Abstract

    In the biological sciences, data from fluorescence and electron microscopy is correlated to allow fluorescence biomolecule identification within the cellular ultrastructure and/or ultrastructural analysis following live-cell imaging. High-accuracy (sub-100 nm) image overlay requires the addition of fiducial markers, which makes overlay accuracy dependent on the number of fiducials present in the region of interest. Here, we report an automated method for light-electron image overlay at high accuracy, i.e. below 5 nm. Our method relies on direct visualization of the electron beam position in the fluorescence detection channel using cathodoluminescence pointers. We show that image overlay using cathodoluminescence pointers corrects for image distortions, is independent of user interpretation, and does not require fiducials, allowing image correlation with molecular precision anywhere on a sample.
    Original languageEnglish
    Article number43621
    JournalScientific Reports
    Volume7
    DOIs
    Publication statusPublished - 2 Mar 2017

    Keywords

    • Fluorescence imaging
    • Microscopy
    • OA-Fund TU Delft

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