The potential of bioorthogonal chemistry for correlative light and electron microscopy: a call to arms
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The potential of bioorthogonal chemistry for correlative light and electron microscopy: a call to arms Daphne M. van Elsland 1,2 & Erik Bos 3 & Herman S. Overkleeft 1,2 & Abraham J. Koster 3 & Sander I. van Kasteren 1,2
Received: 2 December 2014 / Accepted: 27 April 2015 # The Author(s) 2015. This article is published with open access at Springerlink.com
Abstract With correlative light and electron microscopy (CLEM), the ultrastructural cellular location of a biomolecule of interest can be determined using a combination of light microscopy (LM) and electron microscopy (EM). In many cases, the application of CLEM requires the use of markers that need to be attached to a biomolecule of interest to allow its identification and localization. Here, we review the potential of bioorthogonal chemistry to introduce such markers for CLEM. Keywords Bioorthogonal chemistry . Click chemistry . Electron microscopy . CLEM
Introduction Correlative light and electron microscopy (CLEM) is an imaging technique that combines the virtues of light microscopy (LM) with those of electron microscopy (EM). With this technique, specific molecular and cellular structures in a cell can be identified with LM after which ultrastructural information Abraham J. Koster and Sander I. van Kasteren share the corresponding authorship. * Abraham J. Koster [email protected] * Sander I. van Kasteren [email protected] 1
Division of Bio-organic Synthesis, Leiden Institute of Chemistry, Leiden University, Leiden, The Netherlands
2
Institute for Chemical Immunology, Leiden, The Netherlands
3
Molecular Cell Biology, Leiden University Medical Center, Leiden, The Netherlands
about its subcellular location and context can be obtained. CLEM studies that involve fluorescence microscopy may benefit from fluorescent markers that can be attached to molecules of interest to allow their identification and localization. To date, most readily this has been done by fluorescent fusion proteins, by fluorescent antibody labelling or by the chemical modification of a protein with a fluorescent detection group [1–3]. As well as these fluorescent detection moieties, structures must be present in the CLEM sample that are both EM and LM detectable in order to correlate (overlay) the LM image with the EM image. Examples of such EM/LM detectable structures are fluorescently labelled cellular structures that are suitable to be identified by EM through their distinct morphology (e.g. stained nuclei) or fluorescently labelled electron-dense particles (e.g. fluorescent microspheres) [1–4]. The above-mentioned labelling approaches have been very successfully applied to CLEM imaging of specific proteins in their cellular context. However, they carry some limitations. First, the use of fluorescent fusion proteins requires genetic manipulation of the cell, which can be difficult and can affect the function of the protein of interest [5]. An alternative to genetic manipulation is antibody labelling. However, for specimens prepared for CLEM, antibody labell
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