Abstract
A complete portrait of a cell requires a detailed description of its molecular topography: proteins must be linked to particular organelles. Immuno-electron microscopy can reveal locations of proteins with nanometer resolution but is limited by the quality of fixation, the paucity of antibodies, and the inaccessibility of the antigens. Here, we describe correlative fluorescence electron microscopy for the nanoscopic localization of proteins in electron micrographs. Proteins tagged with Citrine or tdEos were expressed in Caenorhabditis elegans, fixed and embedded. Tagged proteins were imaged from ultrathin sections using stimulated emission depletion microscopy (STED) or photoactivated localization microscopy (PALM). Fluorescence was correlated with organelles imaged in electron micrographs from the same sections. These methods were used to successfully localize histones, a mitochondrial protein, and a presynaptic dense projection protein in electron micrographs.
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