Abstract
Dark microglia, a recently described phenotype, are found in high numbers in nonhomeostatic conditions (e.g., Alzheimer’s disease pathology, aging, chronic stress). As a specific protein marker has not yet been defined, they cannot be studied using conventional cellular biology techniques. They are recognized by their unique ultrastructural features visible under electron microscopy. This nanoscale resolution imaging technique allows the identification of cells based on their ultrastructure or immunoreactivity to certain proteins. In this protocol, we describe the steps necessary for the preparation of high-quality brain tissues for transmission electron microscopy, the imaging, the identification of dark microglia, and the ultrastructural analysis of various parameters that can be studied in these cells.
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Acknowledgments
We would like to give a special thanks to Kanchan Bisht and Kaushik Sharma who were the driving force behind the first dark microglia paper. We are grateful to Julie-Christine Lévesque at the Bio-imaging platform of CRCHU de Québec-Université Laval, as well as Nathalie Vernoux, for the training and support provided. We want to highlight the help of Julie Savage for her critical proofreading of the protocol. We also would like to thank Cynthia Lecours and Katherine Picard for contributing images for the figures, as well as Chin Wai (Thomas) Hui for providing the Poly I:C samples. This work was mainly supported by the Canadian Institutes of Health Research (CIHR) foundation grant awarded to M.E.T. as well as through scholarships awarded to the authors. MKSP is recipient of CIHR and Fonds de Recherche du Québec—Santé (FRQS) master scholarships, while M.B. is recipient of a FRQS doctoral scholarship. M.E.T. also holds the Canada Research Chair (Tier II) of Neuroimmune Plasticity in Health and Therapy.
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St-Pierre, MK., Bordeleau, M., Tremblay, MÈ. (2019). Visualizing Dark Microglia. In: Garaschuk, O., Verkhratsky, A. (eds) Microglia. Methods in Molecular Biology, vol 2034. Humana, New York, NY. https://doi.org/10.1007/978-1-4939-9658-2_8
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DOI: https://doi.org/10.1007/978-1-4939-9658-2_8
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