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Isolation of Mitochondria-Associated ER Membranes (MAMs), Synaptic MAMs, and Glycosphingolipid Enriched Microdomains (GEMs) from Brain Tissues and Neuronal Cells

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Mitochondrial Medicine

Part of the book series: Methods in Molecular Biology ((MIMB,volume 2277))

Abstract

Subcellular fractionation is a valuable procedure in cell biology to separate and purify various subcellular constituents from one another, i.e., nucleus, cytosol, membranes/organelles, and cytoskeleton. The procedure relies on the use of differential centrifugation of cell and tissue homogenates. Fractionated subcellular organelles may be subjected to additional purification steps that enable the isolation of specific cellular sub-compartments, including interorganellar membrane contact sites. Here we outline a protocol tailored to the isolation of mitochondria, mitochondria-associated ER membranes (MAMs), and glycosphingolipid enriched microdomains (GEMs) from the adult mouse brain, primary neurospheres, and murine embryonic fibroblasts (MEFs). We also provide a detailed protocol for the purification of synaptosomes and their corresponding MAMs .

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Acknowledgments

We are indebted to Camenzind G. Robinson (Director of Electron Microscopy Shared Resource at SJCRH) for his guidance and skillful help in acquiring the TEM images. A.d’A. holds the Jewelers For Children (JFC) Endowed Chair in Genetics and Gene Therapy. This work was funded in part by the NIH grant GM104981, CA021765, the National Tay-Sachs & Allied Diseases Association (NTSAD), the Assisi Foundation of Memphis, and the American Lebanese Syrian Associated Charities (ALSAC).

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Authors and Affiliations

  1. Department of Genetics, St. Jude Children Research Hospital, Memphis, TN, USA

    Ida Annunziata, Jason Andrew Weesner & Alessandra d’Azzo

  2. Department of Anatomy and Neurobiology, College of Graduate Health Sciences, University of Tennessee Health Science Center, Memphis, TN, USA

    Jason Andrew Weesner

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  1. Ida Annunziata
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  2. Jason Andrew Weesner
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  3. Alessandra d’Azzo
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Correspondence to Alessandra d’Azzo .

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Editors and Affiliations

  1. Department of Pharmaceutical Sciences, Midwestern University, Glendale, AZ, USA

    Volkmar Weissig

  2. Cochin Hospital, Cochin Institute, INSERM U1016, PARIS, France

    Marvin Edeas

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Annunziata, I., Weesner, J.A., d’Azzo, A. (2021). Isolation of Mitochondria-Associated ER Membranes (MAMs), Synaptic MAMs, and Glycosphingolipid Enriched Microdomains (GEMs) from Brain Tissues and Neuronal Cells. In: Weissig, V., Edeas, M. (eds) Mitochondrial Medicine. Methods in Molecular Biology, vol 2277. Humana, New York, NY. https://doi.org/10.1007/978-1-0716-1270-5_22

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  • DOI: https://doi.org/10.1007/978-1-0716-1270-5_22

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  • Publisher Name: Humana, New York, NY

  • Print ISBN: 978-1-0716-1269-9

  • Online ISBN: 978-1-0716-1270-5

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