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Evaluation of Myelin Phagocytosis by Microglia/Macrophages in Nervous Tissue Using Flow Cytometry
Author(s) -
GómezLópez Ariadna Regina,
Manich Gemma,
Recasens Mireia,
Almolda Beatriz,
González Berta,
Castellano Bernardo
Publication year - 2021
Publication title -
current protocols
Language(s) - English
Resource type - Journals
ISSN - 2691-1299
DOI - 10.1002/cpz1.73
Subject(s) - microglia , phagocytosis , flow cytometry , myelin , microbiology and biotechnology , biology , myelin basic protein , in vivo , central nervous system , chemistry , immunology , inflammation , neuroscience , genetics
Determination of microglial phagocytosis of myelin has acquired importance in the study of demyelinating diseases. One strategy to determine microglial phagocytosis capacity consists of assaying microglia with fluorescently labeled myelin; however, most approaches are performed in cell culture, where microglia usually show important phenotypic differences compared with in vivo conditions. In this article we describe an adapted flow cytometry protocol to assay myelin phagocytosis by microglia obtained directly from in vivo tissue of the central nervous system. Key steps for a first analysis of phagocytic microglia are provided. Additionally, we describe how to fluorescently label myelin using a pH‐sensitive tag, pHrodo™ Green STP Ester. © 2021 Wiley Periodicals LLC. Basic Protocol : Assay for determination of myelin phagocytosis by microglia/macrophages using flow cytometry Support Protocol 1 : Conjugation of isolated and purified myelin with pHrodo Green STP Ester Support Protocol 2 : Quantification of phagocytic cell number by flow cytometry