| Pathogen Vacuole Purification from Legionella-Infected Amoeba and Macrophages. | |
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MedLine Citation:
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PMID: 23150404 Owner: NLM Status: Publisher |
Abstract/OtherAbstract:
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Legionella pneumophila replicates intracellularly in environmental and immune phagocytes within a unique membrane-bound compartment, the Legionella-containing vacuole (LCV). Formation of LCVs is strictly dependent on the Icm/Dot type IV secretion system and the translocation of "effector" proteins into the cell. Some effector proteins decorate the LCV membrane and subvert host cell vesicle trafficking pathways. Here we describe a method to purify intact LCVs from Dictyostelium discoideum amoebae and RAW 264.7 murine macrophages. The method comprises a two-step protocol: first, LCVs are enriched by immuno-magnetic separation using an antibody against a bacterial effector protein specifically localizing to the LCV membrane, and second, the LCVs are further purified by density gradient centrifugation. The purified LCVs can be characterized by proteomics and other biochemical approaches. |
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Authors:
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Christine Hoffmann; Ivo Finsel; Hubert Hilbi |
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Publication Detail:
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Type: JOURNAL ARTICLE |
Journal Detail:
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Title: Methods in molecular biology (Clifton, N.J.) Volume: 954 ISSN: 1940-6029 ISO Abbreviation: Methods Mol. Biol. Publication Date: 2013 |
Date Detail:
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Created Date: 2012-11-14 Completed Date: - Revised Date: - |
Medline Journal Info:
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Nlm Unique ID: 9214969 Medline TA: Methods Mol Biol Country: - |
Other Details:
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Languages: ENG Pagination: 309-321 Citation Subset: - |
Affiliation:
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Max von Pettenkofer Institute, Ludwig-Maximilians University, Munich, Germany. |
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From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine
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