title: Separation of magnetically isolated TNF receptosomes from mitochondria.
abstract: We previously demonstrated that tumor necrosis factor receptor-1 (TNF-R1) initiates distinct TNF signaling pathways depending on the localization of the receptor. While TNF-R1 at the plasma membrane transmits proinflammatory and antiapoptotic signals, internalized TNF-R1 forms signaling endosomes (TNF receptosomes) that transmit proapoptotic signals. These findings were obtained by a novel technique for the isolation of morphologically intact endocytic vesicles containing magnetically labeled TNF-R1 complexes using a high-gradient, free-flow magnetic chamber. Since intact mitochondria appeared to be a major contaminating organelle in these preparations, we subsequently included a second purification step by iodixanol density centrifugation to obtain a mitochondria-free receptosome preparation.
authors.affiliation: Institute of Immunology, Christian-Albrechts-University of Kiel, Kiel, Germany.
authors.forename: Vladimir
authors.initials: V
authors.lastname: Tchikov
authors.affiliation: Institute of Immunology, Christian-Albrechts-University of Kiel, Kiel, Germany.
authors.forename: Jürgen
authors.initials: J
authors.lastname: Fritsch
authors.affiliation: Institute of Immunology, Christian-Albrechts-University of Kiel, Kiel, Germany. Electronic address: schuetze@immunologie.uni-kiel.de.
authors.forename: Stefan
authors.initials: S
authors.lastname: Schütze
mesh_terms: D002458:Cell Fractionation / Q000379:methods*; D045744:Cell Line, Tumor; D002499:Centrifugation, Density Gradient; D011992:Endosomes / Q000378:metabolism; D006801:Humans; D018189:Immunomagnetic Separation; D008928:Mitochondria / Q000378:metabolism*; D022161:Transport Vesicles / Q000378:metabolism*; D014409:Tumor Necrosis Factor-alpha / Q000302:isolation & purification / Q000378:metabolism*
chemical_list: C439524:TNF protein, human; D014409:Tumor Necrosis Factor-alpha
keywords: Endosomes; Magnetic separation; Mitochondria; Receptosomes; TNF-receptor 1