biostudies-literatureJeppesen DKNIBIB NIH HHSNIDDK NIH HHSNHLBI NIH HHSVanderbilt Digestive Diseases Research CenterNCI NIH HHSNIHVanderbilt-Ingram Cancer CenterNIGMS NIH HHS428-445.e18https://www.ebi.ac.uk/biostudies/studies/S-EPMC6664447biostudies-literatureUnknown177(2)The heterogeneity of small extracellular vesicles and presence of non-vesicular extracellular matter have led to debate about contents and functional properties of exosomes. Here, we employ high-resolution density gradient fractionation and direct immunoaffinity capture to precisely characterize the RNA, DNA, and protein constituents of exosomes and other non-vesicle material. Extracellular RNA, RNA-binding proteins, and other cellular proteins are differentially expressed in exosomes and non-vesicle compartments. Argonaute 1-4, glycolytic enzymes, and cytoskeletal proteins were not detected in exosomes. We identify annexin A1 as a specific marker for microvesicles that are shed directly from the plasma membrane. We further show that small extracellular vesicles are not vehicles of active CellReassessment of Exosome Composition.PMC6664447UG3 CA241685P30 CA068485F31 HL136081R35 GM125028P30 DK058404P50 CA236733U01-EB021214U01 EB021214U19 CA179514R35 CA197570Ping JCoffey RJPatton JGLiebler DCJeppesen DKLiu QZhang QFranklin JLZimmerman LJRome LHHigginbotham JNEvans RFenix AMFissell WHBurnette DTfalseReassessment of Exosome Composition.The heterogeneity of small extracellular vesicles and presence of non-vesicular extracellular matter have led to debate about contents and functional properties of exosomes. Here, we employ high-resolution density gradient fractionation and direct immunoaffinity capture to precisely characterize the RNA, DNA, and protein constituents of exosomes and other non-vesicle material. Extracellular RNA, RNA-binding proteins, and other cellular proteins are differentially expressed in exosomes and non-vesicle compartments. Argonaute 1-4, glycolytic enzymes, and cytoskeletal proteins were not detected in exosomes. We identify annexin A1 as a specific marker for microvesicles that are shed directly from the plasma membrane. We further show that small extracellular vesicles are not vehicles of active 2019-01-01T00:00:00Z2019 Apr2026-05-04T03:39:20.112Z2020-10-30T08:20:23ZS-EPMC66644473095167010.1016/j.cell.2019.02.029