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dc.contributor.authorClauser, Karl R.
dc.contributor.authorNaba, Alexandra
dc.contributor.authorHynes, Richard O.
dc.date.accessioned2017-01-20T15:11:20Z
dc.date.available2017-01-20T15:11:20Z
dc.date.issued2015-07
dc.identifier.issn1940-087X
dc.identifier.urihttp://hdl.handle.net/1721.1/106549
dc.description.abstractThe extracellular matrix (ECM) is a complex meshwork of cross-linked proteins that provides biophysical and biochemical cues that are major regulators of cell proliferation, survival, migration, etc. The ECM plays important roles in development and in diverse pathologies including cardio-vascular and musculo-skeletal diseases, fibrosis, and cancer. Thus, characterizing the composition of ECMs of normal and diseased tissues could lead to the identification of novel prognostic and diagnostic biomarkers and potential novel therapeutic targets. However, the very nature of ECM proteins (large in size, cross-linked and covalently bound, heavily glycosylated) has rendered biochemical analyses of ECMs challenging. To overcome this challenge, we developed a method to enrich ECMs from fresh or frozen tissues and tumors that takes advantage of the insolubility of ECM proteins. We describe here in detail the decellularization procedure that consists of sequential incubations in buffers of different pH and salt and detergent concentrations and that results in 1) the extraction of intracellular (cytosolic, nuclear, membrane and cytoskeletal) proteins and 2) the enrichment of ECM proteins. We then describe how to deglycosylate and digest ECM-enriched protein preparations into peptides for subsequent analysis by mass spectrometry.en_US
dc.description.sponsorshipUnited States. Department of Defense (DOD Innovator Award W81XWH-14-1-0240)en_US
dc.description.sponsorshipNational Cancer Institute (U.S.) (Grant U54 CA126515/CA163109)en_US
dc.description.sponsorshipBroad Institute of MIT and Harvarden_US
dc.description.sponsorshipHoward Hughes Medical Instituteen_US
dc.description.sponsorshipNational Cancer Institute (U.S.) (David H. Koch Institute for Integrative Cancer Research at MIT. Grant P30-CA14051)en_US
dc.language.isoen_US
dc.publisherMyJoVE Corporationen_US
dc.relation.isversionofhttp://dx.doi.org/10.3791/53057en_US
dc.rightsCreative Commons Attribution-NonCommercial-NoDerivs Licenseen_US
dc.rights.urihttp://creativecommons.org/licenses/by-nc-nd/3.0/en_US
dc.sourceJournal of Visualized Experiments (JoVE)en_US
dc.titleEnrichment of Extracellular Matrix Proteins from Tissues and Digestion into Peptides for Mass Spectrometry Analysisen_US
dc.typeArticleen_US
dc.identifier.citationNaba, Alexandra, Karl R. Clauser, and Richard O. Hynes. “Enrichment of Extracellular Matrix Proteins from Tissues and Digestion into Peptides for Mass Spectrometry Analysis.” Journal of Visualized Experiments 101 (2015): n. pag.en_US
dc.contributor.departmentMassachusetts Institute of Technology. Department of Biologyen_US
dc.contributor.departmentKoch Institute for Integrative Cancer Research at MITen_US
dc.contributor.mitauthorNaba, Alexandra
dc.contributor.mitauthorHynes, Richard O.
dc.relation.journalJournal of Visualized Experimentsen_US
dc.eprint.versionFinal published versionen_US
dc.type.urihttp://purl.org/eprint/type/JournalArticleen_US
eprint.statushttp://purl.org/eprint/status/PeerRevieweden_US
dspace.orderedauthorsNaba, Alexandra; Clauser, Karl R.; Hynes, Richard O.en_US
dspace.embargo.termsNen_US
dc.identifier.orcidhttps://orcid.org/0000-0001-7603-8396
dspace.mitauthor.errortrue
mit.licensePUBLISHER_CCen_US


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