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1、MolecularBioSystemsViewArticleOnlineREVIEWViewJournalProteomicanalysisofexosomalcargo:thechallengeofhighpurityvesicleisolationCitethis:DOI:10.1039/c6mb00082gAgataAbramowicz,PiotrWidlakandMonikaPietrowska*There-discoveryofexosomesasintercellularmessengerswit
2、hhighpotentialfordiagnosticandtherapeuticutilityhasledtothembecomingapopulartopicofresearchinrecentyears.Oneoftheessentialresearchareasinthisfieldisthecharacterizationofexosomalcargo,whichincludesnumerousnon-randomlypackedproteinsandnucleicacids.Unexpectedl
3、y,averychallengingaspectofexplorationofextracellularvesicleshasturnedouttobetheireffectiveandselectiveisolation.Thepluralityofdevelopedprotocolsleadstoqualitativeandquantitativevariabilityintermsoftheobtainedexosomes,whichsignificantlyaffectstheresultsofdowns
4、treamanalysesandmakesthemdifficulttocompare,reproduceandinterpretbetweenresearchgroups.Currently,thereisageneralconsensusamongtheexosome-orientedcommunityconcerningtheurgentneedfortheoptimizationandstandardizationofmethodsemployedforthepurificationofthesevesi
5、cles.Hence,wereviewhereseveralstrategiesforexosomepreparationincludingultracentrifugation,chemicalprecipitation,affinitycapturingandfiltrationReceived1stFebruary2016,techniques.TheadvantagesanddisadvantagesofdifferentapproachesarediscussedwithspecialAccepted14
6、thMarch2016emphasisbeingplacedontheiradequacyforproteomicsapplications,whichareparticularlysensitivetoDOI:10.1039/c6mb00082gsamplequality.Weconcludethatcertainmethods,exemplifiedbyultracentrifugationcombinedwithiodixanoldensitygradientcentrifugationorgelfil
7、tration,althoughlabor-intensive,providesuperiorwww.rsc.org/molecularbiosystemsqualityexosomepreparationssuitableforreliableanalysisbymassspectrometry.Introductioninwardbuddingoflateendosomes/multivesicularbodiesandthataresecretedduringtheirfusionwiththeplas
8、maExosomes,verysmallvesicleswithgreatpotential,areoneofmembrane(reviewedindetailinref.2and3).However,afterthecomponentsoftheextracellularmilieuinvolvedininter-overthirtyyearsofintenseexosome-relatedres