Selected Reaction Monitoring (SRM) Analysis of Epidermal Growth Factor Receptor (EGFR) in Formalin Fixed Tumor Tissue 英文参考文献.docVIP

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Selected Reaction Monitoring (SRM) Analysis of Epidermal Growth Factor Receptor (EGFR) in Formalin Fixed Tumor Tissue 英文参考文献.doc

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Selected Reaction Monitoring (SRM) Analysis of Epidermal Growth Factor Receptor (EGFR) in Formalin Fixed Tumor Tissue 英文参考文献

Hembroughetal.ClinicalProteomics2012,9:5 /content/9/1/5 CLINICAL PROTEOMICS RESEARCH OpenAccess SelectedReactionMonitoring(SRM)Analysisof EpidermalGrowthFactorReceptor(EGFR)in FormalinFixedTumorTissue ToddHembrough1,SheenoThyparambil1,Wei-LiLiao1,MarleneM.Darfler1,JosephAbdo1,KathleenM.Bengali1, PaulTaylor2,JiefeiTong2,HumbertoLara-Guerra3,ThomasK.Waddell3,MichaelF.Moran2,4,5,6, Ming-SoundTsao4,7,8,DavidB.Krizman1,9*andJonBurrows1 Abstract Background:Analysisofkeytherapeutictargetssuchasepidermalgrowthfactorreceptor(EGFR)inclinicaltissue samplesistypicallydonebyimmunohistochemistry(IHC)andisonlysubjectivelyquantitativethroughanarrow dynamicrange.Thedevelopmentofastandardized,highly-sensitive,linear,andquantitativeassayforEGFRforuse inpatienttumortissuecarrieshighpotentialforidentifyingthosepatientsmostlikelytobenefitfromEGFR-targeted therapies. Methods:Amassspectrometry-basedSelectedReactionMonitoring(SRM)assayfortheEGFRprotein(EGFR-SRM) wasdevelopedutilizingtheLiquidTissue?-SRMtechnologyplatform.Tissueculturecells(n=4)wereanalyzedby enzyme-linkedimmunosorbentassay(ELISA)toestablishquantitativeEGFRlevels.Matchingformalinfixedcultures wereanalyzedbytheEGFR-SRMassayandbenchmarkedagainstimmunoassayofthenon-fixedculturedcells. Xenografthumantumortissue(n=10)ofnon-smallcelllungcancer(NSCLC)originandNSCLCpatienttumortissue samples(n=23)weremicrodissectedandtheEGFR-SRMassayperformedonLiquidTissuelysatespreparedfrom microdissectedtissue.Quantitativecurvesandlinearregressioncurvesforcorrelationbetweenimmunoassayand SRMmethodologyweredevelopedinExcel. Results:TheassaywasdevelopedforquantitationofasingleEGFRtrypticpeptideforuseinFFPEpatienttissue withabsolutespecificitytouniquelydistinguishEGFRfromallotherproteinsincludingthereceptortyrosinekinases, IGF-1R,cMet,Her2,Her3,andHer4.Theassaywasanalyticallyvalidatedagainstacollectionoftissueculturecell lineswhereSRManalysisoftheformalinfixedcellsaccuratelyreflectsEGFRproteinlevelsinmatchingnon-formalin fixedculturesasestablishedb