EN 18057 2025.docx

EN18057June2025EuropeanstandardNormeeuropeenneEuropaischenormICS67.120.10EnglishVersionFoodauthenticity-QuantitationofroedeerDNArelativetomammalianDNAinmeatandmeatproductsbyreal-timePCRAuthenticitedesaliments-QuantificationdeADNdechevreuilparrapportaADNdemammiferedanslaviandeetlesproduitscarriesparPCRentempsreelLebensmittelauthentizitat-QuantifizierungvonReh-DNAimVerhaltniszuSaugetier-DNAinFleischundFleischproduktenmittelsReal-time-PCRThisEuropeanStandardwasapprovedbyCENon4May2025.CENmembersareboundtocomplywiththeCEN/CENELECInternalRegulationswhichstipulatetheconditionsforgivingthisEuropeanStandardthestatusofanationalstandardwithoutanyalteration.Up-to-datelistsandbibliographicalreferencesconcerningsuchnationalstandardsmaybeobtainedonapplicationtotheCEN-CENELECManagementCentreortoanyCENmember.ThisEuropeanStandardexistsinthreeofficialversions(English,French,German).AversioninanyotherlanguagemadebytranslationundertheresponsibilityofaCENmemberintoitsownlanguageandnotifiedtotheCEN-CENELECManagementCentrehasthesamestatusastheofficialversions.CENmembersarethenationalstandardsbodiesofAustria,Belgium,Bulgaria,Croatia,Cyprus,CzechRepublic,Denmark,Estonia,Finland,France,Germany,Greece,Hungary,Iceland,Ireland,Italy,Latvia,Lithuania,Luxembourg,Malta,Netherlands,Norway,Poland,Portugal,RepublicofNorthMacedonia,Romania,Serbia,Slovakia,Slovenia,Spain,Sweden,Switzerland,TiirkiyeandUnitedKingdom.EUROPEANCOMMITTEEFORSTANDARDIZATIONCOMITEEUROPEENDENORMALISATIONEuropaischeskomiteefurnormungCEN-CENELECManagementCentre:RuedelaScience23,B-1040Brussels©2025CENAllrightsofexploitationinanyformandbyanymeansreservedRef.No.EN18057:2025EworldwideforCENnationalMembers.ContentsPageEuropeanforeword3Introduction41 Scope52 Normativereferences53 Termsanddefinitions54 Symbolsandabbreviations65 Principle66 Reagentsandmaterials67 Apparatus88 Procedure88.1 Preparationofthetestsampleandtestportion88.2 PreparationofDNAextracts88.3 Preparationofroedeerandmammaliancalibrationstandards88.4 PCRsetup98.4.1 Samplesandcontrols98.4.2 Reactionmixes98.4.3 Thermalcycling109 Accept/rejectcriteria109.1 General109.2 Dataanalysis1110 Validationstatusandperformancecriteria1110.1 General1110.2 Repeatability1410.3 Reproducibility1410.4 Recovery1410.5 LimitofDetection(LOD)1410.6 Specificity1511 Testreport15Bibliography16EuropeanforewordThisdocument(ENl8057:2025)hasbeenpreparedbyTechnicalCommitteeCEN/TC460"Foodauthenticity”,thesecretariatofwhichisheldbyDIN.ThisEuropeanStandardshallbegiventhestatusofanationalstandard,eitherbypublicationofanidenticaltextorbyendorsement,atthelatestbyDecember2025,andcon11ictingnationalstandardsshallbewithdrawnatthelatestbyDecember2025.Attentionisdrawntothepossibilitythatsomeoftheelementsofthisdocumentmaybethesubjectofpatentrights.CENshallnotbeheldresponsibleforidentifyinganyorallsuchpatentrights.Anyfeedbackandquestionsonthisdocumentshouldbedirectedtotheusers'nationalstandardsbody.AcompletelistingofthesebodiescanbefoundontheCENwebsite.AccordingtotheCEN-CENELECInternalRegulations,thenationalstandardsorganisationsofthefollowingcountriesareboundtoimplementthisEuropeanStandard:Austria,Belgium,Bulgaria,Croatia,Cyprus,CzechRepublic,Denmark,Estonia,Finland,France,Germany,Greece,Hungary,Iceland,Ireland,Italy,Latvia,Lithuania,Luxembourg,Malta,Netherlands,Norway,Poland,Portugal,RepublicofNorthMacedonia,Romania,Serbia,Slovakia,Slovenia7Spain,Sweden,Switzerland,TiirkiyeandtheUnitedKingdom.IntroductionFoodauthenticityandintegrityarekeyaspectsintermsofconsumerprotection.Sincethelate1980s,globalizationhastakenplaceinthetradeoffood.Duringthelastdecades,manyofmethodsapplyingPCR,particularlyreal-timePCRprotocols,havebeenestablishedfortheidentificationofanimalspeciesusedforfoodconsumption(e.g.pig,cattle,sheep,horse,chickenandturkey).Gamemeatisparticularlysusceptibletofraudulentlabellingsinceitismorevaluablethanmeatfromdomesticanimals.Avarietyofgamemeatproducts,commonlycontainingreddeerorroedeer,iscommerciallyavailable.Forverifyingthecorrectlabellingofcommercialgamemeatproducts,thedevelopmentofharmonizedandstandardizedprotocolsfortheauthenticationofmeatproductsisimportanttoestablishreliablemethodsforthedetectionofpotentialfoodfraud.1 ScopeThisdocumentspecifiesareal-timePCRprocedureforthequantitationoftheamountofroedeerDNArelativetototalmammalianDNAinmeatandmeatproducts.Resultsofthisroedeerassayareexpressedintermsofroedeer(CaPreOlUSCaPreOlUS)haploidgenomecopynumbersrelativetototalmammalianhaploidgenomecopynumbers.Thecontentofroedeercanalsobeexpressedasmassfractionin%usinggravimetricallypreparedcalibrationmaterialfrommeatmixturesormodelsamples.ThemethodhasbeenpreviouslyvalidatedinacollaborativestudyandappliedtoDNAextractedfromsamplesthatconsistofrawroedeermeatinarawpigmeatbackgroundaswellasrawandboiledsausages.ThelimitofdetectionoftheroedeerPCRhasbeendeterminedexperimentallytobeatleast5t