研究用腫瘤細胞系的質(zhì)量控制和要求課件

研究用腫瘤細胞系的質(zhì)量

控制和要求

中國醫(yī)學科學院基礎醫(yī)學研究所劉玉琴研究用腫瘤細胞系的質(zhì)量控制和要求SomeFactsabouttheuseofculturedcells--Nobelprize2001，Cellcycle2002,apoptosis2006,RNAInterference2007,homologousrecombinationandgenetargeting2008,HPV—cervicalca,HIV--aids2009,telomere&telomeraseNobelprizewinner,useinvitroculturedcellsfortheirresearches.研究用腫瘤細胞系的質(zhì)量控制和要求現(xiàn)代生命科學研究中培養(yǎng)細胞的使用越來越廣泛1969~2004：celluseincreasingsteadilyPapersusecellsincrease2~2.5Xbiology,pharmacologyandmedicine----tumorresearch不可少的研究材料、對象、工具Useofcontaminatedormisidentifiedcellsincrease10X研究用腫瘤細胞系的質(zhì)量控制和要求Commonproblemsmicrobiologicalcontaminationwithparticularrespecttomycoplasmainfections(incidenceofca.25%)cross-contaminationbyothercelllinesandmisidentificationormisclassification(incidenceofmorethan15%)acquisitionofgeneticandepigeneticvariationsduetoover-passagingofcelllinesleadingtoageneticdriftofcellculturesandtotheselectionofalteredsubclones.研究用腫瘤細胞系的質(zhì)量控制和要求Wherefrom?faultycellculturetechniques:sharedreagents,repeateduseofthesamepipetteduringre-feedingoperationsandmanipulationofmultipleculturesatthesametimewithoutadequateisolationofonecelltypefromanother.theintroductionoffalse,contaminatedorover-passagedcelllinesfromothersourcesintothecellculturelaboratory(35%)Mislabeling,lackofawarenessIntentionalco-cultivationduringpropagationofhumanstemorprimarycellsusingafeederlayerderivedfromanotherspecies研究用腫瘤細胞系的質(zhì)量控制和要求細胞交叉污染歷史回顧1968,ApparentHeLacellcontaminationofhumanheteroploidcelllines.Nature.217:750-7511974，HeLacesscontaminatingculturesaroundtheworld,Sciences184:10591981,Contaminatedcellcultures.Nature289:2271981,cross-contaminationofcellsinculture.Sciences.212(4493):226-52研究用腫瘤細胞系的質(zhì)量控制和要求細胞交叉污染歷史回顧1999,Falsehumanhematopoieticcelllins:cross-cantaminationsandmisinterpretations.Leukemia.13(10):1601-72003,Falseleukmia–lymphomacelllines:anupdateonover500celllines.Leukemia17,416-4262004,celllinecross-comtamination:howawareare…invitrocelldevbiolanim.40(7):211-52007,Casesofmistakenidentity.Sciences315:928-9312009,NaturecallsforglobaldatabaseofcelllineSTRprofiles研究用腫瘤細胞系的質(zhì)量控制和要求保藏機構公布的錯誤細胞1細胞系種屬細胞類型污染源種屬細胞類型數(shù)據(jù)來源207人前B白血病REH人前B白血病DSMZ2474/90人胃癌HT-29人結直腸癌DSMZ2957/90人胃癌HT-29人結直腸癌DSMZ3051/80人胃癌HT-29人結直腸癌DSMZADLC-5M人肺癌HELA/-S3人宮頸癌DSMZAV3人羊膜HeLa人宮頸癌ATCCBCC-1/KMC人基底細胞癌HELA/-S3人宮頸癌DSMZBM-1640人前列腺癌DU-145人前列腺癌DSMZC16人胚胎肺成纖維細胞（MRC-5克?。〩eLa人宮頸癌ECACC研究用腫瘤細胞系的質(zhì)量控制和要求保藏機構公布的錯誤細胞2CHANGLiver人胚胎肝上皮HeLa人宮頸癌ATCC;JCRBCOLO-818人黑色素瘤COLO800人黑色素瘤DSMZDAMI人巨核細胞HEL人紅系白血病DSMZECV304人內(nèi)皮T24人膀胱癌ATCCECV-304人正常內(nèi)皮T24人膀胱癌DSMZEJ人膀胱癌T24人膀胱癌ATCC;JCRBEPLC3-2M1人肺癌HELA/-S3人宮頸癌DSMZ;JCRBEPLC-65人肺癌HELA/-S3人宮頸癌DSMZF2-4E5人胸腺上皮SK-HEP-1人肝癌DSMZF2-5B6人胸腺上皮SH-HEP-1人肝癌DSMZFL人羊膜HeLa人宮頸癌ATCCGHE人星狀細胞瘤T-24人膀胱癌DSMZGirardiHeart豬成年心臟HeLa人宮頸癌ATCC研究用腫瘤細胞系的質(zhì)量控制和要求HAG人T-24人膀胱癌DSMZHEp-2人成年喉上皮HeLa人宮頸癌ATCCHMV-1人黑色素瘤HeLa-S3人宮頸癌DSMZHuL-1HeLa人宮頸癌JCRBIMC-2人顎骨癌HeLa-S3人宮頸癌DSMZIntestine407人小腸上皮HeLa人宮頸癌ATCCJ-111HeLa人宮頸癌JCRBJOSK-I人單核細胞白血病U-937人組織細胞淋巴瘤DSMZJOSK-K人單核細胞白血病U-937人組織細胞淋巴瘤DSMZJOSK-M人單核細胞白血病U-937人組織細胞淋巴瘤DSMZ保藏機構公布的錯誤細胞3研究用腫瘤細胞系的質(zhì)量控制和要求JOSK-S人單核細胞白血病U-937人組織細胞淋巴瘤DSMZJTC-17

HeLa人宮頸癌Yamakage(JCRB)KB人HeLa人宮頸癌ATCC;JCRBKO51K562人髓系白血病DSMZ;JCRBKOSC-3Ca9-22人JCRBL132人胚胎肺上皮HeLa人宮頸癌ATCCLR10.6人前B白血病NALM-6人前B白血病DSMZMaTu人乳腺癌HELA/-S3人宮頸癌DSMZMC-4000人乳腺癌HELA/-S3人宮頸癌DSMZMKB-1人T細胞白血病CCRF-CEM人T細胞白血病DSMZ保藏機構公布的錯誤細胞4研究用腫瘤細胞系的質(zhì)量控制和要求MKN28MKN74人JCRBMOLT-15人T細胞白血病CTV-1人單核細胞白血病DSMZMT-1人乳腺癌HELA/-S3人宮頸癌DSMZNCC16PHK16-0b人JCRBP1-1A3人胸腺上皮SK-HEP-1人肝癌DSMZP1-4D6人胸腺上皮SK-HEP-1人肝癌DSMZP39TSU人HL60人髓系白血病JCRBPBEI人前B白血病NALM-6人前B白血病DSMZPSV811人成纖維WI38人成纖維JCRBRAMAK-1人肌肉滑膜T-24人膀胱癌DSMZ保藏機構公布的錯誤細胞5研究用腫瘤細胞系的質(zhì)量控制和要求SBC-2人膀胱癌HELA/-S3人宮頸癌DSMZSBC-7人膀胱癌HELA/-S3人宮頸癌DSMZSCLC-16H人SCLCSCLC-21/22H人SCLCDSMZSCLC-24H人SCLCSCLC-21/22H人SCLCDSMZSNB-19U251MG人膠質(zhì)瘤ATCCSPI-801人T細胞白血病K562人髓系白血病DSMZSPI-802人T細胞白血病K562人髓系白血病DSMZTK-1人?U251MG人膠質(zhì)瘤JCRBTMH-1IHH-4人JCRBWISH人新生羊膜上皮HeLa人宮頸癌ATCC保藏機構公布的錯誤細胞6研究用腫瘤細胞系的質(zhì)量控制和要求DSMZ德國細胞庫550leukemiaandlymphomacelllinessubmitted59/395(15%)byoriginators23 /155(15%)bysecondarysources研究用腫瘤細胞系的質(zhì)量控制和要求交叉污染細胞使用的后果Erroneousresults:WastedresearchmoneyPotentialdelayindiscovery研究用腫瘤細胞系的質(zhì)量控制和要求錯誤細胞使用2000-2004：19/Int407;45/WISH;59/Changliver;470/Hep-2;556/KB;2004(Buehring),pubmedsearch220publicationsinvolved,32%useHeLacells,33%testedforauthenticity,35%getcellsfromotherlabs.NCIpanelof60celllineswereusedforanticancerdrugevaluation.AmongthemADR-RESisacutallyOVCAR-8.Ithadbeenusedinabout300papers.研究用腫瘤細胞系的質(zhì)量控制和要求HeLacervicalcancercellsInt-407(describedas“non-transformedintestinalepithelialcells”)inBr.J.Cancer101,1596(2009),EMBOJ.22,5003(2003)andJ.Biol.Chem.280,13538(2005)WISH(describedas“non-transformedamnioticepithelialcells”)inMol.Pharmacol.69,796(2006),Endocrinology147,2490(2006)andJ.Biol.Chem.278,31731(2003)Changliver(describedas“normallivercells”)inOncogene28,3526(2009),Proteomics14,2885(2008)andJ.Biol.Chem.279,28106(2000)HEp-2(describedas“l(fā)aryngealcancer”)inInvestig.NewDrugs26,111–118(2008),Carcinogenesis29,1519(2008)andJ.Biol.Chem.283,36272(2008)KB(describedas“oralcancer”)inBiochem.Pharmacol.73,1901–1909(2007),Clin.CancerRes.14,8161(2008)andJ.Biol.Chem.280,23829(2005)HeLa,Int-407andHEp-2cellswereusedasthreedistinctcelllinesinthesamestudyinCancerRes.69,632(2009)研究用腫瘤細胞系的質(zhì)量控制和要求Ecv-304,“人臍靜脈內(nèi)皮細胞”日本實驗室建立，ATCC、DSMZ等機構有保藏，在1999—2000年，經(jīng)STR檢測，證明為T24(膀胱癌)，各自網(wǎng)站有公布。在實驗室間廣泛傳播，2006y,全世界600余論文，利用該細胞進行研究，2/3在中國。Why?Donotknow?NoothersuitablecellularmodelExpensieve,cannotafford,havetocompromise研究用腫瘤細胞系的質(zhì)量控制和要求Ecv-304/T24研究用腫瘤細胞系的質(zhì)量控制和要求對細胞質(zhì)量的要求2007.10,Goslar,Germany，11ICCCScientistfromATCCproposed:(1)fundingagencies,gov.orprivaterequirecelllineauthenticationasaconditionfortheawardofgrantandcontractsfunds.(2)keyscientificJ…asaconditionforpublication.(3)conference,workshopand/ortraining.(4)labheadsbeencouragedtoensureQCmeasures.Authentic,WellcharacterizedTheproblemofSub-culturing研究用腫瘤細胞系的質(zhì)量控制和要求NIHNOTICE2007.12,RequirecellanthenticationForgrantForpaper研究用腫瘤細胞系的質(zhì)量控制和要求雜志及細胞使用者的要求

Examples1Thp-1,2009-1-5celllineauthentication,characterizationandfreedomfromcontaminationThisisanimportantissue,towhichauthorsshouldpayparticularattention.Itistheauthors'responsibilitytoensurethatcelllinesusedinanystudy(andthemajoritydoinvolvethem)arecertifiedasbeingthedesignatedtype,thattheyhavebeencheckedtoensuretheyarefreeofcontamination(e.g.mycoplasma),andthattheyhavebeenusedfromyoungstock.Thesearethethreecriteriauponwhichgoodpracticeisbased,assetoutintheUKCCCGuidelinesontheuseofcelllines().AttentionshouldbepaidtothisintheMaterialandMethodssection,andauthorsomittingtoincludeevidenceofgoodpracticewillfindthattheirarticlesmaybedelayeduntilsuchguaranteesaregiven.研究用腫瘤細胞系的質(zhì)量控制和要求Example2,ReportingonCellLineUseinAACRJournalsNEW!EFFECTIVEJULY1,2009

AACRstronglyencouragestheauthenticationofcelllinesusedintheresearchreportedinitsjournals.Ifcelllineswereusedintheresearch,astatementaddressingthefollowingpointsmustbeincludedinthe"MaterialsandMethods"section:1.

Fromwhereandwhenthecellswereobtained2.

Whetherthecelllineshavebeentestedandauthenticated3.

Themethodbywhichthecellsweretested4.

Howandwhenthecellswerelasttested研究用腫瘤細胞系的質(zhì)量控制和要求AACRJournalsCancerResearchClinicalCancerResearchCancerEpidemiology,Biomarkers&Prevention，

CEBPCancerPreventionResearchMolecularCancerResearchMolecularCancerTherapeuticsCancerReviewsOnlinePreventionPortal研究用腫瘤細胞系的質(zhì)量控制和要求Example2—shelflifeIfcellswereobtaineddirectlyfromacellbankthatperformscelllinecharacterizationsandpassagedintheuser'slaboratoryforfewerthan6

monthsafterreceiptorresuscitation,re-authorizationisnotrequired.Inthesecases,pleaseincludethemethodofcharacterizationusedbythecellbank.Ifcellswereobtaineddirectlyfromacellbankthatperformscelllinecharacterizationsandpassagedintheuser'slaboratoryforfewerthan6

monthsafterreceiptorresuscitation,re-authorizationisnotrequired.Inthesecases,pleaseincludethemethodofcharacterizationusedbythecellbank.研究用腫瘤細胞系的質(zhì)量控制和要求KeymilestonesintheefforttoaddresscelllinemisidentificationHelacelllineestablishedfromHenriettaLacksATCC,AmericanTypeCultureCollection.NIHNationalInstitutesofHealth.STR,ShortTandemRepeatInterspeciescross-ContaminationshownATCCstartsCuratingcelllinesStanGartlershowsintraspeciesCross-contaminationbetweenHumancelllinesNelson-Rossconfirms&estendsStanGartler’sfindingsJohnMaddoxdescribesIndividuallikeNelson-RossAsself-appointedvigilantes1981-2005crossContaminationspreedsBeyongdHeLacellsNIHterminatescontractofNelson-RossNIHissuesguidelinesToavoidtheuseofmisidentifiedcelllinesRolandNardonestartstheSecondcrusadeagsinstCorsscontaminationNaturecallsforaGlobaldatabaseofCelllinesSTRprofies1952195819621966197419801981200520072009HelacelllineestablishedfromHenriettaLacksATCCstartsCuratingcelllinesHelacelllineestablishedfromHenriettaLacksATCCstartsCuratingcelllinesHelacelllineestablishedfromHenriettaLacksATCCstartsCuratingcelllinesNelson-Rossconfirms&estendsStanGartler’sfindingsHelacelllineestablishedfromHenriettaLacksATCCstartsCuratingcelllinesNelson-Rossconfirms&estendsStanGartler’sfindingsHelacelllineestablishedfromHenriettaLacksATCCstartsCuratingcelllinesNelson-Rossconfirms&estendsStanGartler’sfindingsHelacelllineestablishedfromHenriettaLacksATCCstartsCuratingcelllines1981-2005crossContaminationspreedsBeyongdHeLacellsNelson-Rossconfirms&estendsStanGartler’sfindingsHelacelllineestablishedfromHenriettaLacksATCCstartsCuratingcelllinesNIHissuesguidelinesToavoidtheuseofmisidentifiedcelllines1981-2005crossContaminationspreedsBeyongdHeLacellsNelson-Rossconfirms&estendsStanGartler’sfindingsHelacelllineestablishedfromHenriettaLacksATCCstartsCuratingcelllines1952195819621966197419801981200520072009Interspeciescross-Contaminationshown1952195819621966197419801981200520072009StanGartlershowsintraspeciesCross-contaminationbetweenHumancelllinesInterspeciescross-Contaminationshown1952195819621966197419801981200520072009StanGartlershowsintraspeciesCross-contaminationbetweenHumancelllinesInterspeciescross-Contaminationshown1952195819621966197419801981200520072009JohnMaddoxdescribesIndividuallikeNelson-RossAsself-appointedvigilantesStanGartlershowsintraspeciesCross-contaminationbetweenHumancelllinesInterspeciescross-Contaminationshown1952195819621966197419801981200520072009JohnMaddoxdescribesIndividuallikeNelson-RossAsself-appointedvigilantesStanGartlershowsintraspeciesCross-contaminationbetweenHumancelllinesInterspeciescross-Contaminationshown1952195819621966197419801981200520072009NIHterminatescontractofNelson-RossJohnMaddoxdescribesIndividuallikeNelson-RossAsself-appointedvigilantesStanGartlershowsintraspeciesCross-contaminationbetweenHumancelllinesInterspeciescross-Contaminationshown1952195819621966197419801981200520072009NIHterminatescontractofNelson-RossJohnMaddoxdescribesIndividuallikeNelson-RossAsself-appointedvigilantesStanGartlershowsintraspeciesCross-contaminationbetweenHumancelllinesInterspeciescross-Contaminationshown1952195819621966197419801981200520072009RolandNardonestartstheSecondcrusadeagsinstCorsscontaminationNIHterminatescontractofNelson-RossJohnMaddoxdescribesIndividuallikeNelson-RossAsself-appointedvigilantesStanGartlershowsintraspeciesCross-contaminationbetweenHumancelllinesInterspeciescross-Contaminationshown1952195819621966197419801981200520072009RolandNardonestartstheSecondcrusadeagsinstCorsscontaminationNIHterminatescontractofNelson-RossJohnMaddoxdescribesIndividuallikeNelson-RossAsself-appointedvigilantesStanGartlershowsintraspeciesCross-contaminationbetweenHumancelllinesInterspeciescross-Contaminationshown1952195819621966197419801981200520072009NaturecallsforaGlobaldatabaseofCelllinesSTRprofiesRolandNardonestartstheSecondcrusadeagsinstCorsscontaminationNIHterminatescontractofNelson-RossJohnMaddoxdescribesIndividuallikeNelson-RossAsself-appointedvigilantesStanGartlershowsintraspeciesCross-contaminationbetweenHumancelllinesInterspeciescross-Contaminationshown1952195819621966197419801981200520072009NIHissuesguidelinesToavoidtheuseofmisidentifiedcelllines1981-2005

crossContaminationspreedsBeyongdHeLacellsNelson-Rossconfirms&estendsStanGartler’sfindingsHelacelllineestablishedfromHenriettaLacksATCCstartsCuratingcelllinesNaturecallsforaGlobaldatabaseofCelllinesSTRprofiesRolandNardonestartstheSecondcrusadeagsinstCorsscontaminationNIHterminatescontractofNelson-RossJohnMaddoxdescribesIndividuallikeNelson-RossAsself-appointedvigilantesStanGartlershowsintraspeciesCross-contaminationbetweenHumancelllinesInterspeciescross-Contaminationshown1952195819621966197419801981