毛细管电泳对异常球蛋白的检测.ppt

InterestofserumproteincapillaryelectrophoresisandimmunotypingforthediagnosisofpathologicalconditionsandgammopathiesELPpresentationforJoinscienceShanghaiwithChineseByGenevieveHENNACHEDirectorofSEBIADiagnosticDepartment,毛细管电泳对异常球蛋白的检测,Theelectrophoresisassayiseasytooperate,notveryexpensive,andcanbeperformedroutinelyinmostpathologylaboratoriestoseparate,quantifyoridentifyproteinsindifferentfluids(serum,urine,CSF)电泳方法简便，廉价。在各级实验室都可实现对不同的体液蛋白的分析Electrophoresisisamultipurposetestthatgivesessentialinformationforthediagnosisofvariouspathologies(cirrhosis,nephroticsyndrome,multiplesclerosis,hemoglobinopathies,MC)电泳是一种多目的检测方法，可对多种疾病提供重要的信息（肝硬化，多发性硬化症.）Thereiscurrentlynoalternateassayfordeterminingtheimmunoglobulinmonoclonality,evidenceofabonemyeloma目前对于单克隆免疫球蛋白的检测是唯一选择,Capillaryelectrophoresis,Agarosegelelectrophoresis,ProteinElectrophoresis（蛋白电泳技术）,CAPILLARYS,8capillaryfullyautomatedsystem,Continuoussampleloading(upto13racks)Upto78samples/hourProtein6assay,Upto53samples/hourUrineassay,Upto8samples/hourITorUrineITassay,Upto33samples/hourHbassay,Upto38samples/hourCDTassay,Upto53samples/hourHRassay,Upto24samples/hourNeonatassay,Upto48samples/hourFastNeonatassay,ManagedbyPhoresis.,CapillarysSpecifications毛细管电泳参数,MiniCapSpecifications两管毛细管参数,Twocapillaryfullyautomatedsystem,28positionsonacarouselforcontinuoussampleloadingUpto19samples/hourProtein6assay,Upto12samples/hourUrineassayUpto2samples/hourITorUrineITassay,Upto9samples/hourHbassay,Upto10samples/hourCDTassayManagedbyPhoresis.,MINICAPOperation操作,VerysimpletoOperatePlacepatienttubesonthecarouselClosethedoorMINICAPwilllocatesamplesSearchforabarcodeifpresentPerformautomaticallythedilutionandtheanalysis,7,AdvantagesofCapillaryTechnology毛细管电泳技术优势(handson,workflow),AdvantagesofCapillaryTechnology毛细管电泳技术优势(results)（电泳结果）,Useofcapillaryelectrophoresisfortheanalysisofserumproteinelectrophoresis毛细管电泳血清蛋白分析应用,Serumproteinelectrophoresis(SPE)istheonlyproceduretoconfirmanimmunoglobulinmonoclonalityinthepatientswithasuspectedbonemyeloma对于疑似骨髓瘤患者，血清蛋白电泳是唯一能确认单克隆免疫球蛋白的手段,Capillaryselectrophoresisallowstoseparateandquantifyserumproteinsin6fractions毛细管电泳对血清蛋白分成6组份,a-1antitrypsin,Haptoglobin,Transferrin,a-1acidglycoprotein,a-2macroglobulin,Hemopexin,Albumin,Gamma,Beta-2,Beta-1,Alpha-2,Alpha-1,C3complement,Gammaglobulins,SEBIACAPILLARYSPROTEIN,Confirmapathologicaldiagnosisinassociationwithotherparameters:inflammation,hepatitis,cirrhoticornephroticprofile和其他检测方法配合，可确定相关疾病：炎症，肝炎，肝硬化或肾病Identifythepresenceofamonoclonalimmunoglobulinwhenabonemyelomaissuspectedwithclinicalsigns当出现临床疑似骨髓瘤症状，可对其辨认是否出现单克隆免疫球蛋白Allowanearlydetectionofasymptomaticmyeloma对无症状的骨髓瘤进行早期检测Tomonitortheevolutionofmalignantdiseaseorfollowtheefficiencyofimmunosuppressivetreatment,justbythepeakquantification通过简单的单克隆峰定量，对此恶性疾病的发展或免疫抑制治疗进行观察,Variousquestionscanbeansweredwithaserumproteinelectrophoresis血清蛋白电泳可解决不少问题,Gaussianaspectingamma&Noincreaseoradditionaldeformation/peakingamma,beta1,beta2andalpha2,NormalSerum正常血清,A1ATORO,HPT,InflammatoryPattern炎症,Alb,a1,a2,b1,b2,g,A1ATORO,HPT,IncreaseofpolyclonalIgAinbeta2andGingamma,InflammatoryPattern炎症,Totalproteinconcentrationisdecreased(under60g/lor6g/dl),a2macroglobulin,lipoproteins,Hypogamma,Hypoalbuminemia,NephroticPattern肾病型,PolyclonalincreaseofIgAinbeta2andIgGingamma,BetaGammaBridge-Gamma桥,Causes:immunodeficiency,immunosuppressivetreatment,freelightchainsdisease,IfnoabnormalityispresentonserumELPandthecausesofhypogammaareunknown,aBJsearchintheurineisrecommended,Hypogammaglobulinemia低免疫球蛋白,Symptomaticgammopathies:有症状免疫球蛋白疾病MultipleMyeloma(MM)WaldenstrmdiseaseNonHodgkinsLymphomaChronicLymphodLeukemiaHeavychaindiseaseAmyloidosisAsymptomaticgammopathies:无症状的免疫球蛋白疾病Smolderingmyeloma(SM)MonoclonalGammopathiesofUndeterminedSignificance(MGUS),Gammopathiesarecharacterizedbythepresenceofamonoclonalimmunoglobulininserumproteinelectrophoresis出现单克隆特征的免疫球蛋白病,Normalserum,SerumwithMonoclonalIg,DiscoveryofamonoclonalimmunoglobulinbyserumproteinelectrophoresisusingCapillarys通过毛细管血清蛋白电泳发现单克隆,PositionofamonoclonalIg单克隆的电泳位置,Mainlyinthegammazone主要在GAMMA区Lessfrequentlyinanyotherzones(beta1,beta2andalpha)少见于在其他区带,ITorIFrequiredtocharacterizethemonoclonalIginserum(BJsearchintheurine),StrongMonoclonalPeakinGammaGAMMA出现高单克隆峰,ITorIFrequiredtocharacterizethemonoclonalIginserum(BJsearchintheurine),WeakMonoclonalPeakinGammaGAMMA出现弱单克隆峰,ITorIFrequiredtocharacterizethemonoclonalIginserum(BJsearchintheurine),MonoclonalIginbeta1beta1区出现单克隆峰,0.18g/L,Between0.20.6g/Laccordingtothemobilityandthepolyclonalbackground,LimitofsensitivityforthedetectionofmonoclonalIg,单克隆蛋白的检测灵敏度,Zoom+smoothing0,HighresolutionofCapillarys/MinicaptodetectaweakmonoclonalIg毛细管电泳能对弱单克隆峰高分辨检测,Quantificationofmonoclonalpeak单克隆峰定量,Todaythespikeconcentrationisusedtofollowupthedisease目前检测单克隆浓度被用于观察疾病发展,PeakQuantification单克隆峰定量,ITorIFisalwaysrequiredtoconfirmanoligoclonalprofile,OligoclonalProfileinGammaGamma区出现寡克隆,CAPILLARYSHYDRAGEL+HYRYS,HighresolutionofCapillarys/Minicapallowsaneasydetectionofoligoclonalprofile相对于琼脂糖电泳，高分辨的毛细管电泳对寡克隆更易检测,OligoclonalProfileinGamma,Inconclusion,总结,Everyzoneonelectrophoresisshouldbeanalyzedforqualitativeand/orquantitativeabnormalities应对电泳各区带出现定性或定量异常的情况进行分析*Increasedzones区带上升*Decreasedzones区带下降*Presenceofunusualfractionorunusualshapeofafraction(distortionofonefraction)出现非常见组份或条带形态异常,Informationstoconsiderfortheinterpretation结果解读应考虑的因素*Haveagloballookatthecompletepattern整体检测情况*Lookatthecolorandaspectoftheserum(hemolyzed,icteric,turbidorprecipitatedserum(cryoglobulin)观察血清颜色和外观*Totalproteinconcentration蛋白总浓度*Lookattheageofthepatient.Anypatientover45yearsoldmaypotentiallyhaveaMC患者的年龄。超过45岁，有出现单克隆潜在危险*Theclinicalinformation,ifpossible:bonepains,lossofweight,anemia,increasedESR,hypercalcemia.是否有相关临床症状，如骨痛，贫血，血沉加快，高钙血*Measureandfollowthepeakconcentration测定和跟踪单克隆峰浓度,Whenaclearpeakoraslightdistortionisvisibleinthegammazone.Gamma区中出现明显的峰或区带变形Whenanadditionalpeakisvisibleinthealpha2orbetazones(innonhemolysedsamples)在2和区有增加峰出现Whenthebetavalueisincreasedalonewithoutanyassociatedincreasesofalpha1andalpha2zones值单独上升，12区无相应变化,Iftheseabnormalitiesareobservedontheserumproteinelectrophoresis:血清电泳中出现以下异常情况：,Acomplementaryimmunotyping(IT)isrequired需要进行免疫分型实验,CAPILLARYS2,IT:8sample/hr,MINICAP,IT:2sample/hr,CharacterizationofamonoclonalIgbyimmunotyping(IT)用IT对单克隆定性分析,RequestanImmunotypinginProtein6血清模式下IT分析请求,ImmunotypingonCapillarys毛细管上免疫分型,ELP,G,A,M,k,l,DILUENT,*6barcodedVialsofsamplediluant*1barcodedELPvial*Barcodedantiserum(G,A,M,K,L)vials,ImmunotypingonMinicap两管毛细管免疫分型,Animmunecomplexisformedwhichmigratesinanodicposition:moreanodicthanalbuminorbetweenalbuminandalpha1fractionThemonoclonalpeakisidentifiedwhenthepeakdisappearswithagivenantibodyagainstanheavychain(G,AorM)andwithanantibodyagainstkappaorlambdalightchain,Immunotyping:Interpretation免疫分型结果判断,Referencecurve,Immunosubtractionwithanti-IgGantiserum,Immunosubtractionwithanti-IgAantiserum,Immunosubtractionwithanti-IgMantiserum,Immunosubtractionwithanti-kappaantiserum,Immunosubtractionwithanti-lambdaantiserum,AllAg-Accomplexesmigrateinthezonealbumin-1,Interpretation解读,IgGIgAIgMKappaLambda,Interpretation解读,80%IgG,15%IgA,5%IgM,2/3Kappa,1/3Lambda,ELP,G,A,M,K,L,MonoclonalpeakinGamma,ITrequired,Patient1:SerumProteinElectrophoresis,Zoom,Patient1:Immunotyping,IgGLambda,MonoclonaldistorsioninGamma,ITrequired,Patient2:SerumProteinElectrophoresis,Zoom,Patient2:Immunotyping,IgMKappa,IsolatedincreaseinBeta2,ITrequired,Patient3:SerumProteinElectrophoresis,Zoom,Patient3:Immunotyping,IgAKappa,Inconclusion,总结,SerumproteinelectrophoresisandimmunotypingusingSEBIAtechnologies赛比亚血清电泳技术和免疫分型分析,SEPcanallowtoconfirmapathologicaldiagnosis(inflammation,cirrhoticornephroticprofile,MCdetection)血清电泳可对一些疾病的诊断进一步确定SEP+ITcanallowtoidentify,quantifyandtypethemonoclonalimmunoglobulinassociatedwithasymptomaticbonemyelomaforitsstaginganditsfollow-upduringanimmunosuppressivetreatment血清电泳+IT可对单克隆辨认及定量，并可用于有症状的骨髓瘤分级和免疫抑制治疗的跟踪观察SEP+ITcanallowtheearlydetectionofasymptomaticmyeloma(SM