监测ABO不相容性肾移植的ABO血型抗体滴

监测ABO不相容性肾移植的ABO血型抗体滴第1页/共61页

监测ABO不相容性肾移植的

ABO血型抗体滴度

第2页/共61页BackgroundInformationJohnsHopkinsHospitalDepartmentofPathologyTransfusionMedicineDivisionSpecialistinBloodBankTechnologyAABB(AmericanAssociationofBloodBanks)ABOIncompatibleKidneyTransplantProgram

第3页/共61页背景知识约翰霍伯金斯大学医院病理系输血医学部美国血库协会血库技术专家ABO血型不相容性肾移植计划第4页/共61页CrossingtheABOBarrierforKidneyTransplantsAsofMay25,2007:75,711patientsawaitingkidneytransplantationIn2006,17,092kidneytransplantswereperformed(6,433or37.6%livingdonors): GroupO 7,662 A 6,354 B 2,255 AB 821 第5页/共61页至2007年5月25日止，

有75,711名病人在等待进行肾移植

2006年

进行了17,902个肾移植，其中6,433约37.6%是活体肾移植

血型肾移植

O7,662

A6,354

B2,255

AB821

克服ABO血型障碍进行的肾移植第6页/共61页DeathsonWaitingList

In2006,4,056patientsdiedwaitingforakidneytransplant,or11.1patientsdiedperday

2,049GroupO1,250GroupA629GroupB128GroupAB第7页/共61页在等待中死去的病人

2006年，共有4,056名等待肾移植的病人死亡，即每天有11.1名病人在等待中死去。第8页/共61页CadavericKidneyTransplantWaitAsof5/25/07Registered5orMoreYearsAllABO75,7118,079O39,7134,459A21,4571,844B12,3791,592AB2,162184第9页/共61页等待尸肾移植的病人数及等待时间至07年5月25日登记等待5年或5年以上所有血型 75,711 8,079O

39,7134,459A 21,4571,844B 12,3791,592AB2,162184第10页/共61页CadavericKidneyTransplantWaitIfregisteredin2001-2002,medianwaittimesasofMay25,2007

BloodGroupDaysO1836A1135B2032AB732第11页/共61页2001-2002年间登记的病人，到2007年5月25日平均等待时间是血型天数O1836A1135B2032AB732第12页/共61页ObjectivesExpanddonorpoolandavailabilityoforgansfortransplantationDecreasetimeonorganwaitinglistCircumventhyperacuterejectionand/orAMR(duetoABOand/orHLAantibodies)bydevelopingimmunomodulatoryprotocolsABOincompatibleDonor-recipientwithpositivecrossmatches(HLA)PosttransplantpatientswhodevelopAMR(HLA)第13页/共61页目的扩大器官捐献群体及移植器官的来源减少器官移植的等待时间通过免疫调节方案，防止超急性排斥反应及/或因ABO血型或HLA抗体介导的排斥反应ABO不相容性供-受者交配反应阳性（HLA）病人移植后出现抗体介导的排斥反应(HLA)第14页/共61页ABOIncompatibleTransplantationBasedonJapaneseexperienceBasedonourexperience/protocolforcrossmatchpositivedonor/recipientpairsandantibody-mediatedrejectionFacilitatedbylaparoscopicdonornephrectomyPreparativeregimenwithintensivefollowuppre/postoperativeplasmapheresisIVIgGandimmunosuppression第15页/共61页ABO不相容的移植基于日本的经验基于我们对交配阳性的供-受者和抗体介导的排斥反应的经验/方案腹腔镜捐献者取肾促进了该类移植强化随访的予处理方案术前/后血浆置换静脉注射IgG和免疫抑制第16页/共61页Reference第17页/共61页ReferenceABO不相容性肾移植的长期结果第18页/共61页第19页/共61页ABO不相容性病人在亲属活体肾移植后抗-A、B的滴度变化第20页/共61页OurProgram-InkTpSpecializedIncompatibleKidneyTransplantserviceandclinicWeeklyinterdisciplinaryrounds,composedofbloodbank,apheresisservice,HLAlab,nephrology,transplantsurgery,pharmacist,psychologist,andsocialworker

第21页/共61页我们的不相容肾移植项目专业化的服务和临床每周跨科的综合查房，包括血库、单采、HLA实验室、肾科、移植外科、药剂师、心理学家和社会工作者第22页/共61页OurProtocolModifiedfromtheJapaneseprotocolInitialevaluationdonorandrecipientmedical,psychologicalandsocialworkevaluationsHLAandABOtypingandtitersReviewatweeklymeetingdeterminationofoptimaldonor(HLAvsABO)treatmentplanestablished第23页/共61页我们的方案根据日本方案改进初步评价捐献者和受者进行医学、心理和社会服务的评估

HLA和ABO分型及抗体滴度分析每周会议的评述根据HLA与ABO相配情况决定最佳捐献者建立治疗方案第24页/共61页TreatmentPlanImmunosuppression,mayincludesplenectomyAllpatientsvaccinated(pneumococcus,meningococcus,haemophilusinfluenzae),regardlessofsplenectomystatusPheresisplanestablished(numberofprocedurespreandpostop)第25页/共61页治疗方案

免疫抑制，可能包括脾切除不管是否脾切除，所有病人接种疫苗（肺炎球菌、脑膜炎球菌、流感嗜血杆菌）建立置换计划（术前/术后的置换程序和次数）第26页/共61页PlasmapheresisRemovesnaturalanti-Aand/oranti-BantibodisBothpreopandpostopproceduresRemove1plasmavolumeReplaceat100%ofvolumeremovedReplacewith5%albuminexceptproceduresimmediatelypre-andpost-surgery,replacewithplasma第27页/共61页血浆置换去除天然的抗-A/抗-B抗体术前/后都进行置换去除一个血浆容量等量替换用5%的白蛋白置换除术前/后立即的置换外，只用血浆置换第28页/共61页ABOAntibodyTiterGoalsPretransplantgoal:

titer16orlessPosttransplantContinuetomonitorABOantibodytitersSignificanceinpredictionofantibodymediatedrejectionisuncertain第29页/共61页ABO抗体滴度目标移植前目标：滴度≤16移植后继续监测ABO抗体滴度对于预测抗体介导的排斥反应的意义仍不确定第30页/共61页PlasmapheresisProceduresABOAntibodyTiter(AHG)PretransplantTreatments(#)PosttransplantTreatments(#)<16221622323364431285-632567-845129-104102410-124>1024>155第31页/共61页血浆置换过程ABO抗体滴度移植前治疗次数移植后治疗次数<16221622323364431285-632567-845129-104102410-124>1024>155第32页/共61页ABOAntibodyTitersConventionaltesttubemethodAABBtechnicalmanual

15thed.Bethesda:AmericanAssociationofBloodBanks,2005.Serialdilutionsofeachsamplewerepreparedin0.9%salinePooledindicatorcells(Immucor,Norcross,GA)oftheappropriateABOtypewereadded30minutes22Cincubationtestphase30minutes37CincubationtestphaseAHGtestphaseusingmonospecificanti-IgG(Immucor,Norcross,GA)AgglutinationwasscoredusingtheMarsh0to12scoringsystemAHGtiterendpointwasthereciprocalofthehighestdilutiondemonstratingmacropscopic(score3)agglutination

第33页/共61页ABO抗体滴定常规的试管法AABB技术手册样品用0.9%的盐水进行连续稀释加入适量的混合ABO血型指示细胞（Immucor，Norcross,GA）22度孵育30分钟37度孵育30分钟单特异性抗-IgG(Immucor,Norcross,GA)的抗人球蛋白（AHG）实验阶段用Marsh0到12打分系统给凝集打分AHG最终滴度是样品出现肉眼可见的（3分）凝集的最高稀释倍数第34页/共61页ABOAntibodyTitersItistimeconsumingTurnaroundtimeiscriticalOurexperienceindicatesthatonlytheAHGtitervaluesarecriticalTiterendpointisdemonstratedwithamacroscopicagglutination第35页/共61页ABO抗体滴度这是一个花费时间的实验关键是出现结果的时间我们的经验表明只有AHG滴度是关键值最终滴度是由肉眼可见的凝集决定第36页/共61页ObjectivesThegoalofthepresentstudywastostreamlineABOantibodytiterdeterminationsbyassessingAHGtitersusing

1)Arevisedtesttubemethodwheretheroomtemperatureincubationphasewaseliminated2)Ananti-IgGgelmicrocolumnmethod第37页/共61页目的本研究的目的是通过评估AHG滴度，系列测定ABO抗体

1)应用不需要室温孵育过程的改良试管法

2)应用抗-IgG凝胶微柱法第38页/共61页Materialandmethods

RevisedtesttubetitrationmethodSerialdilutionsofeachsamplewerepreparedin0.9%salinePooledindicatorcells(Immucor,Norcross,GA)oftheappropriateABOtypewereadded30minutesroomtemperaturewasomittedTiterswereincubatedfor30minutes37CandthenconvertedtotheAHGtestphaseusingmonospecificanti-IgG(Immucor,Norcross,GA)AgglutinationwasscoredusingtheMarsh0to12scoringsystemAHGtiterendpointwasthereciprocalofthehighestdilutiondemonstratingmacropscopic(score3)agglutination第39页/共61页实验材料和方法

改良的试管法每个样品用0.9%的盐水进行系列稀释加入适当的ABO血型的混合指示细胞（Immucor，Norcross,GA）省略室温30分钟孵育37度孵育30分钟后用单特异性抗-IgG进行AHG实验用Marsh打分系统对凝集进行打分AHG最终滴度是样品出现肉眼可见的（3分）凝集的最高稀释度的倒数第40页/共61页MaterialandmethodsAnti-IgGmicrocolumngelmethodSerialdilutionsofeachsamplewerepreparedTwenty-fivemicrolitersofeachplasmadilutionand50microlitersof0.8%indicatorredcellspreparedinMTS™Diluent2wereaddedtothegelcardmicrocolumnsAfterincubationat37Cfor15minutes,thegelcardswerecentrifugedfor10minutesThetiterendpointwasthereciprocalofthehighestdilutiondemonstrating1+第41页/共61页实验材料和方法

抗-IgG微柱凝胶法每个样品系列稀释在微柱凝胶卡中加入25微升每一稀释度的血浆和50微升应用MTS™稀释液2制备的0.8%的指示红细胞。37℃孵育15分钟后将凝胶卡离心10分钟最终滴度是显示1+凝集的最高稀释度的倒数第42页/共61页Anti-IgGMicrocolumnGelCards

MicroTypingSystems,MTS™,OrthoClinicalDiagnostics,Raritan,NJ,USA第43页/共61页抗-IgG微柱凝胶卡片MicroTypingSystems,MTS™,OrthoClinicalDiagnostics,Raritan,NJ,USA

微型血型仪第44页/共61页PatientsamplesSerialEDTAplasmasamplesfrompatientsinourABOINKTprogramFiftysampleswithanti-Aand/orAnti-BantibodiesAHGtitersrangingfrom2to51218ofthe50(36%)sampleswereselectedwithtitersof16第45页/共61页病人样品我们ABO不相容的肾移植项目的病人EDTA血浆标本50个有抗-A/抗-B抗体的标本AHG滴度在2-512之间50个中有18个样本滴度为16第46页/共61页Patientsamples

NumberofSamplesTitersAnti-AAnti-BAnti-A/Anti-BTotal2-8622101612511832-649312128-25672951211Total3413350第47页/共61页病人样本样本数滴度抗-A抗-B抗-A/抗-B总计2-8622101612511832-649312128-25672951211总计3413350第48页/共61页Results第49页/共61页结果第50页/共61页Results(table1)Comparisonofantiglobulin(AHG)titerresultsbytesttubemethodswithandwithoutaroomtemperature(RT)incubationphaseConventionaltesttubemethodRevisedTesttubemethod(RTT)N=IdenticaltoCTTAHGtiter>CTTAHGtiter<CTTAHGtiter211433864216181323327412645321286321256311151211Total5032(64%)7(14%)11(22%)第51页/共61页结果（表1）有、无室温孵育的试管法抗人球（AHG）滴度实验结果的比较(CTT)传统试管法改良的试管法N=与传统试管法滴度相等>传统AHG滴度<传统AHG滴度的211433864216181323327412645321286321256311151211总计5032(64%)7(14%)11(22%)第52页/共61页Results(table2)ComparisonofconventionaltesttubeAHGtitersthatincludedaroomtemperaturetestphasetothetitersperformedwithanti-IgGgelcards(IgGGEL)ConventionaltesttubemethodIgGGelTesttubemethodN=IdenticaltoCTTAHGtiter>CTTAHGtiter<CTTAHGtiter211432186421618133232723264532128612325632151211Total5026(52%)12(24%)12(24%)第53页/共61页结果（表2）有室温孵育的传统试管法与抗-IgG凝胶卡法AHG滴度的比较(CTT)传统试管法IgG凝胶卡法N=与传统试管法滴度相等>传统AHG滴度<传统AHG滴度211432186421618133232723264532128612325632151211总计5026(52%)12(24%)12(24%)第54页/共61页Results(table3)ComparisonofrevisedtesttubeAHGtiterresultswiththeIgGGELtitersRevisedtesttubemethod(RTT)IgGGelTesttubemethodN=IdenticaltoRTTAHGtiter>RTTAHGtiter<RTTAHGtiter2114321866161816232761645512864225632151211Total5043(86%)5(10%)2(4%)第55页/共61页结果（表3）改良试管法和IgG凝胶法AHG滴度的比较改良的试管法IgG凝胶试管法N=与改良的试管法滴度相等>改良试管法滴度<改良试管法滴度2114321866161816232761645512864225632151