監(jiān)測ABO不相容性腎移植的ABO血型抗體滴

監(jiān)測ABO不相容性腎移植的ABO血型抗體滴第1頁/共61頁

監(jiān)測ABO不相容性腎移植的

ABO血型抗體滴度

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

第3頁/共61頁背景知識約翰霍伯金斯大學(xué)醫(yī)院病理系輸血醫(yī)學(xué)部美國血庫協(xié)會血庫技術(shù)專家ABO血型不相容性腎移植計劃第4頁/共61頁CrossingtheABOBarrierforKidneyTransplantsAsofMay25,2007:75,711patientsawaitingkidneytransplantationIn2006,17,092kidneytransplantswereperformed(6,433or37.6%livingdonors): GroupO 7,662 A 6,354 B 2,255 AB 821 第5頁/共61頁至2007年5月25日止，

有75,711名病人在等待進行腎移植

2006年

進行了17,902個腎移植，其中6,433約37.6%是活體腎移植

血型腎移植

O7,662

A6,354

B2,255

AB821

克服ABO血型障礙進行的腎移植第6頁/共61頁DeathsonWaitingList

In2006,4,056patientsdiedwaitingforakidneytransplant,or11.1patientsdiedperday

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

2006年，共有4,056名等待腎移植的病人死亡，即每天有11.1名病人在等待中死去。第8頁/共61頁CadavericKidneyTransplantWaitAsof5/25/07Registered5orMoreYearsAllABO75,7118,079O39,7134,459A21,4571,844B12,3791,592AB2,162184第9頁/共61頁等待尸腎移植的病人數(shù)及等待時間至07年5月25日登記等待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頁/共61頁CadavericKidneyTransplantWaitIfregisteredin2001-2002,medianwaittimesasofMay25,2007

BloodGroupDaysO1836A1135B2032AB732第11頁/共61頁2001-2002年間登記的病人，到2007年5月25日平均等待時間是血型天數(shù)O1836A1135B2032AB732第12頁/共61頁ObjectivesExpanddonorpoolandavailabilityoforgansfortransplantationDecreasetimeonorganwaitinglistCircumventhyperacuterejectionand/orAMR(duetoABOand/orHLAantibodies)bydevelopingimmunomodulatoryprotocolsABOincompatibleDonor-recipientwithpositivecrossmatches(HLA)PosttransplantpatientswhodevelopAMR(HLA)第13頁/共61頁目的擴大器官捐獻群體及移植器官的來源減少器官移植的等待時間通過免疫調(diào)節(jié)方案，防止超急性排斥反應(yīng)及/或因ABO血型或HLA抗體介導(dǎo)的排斥反應(yīng)ABO不相容性供-受者交配反應(yīng)陽性（HLA）病人移植后出現(xiàn)抗體介導(dǎo)的排斥反應(yīng)(HLA)第14頁/共61頁ABOIncompatibleTransplantationBasedonJapaneseexperienceBasedonourexperience/protocolforcrossmatchpositivedonor/recipientpairsandantibody-mediatedrejectionFacilitatedbylaparoscopicdonornephrectomyPreparativeregimenwithintensivefollowuppre/postoperativeplasmapheresisIVIgGandimmunosuppression第15頁/共61頁ABO不相容的移植基于日本的經(jīng)驗基于我們對交配陽性的供-受者和抗體介導(dǎo)的排斥反應(yīng)的經(jīng)驗/方案腹腔鏡捐獻者取腎促進了該類移植強化隨訪的予處理方案術(shù)前/后血漿置換靜脈注射IgG和免疫抑制第16頁/共61頁Reference第17頁/共61頁ReferenceABO不相容性腎移植的長期結(jié)果第18頁/共61頁第19頁/共61頁ABO不相容性病人在親屬活體腎移植后抗-A、B的滴度變化第20頁/共61頁OurProgram-InkTpSpecializedIncompatibleKidneyTransplantserviceandclinicWeeklyinterdisciplinaryrounds,composedofbloodbank,apheresisservice,HLAlab,nephrology,transplantsurgery,pharmacist,psychologist,andsocialworker

第21頁/共61頁我們的不相容腎移植項目專業(yè)化的服務(wù)和臨床每周跨科的綜合查房，包括血庫、單采、HLA實驗室、腎科、移植外科、藥劑師、心理學(xué)家和社會工作者第22頁/共61頁OurProtocolModifiedfromtheJapaneseprotocolInitialevaluationdonorandrecipientmedical,psychologicalandsocialworkevaluationsHLAandABOtypingandtitersReviewatweeklymeetingdeterminationofoptimaldonor(HLAvsABO)treatmentplanestablished第23頁/共61頁我們的方案根據(jù)日本方案改進初步評價捐獻者和受者進行醫(yī)學(xué)、心理和社會服務(wù)的評估

HLA和ABO分型及抗體滴度分析每周會議的評述根據(jù)HLA與ABO相配情況決定最佳捐獻者建立治療方案第24頁/共61頁TreatmentPlanImmunosuppression,mayincludesplenectomyAllpatientsvaccinated(pneumococcus,meningococcus,haemophilusinfluenzae),regardlessofsplenectomystatusPheresisplanestablished(numberofprocedurespreandpostop)第25頁/共61頁治療方案

免疫抑制，可能包括脾切除不管是否脾切除，所有病人接種疫苗（肺炎球菌、腦膜炎球菌、流感嗜血桿菌）建立置換計劃（術(shù)前/術(shù)后的置換程序和次數(shù)）第26頁/共61頁PlasmapheresisRemovesnaturalanti-Aand/oranti-BantibodisBothpreopandpostopproceduresRemove1plasmavolumeReplaceat100%ofvolumeremovedReplacewith5%albuminexceptproceduresimmediatelypre-andpost-surgery,replacewithplasma第27頁/共61頁血漿置換去除天然的抗-A/抗-B抗體術(shù)前/后都進行置換去除一個血漿容量等量替換用5%的白蛋白置換除術(shù)前/后立即的置換外，只用血漿置換第28頁/共61頁ABOAntibodyTiterGoalsPretransplantgoal:

titer16orlessPosttransplantContinuetomonitorABOantibodytitersSignificanceinpredictionofantibodymediatedrejectionisuncertain第29頁/共61頁ABO抗體滴度目標移植前目標：滴度≤16移植后繼續(xù)監(jiān)測ABO抗體滴度對于預(yù)測抗體介導(dǎo)的排斥反應(yīng)的意義仍不確定第30頁/共61頁PlasmapheresisProceduresABOAntibodyTiter(AHG)PretransplantTreatments(#)PosttransplantTreatments(#)<16221622323364431285-632567-845129-104102410-124>1024>155第31頁/共61頁血漿置換過程ABO抗體滴度移植前治療次數(shù)移植后治療次數(shù)<16221622323364431285-632567-845129-104102410-124>1024>155第32頁/共61頁ABOAntibodyTitersConventionaltesttubemethodAABBtechnicalmanual

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

第33頁/共61頁ABO抗體滴定常規(guī)的試管法AABB技術(shù)手冊樣品用0.9%的鹽水進行連續(xù)稀釋加入適量的混合ABO血型指示細胞（Immucor，Norcross,GA）22度孵育30分鐘37度孵育30分鐘單特異性抗-IgG(Immucor,Norcross,GA)的抗人球蛋白（AHG）實驗階段用Marsh0到12打分系統(tǒng)給凝集打分AHG最終滴度是樣品出現(xiàn)肉眼可見的（3分）凝集的最高稀釋倍數(shù)第34頁/共61頁ABOAntibodyTitersItistimeconsumingTurnaroundtimeiscriticalOurexperienceindicatesthatonlytheAHGtitervaluesarecriticalTiterendpointisdemonstratedwithamacroscopicagglutination第35頁/共61頁ABO抗體滴度這是一個花費時間的實驗關(guān)鍵是出現(xiàn)結(jié)果的時間我們的經(jīng)驗表明只有AHG滴度是關(guān)鍵值最終滴度是由肉眼可見的凝集決定第36頁/共61頁ObjectivesThegoalofthepresentstudywastostreamlineABOantibodytiterdeterminationsbyassessingAHGtitersusing

1)Arevisedtesttubemethodwheretheroomtemperatureincubationphasewaseliminated2)Ananti-IgGgelmicrocolumnmethod第37頁/共61頁目的本研究的目的是通過評估AHG滴度，系列測定ABO抗體

1)應(yīng)用不需要室溫孵育過程的改良試管法

2)應(yīng)用抗-IgG凝膠微柱法第38頁/共61頁Materialandmethods

RevisedtesttubetitrationmethodSerialdilutionsofeachsamplewerepreparedin0.9%salinePooledindicatorcells(Immucor,Norcross,GA)oftheappropriateABOtypewereadded30minutesroomtemperaturewasomittedTiterswereincubatedfor30minutes37CandthenconvertedtotheAHGtestphaseusingmonospecificanti-IgG(Immucor,Norcross,GA)AgglutinationwasscoredusingtheMarsh0to12scoringsystemAHGtiterendpointwasthereciprocalofthehighestdilutiondemonstratingmacropscopic(score3)agglutination第39頁/共61頁實驗材料和方法

改良的試管法每個樣品用0.9%的鹽水進行系列稀釋加入適當?shù)腁BO血型的混合指示細胞（Immucor，Norcross,GA）省略室溫30分鐘孵育37度孵育30分鐘后用單特異性抗-IgG進行AHG實驗用Marsh打分系統(tǒng)對凝集進行打分AHG最終滴度是樣品出現(xiàn)肉眼可見的（3分）凝集的最高稀釋度的倒數(shù)第40頁/共61頁MaterialandmethodsAnti-IgGmicrocolumngelmethodSerialdilutionsofeachsamplewerepreparedTwenty-fivemicrolitersofeachplasmadilutionand50microlitersof0.8%indicatorredcellspreparedinMTS?Diluent2wereaddedtothegelcardmicrocolumnsAfterincubationat37Cfor15minutes,thegelcardswerecentrifugedfor10minutesThetiterendpointwasthereciprocalofthehighestdilutiondemonstrating1+第41頁/共61頁實驗材料和方法

抗-IgG微柱凝膠法每個樣品系列稀釋在微柱凝膠卡中加入25微升每一稀釋度的血漿和50微升應(yīng)用MTS?稀釋液2制備的0.8%的指示紅細胞。37℃孵育15分鐘后將凝膠卡離心10分鐘最終滴度是顯示1+凝集的最高稀釋度的倒數(shù)第42頁/共61頁Anti-IgGMicrocolumnGelCards

MicroTypingSystems,MTS?,OrthoClinicalDiagnostics,Raritan,NJ,USA第43頁/共61頁抗-IgG微柱凝膠卡片MicroTypingSystems,MTS?,OrthoClinicalDiagnostics,Raritan,NJ,USA

微型血型儀第44頁/共61頁PatientsamplesSerialEDTAplasmasamplesfrompatientsinourABOINKTprogramFiftysampleswithanti-Aand/orAnti-BantibodiesAHGtitersrangingfrom2to51218ofthe50(36%)sampleswereselectedwithtitersof16第45頁/共61頁病人樣品我們ABO不相容的腎移植項目的病人EDTA血漿標本50個有抗-A/抗-B抗體的標本AHG滴度在2-512之間50個中有18個樣本滴度為16第46頁/共61頁Patientsamples

NumberofSamplesTitersAnti-AAnti-BAnti-A/Anti-BTotal2-8622101612511832-649312128-25672951211Total3413350第47頁/共61頁病人樣本樣本數(shù)滴度抗-A抗-B抗-A/抗-B總計2-8622101612511832-649312128-25672951211總計3413350第48頁/共61頁Results第49頁/共61頁結(jié)果第50頁/共61頁Results(table1)Comparisonofantiglobulin(AHG)titerresultsbytesttubemethodswithandwithoutaroomtemperature(RT)incubationphaseConventionaltesttubemethodRevisedTesttubemethod(RTT)N=IdenticaltoCTTAHGtiter>CTTAHGtiter<CTTAHGtiter211433864216181323327412645321286321256311151211Total5032(64%)7(14%)11(22%)第51頁/共61頁結(jié)果（表1）有、無室溫孵育的試管法抗人球（AHG）滴度實驗結(jié)果的比較(CTT)傳統(tǒng)試管法改良的試管法N=與傳統(tǒng)試管法滴度相等>傳統(tǒng)AHG滴度<傳統(tǒng)AHG滴度的211433864216181323327412645321286321256311151211總計5032(64%)7(14%)11(22%)第52頁/共61頁Results(table2)ComparisonofconventionaltesttubeAHGtitersthatincludedaroomtemperaturetestphasetothetitersperformedwithanti-IgGgelcards(IgGGEL)ConventionaltesttubemethodIgGGelTesttubemethodN=IdenticaltoCTTAHGtiter>CTTAHGtiter<CTTAHGtiter211432186421618133232723264532128612325632151211Total5026(52%)12(24%)12(24%)第53頁/共61頁結(jié)果（表2）有室溫孵育的傳統(tǒng)試管法與抗-IgG凝膠卡法AHG滴度的比較(CTT)傳統(tǒng)試管法IgG凝膠卡法N=與傳統(tǒng)試管法滴度相等>傳統(tǒng)AHG滴度<傳統(tǒng)AHG滴度211432186421618133232723264532128612325632151211總計5026(52%)12(24%)12(24%)第54頁/共61頁Results(table3)ComparisonofrevisedtesttubeAHGtiterresultswiththeIgGGELtitersRevisedtesttubemethod(RTT)IgGGelTesttubemethodN=IdenticaltoRTTAHGtiter>RTTAHGtiter<RTTAHGtiter2114321866161816232761645512864225632151211Total5043(86%)5(10%)2(4%)第55頁/共61頁結(jié)果（表3）改良試管法和IgG凝膠法AHG滴度的比較改良的試管法IgG凝膠試管法N=與改良的試管法滴度相等>改良試管法滴度<改良試管法滴度2114321866161816232761645512864225632151