重组表达糖蛋白GPIIIa片段偶联荧光微球分析人血小板抗原-1系统同种抗体  

Analysis of Human Platelet Antigen-1 System Alloantibodies Using Recombinant GPIIIa Fragments Coupled to Luminex Beads

作　　者：许先国[1] 刘瑛[1] 陈舒[1] 洪小珍[1] 陶苏丹[1] 马开荣[1] 蓝小飞[1] 何吉[1] 朱发明[1] 吕杭军[1] 

机构地区：[1]浙江省血液中心输血研究所,卫生部血液安全研究重点实验室,浙江杭州310006

出　　处：《中国实验血液学杂志》2015年第5期1386-1390,共5页Journal of Experimental Hematology

基　　金：国家自然科学基金(81371905和81570170);浙江省科技厅公益技术项目(2013C33193);浙江省医药卫生科研项目(2011ZDA005和2014KYB066);浙江省卫生高层次创新人才培养项目

摘　　要：目的:应用偶联血小板GPIIIa重组表达片段的Luminex xMAP荧光微球,检测血小板抗HPA-1a和-1b抗体。方法:以倍比稀释成12个浓度(原液~1/2048)的抗HPA-1a WHO国际标准品,比较MAIPA和Luminex xMAP技术检测抗HPA-1a的灵敏度。以8份抗HPA-1a/b阴、阳性对照,36份WHO血小板抗体质控盲样,评估流式荧光微球技术检测抗HPA-1a和-1b的特异性。结果:MAIPA检测抗HPA-1a的灵敏度为滴度1/64(抗体浓度1.56 IU/ml),流式荧光微球检测的灵敏度远高于滴度1/2048(抗体浓度0.049 IU/ml)。微球分析法能特异性鉴别抗HPA-1a和-1b阴、阳性对照。36份盲样中,有1份样本,在高浓度对偶抗体的影响下,产生抗HPA-1b假阳性结果。其余样本的微球分析结果与预期结果相符,能准确检出抗HPA-1a和-1b抗体,而且与HLA和ABO抗体以及其它的血小板抗体(包括抗HPA-3b、-5b、-15b、-GPIb/IX和非HPA-1特异性的抗-GPIIb/IIIa)未发生交叉反应。结论:基于重组偶联血小板GPIIIa的流式荧光微球技术可灵敏、特异性检测血小板HPA-1系统抗体,适合于临床免疫性血小板减少症患者的抗体特异性鉴定。Objective: To detect platelet anti-HPA-1a and-1b antibodies using recombinant GPIIIa fragments coupled to Luminex beads. Methods: The sensitivity of 2 techniques,monoclonal antibody specific immobilization of platelet antigen( MAIPA) and Luminex bead assay,was compared using 12 twofold-serial dilutions( from neat to 1 in 2048) of an anti-HPA-1a WHO international standard. The specificity of Luminex assay to identify anti-HPA-1a and-1b antibodies was assessed using 8 negative or positive controls and 36 blinded samples provided by WHO Platelet Workshop. Results: The sensitivity of MAIPA and Luminex bead assay to detect anti-HPA-1a was dilution 1 /64( i. e.1. 56 IU /ml) and far more than dilution 1 /2048( i. e. 0. 049 IU /mL),respectively. The Luminex bead assay could specifically identify negative and positive controls of anti-HPA-1a and-1b. All results of 36 blinded samples by Luminex assay were accordant to reference results except one sample which contained high concentration antithetical antibody and resulted in false positive of anti-HPA-1b. Cross-reactivity was also not observed with the samples containing HLA,ABO or other platelet antibodies. Conclusion: The Luminex beads coupled with recombinant GPIIIa fragments can be used to detect HPA-1 system antibodies with sufficient sensitivity and specificity,that is suitable for the detection of platelet alloantibodies in clinical alloimmune thrombocytopenia.

关 键 词：人类血小板抗原 同种抗体 GPIIIa片段 MAIPA 荧光微球技术 

分 类 号：R558.2[医药卫生—血液循环系统疾病]