HPA-1a抗体MAIPA定量检测标准体系的建立  

The development of quantitative MAIPA assay for antibodies against human platelet antigen-1a

作　　者：李睿书[1] 乔郑磊 凌冰[1] 陆萍[1] 

机构地区：[1]上海市血液中心上海市输血研究所,上海200051 [2]华东师范大学生命科学学院

出　　处：《中国输血杂志》2015年第6期642-644,共3页Chinese Journal of Blood Transfusion

摘　　要：目的建立针对人类血小板抗原HPA-1a抗体的血小板抗原单克隆抗体特异性免疫固定定量检测(MAIPA)的标准体系。方法用MAIPA方法检测不同稀释度的标准HPA-1a抗血清,优化该方法的最佳实验条件,绘制线性标准曲线,并对该方法的敏感度、特异性、准确度和精密度进行了分析。结果经优化,确定MAIPA定量检测方法的最佳实验条件如下:铺板抗体羊抗鼠Ig G单抗:3μg/m L,血小板每孔2×107个,检测抗体小鼠抗人CD61单抗:20μg/m L,酶标羊抗人Ig G:稀释度1:10 000。该方法的线性检测范围是0-4 IU。该方法最低检测限至少为0.3 IU,批内变异系数为1.3%-3.6%,批间变异系数为4.0%-5.6%。特异性检测:用抗-A、抗-B、抗-H、抗-I、抗-P1、抗-Lewisa等试剂以及含其他抗体的抗血清进行试验,均无明显阳性。结论本研究建立的针对HPA-1a抗体的MAIPA定量检测的标准方法具有较高的敏感度、特异性、准确度和精密度,对于临床相关疾病的诊断和血小板安全输注具有重要的应用价值。Objective To develop a standardized quantitative monoelonal antibody-specific immobilization of platelet antigen （MAIPA） assay for antibodies against human ptatelet antigen-1a （HPA-1a）. Methods MAIPA protocol was applied on serial dilutions of standard reagents containing antibodies against HPA-la. The optimal experimental parameters were determined and standard curves were constructed with a linear regression. The sensitivity, specificity, accuracy and precision of the assay were also evaluated. Results The optimum conditions for the MAIPA assay were as followed ： coating antibody AffiniPure Goat Anti-Mouse IgG Fc~/ Fragment Speeifie： 3 μg/mL, platelets： 2 × 10^7/well, detecting antibodies： Purified Mouse Anti-Human CD61：20 μg/mL, and Peroxidase-AffiniPure Goat Anti-Human IgG Fcγ Fragment Speeific： 1 ： 10 000 dilution. The linear range of the assay was 0 -4 IU; the intra-assay eoeffieients of variation （CVs） were 1.3% - 3.6% ; the inter-assay CVs were 4. 0% - 5.6%. In specificity tests, some monoclonal antibodies, such as anti-A, anti-B, anti-H, anti-I, anti-P1 and anti-Lewisa were tested and no obvious positive result was observed. Conclusion The standardized quantitative MAIPA assay for antibodies against HPA-1a has good sensitivity, specificity, aeeuracy and precision. It has important application value in clinical diagnosis of alloimmunization disease caused by HPA antibodies and safe transfusion of platelets.

关 键 词：MAIPA 标准体系 HPA-1a 安全输注 

分 类 号：R457.11[医药卫生—治疗学] R331.143[医药卫生—临床医学]