流式免疫微球芯片技术检测血小板膜糖蛋白特异性自身抗体的方法及其临床应用

Establishment and clinical application of flow cytometric immuno-bead array assay in detecting platelet-associate autoantibody against platelate-specific glycoproteins

目的建立流式免疫微球芯片技术(FCIBA)检测血小板膜糖蛋白(GP)特异性自身抗体的方法及其临床应用。方法使用流式细胞仪检测包被不同GP的单克隆抗体,抗GPⅨ(SZ1)、抗GPⅠb(SZ2)、抗GPⅢa(SZ21)、抗GPⅡb(SZ22)、抗P-选择素(SZ51)的不同荧光强度的微球,检测结果与单克隆抗体俘获血小板抗原技术(MAI-PA)进行比较。选取特发性血小板减少性紫癜(ITP)组、非ITP(NITP)组和健康人对照组样本用FCIBA分析SZ1、SZ2、SZ21、SZ22、SZ51共5种特异性血小板自身抗体。结果批内批间差异小;ITP组与NITP组和健康人对照组比较,差异均有高度统计学意义(均P<0.01);对ITP诊断的总特异度为81.4%(70/86),MAIPA总特异度为80.2%(69/86),差异无统计学意义(P>0.05)。敏感度为62.00%(31/50),明显高于改良间接MAIPA的31.00%(19/50,P<0.05)。结论 FCIBA可以一次同时进行5种自身抗体的联合检测,为ITP的临床诊断提供了一种快速测定的新方法。

Objective To establish a new method to detect platelet-associate autoantibodies against platelate-specific glycoproteinsss by flow cytometric immuno-bead array assay （FCIBA） and study its clinical application. Methods A series of beads were detected by the flow cytometry. The beads with different fluorescent intensity were used for coating different anti-platelet glycoproteins monoclonal antibodies, anti- GPIX （ SZ1 ）, anti-GP I b （ SZ2 ）, anti-GP ma （ sz215, anti-GP lib （ SZ22 ）, anti-P- selectin （SZ51）. In addition, its results were compared with monoclonal antibody immobilization of platelet antigens （MAIPA） technology. The new method was used for detecting five autoantibodies SZ1, SZ2,SZ21, SZ22 and SZS1. Results The coefficient of variation （CV） of intra-repetition and inter- repetition were small in difference. ITP group was compared with NITP group and the healthy control, the difference in the MFI was significant （P 〈 0.01 ）. The specificity of FCIBA was 80.2% （70/86）, and the specificity of MAIPA was 80.2 % （69/86）, the difference was not significant （P 〉 0.05 ）. The sensitivity was 62.00% （31/50）, which was higher than that of MAIPA 31.00% （19/50）, the difference was significant （P 〈 0.05 ）. Conclusion FCIBA can be used to detect platelet-associate autoantibodies, particularly in conjunction with five antibodies can improve the positivity rate, which might be a useful method to improve the diagnosis of ITP.