有核细胞嗜水气单胞菌溶素变异体流式法:一种更敏感稳定的阵发性睡眠性血红蛋白尿症检测方法

摘要

目的 建立一种以有核细胞为研究对象,用Alexa-488标记嗜水气单胞菌溶素变异体(FLAER)检测阵发性睡眠性血红蛋白尿症(PNH)的方法,并与传统CD55、CD59检测比较.方法 联合FLAER和CD45、CD33直接检测PNH患者和非PNH患者外周血单核细胞和粒细胞表面糖化肌醇磷脂(GPI)锚的缺失率,同时检测红细胞CD55、CD59表达水平,并进行比较.结果 成功建立了基于有核细胞的FLAER流式检测法.与传统的CD55、CD59相比,FLAER检测更敏感,在流式图上更清晰明显；对于微小PNH异常克隆,单核细胞FLAER法更敏感(P＜0.05).结论 有核细胞FLAER多参数流式法检测PNH异常克隆更直接更灵敏、稳定.%Objective To develop a karyocyte-base flow cytometry to detect paroxysmal nocturnal hemoglobinuria( PNH) using Alexa488-labeled variant of aerolysin, then comparison with traditional CD55 and CD59 analysis. Methods We combined fluorescent aerolysin (FLAER) with CD45,CD33 allowing the direct analysis of the deletion rate of GPI anchor on granulocyte and and monocyte lineages in 45 patients including PNH and non-PNH. Meanwhile, loss of the GPI-linked structures CD55 and CD59 from e-rythrocyte were analysed. Results A karyocyte-base FLAER flow cytometry of detecting PNH was established successfully. Comparison to traditional CD55 and CD59 analysis, karyocyte-base flow cytometry showed excellent sensitivity and clearly. The tiny PNH abnormal clones was found more easily by FLAER assay than traditional CD55 and CD59 analysis(P <0. 05). Conclusion Karyocyte-base FLAER flow cytometry offers a more stable, direct and sensitive assay for dignosis and monitoring PNH clones.