流式细胞术诊断X连锁高IgM血症的临床应用

目的 建立流式细胞术诊断X连锁高IgM血症(X-linked hyper-lgM syndrome,XHIM)的方法.方法 取患者、患者母亲和健康对照者外周血,肝素抗凝,用RPMI-1640培养基稀释,以15μl的佛波醇酯(1 ng/μl)和6 μl的离子霉素(50 ng/μl)作刺激剂,从而将外周血标本分为刺激组和非刺激组,运用荧光标记的抗细胞表面抗原单抗检测CD3+CD8-T淋巴细胞表而CD40配体的表达情况,同时检测CD3+淋巴细胞表面的CD69分子作为激活对照,以确定细胞活化的程度.结果 刺激组细胞表面CD69的表达率均在96%以上;患者母亲及健康对照者CD3+CD8-T淋巴细胞表面CD40的表达率分别为60.04%和62.87%,而患者仅为0.8%.非刺激组CD69的表达率均小于3%;患者、患者母亲和健康对照者CD40L的表达率分别为0.88%、4.15%和5.51%.结论 流式细胞术可以快速诊断X连锁高IgM血症,有可能成为该类疾病的初筛方法.

Application of flow cytometry in the diagnosis of X-linked hyper-IgM syndrome

Objective To develop a flow cytometrie assay to for diagnosis of X-Linked Hyper.1gM Syndrome(XHIM).Methods Heparinized peripheral blood obtained from patient,mother and a healthy control was diluted with RPMI- 1640(unstimulated control)or with RPMI-1640 containing 15μl PMA(1 rig/μl)and 6 trl ionomycin(50 ng/μl)(stimulated cell).Using directly labeled antibodies,we have examined CD40 ligand levels on CD3+ CD8- lymphocyte surface,and CD69 levels on CD;lymphocyte Surface to determine whether the cells were activated.Results CD69 levels on CD3+ lymphocyte surface from stimulated group and from unstimulated group were above 96% and below 3%,respectively.CD40L levels Oil CD3 CDs-T lymphocyte surface from stimulated group were 0.8% (patient),60.04%(mother) and 62.87%(healthy contr01).CD40L levels on CD3+ CD8-T lymphocyte surface from unstimulated group were 0.88% (patient),4.15%(mother)and 5.51%(healthy contr01).Conclusion This flow cytometric assayis accurate and convenient,which Can be used in neonatal screening.