摘要
为了建立一种基于细胞表型异常的流式细胞术以检测急性前体B细胞白血病微小残留病,对35份pre-cursor-B-ALL病人骨髓和19例正常对照骨髓应用BIOMED-1推荐的5种3色抗体组合(TdT/CD10/CD19,CD10/CD20/CD19,CD34/CD38/CD19,CD34/CD22/CD19和CD19/CD34/CD45)进行了流式免疫分型,以确定前体B细胞正常和异常抗原表型流式图形特征。在35例患者中初诊病人13例,完成诱导缓解后的病人15例,完成巩固治疗的患者7例。应用不同比例的正常骨髓单个核细胞和带有CD34/CD38/CD19阳性白血病细胞进行了系列稀释试验。结果显示:在正常对照组中流式细胞术分析显示了3群CD19阳性细胞,代表了B细胞的3个连续成熟阶段。在precursor-B-ALL患者中这3群细胞消失,代之以大量的白血病细胞,而这些白血病细胞的表型特征与正常B细胞不同。当病人获得完全缓解时,这3群细胞会重新出现,而且具有与正常CD19阳性细胞几乎相同的表型特征。用5组3色抗体组合检测病人时,初诊患者12/13(92.3%)可检出抗原表型异常,也即在0.01%的敏感性水平每个患者至少有1种抗体组合的异常。在本研究初诊病人中这些抗体组合的异常频率:CD10/CD20/CD19为8/13(61.5%);CD34/CD38/CD19为5/13(38.5%);CD10/TdT/CD19为4/13(30.8%);CD34/CD22/CD19为3/13(23.1%);CD34/CD45/CD19为2/13(15.4%)。刚获得完全缓解的患者抗原表型异常的检出率为5/15(33.3%),其中初诊和缓解时同时检出异常者3/8(37.5%)。稀释试验表明,从1∶1至1∶400000的范围,流式细胞术检出与已知加入的CD34/CD38/CD19阳性白血病细胞数有良好的线性相关(r=0.80,P<0.05)。结论:BIOMED-1协作组建议的基于细胞表型异常的流式细胞术用于precursor-B-ALL微小残留病的检测在本研究中能较好地实现。在104个正常骨髓细胞中可以有效检出1个precursor-B-ALL白血病细胞。
To test the European BIOMED-1 Concerted Action proposed technique to detect minimal residual disease（MRD） in the chinese patients with precursor-B-acute lymphoblastic leukemia （precursor-B-ALL） by triple-staining flow cytometry and to define both normal and aberrant phenotypic profiles of precursor B cells, a series of bone marrow samples, 35 from precursor-B-ALL（ 13 in newly diagnosed cases, 15 at the end of remission induction therapy and 7 at end of the consolidations）, and 19 from normal controls, were immunophenotyped with the five triple-staining antibodies（TdT/CD10/CD19, CD10/CD20/CD19, CD34/CD38/CD19, CD34/CD22/CD19 and CD19/CD34/CD45） recommended by the BIOMED-1 using common flow cytometric protocols. Further, with different ratios of the leukemic cells with CD34/CD38/CD19 phenotype and normal mononuclear cells, a serial dilution test was analyzed. The results showed that three major CD19^＋ cell subpopulations were identified in the normal controls, representing three consecutive maturation stages. The subpopulations in the precursor-B-ALL cases disappeared and were replaced with a great number of luekemic cells which had different characteristics of phenotypes, and then they reappeared with almost same characteristics as the normal CD19^＋ cells after the patients achieved complete remission. When the five triplestaining antibody combinations were used, the phenotypic aberrancies could be identified in 12/13 （92.3%） cases with newly diagnosed precursor-B-ALL, at least one triple-labeling per case at the level of 0.01% or more. The frequencies of phenotypic aberrations detected with the triple-staining were 8/13 （61.5%）for CD10/CD20/CD19, 5/13 （38.5%）for CD34/CD38/CDI9, 4/13（30.8% ） for CD10/TdT/CD19, 3/13（23.1% ） for CD34/CD22/CD19, and 2/13（ 15.4% ） for CD34/CD45/CD19. At the end of remission induction, the phenotypic aberrancies could be detected in 5/15（33.3%）, of which, 3/8 （37.5%） cases with the leukemic phenotypes detected both at the newly diagnosis and at the end of induction. The dilution test indicated that the cells with CD34/CD38/CD19 detected by flow cytometry correlated well with the leukemic cells added（ r = 0.85, P 〈 0.05 ） over 1 ： 1 to 1 ： 400 000. It is concluded that the flow cytometric detection of precursor-B-ALL-MRD proposed by BIOMED-1 Concerted Action were well realized in this study. The one precursor-B-ALL cell can be effectively detected out of 10^4 normal bone marrow cells.
出处
《中国实验血液学杂志》
CAS
CSCD
2005年第4期557-562,共6页
Journal of Experimental Hematology
基金
深圳市卫生局科技项目基金资助
编号:199806002
