利用微卫星多态性检测造血干细胞移植后嵌合体的研究

EXPERIMENTAL STUDY ON ANALYSIS OF HEMATOPOIETIC CHIMERISM BASED ON DNA POLYMORPHIC MICROSATELLITES

目的利用微卫星DNA的多态性,检测造血干细胞移植后嵌合体。方法用无关个体的单个核细胞混合标本在体外模拟混合嵌合体,采用聚合酶链反应(PCR)结合聚丙烯酰胺凝胶电泳、银染色和凝胶成像分析技术半定量检测嵌合体。结果本方法的敏感性为1.25%～5%,不同位点敏感性不同;混合模拟标本扩增产物分析计算所得嵌合率与扩增前供体细胞比例呈显著直线相关(r=0.97～0.98,P<0.001),不同位点间相关性有所差异,以各位点的平均值进行线性分析更趋于直线相关(r=0.998)。结论STR-PCR结合凝胶电泳成像分析技术是一种敏感、准确、可靠、经济的嵌合体半定量检测方法。

Objective To establish a method for analysis of hematopoietic chimerism by DNA polymorphic microsatellites. Methods Cell dilution experiments were performed by mixing mononuclear cells （MNCs） obtained from peripheral blood samples of unrelated individuals. Semi - quantitative assessment of hematopoietic chimerism was performed by short tandem repeat - polymerase chain reaction （ STR - PCR）. Polyacrylamide gels, silver staining were analyzed by Image Analysis System. Results Sensitivity varied from 1.25 % to 5 % depending on the STR locies being tested. The quantitative results showed a linear correlation between the percent chimerism calculated and the DC proportion mixed samples（ r = 0.97 - 0.98, P 〈 0. 001 ）. As an average of all informative markers, concordance between the calculated and mixed DC percent showed minimal deviation from（ r = 0. 998）. Conclusion STR- PCR provides a sensitive, precise, reliable and economic approach for semi -quantitative analysis of hematopoietic chimerism.