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目的：为了准确地进行造血干细胞移植供-受体的HLA-Ⅱ类配型，对HLA-Ⅱ类基因分型实验的影响因素进行了探讨。方法：采用美国莱姆达公司提供的微量SSPTMHLA-Ⅱ类PCR-SSP分型试剂盒对400例造血干细胞移植供-受体进行基因分型。结果：采用0．5％EDTA抗凝的血标本与肝素抗凝的血标本相比，前者的扩增效果好。DNA终浓度为25~200ng/μl,最佳浓度为100ng/μl,A260/A280比例在1．65~1．80之间。PCR反应参数是影响反应的重要因素之一。DNATaq酶的使用量和活性将直接影响反应结果。DMIX管应保存在-65℃，避免反复冻融，否则将使反应带变弱。结论：HLA-Ⅱ类基因分型标本的处理和反应条件的优化是影响基因分型效果的因素。
OBJECTIVE: To investigate the HLA-Ⅱ genotyping of donor-acceptor cells for hematopoietic stem cell transplantation, the influencing factors of HLA-Ⅱ genotyping experiments were discussed. Methods: 400 donor hematopoietic stem cell transplantation recipients were genotyped by using the SSPTMHLA-Ⅱ PCR-SSP kit provided by the American company LAMBDA. Results: Compared with the heparin-anticoagulated blood samples, blood samples with 0.5% EDTA anticoagulation showed good amplification. DNA final concentration of 25 ~ 200ng / μl, the optimal concentration of 100ng / μl, A260 / A280 ratio between 1.65 ~ 1.80. PCR reaction parameters are one of the important factors affecting the reaction. DNATaq enzyme usage and activity will directly affect the reaction results. DMIX tubes should be stored at -65 ° C to avoid repeated freezing and thawing, otherwise the reaction zone will be weakened. Conclusion: The treatment of HLA-Ⅱ genotyping samples and the optimization of reaction conditions are the factors influencing the genotyping effect.