摘要
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ＳＳＰ＾ＴＭＨＬＡ－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，前者的护增效果好。ＤＮＡ终浓度为２５～２００ｎｇ／ｕｌ，最佳浓度为１００ｎｇ／ｕｌ。
Objective: In order to type HLA class Ⅱ of hematopoietic stem cell transplantation donor-recipient pairs accurately, the authors studied the factors impacting on HLA class Ⅱ genotyping methods. Methods: Genotyping methods of HLA-Ⅱ PCR-SSP (Lambda, Inc)were employed to genotype 400 cases of related hematopoietic stem cell transplant donors and recipients. Results: The results indicated that amplified results were much better in 0. 5% EDTA anticoagulated peripheral bled compared with heparin samples. Final DNA concentration should be 25 ~ 200 ng/μl(100ng/μl is optimal) with A260/A280 ratio between 1 .65 ~ 1.80. PCR reaction parameters is also very important in HLA class Ⅱ genotyping. The usage and the activity of Taq DNA polymerase directly affect typing results. DMIX tube should be stored at -65℃, and avoid frequently freezing and melting, otherwise, it might decrease the output of amplification. Conclusion: The proper processing of specimen and optimization d reaction conditions for HLA class Ⅱ genotyping are the key factors for good results.
出处
《中国免疫学杂志》
CAS
CSCD
北大核心
2000年第7期369-370,共2页
Chinese Journal of Immunology
基金
国家自然科学基金!3990187
关键词
HLA-Ⅱ类
基因分型
造血干细胞移植
HLA class Ⅱ Genotyping Hematopoietic stem cell transplantation
