Development of a new HLA-DNA typing method applicable in the nation-wide renal transplantation network.

在非亲缘身体器官移植中，从公平和效率的角度出发，器官移植中供者和受者的HLA配型都是重要的。我们已经开发了一种新的人类白细胞抗原-DNA分型方法，它使用针对基因组DNA的RNA扩增方法、转录介导的扩增(TMA)和基于化学发光的杂交保护分析(HPA)，这是序列特异性寡核苷酸(SSO)方法的一种。在本研究中，我们将聚合酶链式反应(PCR)和HPA方法相结合，进一步分析DR6抗原组的精细等位基因。用DR356DNA特异性引物对121份标本进行聚合酶链式反应和高效液相色谱(HPA)检测。在DRB1^&lt；**&gt；13等位基因群中，有10个等位基因(DRB1^&lt；**&gt；1301-1307，^&lt；**&gt；1310-1312)，在DRB1^&lt；**&gt；14，11个等位基因中(DRB1^&lt；**&gt；l401-1408，^&lt；**&gt；1411^&lt；，**&gt；1412，^&lt；**&gt；l417)。然而，某些等位基因组合(^&lt；**&gt；1301/02，^&lt；**&gt；1140l/07)没有明确的分型，需要额外的引物组合。PCR-HPA法提取DNA所需时间为1小时，扩增为2小时，检测为1小时，共4小时。因此，用PCR-HPA方法进行人类白细胞抗原DNA配型是一种简便、快速、非同位素、准确的方法，将保证器官移植的公平和效率。

HLA typing of both donor and recipients in organ transplantation is important from the stand point of equity and efficiency in unrelated cadaver organ transplantation. We have already developed a new HLA-DNA typing method using a RNA amplification method targeting at the genomic DNA,Transcription-Mediated Amplification (TMA) and a chemiluminescence-based Hybridization Protection Assay (HPA), which is one type of sequence-specific oligonucleotide (SSO) methods. In this research, we combined polymerase chain reaction (PCR) with HPA method, to analyze further fine alleles of the DR6 antigen group. One-hundred and twenty-one DNA samples were amplified with DR356-specific primers by PCR and detected by HPA method. Among DRB1^<**>13 allele group, 10 alleles (DRB1^<**>130l-1307, ^<**>1310-1312) were derfinitely typed, and among DRB1^<**>14,11 a11eles (DRBl^<**>l401-1408, ^<**>1411^<, **>1412, ^<**>l417). However, a certain combination of alleles (^<**>1301/02, ^<**>1140l/07) were not typed unequivocally and need additional primer combinations. The procedure time required for PCR-HPA method is one hour for extraction of DNA,2 hours for amplification, and one hour for detection, a total of 4 hours. Thus, PCR-HPA method for HLA-DNA typing is a simple, fast, non-isotopic and accurate method, which will guarantee the equity and efficiency in organ transplantation.