PCR-SBT法HLA基因分型模棱两可结果的判读及解决对策

Judgment on the Ambiguous Results of PCR-SBT HLA Genotyping and its Solution

目的研究PCR-SBT法HLA基因分型结果判读中出现的模棱两可问题并提出解决策略。方法对306名随机抽取的组织配型患者的DNA采用PCR-序列特异性引物(Sequence Specific Primer,SSP)低分辨方法检测,同时采用PCR-SBT法进行HLA-A,B,DRB1高分辨基因分型。PCR-SBT法模棱两可结果用PCR-SSP高分辨方法复核确认。结果所有306例检测标本的高低分辨血清学抗原一致,经PCR-SBT基因测序方法检测有111份模棱两可结果,占36.3%(111/306)。其中HLA-A座位有30对等位基因存在模棱两可结果,占所有检测标本等位基因总数的3.3%;HLA-B座位有66对,占7.2%;HLA-DRB1座位有36对,占3.9%。所有模棱两可标本经PCR-SSP HLA基因高分辨分型试剂PCR-SSP等方法复核确认。结论针对PCR-SBT法进行HLA-A,B,DRB1高分辨基因分型模棱两可结果所建立的解决策略,对于提高HLA数据分型的速度和分型准确性有重要意义。

Objective To investigate the frequencies of HLA-alleles ambiguous typings by sequence based typing and propose the way to resolve. Method Of 306 randomly selected patients,their DNAs were genotyped using PCR-sequence specific primer （SSP） for low resolution typing and PCR-SBT for allelic designation of HLA-A, B, DRB1. PCR- SSP high-resolution kit were used to resolve the PCR-SBT ambiguous typings. Results Each of the 306 samples showed the identical serological antigens for both low and high resolution genotyping of HLA-A, B and DRB1. Among them, there were 111 results with ambiguilty （of the total shares of 36.3% for HLA-A, B and DRB1） by sequencing, in which, HLA- A had 30 seats（3.3% of the total shares）, HLA-B 66（7.2% of the total shares）, and HLA-DRB1 36（3.9% of the total shares）. All the ambiguous were reviewed by PCR-SSP high resolution kit and got the exact pair of alleles. Conclusion The settlement for the PCR-SBT ambiguous typings against HLA-A,B,DRB1 high-resolution genotyping is of great significance to enhance HLA typing speed and accuracy.