Chen Yue-Kang, Li Da-Cheng, Wang Da-Ming, Li Qian, Deng Zhi-Hui
Shenzhen Blood Center, Shenzhen, 518035, Guangdong Province, China.
Zhongguo Shi Yan Xue Ye Xue Za Zhi. 2008 Feb;16(1):185-8.
This study was aimed to establish the reliable genotyping method of human platelet antigens of HPA-15 system by PCR-SSP and to use this assay in the further HPA genotyping of volunteer platelet donors. 3 sequence-specific primers recommended by the 11th Platelet Genotyping and Serology Workshop on behalf of International Society of Blood Transfusion (ISBT) were synthesized. The concentration of each primer pair, the concentration of Mg(2+) and the PCR conditions were adjusted to optimize the conditions so that HPA-15 system could be specific amplified. The accuracy and reliability of the developed assay was evaluated and confirmed by typing the coded DNA samples provided by the 11th Platelet Genotyping and Serology Workshop. As a parallel control, a total of 50 volunteer platelet donors in Shenzhen were genotyped by both our assay and the G&T commercial kit at HPA-15 system. 10 coded samples distributed by the 11th Platelet Genotyping and Serology Workshop were genotyped by established PCR-SSP method. The results showed that a concordance rate of 100% was observed between the results obtained by established PCR-SSP method and the results provided by ISBT report. The HPA gene frequencies observed in 50 randomly-selected platelet donors in Shenzhen were 0.5100 and 0.4900 for HPA-15a and HPA-15b respectively. In conclusion, PCR-SSP assay established in our study provides a simple, rapid and accurate method for HPA-15 system genotyping, which assay is suitable for routine clinical HPA genotyping and shows a broad prospect in its further applications.
本研究旨在通过聚合酶链反应-序列特异性引物（PCR-SSP）建立可靠的人类血小板抗原HPA-15系统基因分型方法，并将该检测方法用于进一步对志愿血小板捐献者进行HPA基因分型。合成了第11届血小板基因分型和血清学研讨会代表国际输血协会（ISBT）推荐的3条序列特异性引物。调整各引物对的浓度、Mg(2+)浓度及PCR条件以优化条件，使HPA-15系统能够被特异性扩增。通过对第11届血小板基因分型和血清学研讨会提供的编码DNA样本进行分型，评估并确认了所建立检测方法的准确性和可靠性。作为平行对照，采用我们的检测方法和G&T商业试剂盒对深圳的50名志愿血小板捐献者进行HPA-15系统基因分型。采用所建立的PCR-SSP方法对第11届血小板基因分型和血清学研讨会分发的10个编码样本进行基因分型。结果显示，所建立的PCR-SSP方法获得的结果与ISBT报告提供的结果之间的一致性率为100%。在深圳随机选择的50名血小板捐献者中观察到的HPA-15a和HPA-15b的HPA基因频率分别为0.5100和0.4900。总之，我们研究中建立的PCR-SSP检测方法为HPA-15系统基因分型提供了一种简单、快速且准确的方法，该检测方法适用于常规临床HPA基因分型，并在其进一步应用中显示出广阔前景。
