HLA-B locus genotyping with reference strand mediated conformation analysis (RSCA) system in clinical usage / 中华器官移植杂志

ABSTRACT

Objective To establish and stabilize a new HLA-B locus typing strategy, reference strand mediated conformation analysis(RSCA)system and to conform its advantages.Methods Of 27 hematopoitic stem cell transplant patients and 63 potential donors DNA samples were extracted from peripheral blood cells and HLA-B loci were both typed by RSCA and PCR-SSP methods. The (ambi)-(guous) results were identified by using DNA sequencing. Results Among 90 samples, 87/90 ((96.7) %) cases could be designated definitely, but one of them was disagreement with the SSP result which was confirmed by sequencing, and 67/90 ((74.4) %) cases could be typed to allelic level. 1/90 ((1.1) %) case could not be identified by RSCA for its bad PCR results. Only one allele of each 2 samples could be designated and the other could not be identified by RSCA, which were further confirmed by sequencing method, and the results confessed which were known as alleles, but there were no corresponding data in RSCA database. Twenty samples were randomly selected to identify the replication rate of RSCA, and the results demonstrated that the replication rate was 100 %. Among PCR-SSP typing results, about 10 % samples might be typed for 1-3 times to confirm their types. Conclusion RSCA had some advantages such as high resolution, high sensitivity, high accuracy, high replication, finding new alleles, large scale and low cost, and was especially suitable for unrelated donor-recipient (screening).