การศึกษาเพื่อประเมินน้ำยาตรวจหาอัลลีล HLA ด้วยเทคนิค Next Generation Sequencing

Abstract

Abstract:

Background: Currently, many Thai patients who have registered for unrelated stem cell donors from the Thai Stem Cell Donor Registry (TSCDR) but could not find human leukocyte antigens (HLA) - matched donors. One factor that influenced the selection was donors’ HLA allele typing results reported as low/intermediate resolution. Thus, more time was needed to perform high resolution typing in order to identify HLA-A, -B, -C and -DRB1 alleles. Objective: To evaluate next generation sequencing (NGS) platforms for high resolution HLA allele typing. Materials and Methods: HLA allele typing was performed in 24 known DNA samples using commercial reagent kits. The HLA typing results of 5 loci (HLA-A, -B, -C, -DRB1 and -DQB1) were compared with the results obtained from the proficiency testing programs. Results: The three commercial kits could identify 4 digits resolution typing of each locus and the concordance with the known alleles was 99.58%, 99.16% and 99.58% for Holotype HLA, NGSgo, and AllTypeNGS assay, respectively. Times requiring for the whole process of typing varied from 31 hours (AllTypeNGS assay), 32 hours (NGSgo) and 41 hours and 30 minutes (Holotype HLA), respectively. Test limitations were due to different primer design and software analysis. Conclusion: In this study, we evaluated NGS platforms for high resolution HLA typing. All kits were able to give HLA typing results as the same alleles with more than 99% accuracy. Differences in testing time, procedures and program analysis were found. However, the analysis for cost of reagents and number of samples tested are needed to be considered before HLA typing by NGS can be implemented in TSCDR.