No abstract

BackgroundNext‐generation sequencing (NGS) is the most modern sequencing technique that has revolutionized HLA typing, providing high‐resolution results with low ambiguity rates. This study aimed to show the experiences and challenges of an HLA laboratory in the validation process of the NGS methodology for HLA typing and show the use of this method for the study of HLA genetic diversity.MethodsWe used 115 samples that comprised a comprehensive testing panel for validation of the NGS methodology using the AllType kit (One Lambda, Canoga Park, California) on the Ion Torrent S5 NGS platform. All quality metrics were analyzed. During validation, two new HLA sequences were identified and named by the HLA Nomenclature Committee.ResultsA total of 1380 alleles from the HLA‐A, ‐B, ‐C, ‐DRB1, ‐DQB1, and ‐DPB1 loci were examined by NGS. This validation panel provided a wide range of HLA sequence variations, including non‐CWD HLA alleles, new variants, and homozygous alleles. The concordance rate with Sanger sequencing‐based typing was 100.0% for HLA‐A, ‐B, ‐C, ‐DRB1, ‐DQB1, and 99.93% for HLA‐DPB1. The newly identified HLA alleles were HLA‐B*14:69N and HLA‐DQB1*02:145.ConclusionWe have successfully validated NGS HLA typing despite numerous challenges, contributing to the identification of novel alleles that impact on HLA matching and antibody evaluation in organ and tissue transplantation.

The novel HLA‐A*11:379, B*45:01:11, B*15:571, B*57:137, C*07:893 alleles were identified in Brazilian individuals.

Fifteen novel HLA alleles in the HLA‐A, ‐B, ‐C, DRB1, and DQB1 loci were described.

The novel HLA‐DRB1*03:178, ‐DRB1*03:179, ‐DRB1*11:276 alleles were identified in healthy Brazilian individuals.