Validation of Sequence-Specific Oligonucleotide Probe-Based HLA Homozygosity Classification Using 11-Locus Next-Generation Sequencing in Korean Cord Blood Units: Extended Haplotype Architecture and Implications for Induced Pluripotent Stem Cell Haplobanking.

BACKGROUND: Induced pluripotent stem cell (iPSC) haplobanks require accurate identification of HLA-homozygous cord blood units (CBUs). We validated sequence-specific oligonucleotide probe (SSOP)-based homozygosity classification using high-resolution next-generation sequencing (NGS). METHODS: We performed 11-locus NGS (HLA-A, HLA-B, HLA-C, HLA-DRB1, HLA-DRB3, HLA-DRB4, HLA-DRB5, HLA-DQA1, HLA-DQB1, HLA-DPA1, HLA-DPB1) on 179 CBUs classified as HLA-A, HLA-B, and HLA-DRB1 homozygous through SSOP typing at a Korean public cord blood bank. Concordance was evaluated at two-digit and four-digit resolutions, and extended homozygosity patterns were analyzed. RESULTS: At the two-digit level, SSOP and NGS showed 100% concordance. At the four-digit level, true homozygosity was confirmed in 96.6% (HLA-A), 100% (HLA-B), and 97.8% (HLA-DRB1) of homozygous classifications. All 10 discordant cases involved intra-group allelic divergence. Among 169 confirmed homozygous CBUs, extended homozygosity was observed at HLA-C (97.6%), HLA-DQA1 (100%), and HLA-DQB1 (100%), whereas HLA-DPA1 (56.2%) and HLA-DPB1 (42.6%) showed lower rates reflecting the DQ-DP recombination hotspot. Two previously unreported alleles, subsequently designated HLA-DPA1*01:106 and HLA-DPB1*1344:01, were identified at the DP loci. CONCLUSIONS: SSOP reliably identified HLA-A, HLA-B, HLA-DRB1 homozygous CBUs and frequently resulted in effective five-locus matching owing to conserved extended haplotype structures. This two-tiered SSOP-to-NGS strategy offers a cost-effective framework for iPSC haplobanking in the Korean population.