We reported herein seven new HLA alleles, A*0259, A*02017, B*5609, B*5131, B*5205, DRB1*1444 and DRB1*1445, discovered during routine typing and identified by the sequencing based typing method using specific primers. The A*0259 allele showed a single nucleotide difference at position 125 (guanine vs adenine). This was a novel difference in nucleotide position in the HLA-A, B and C alleles. This nonsynonymous difference may lead to an amino acid change at residue 18 (glycine vs serine). The A*020107 allele was a synonymous substitution by a single nucleotide change at position 123 (cytosine vs thymine). These alleles may be generated by the nucleotide changes from the A*020101 allele. A new B*5609 allele may be generated by a gene conversion of two segments originating from exon 2 of the B*5502 or B*5601 allele and exon 3 of the B*350101 allele. The B*5131 and B*5205 alleles showed a single nucleotide difference at position 527 (adenine vs thymine) and at 134 (guanine vs cytosine). These nonsynonymous differences caused to amino acid mutations at residue 152 (glutamic acid vs valine) in the B*5131 allele and at 21 (arginene vs proline) in the B*5205 allele. The B*5131 allele was identical to the B*5116 allele, and the B*5205 allele was to the B*5201 allele except for the nucleotide differences. The B*5131 allele was found in a non-Japanese blood donor. The DRB 1 *1444 allele had two nucleotide differences at positions 344 (thymine vs guanine) and 345 (guanine vs thymine), and the DRB1*1445 allele had that at 286 (cytosine vs adenine) compared with the DRB1*140501 allele. These nonsynonymous differences caused amino acid mutations at residue 86 (valine vs glycine) in the DRB1*1444 allele and at 67 (leucine vs isoleucine) in the DRB1*1445 allele.

The PCR-Luminex method using fluorescence microsphere was developed for high-resolution HLAA, HLA-B and HLA-DRB1 genotyping in the Japanese population. This genotyping method allowing to define all the possible combinations of alleles at each loci existing in Japanese at the four-digital level except for A*2402, 2601and A*2404, 2605 was accomplished using 33 probes for HLA-A, 46 probes for HLA-B and 44 probes for HLA-DRB1. This method was evaluated with 138 Japanese samples and concordance rates of this typing to the predetermined assignments were 99.3% at the HLA-A and HLA-B loci, and 100% at the HLA-DRB1 locus. Collectively, the PCR-Luminex method is a simple, accurate and high-through put (HLA-A, -B and DRB1 typing of 47 samples for 5 hours) genotyping system offering the efficient routine platform in the HLA field.