The permanently increasing number of human leukocyte antigen (HLA)-alleles and the growing list of ambiguities require continuous updating of high-resolution HLA typing results. Two different kinds of ambiguities exist: the first, when two or more allele combinations have identical heterozygous sequences, and the second, when differences are located outside the analyzed region. The number of HLA-A, B and C alleles recognized in 1999 was almost tripled in 2006. Two hundred individuals, sequence-based typing (SBT) typed in the period from 1999 to 2002, were reanalyzed using the 2006 database. A final allele typing result of at least four digits was obtained for HLA-A, -B and -C by heterozygous sequencing of exons 2 and 3 and, if necessary, additional exons and/or allele-specific sequencing. Storage of the individual sequences in a specially developed database enabled reanalysis with all present and future HLA releases. In the 5-year period HLA-A, -B and -C typing results became ambiguous in 37%, 46% and 41% of the cases. Most were because of differences outside the analyzed region; ambiguities because of different allele combinations with identical heterozygous sequences were present in 7%, 8% and 13% of the HLA-A, -B and -C typings. These results indicate that within 5 years, approximately half of the HLA SBT typings become ambiguous.

In our recent study using high-resolution HLA-B locus typing by sequence-based typing (SBT) we identified 9 new alleles in a total of 355 unrelated individuals (4). Three of them concerned an allele belonging to the B22 group. One of them, B*5607, showed the unusual presence of a Bw4 sequence motif, as described previously (5). In this report the other two B22 variants are described; one belonging to the B55 specificity and named B*5509; the other one being a B*56 allele and assigned B*5606, which brings the total number of alleles belonging to the B22 group to 18.

A new B*35 allele, B*3556, was identified in a Caucasian individual. Direct sequencing of exons 2 and 3 revealed that B*3556 was identical to B*3503, except for a nucleotide substitution from G to A at position 302, which causes a change from AGC to AAC at codon 101. This results in an amino acid change from Ser to Asn at position 77 of the mature protein. Two B*44 alleles, B*4420 and B*4427, were detected in a liver recipient and in an unrelated bone marrow donor, respectively, and their sequences were confirmed. The sequences of exons 1,4 and 5 were also elucidated; for B*4420, these exons proved to be identical to those of B*440201. The sequence of exon 5 of B*4427 showed three mismatches with that of B*440201, implicating that it probably arose from B*440201 by allele conversion with an allele of the B*15, 45, 46, 49 or 50 allele group. Elucidation of the sequence of introns 3 and 4 indicated that the breakpoint for allele conversion has to be located in intron 3 or exon 4.

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