A novel HLA-C null allele, HLA-C*05:99NB. Cauwelier1, F. Nollet1, A. Gadisseur2, W. Schroyens2 & Z. Berneman2

1 Laboratory Medicin, AZ St Jan Brugge-Oostende, Brugge, Belgium2 Department of Hematology, Antwerp University Hospital, Antwerp, Belgium

Key words: HLA-C*05:99N ; new human leukocyte antigen-C; null allele; sequence-based typing

HLA-C*05:99N results from a single nucleotide loss com-pared with its closest allele HLA-C*05:01:01:01.

Here we describe a new HLA-C null allele observed duringroutine HLA typing of a Caucasian hematopoietic stem cellrecipient and her offspring. Blood samples were extractedusing the QiaSymphony DSP DNA extraction kit on theQiasymphony instrument (Qiagen, Hilden, Germany). Lowresolution typing was performed for the human leukocyte anti-gen (HLA)-A, -B, -C, -DRB1 and -DQB1 loci using LifecodesHLA SSO typing kits (Lifecodes, Immucor©, Heppignies,

Belgium) on a Luminex® platform (Luminex Corporation,Oosterhout, the Netherlands). The C-locus was typed asa HLA-C*05,07. Subsequent high resolution confirmatorytyping was performed by sequence-based typing (SBT) ofexons 1–7 using Gendx© SBT reagents (GenDx, Utrecht,the Netherlands) on a 3500xL Genetic analyzer (ThermoFisherScientific Inc., Erembodegem, Belgium). The follow-ing genotypic ambiguities were found for the HLA-C locus:C*05:05,07:01:01:01/C*05:01:01:01,07:18/C*05:01:03,07:166. No gene-specific sequence primers (GSSPs) wereavailable to resolve these ambiguities.

420 © 2014 John Wiley & Sons A/S. Published by John Wiley & Sons LtdTissue Antigens, 2014, 84, 405–437

Figure 1 Nucleotide (A) and protein sequence (B) of HLA-C*05:99N compared to its closest allele HLA-C*05:01:01:01.

Subsequent allele-specific sequencing of the HLA-C locusexons 1–4 was performed using the Protrans© HLA SBT kit(Protrans, Hockenheim, Germany). For the HLA-C*07 allelewe identified an ambiguous result: HLA-C*07:01:01:01/07:18.Polymerase chain reaction-sequence specific primer (PCR-SSP,OleRup, Vienna, Austria) was used to determine the uniquenucleotide position at exon 6, resulting in a typing ofHLA-C*07:18.

For the HLA-C*05 allele, homozygous sequencing identifieda best but not complete match with HLA-C*05:01:01:01. Fur-ther analysis of exon 3 sequences of the obtained sequences incomparison with allele C*05:01:01:01 (Figure 1A) showed asingle nucleotide loss of a G nucleotide at position 381 (codon104) (IMGT/HLA database, version 3.11). This deletion causesa frameshift and premature TGA stop codon at codon 126(Figure 1B), resulting in a truncated protein sequence with par-tial loss of the antigen-binding domain thus assigned as a HLAnull allele.

Sequences for exons 2 and 3 were submitted to GenBank(accession number KF806553). The name C*05:99N has beenofficially assigned by the World Health Organization (WHO)Nomenclature Committee in November 2013. This follows theagreed policy that, subject to the conditions stated in the mostrecent Nomenclature Report (1), names will be assigned to newsequences as they are identified.

The complete typing of the individual possessing the newallele is: A*02:05, A*02:20, B*44:02, B*58:01, C*05:99N,

C*07:18, DRB1*04:07/04:92, DRB1*14:54, DQB1*03:01,DQB1*05:03, DPB1*03:01/124:01, and DPB1*04:01. Thesame C*05:99N null allele was observed in the HLA typing ofthe daughter of this patient.

Correspondence

Barbara CauwelierLaboratory MedicinAZ St Jan Brugge-OostendeBruggeBelgiumTel: 003250452610Fax: 003250452619e-mail: barbara.cauwelier@azsintjan.be

doi: 10.1111/tan.12404

Conflict of Interests

The authors have declared no conflicting interests.

Reference

1. Marsh SGE, Albert ED, Bodmer WF et al. Nomenclature for factors ofthe HLA system, 2010. Tissue Antigens 2010: 75: 291–455.

© 2014 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd 421Tissue Antigens, 2014, 84, 405–437