Improvement in HLA‐typing by new sequence‐specific oligonucleotides kits for HLA‐A, ‐B, and ‐DRB1 loci

Abstract

Polymerase chain reaction sequence-specific oligonucleotide is commonly used for HLA-typing. We replaced our LabType SSO HD (HD) kits with LabType SSO XR (XR) kits (One Lambda, Inc., Canoga Park, California) for HLA-A, -B, and -DRB1 following acquisition of a LABScan3D analyzer. The XR kits have more bead regions than the HD kits, allowing for an extended number of probes and exon coverage. They are claimed to improve typing resolution and to diminish the number of allele ambiguities, including common and well-documented (CWD) and null alleles to be resolved. We retrospectively selected patients who had their first HLA-typing performed with the HD kits and their second determination with the XR kits between 2015 and 2016. Forty-two patients were selected for HLA-A typing comparison, and 48 for HLA-B and 41 for HLA-DRB1. XR kits significantly decreased the number of allele ambiguities for HLA-A and -B. On the other hand, the improvement was limited for the HLA-DRB1 locus. The XR kits did not resolve all the CWD HLA allele ambiguities, which may be important for organ and/or hematopoietic stem cell transplantations. The XR kits eliminated 88%, 62%, and 27% of null allele ambiguities for HLA-A, -B, and -DRB1 loci, respectively. In conclusion, the XR kits allow for a significant improvement of HLA-typing resolution for HLA-A and -B loci in comparison with HD kits. In contrast, the number of oligonucleotides in the XR HLA-DRB1 kit should be extended to include exon 3 at the very least. It could also be interesting to include oligonucleotides allowing HLA-DRB3, 4, and 5 typing.

Characterization of the novel HLA‐C*07:708 allele by sequencing‐based typing

“…The new T base was attributed 511 times to the new HLA‐C*07 allele, and can be only attributed to this allele because it was possible to discriminate from the associated HLA‐C*16:01:01:01 allele by virtue of 12 variants positions each distant by less than 100 base pairs. HLA typing by Luminex reverse sequence‐specific oligonucleotide (SSO) was performed (OneLambda Labtype XR) . With this assay (lot 001, catalog RSSOX1C_001_08), the HLA‐typing of the HLA‐C*07 allele was HLA‐C*07:01/07:78/07:103/07:153/07:166/07:408/07:417/07:418/07:419/07:422/07:424/07:442/07:443/07:448/07:453/07:458/07:461/07:462/07:463/07:469/07:470/07:471/07:479/07:491/07:493/07:502/07:506/07:508/07:522/07:536/07:545/07:547/07:549/07:554/07:555/07:557/07:561/07:570/07:588/07:591/07:603N/07:610/07:617/07:619/07:623/07:624/07:627/07:633N without any bead modification.…”

Characterization of the novel HLA‐A*33:170 allele by sequencing‐based typing

“…We were very confident in the phasing as the sample displayed a mean read length of 309 base pairs over all the loci, the mismatched C base was attributed 1060 times to the new HLA‐A*33 allele, and can be only attributed to this allele because it was possible to discriminate from the associated HLA‐A*31:01:02:01 allele by virtue of one variant position distant by less than 100 base pairs. HLA typing by Luminex reverse sequence‐specific oligonucleotide (SSO) was performed (OneLambda Labtype XR) . With this assay (lot 002, catalog RSSOX1A_002_05), the HLA‐typing of the HLA‐A*33 allele was HLA‐A* 33:01/03/18/69/92/95/97/109/111/123N without any bead modification.…”

Characterization of the novel HLA‐DQA1*05:14 allele by sequencing‐based typing

“…We were very confident in the phasing as the sample displayed a mean read length of 352 base pairs over all the loci, the mismatched T base was attributed 285 times to the new HLA‐DQA1*05 allele, and can be only attributed to this allele because it was possible to discriminate from the associated HLA‐DQA1*01:01:01 allele by virtue of six variants positions each distant by less than 100 base pairs. HLA typing by Luminex reverse sequence‐specific oligonucleotide (SSO) was performed (OneLambda Labtype HD) . With this assay (lot 011, catalog RSSO2Q_011_02), the HLA‐typing of the HLA‐DQA1*05 allele was HLA‐DQA1*05:05/09/11 without any bead modification.…”