Authors: Efi Melista, Krisztina Rigó, Szilveszter Juhos, György Horváth, Peter Meintjes, Tim Hague
Abstract:
NGS-based (Next Generation Sequencing) HLA typing can offer accurate and unambiguous genotyping results with minimal manual intervention, eliminating the requirement for reflexive testing. In a clinical setting an easy-to-follow protocol with minimal hands-on time is also essential. Holotype HLA is an optimised, pre-configured assay and software combination product that provides comprehensive gene characterisation of multiple HLA loci for sequencing on the Illumina MiSeq. The assay provides a streamlined, high-throughput, fully automatable protocol that is designed specifically for clinical lab implementation.
The software, HLA Twin, uses two independent algorithms and 15 informative Quality Control metrics for confident allelic determination. The Early Access Program was launched in October 2014 and run for 9 months.
Overall, 96.7% concordance with reference typings was identified. Of the discordant samples, 1.7% were due to an incorrect reference typing, 0.7% were due to a novel allele and the remaining were due to low quality data indicated by QC failures. No ambiguities were identified for results up to the 3rd field, except in 6 instances for HLA-DRB1. The cause of these ambiguities was due to variations located in off-target amplification regions. Finally, the accuracy of the results was more than 99.9% for HLA-A, HLA-B, HLA-C and HLA-DRB1 and 99.76% for HLA-DQB1. The cause of the lower accuracy rate for HLA-DQB1 has been identified and successfully improved since the end of the EAP.
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