(607) Validation of r-SSO HLA Typing Reagents and Analysis Software: We Must do Better to Avoid Erroneous Typing Results - Sometimes You Pay More and Get Less!

Aim: As rSSO kits evolving we examined the most advanced and more expensive LABType™-XR kit, in comparison to the generic LABType™ kit, to identify issues that without validation by laboratories can result in erroneous HLA typing data.

Methods: Among many in-house samples typing for allogeneic HSCT, using LABType™-XR reagents analysed on FUSION software v.4.6, three different samples in different HLA-typed persons erroneous typing flagged for A*30:02. The three samples had different combinations of A*30:02 (i.e., A*02:01, 30:02 and A*03:02, 30:02).

Results: All 30:02 samples were erroneously typed as A*30:10, due to false positive (FP) signal by bead #821, which is an “Specific” bead for the following alleles: A*02:548, A*03:311/397, A*11:151/401, A*30:10/88, A*31:60N, A*32:132N and A*33:126. Bead #821 probe reactivity is: [98-H----103 / 97---S99 / 96-M---Z101]. Only when bead #821 was converted into negative, rather than FP, A*30:10 ambiguity resolved to A*30:02. Bead #821 does not have a positive control on the manufacturer's quality control (QC) data but the highest normal value for negative samples is 6 and an arbitrary cutoff of 15 was set by the manufacturer for this bead. All samples produced reactivity with this bead at a normal value of 30. Sequence analysis using IMGT reveals that while A*30:10 can cause this bead to be positive, the A*30:02 allele sequence can also cause a positive reaction on this bead but this information is not provided by the software. In two cases bead #821 was associated with total absence of bead #983reactivity, a “Generic” bead that has many A*30:XX alleles, but A*30:10. In such sample the software excluded bead #983 automatically.

Conclusion: Our data clearly shows the need for assay validation in the laboratory and more thorough validation by the manufacturers to avoid erroneous HLA typing results. For the LABType™-XR product, we also found several coding errors in the probe pattern file and found that several beads have no positive control samples in the manufacturer's QC data. We believe that issues with HLA typing reagents and software are not isolated to one product/platform. To enhance the quality and accuracy of HLA typing reagents and software a partnership between laboratories and manufacturers is necessary.