Historically, HLA-DQ antigens and antibodies (Ab) were difficult to classify due to the lack of serological typing reagents. However, with the advent of molecular typing and improved Ab detection reagents there is growing awareness of the prevalence and detrimental effect of donor-specific antibodies to HLA-DQ. Still, despite improvement of reagents, HLA-DQ DSA remain underestimated and underappreciated. We and others have shown that inhibitory factors in some patients' serum can limit the ability to detect HLA Ab using the SAB assay. In analyzing the results per HLA locus, we observed this effect mostly for HLA-DQ Ab. While many laboratories now use techniques to remove at least some inhibition, it is important to realize that in historical studies that did not address inhibition, HLA-DQ Ab were the most likely to be undetected/underappreciated.

Performing Ab strength assessment using titration studies allowed us to recognize an additional limitation of the SAB assay. Studying 55 highly sensitized patients (total 7087 positive beads), we saw that in patients with high levels of Ab, the antigens coating the beads reached saturation, leaving the excess Ab in the serum undetected. This can be identified only by serial dilution of the serum. Moreover, while statistically we observed a good correlation between Ab strength as measured by titers and median MFI values, this held true only for Ab with a titer <1:256. A lack of increase in MFI values confirmed that higher titer Ab saturated the beads, affecting HLA-DQ Ab the most. In this study, 7.4% of beads had titer ≥1:256, and 39.9% of those beads were specific for HLA-DQ antigens. Importantly, some patients exhibited titers as high as 1:65,516.

Additional analysis indicated that HLA-DQ Ab also had the highest variability of MFI in a given Ab titer group. While HLA-A, -B, and –DR Ab MFIs had an average standard deviation of 3416, 2796, and 3328 per titer group, respectively, HLA-DQ average SD was 4435, which increased to 5957 when looking only at Ab with a titer ≥1:256.

Overall, these results indicate that HLA-DQ Ab are more susceptible to inhibition, saturation and variability in binding compared with other HLA Ab, and thus are the most likely to be underappreciated. This in turn can lead to the false impression that HLA-DQ Ab are less responsive to therapy during pre-transplant desensitization or treatment for AMR. It can also affect our ability to understand the immunogenicity of HLA-DQ Ab. Since HLA-DQ DSA are considered the most detrimental to transplant outcome, it is important to appreciate potential limitations of current approaches.

CITATION INFORMATION: Pinelli D., Tambur A. The Complexity of HLA-DQ Am J Transplant. 2017;17 (suppl 3).

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