15-OR

Abstract

Aim Complement-fixing (CF) HLA antibodies (Ab) play an important role in transplantation because they can initiate inflammatory processes within the graft that lead to rejection. The Luminex Ig-binding assay (LUM-Ig) is now the principal method for HLA Ab testing. The C1q-binding assasy (LUM-C1q) can distinguish between CF and non-CF Abs. We show here evidence that the differential reactivity of CF HLA Ab depends on the structural composition of epitope (Ep)-carrying alleles (Al). Methods Human cytotoxic class I Ep-specific monoclonal antibodies (mAb) were tested in LUM-Ig and LUM-C1q (OneLambda). Results were compared to CDC reactivity with informative cells from the 13th International Workshop. Results Two reactivity patterns were observed. For some mAb, Ep-carrying Al reacted similarly in all assays. But certain mAb reacted with more Al in LUM-Ig than in LUM-C1q and CDC. Reactivity was analyzed with HLAMatchmaker, incorporating the concept that eplets are essential parts of structural Eps which can contact the 6 Complementarity Determining Regions (CDRs) of Ab. The data show that LUM-Ig mAb are specific for eplets but that LUM-C1q and CDC reactivity require an additional polymorphic amino acid configuration on eplet-carrying Al. Conclusions The findings must be viewed in context of formation of an antigen-Ab complex resulting in release of free energy necessary to stabilize binding and to induce conformational change in the Fc part of the Ab structure to expose the C1q receptor, the first step of Complement activation. The CF properties of HLA Ab require not only specific recognition of an eplet but also depend on interactions of other CDRs with critical amino acid configurations within the structural Ep. Eplet-carrying Al that lack such critical configurations will bind with Ab but the amount of free energy is insufficient to expose the C1q-binding site and there is no CF. This concept is important to our understanding whether or not CF donor-specific HLA Ab can initiate Ab-mediated rejection.