[Background] Non-human primate models of sensitization provide great preclinical models to investigate the mechanisms of antibody-mediated rejection or desensitization. Now there a growing interest on the use of luminex beads assays for the nonhuman primate model. In this study, we compared serum DSA level to HLA FlowPRA bead assay and HLA single antigen bead assay (SAB).

[Method] Rhesus macaques received skin transplantation (n=6) or peripheral blood mononuclear cells (n=6) intradermally (with adjuvant) from full MHC mismatched donor. DSA was measured by T- and B-cell flow crossmatches (FXM). FXM were considered positive when reaching MFI-ratio >2 (two fold MFI value compared to baseline). Pre- and Post-transplant sera of all animals were tested retrospectively for anti-HLA class I and class II antibodies using multianalyte bead assays and for anti-HLA binding level of DSA using SAB assay performed on the Luminex platform.

[Results] Animals sensitized by the skin transplantation showed a positive T-and B-cell FXM at peak (MFI-ratio, 9.2±7.1 and 6.5±3.2 respectively). However, animals sensitized with PBMC showed negative T cell FXM at all time points (1.49±0.3 at peak), although all had a strongly positive B-cell FXM (9.19±4.9 at peak) (Figure 1A). Concurring with this finding, all skin recipients were positive for both MHC class I and II while PBMC sensitized animals were positive only for MHC class II (Figure 1B). Furthermore, the SAB assay also showed positive binding level of DSA for both class I and II HLA antigens after skin transplantation but only class II HLA antigens after PBMC immunization (Figure 1C).

[Conclusion] Anti-MAMU class I and class II antibodies were able to bind against class I (HLA-A and B) and class II (HLA-DP, DQ and DR) HLA antigens respectively despite of MHC polymorphism in between species. Class I/II bead assays may be useful in affirming FXM results in NHP. Alloantibody trends in NHP may be tracked using SAB assays, although the relevance to MAMU antigens is unclear.

CITATION INFORMATION: Kwun J, Burghuber C, Manook M, Clark A, Chen D, Knechtle S. Utilizing HLA Single Antigen Flow Beads Assays to Evaluate Anti-MAMU Antibody in Sensitized Nonhuman Primate Model. Am J Transplant. 2016;16 (suppl 3).

« Back to 2016 American Transplant Congress