1. Measuring anti-HLA antibody active concentration and affinity by surface plasmon resonance: Comparison with the luminex single antigen flow beads and T-cell flow cytometry crossmatch results.
- Author
-
Visentin J, Leu DL, Mulder A, Jambon F, Badier L, Lee JH, Guidicelli G, Bouthemy C, Ralazamahaleo M, Claas F, Di Primo C, and Taupin JL
- Subjects
- Alleles, Epitopes chemistry, Humans, Kinetics, Antibodies, Monoclonal analysis, Flow Cytometry methods, HLA Antigens immunology, Histocompatibility Testing, Rheology, Surface Plasmon Resonance methods, T-Lymphocytes metabolism
- Abstract
Background: Although the Luminex single antigen flow beads (SAFB) and the flow cytometry crossmatch (FCXM) are the most sensitive assays used for anti-HLA antibodies characterization in transplant recipients, their semi-quantitative fluorescence read-out is not closely linked to graft outcome., Methods: Surface plasmon resonance (SPR) was implemented to determine truly quantitative parameters of five human monoclonal anti-class I HLA antibodies (mAbs): first the active concentration and then the binding constants. The results were compared to those obtained with SAFB and T-cell FCXM (T-FCXM)., Results: The five mAbs displayed different rate and equilibrium constants for their cognate antigens. No correlation was evidenced between SAFB MFI or T-FCXM ratio and the binding parameters measured by SPR. Some mAbs amino acid substitutions within the epitope that influenced SAFB MFI resulted in affinity variations evidenced by SPR., Conclusion: The SAFB MFI and T-FCXM ratio, both semi-quantitative parameters, only partially reflected the subtlety of the anti-HLA antibody/antigen interaction as it can be analyzed by SPR. Future clinical studies using SPR for anti-HLA antibodies characterization could bring novel insights into the understanding of HLA/anti-HLA interaction and therefore anti-HLA antibodies pathogenicity., (Copyright © 2019 Elsevier Ltd. All rights reserved.)
- Published
- 2019
- Full Text
- View/download PDF