Determining the Binding Affinity in Monoclonal Antibody-Alloantigen Interactions.

Humoral immunity, a significant facet of the adaptive immune system, neutralizes targets when circulating antigen-specific antibodies recognize a foreign antigen and recruit antibody-dependent effector systems. Antibody-mediated complement deposition and cellular removal through engagement of Fc receptors can mediate intravascular and extravascular hemolysis following an incompatible red blood cell (RBC) transfusion. Efficiency of hemolysis is influenced by the affinity of the antibody-antigen reaction. Therefore, determining the affinities of alloantibodies can be important when investigating the potential consequences of antibody engagement on RBC clearance and possible changes to the target antigen. Here, we describe three techniques, surface plasmon resonance, protein microarray, and flow cytometry, to distinguish affinities of monoclonal antibodies specific to the Hen Egg Lysozyme, Ovalbumin, and Duffy (HOD) fusion model antigen.