Related Experiment Video
Updated: Sep 3, 2026

Determining Binding Affinity (KD) of Radiolabeled Antibodies to Immobilized Antigens
Published on: June 23, 2022
Determining the Binding Affinity in Monoclonal Antibody-Alloantigen Interactions
Kashyap R Patel1, Ryan P Jajosky1, Victoria M Vance1
1Division of Transfusion Medicine, Mass General Brigham, Harvard Medical School, Boston, MA, USA.
Abstract:
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.
Related Concept Videos
Affinity and Avidity
The Equilibrium Binding Constant and Binding Strength
Antibody Actions
Neutralization
Antibodies can bind to pathogens, preventing them from infecting host cells. This process...
Protein-Drug Binding: Determination Methods
Indirect methods involve isolating the bound drug from its free form in biological samples such as blood, serum, or plasma. These techniques aim to measure the percentage of drugs bound to proteins. Equilibrium dialysis is a commonly used method where the free drug concentration at equilibrium is measured by separating the bound...
Cooperative Allosteric Transitions
Ligand Binding and Linkage

