Related Experiment Video
Updated: Jul 13, 2026

Rapid Determination of Antibody-Antigen Affinity by Mass Photometry
Published on: February 8, 2021
A one-step method for determining the maximum number of bound antibodies, and the affinity and association rate
Jostein Dahle1, Cecilie Krogh, Katrine B Melhus
1Department of Radiation Biology, Institute for Cancer Research, The Norwegian Radium Hospital, Oslo, Norway. jostein.dahle@rr-research.no
Objective:
A reliable analysis of antibody binding may lead to more successful selection of the optimal antibodies. The most important parameters are affinity (equilibrium dissociation constant, Kd), the number of antigen sites on the cells (Bmax) and the on (ka) and off (kd) rate constants of binding. The affinity and the number of cellular binding sites are usually determined by equilibrium binding experiments and subsequent Scatchard analysis. The on and off rate constants are determined by kinetic binding experiments. However, it is necessary to perform two to three different types of experiment in order to determine these parameters.
Methods:
We have developed an alternative one-step method based on a kinetic binding experiment and a mathematical description of antibody binding to antigen. The method was compared with kinetic and equilibrium binding methods.
Results:
The results obtained using two different cell lines were in good agreement with results obtained with Scatchard analysis and kinetic binding experiments.
Conclusion:
An alternative one-step method for determination of parameters describing binding of antibodies to antigens on cells has been developed. The method gives reliable estimates of affinity and number of antigens and in addition gives information on the kinetics of binding.
Related Concept Videos
The Equilibrium Binding Constant and Binding Strength
The Equilibrium Binding Constant and Binding Strength
Affinity and Avidity
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...
Quantitative Aspects of Drug-Receptor Interaction

