Antibody affinity is defined as strength of the binding interaction between antigen and antibody. It depends on the closeness of the stereochemical fit between antibody sites and antigen determinants, on the size of the area of contact between them, and on the distribution of charged and hydrophobic groups. Antibody affinity is important in the activation of B cells in the elimination of antigens, and in the regulation of the immune response. Measuring the affinity of the antibody or protein for its ligand is a key part of understanding the molecule. While the affinity of monoclonal antibodies can be characterized, the affinity of polyclonal antibodies cannot be determined, because polyclonal serum contains a combination of antibodies with different affinities. Antibody affinity can be measured by ELISA, which was shown that not only more precise and more convenient in comparison to that suggested earlier, but also more informative graphical representation of the experimental data in the appropriate coordinate could be used for evaluation of antibody affinity. Besides, Surface Plasmon Resonance (SPR) technology can be used for determining antibody. Abvigen can provide you with antibody affinity determination service.
Abvigen can provide customers with accurate, fast and high-throughput antibody affinity determination services, based on ELISA, SP-RIA, BLI, RIA, et al.
Surface Plasmon Resonance (SPR) technology can be used for real-time measurement of label-free interactions and remains the gold standard for determining antibody : antigen affinity constants. Using SPR-based methods including single or multicycle kinetics, a range of binding parameters such as KD, ka and kd can be determined.
Fig. 1 Schematic ilustration of SPR measurement device
BLI using the optical interference principle with Fortebio Octet, to detect antibody affinity achieved by detecting changes in the optical interference signal in real time.
The amount of free antibody was determined by indirect ELISA. The affinity constant KD was calculated from the OD value. The calculation was performed as follows:
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