The Kinetics of Binding of Peptide/MHC Complexes to T-Cell Receptors: Application of Surface Plasmon Resonance to a Low-Affinity Measurement

• Published in: Methods (San Diego, Calif.) Vol. 6; no. 2; pp. 168 - 176
• Main Authors: Boniface, J.Jay, Davis, Mark M.
• Format: Journal Article
• Language: English
• Published: Elsevier Inc 01.06.1994
• ISSN: 1046-2023; 1095-9130
• DOI: 10.1006/meth.1994.1019

The BIAcore is a powerful tool for the analysis of biomolecular interactions and its applications are expanding rapidly. At its inception the BIAcore was shown to be useful for qualitative comparisons of high-affinity interactions, such as the mapping of antibody epitopes on antigens [L. G. Fagerstam et al. (1990) J. Mol. Recogn. 3, 208-214]. Soon thereafter, the instrument was applied to more quantitative measurements, such as the determination of affinities and rate constants for antibody/antigen interactions [R. Karlsson et al. (1991) J. Immunol. Methods145, 229-240]. Currently, the instrument is being applied to the analyses of many diverse biological interactions, including the binding of transcription factors to DNA, peptide/protein interactions, DNA hybridization, and protein/proteoglycan binding. The application of the BIAcore to these measurements is typically more convenient and faster than the technologies it replaces, but similar data can be obtained with conventional methodology. In contrast, until the advent of the BIAcore, no single, easily adaptable technique for the analysis of low-affinity macromolecular associations was available. For example, radioligand binding assays have seen universal and diverse use but often fail to measure interactions with equilibrium dissociation constants (Kd) greater than 10−7 M. Using the BIAcore to investigate T-cell receptor (TCR) binding to peptide/MHC molecules, we find that we can measure interactions that are 1000-fold weaker (10−4M). Thus, an extremely important application of the BIAcore technology is the characterization of transient, low-affinity interactions. Here, we concentrate on this application and use the binding of peptide/MHC complexes to T-cell receptors as an example.