Methods for quantifying T cell receptor binding affinities and thermodynamics

αβ T cell receptors (TCRs) recognize peptide antigens bound and presented by class I or class II major histocompatibility complex (MHC) proteins. Recognition of a peptide/MHC complex is required for initiation and propagation of a cellular immune response, as well as the development and maintenance of the T cell repertoire. Here, we discuss methods to quantify the affinities and thermodynamics of interactions between soluble ectodomains of TCRs and their peptide/MHC ligands, focusing on titration calorimetry, surface plasmon resonance, and fluorescence anisotropy. As TCRs typically bind ligand with weak-to-moderate affinities, we focus the discussion on means to enhance the accuracy and precision of low-affinity measurements. In addition to further elucidating the biology of the T cell mediated immune response, more reliable low-affinity measurements will aid with more probing studies with mutants or altered peptides that can help illuminate the physical underpinnings of how TCRs achieve their remarkable recognition properties.