Focal Molography Allows for Affinity and Concentration Measurements of Proteins in Complex Matrices with High Accuracy.

ABSTRACT: Characterizing biomolecular receptor-ligand interactions is critical for research and development. However, performing analyses in complex, biologically relevant matrices, such as serum, remains challenging due to non-specific binding that often impairs measurements. Here, we evaluated Focal Molography (FM) for determining K_D and kinetic constants in comparison to gold-standard methods using single-domain heavy-chain antibodies in various systems. FM provided kinetic constants highly comparable to SPR and BLI in standard buffers containing blocking proteins, with K_Ds of soluble CD4 (sCD4) interactions within a 2.4-fold range across technologies. In buffers lacking blocking proteins, FM demonstrated greater robustness against non-specific binding and rebi

SUBMITTER: Dirscherl L 

PROVIDER: S-EPMC11853488 | biostudies-literature | 2025 Jan

REPOSITORIES: biostudies-literature