Insights into weak affinity antibody-antigen interactionsStudies using affinity chromatography and optical biosensor

Molecular recognition and interaction is fundamental for the function of biological systems. The properties of the interacting biomolecules dictate the type of forces involved and the strength and dynamics of the interaction. Many interactions are very strong whereas others exhibit weak affinity. Typically, weak interactions work in concert to trigger a biological response. The advantage with this approach is the inherent dynamics. It has been shown that this approach can be successful for in vitro applications as well. By exposing analytes to a multitude of specific, weak affinity interactions, which are governed by fast association and dissociation rates, separation based on small differences in affinity is possible. The same principles can also be used to characterize biological weak affinity binders and for analytical purposes. This thesis has discussed biomolecular interactions in the weak affinity range (defined in this investigation as dissociation constants (KD) larger than 0.01 mM) in general and has focuses on how they can be studied and exploited in vitro. Weak affinity monoclonal antibodies were used as model systems in three different applications: (i) to explore how weak affinity chromatography based on monoclonal IgM can be employed to separate structurally related steroids under non-denaturing conditions. (ii) to investigate the possibilities of studying weak interactions between antibodies and haptens with a real-time optical biosensor based on surface plasmon resonance. A rationale for the design of such experiments to avoid inaccurate results was suggested (iii) to introduce continuous real-time immunosensing for monitoring fluctuating concentrations of biomolecules in a flow