SURFACE ENGINEERING - OPTIMIZATION OF ANTIGEN PRESENTATION IN SELF-ASSEMBLED MONOLAYERS

The formation of self-assembled monolayers (SAMs) on gold surfaces con taining an antigenic peptide (NANP)(6) and HS(CH2)(11)OH, and the spec ific binding of a monoclonal antibody to these layers were investigate d by surface plasmon resonance (SPR). Peptides were synthesized by sol id-state phase synthesis and were linked either to cysteine or to an a lkyl-thiol to allow covalent attachment to gold. The content of the pe ptide in the SAMs was systematically varied, and the binding propertie s of the monoclonal antibody were compared with those measured by micr ocalorimetry in solution. At a critical peptide concentration in the S AM an optimal antibody binding and complete surface coverage was attai ned, At lower peptide concentrations, the amount of adsorbed antibody decreased; at higher peptide concentrations, the binding constant decr eased. These effects can be explained if the accessibility of the anti genic epitopes depends on the peptide density. Addition of free antige n induced the desorption of bound antibodies and allowed accurate meas urements of the dissociation rate constant. Binding constants obtained from steady-state measurements and from measurements of the kinetic r ate constants were compared.