To investigate the effect of antibody orientation on its immunological activities, we developed a novel and versatile platform consisting of a well-defined phospholipid polymer surface on which staphylococcal protein A (SpA) was site-selectively immobilized. The phospholipid polymer platform was prepared on silicon substrates using the surface-initiated atom transfer radical polymerization technique. An enzymatic reaction was performed for orientation-selective coupling of SpA molecules to the polymer brush surface. Orientation-controlled antibodies were achieved using enzymatic reactions, and these antibodies captured 1.8 antigens on average. Theoretical multivalent binding analysis further revealed that orientation-controlled antibodies had antigen-antibody reaction equilibrium dissociation constants (Kd) as low as 8.6 x 10-10 mol/L, whereas randomly oriented and partially oriented antibodies showed Kd values of 2.0 x 10-7 and 1.2 x 10-7 mol/L, respectively. These findings support the significance of antibody orientation because controlling the orientation resulted in high reactivity and theoretical binding capacity.

Well-oriented antibodies on phospholipid polymer platform recorded high binding affinity

243rd National Meeting (2012)

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