title: The use of Fab-masking antigens to enhance the activity of immobilized antibodies.
abstract: This study demonstrates that masking the Feb regions of a monoclonal antibody (Mab) with synthetic antigens prior to covalent immobilization efficiency. Water-soluble adducts of poly(2-methyloxazoline) polymers and a synthetic-peptide epitope for the Mab were constructed. These synthetic antigens are referred to as Fab-masking antigents (FMAs). The antibody used in this study is a Ca(2+)-dependent murine monoclonal lgG directed against the plasma protein, human protein C (hPC). The FMAs were pre-equilibrated with Mab in the presence of calcium prior to immobilization and were then removed by EDTA, which destabilized the FMA-Mab complexes. The antigen binding efficiency and accessibility of the Fab domain of the immobilized antibody was significantly increased for Mab immobilized in the presence of FMA relative to those Mab immobilized without FMA. The increase in binding efficiency was most pronounced for the largest FMA employed. No appreciable differences were detected in the avidity of hPC-Mab complexes formed by immunosorbents produced by either masked or unmasked antibody. These results provide evidence that orientation may play an important role in the binding activity of immobilized antibodies.
authors.affiliation: Department of Chemical Engineering, Virginia Polytechnic Institute and State University, Blacksburg, Virginia 24061, USA.
authors.forename: W H
authors.initials: WH
authors.lastname: Velander
authors.forename: A
authors.initials: A
authors.lastname: Subramanian
authors.forename: R D
authors.initials: RD
authors.lastname: Madurawe
authors.forename: C L
authors.initials: CL
authors.lastname: Orthner