title: Assessment of heterogeneity in antibody-antigen displacement reactions.
abstract: The intrinsic binding characteristics of monoclonal antibodies are modified upon immobilization onto a solid-phase matrix. Factors such as the distribution in affinity must therefore be taken into consideration in order to predict the kinetics of antibody binding at solid-liquid interfaces. A mathematical analysis is presented herein that allows the assessment of heterogeneity in the affinity of monoclonal antibodies immobilized onto a solid support. This model is based on a modified version of the Sips distribution function adapted to the conditions of a solid-phase displacement assay in flow. An assay for trinitrotoluene (TNT) provides the data to evaluate the extent of heterogeneity introduced by immobilization of antibodies in a flow immunoassay. We determined the index of antibody heterogeneity on two solid supports, controlled-pore glass beads and agarose beads, coated with a monoclonal anti-TNT antibody at varying densities. The data confirm that the threshold for crossover from homogeneous to heterogeneous forms of the reaction isotherm is different in displacement reactions than in association-dissociation reactions. Our analysis shows that the measured displacement isotherm is consistent with a homogeneous or only moderately heterogenous distribution of relative affinities.
authors.affiliation: Bioengineering Program, Hofstra University, Hempstead, New York 11550, USA.
authors.forename: S Y
authors.initials: SY
authors.lastname: Rabbany
authors.forename: R
authors.initials: R
authors.lastname: Piervincenzi
authors.forename: L
authors.initials: L
authors.lastname: Judd
authors.forename: A W
authors.initials: AW
authors.lastname: Kusterbeck
authors.forename: R
authors.initials: R
authors.lastname: Bredehorst
authors.forename: K
authors.initials: K
authors.lastname: Hakansson
authors.forename: F S
authors.initials: FS
authors.lastname: Ligler
mesh_terms: D000911:Antibodies, Monoclonal / Q000737:chemistry*; D000937:Antigen-Antibody Reactions; D007118:Immunoassay; D007700:Kinetics
chemical_list: D000911:Antibodies, Monoclonal