title: Novel approach to affinity chromatography using "weak" monoclonal antibodies.
abstract: Affinity purification generally relies on specific high-affinity recognition between two species of biological molecules: one molecular species (the ligate) dissolved in a mobile phase is selectively adsorbed to the other species (the ligand) coupled to a solid support. Desorption of the ligate often requires harsh conditions that degrade biological activity of the purified product. As an alternative to this general procedure, we have studied affinity chromatography in a weak affinity mode, where ligand-ligate interactions are in dynamic equilibrium. Ligates recognized with low affinities (dissociation constant greater than 10(-4) M) elute from affinity columns under mild, isocratic conditions as retarded peaks, separated from noninteracting solutes that elute in the void volume. To illustrate the procedure, we report chromatography of an oligosaccharide on a 2-ml column containing 86 mg of a monoclonal antibody coupled to 10-micron microparticulate silica particles. Using a temperature-sensitive antibody, we observed that when the ligand-ligate dissociation constant is greater than 10(-3) M, performance of the system exceeds 300 theoretical plates/10 cm column length and approaches the efficiencies generally associated with high-performance liquid chromatography.
authors: Perstorp Biolytica AB, Lund, Sweden.
authors: S
authors: S
authors: Ohlson
authors: A
authors: A
authors: Lundblad
authors: D
authors: D
authors: Zopf
mesh_terms: D000911:Antibodies, Monoclonal; D002241:Carbohydrates / Q000302:isolation & purification; D002846:Chromatography, Affinity / Q000379:methods*; D008024:Ligands; D013696:Temperature
chemical_list: D000911:Antibodies, Monoclonal; D002241:Carbohydrates; D008024:Ligands