title: Use of thiol-terminal silanes and heterobifunctional crosslinkers for immobilization of antibodies on silica surfaces.
abstract: A procedure for covalent immobilization of functional proteins on silica substrates was developed using thiol-terminal silanes and heterobifunctional cross-linkers. Using this procedure, a high density of functional antibodies was bound to glass cover slips and silica fibers. The amount of anti-IgG antibody immobilized was determined to be in the range of 0.66 to 0.96 ng/mm2 using radiolabeled antibody. The relative amount of IgG antigen bound by the immobilized antibody (0.37 to 0.55 mol antigen/mol antibody) was three to five times greater than other investigators have reported. In addition, the amount of protein nonspecifically adsorbed to the antibody-coated surface was further reduced by the addition of blocking agents so that nonspecific adsorption of protein antigens represented only 2-6% of the total antigen binding. With this low background, IgG antigen binding could be measured at levels as low as 150 fmol when an antigen concentration of 3 pmol/ml was applied. The process for antibody immobilization is straightforward, easy to perform, and adaptable for modifying mass quantities of biosensor components.
authors.affiliation: Geo-Centers, Inc., Newton Centre, Massachusetts 02159.
authors.forename: S K
authors.initials: SK
authors.lastname: Bhatia
authors.forename: L C
authors.initials: LC
authors.lastname: Shriver-Lake
authors.forename: K J
authors.initials: KJ
authors.lastname: Prior
authors.forename: J H
authors.initials: JH
authors.lastname: Georger
authors.forename: J M
authors.initials: JM
authors.lastname: Calvert
authors.forename: R
authors.initials: R
authors.lastname: Bredehorst
authors.forename: F S
authors.initials: FS
authors.lastname: Ligler
mesh_terms: D000906:Antibodies; D000937:Antigen-Antibody Reactions; D003432:Cross-Linking Reagents; D012822:Silicon Dioxide; D013438:Sulfhydryl Compounds
chemical_list: D000906:Antibodies; D003432:Cross-Linking Reagents; D013438:Sulfhydryl Compounds; D012822:Silicon Dioxide