Direct oriented immobilization of F(ab′) antibody fragments on gold

Abstract The preparation of oriented immunosurfaces by the direct immobilization of F(ab′) fragments of rabbit anti-calf alkaline phosphatase (RACAP) Immunoglobulin G (IgG) onto gold (Au) surfaces via the formation of a Au-thiolate bond is described. In order to compare specific and random immobilization methods, F(ab′)-modified Au is compared to immunosurfaces of non-specifically adsorbed F(ab) fragments of RACAP IgG. The immobilization chemistries of the resulting antibody-modified Au surfaces are characterized with X-ray photoelectron spectroscopy (XPS). Antigen binding is characterized using both a quartz crystal microbalance (QCM) and a solid phase enzyme linked immunosorbent assay (ELISA). We demonstrate that specifically immobilized F(ab′) fragments can be used to produce immunosurfaces with higher antigen-binding efficiencies than corresponding surfaces modified with randomly immobilized antibody fragments. The relationship between antigen-binding efficiency and the surface coverage of immobilized antibody fragments is also discussed.

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