Functional characterisation of Fab′-fragments self-assembled onto hydrophilic gold surfaces

Antibody Fab'-fragments have been immobilised on hydrophilic gold by direct self-assembly, and embedded in a matrix of non-ionic hydrophilic polymers, tris(hydroxymethyl)methylacrylamide, carrying lipoate terminal linking groups. Different polymers were synthesised, and co-adsorbed or post-adsorbed between the antibody fragments in order to optimise the antigen binding. Various factors were investigated that influence the activity of the immobilised Fab'-fragments for binding of the antigen, human IgG. The Fab'-fragments were immobilised in dense layers close to monolayer coverage, and the stoichiometric efficiency of immobilisation was up to 30%, with the human IgG also approaching monolayer coverage. The cleaning of the gold surface was a crucial factor in preservation of activity. Besides the usual treatment in hot ammonia/peroxide Solution, hot DMSO appeared to be highly effective as a cleaning agent. (C) 2008 Elsevier B.V. All rights reserved.