IMMOBILIZATION OF ANTIBODIES ON A PHOTOACTIVE SELF-ASSEMBLED MONOLAYER ON GOLD

This paper presents a strategy for immobilizing biomolecules on a phot oactivable surface. A self-assembled monolayer is prepared by adsorbin g an omega-functionalized dialkyl disulfide on gold. Functional groups of this monolayer are converted in two steps into a benzophenone deri vative with an overall yield of 50 +/- 10%. Several independent techni ques (ellipsometry, X-ray photoelectron spectroscopy, scanning electro n microscopy, atomic force microscopy, radiolabel assay, and autoradio graphy) characterize the reaction and photoimmobilization of antibodie s on this surface. The photoimmobilized antibodies cover the surface a s a homogeneous and dense monolayer that could not be disrupted by vig orous washing with the detergent Tween 20. Immunoassays demonstrated s pecific recognition of the immobilized immunoglobulins as measured by their complexation with alkaline phosphatase-linked antibodies. The me thod of photoimmobilization used here leads to a homogeneous single la yer of IgGs, in which the proteins maximize their contact with the sur face. Residual adsorption of IgG on the nonirradiated surface of benzo phenone remains one limitation of this approach. Progressively higher coverages of IgGs on the surface did not lead to strictly proportional changes of the biological activity of these surfaces, probably becaus e of interactions between the IgGs in the film. This method of photoim mobilization is nonetheless useful as an experimental system to immobi lize other proteins because it is simple, flexible, and efficient.