MICROMINIATURIZED IMMUNOASSAYS USING ATOMIC-FORCE MICROSCOPY AND COMPOSITIONALLY PATTERNED ANTIGEN ARRAYS

This paper combines the topographic imaging capability of the atomic f orce microscope (AFM) with a compositionally patterned array of immobi lized antigenic rabbit IgG on gold as an approach to performing immuno assays, The substrates are composed of micrometer-sized domains of IgG that are covalently linked to a photolithagraphically patterned array of a monolayer-based coupling agent. The immobilized coupling agent, which is prepared by the chemisorption of dithiobis(succinimidyl undec anoate) on gold, is separated by micrometer-sized grids of a monolayer formed from octadecanethiol (ODT), The strong hydrophobicity of the O DT adlayer, combined with the addition of the surfactant Tween 80 to t he buffer solution that is used in forming the antibody-antigen pairs, minimizes the nonspecific adsorption of proteinaceous materials to th e grid regions, This minimization allows the grids to function as a re ference plane for the AFM detection of the height increase when a comp lementary antibody-antigen pair is formed, The advantageous features o f this strategy, which include ease of sample preparation, an internal reference plane for the detection of topographic changes, and the pot ential for regeneration and reuse, are demonstrated using rabbit IgG a s an immobilized antigen and goat anti-rabbit IgG as the complementary antibody. The prospects for further miniaturization are discussed.