This paper investigates three different approaches to patterning proteins within ultrathin resist layers formed from self-assembled monolayers using scanning probe lithography (SPL) at the submicrometer length scale. The first approach uses a “nanografting” method to pattern a reactive carboxylic acid terminated thiol into a resist composed of a methyl-terminated monolayer. Rabbit IgG antigen is bound to the patterned region, and an immunoassay utilizing direct readout of the topographic change resulting from specific binding of anti-rabbit IgG antibody is performed using scanning force microscopy. To address issues related to nonspecific protein adsorption, the other two approaches investigated the patterned removal of glycol-terminated monolayers by mechanically “scraping” patterns at high tip−sample forces by SPL. Protein attachment to the scraped regions was achieved either through the chemisorption of a disulfide coupling agent or by the direct adsorption of Fab‘-SH antibody fragments. Results obtained from all approaches are presented and compared, and the strengths and weaknesses of each toward fabricating high-density, multiple protein arrays are discussed.

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This paper describes a new strategy for the creation of chemically modified carbonaceous stationary phases. The strategy exploits the electroreduction of arenediazonium salts as a means for functionalizing the surface of glassy carbon (GC) and porous graphitic carbon (PGC) stationary phases. The one-electron reduction of these salts forms an arene radical which then couples via a carbon-carbon linkage to the carbon framework at the surface of the stationary phase. Two arenediazonium-based modifiers were used in evaluating the potential utility of this strategy: 4-nitrobenzenediazonium tetrafluoroborate for the GC and PGC phases and 4-hexylbenzenediazonium tetrafluoroborate for only the PGC phases. Modifications were carried out by packing the phases into a column used for electrochemically modulated liquid chromatography. The effectiveness of the modifications was assessed by X-ray photoelectron spectroscopy and by comparing the liquid separation of a series of mixtures before and after coating deposition. For the nitrobenzyl-modified GC phase, the test mixture contained both anisole and fluoranthene. The performance of the nitrobenzyl- and hexylbenzyl-modified PGC stationary phases was characterized by the separations of substituted phenols (i.e., nitrophenol and resorcinol) and a few important pharmaceutical agents (i.e., hexobarbital, oxazepam, and nitrazepam). The potential utility of this modification procedure to form stationary phases that are stable upon extended exposure to aggressive mobile phases is discussed and briefly examined.