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The reaction between the arginine's guanidino group and α-dicarbonyl functionalities was used to develop a novel surface modification chemistry. The first modification step consists of the electrostatic adsorption of a poly-l-arginine layer onto ionizable alkanethiol modified gold surfaces. The strongly basic character of the guanidino group of the arginine residues (pKa > 12) guarantees the robust attachment of the polypeptide to negatively charged gold surfaces until very high pH. By varying the pH of the solution from which poly-l-arginine is electrostatically adsorbed, it is possible to control the amount deposited. The availability of the surface guanidino groups of the poly-l-arginine layer for further derivatization with α-dicarbonyl reaction probes, yielding stable heterocyclic condensation adducts, is demonstrated. In addition, the reaction with the heterobifunctional reagent p-azidophenyl glyoxal (APG) provides a surface terminated with a photosensitive aryl azide group which was employed for the photochemical immobilization of proteins to the surface. The application of this surface modification chemistry to immobilize antibodies is demonstrated.

A novel surface modification procedure for the creation of sulfhydryl-terminated alkanethiol monolayers that can be used for the attachment of biomolecules onto gold surfaces is described. A self-assembled monolayer of the amine-terminated alkanethiol 11-mercaptoundecylamine (MUAM) is reacted with the heterobifunctional cross-linker N-succinimidyl S-acetylthiopropionate (SATP) in order to create a protected sulfhydryl-terminated monolayer. This monolayer can then be deprotected in an alkaline solution to create an active sulfhydryl surface. Compounds that have been modified to contain a maleimide moiety can be easily attached onto the sulfhydryl-derivatized gold surface. In a second attachment strategy, the sulfhydryl-terminated monolayer is reacted with 2,2‘-dipyridyl disulfide to form disulfide bonds on the surface. These disulfide bonds are then used in a thiol−disulfide exchange reaction with free sulfhydryls in order to attach biomolecules, such as thiol-modified DNA or cysteine-containing polypeptides, onto the surface. In contrast to the maleimide-attached monolayers, the disulfide-immobilized species can be cleaved in the presence of dithiothreitol (DTT) in order to regenerate the free sulfhydryl surface. Polarization modulation FTIR reflection−absorption spectroscopy (PM-FTIRRAS) has been used to characterize these surface reactions, and fluorescence “wash off” measurements provided an estimate of 1.5 × 10¹² molecules/cm² for the surface coverage of DNA immobilized using a thiol−disulfide exchange reaction. Surface plasmon resonance (SPR) imaging measurements were employed to monitor in situ hybridization onto DNA arrays fabricated using this surface immobilization reaction.