COVALENT IMMOBILIZATION OF NATIVE BIOMOLECULES ONTO AU(111) VIA N-HYDROXYSUCCINIMIDE ESTER FUNCTIONALIZED SELF-ASSEMBLED MONOLAYERS FOR SCANNING PROBE MICROSCOPY

We have worked out a procedure for covalent binding of native biomacro molecules on flat gold surfaces for scanning probe microscopy in aqueo us buffer solutions and for other nanotechnological applications, such as the direct measurement of interaction forces between immobilized m acromolecules, of their elastomechanical properties, etc. It is based on the covalent immobilization of amino group-containing biomolecules (e.g., proteins, phospholipids) onto atomically flat gold surfaces via omega-functionalized self-assembled monolayers. We present the synthe sis of the parent compound, dithio-bis(succinimidylundecanoate) (DSU), and a detailed study of the chemical and physical properties of the m onolayer it forms spontaneously on Au(lll), Scanning tunneling microsc opy and atomic force microscopy (AFM) revealed a monolayer arrangement with the well-known depressions that are known to stem from an etch p rocess during the self-assembly. The total density of the omega-N-hydr oxysuccinimidyl groups on atomically flat gold was 585 pmol/cm(2), as determined by chemisorption of C-14-labeled DSU. This corresponded to approximately 75% of the maximum density of the omega-unsubstituted al kanethiol. Measurements of the kinetics of monolayer formation showed a very fast initial phase, with total coverage within 30 s. A subseque nt slower rearrangement of the chemisorbed molecules, as indicated by AFM, led to a decrease in the number of monolayer depressions in appro ximately 60 min. The rate of hydrolysis of the omega-N-hydroxysuccinim ide groups at the monolayer/water interface was found to be very slow, even at moderately alkaline pH values. Furthermore, the binding of lo w-molecular-weight amines and of a model protein was investigated in d etail.