Abstract
We show that sequential protein deposition is possible by photodeprotection of films formed from a tetraethylene-glycol functionalized nitrophenylethoxycarbonyl-protected aminopropyltriethoxysilane (NPEOC-APTES). Exposure to near-UV irradiation removes the protein-resistant protecting group, and allows protein adsorption onto the resulting aminated surface. The protein resistance was tested using proteins with fluorescent labels and microspectroscopy of two-component structures formed by micro- and nanopatterning and deposition of yellow and green fluorescent proteins (YFP/GFP). Nonspecific adsorption onto regions where the protecting group remained intact was negligible. Multiple component patterns were also formed by near-field methods. Because reading and writing can be decoupled in a near-field microscope, it is possible to carry out sequential patterning steps at a single location involving different proteins. Up to four different proteins were formed into geometric patterns using near-field lithography. Interferometric lithography facilitates the organization of proteins over square cm areas. Two-component patterns consisting of 150 nm streptavidin dots formed within an orthogonal grid of bars of GFP at a period of ca. 500 nm could just be resolved by fluorescence microscopy.
Highlights
Patterned assemblies of biomolecules have attracted a great deal of interest for applications in biosensors and in clinical diagnostics.[1−4] For example, arrays of antibodies are selected to bind antigens that are markers for disease
In the present work we describe how sequential deprotection of OEGNPEOC-APTES using both interferometric and near-field methods followed by protein adsorption facilitates the fabrication of multiple component submicrometer protein patterns
Films formed by the adsorption of OEG-NPEOC-APTES demonstrate exceptional protein-resistance, enabling the execution of multiple lithographic processing steps with little diminution of their performance
Summary
Patterned assemblies of biomolecules have attracted a great deal of interest for applications in biosensors and in clinical diagnostics.[1−4] For example, arrays of antibodies are selected to bind antigens that are markers for disease. Photochemical methods are an attractive route to the patterning of biological interactions at surfaces,[46−55] offering the capacity to execute specific chemical transformations through strategies such as the use of nitrophenyl protecting groups,[56−64] and they have been found to facilitate the formation of multiple-component biological assemblies at micrometer length scales.[51] Photopatterning at the nanometer scale is feasible through the use of near-field optical methods[31,41,42,65] and through the use of interferometric lithography.[26,66] In the present study we demonstrate that both methods may be applied to enable selective deprotection of (methoxyheptaethylene glycol)nitrophenylethoxycarbonylprotected aminopropyltriethoxysilane ( OEGNPEOC-APTES), an aminosilane bearing a photoremovable nitrophenyl group derivatized with an oligo(ethylene glycol) adduct.[61] Intact OEG-NPEOC-APTES has been shown previously to be highly protein resistant.[31,61] on exposure to UV light, the nitrophenyl protecting group is removed, exposing an amine group (Scheme 1). Samples stored in HEPES buffer (pH 7.4) were rinsed with 100 mM ammonium acetate solution (pH 7.4) and dried by nitrogen gas, to eliminate deposits from the buffer remaining at the surface after protein adsorption. Ammonium acetate is volatile and leaves no residue, and effectively displaces materials deposited from buffer solutions
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