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Please use this identifier to cite or link to this item: http://hdl.handle.net/1942/8438

Title: Topographical and functional characterization of the ssDNA probe layer generated through EDC-mediated covalent attachment to nanocrystalline diamond using fluorescence microscopy
Authors: VERMEEREN, Veronique
DAENEN, Michael
VAN DE VEN, Martin
WAGNER, Patrick
Issue Date: 2008
Citation: LANGMUIR, 24(16). p. 9125-9134
Abstract: The covalent attachment method for DNA on nanocrystalline diamond (NCD), involving the introduction of COOH functionalities on the surface by photoattachment of 10-undecenoic acid (10-UDA), followed by the 1-ethyl-3-(3-dimethylaminopropyl)-carbodiimide (EDC)-mediated coupling to NH2-labeled ssDNA, is evaluated in terms of stability, density, and functionality of the resulting biological interface. This is of crucial importance in DNA biosensor development. The covalent nature of DNA attachment will infer the necessary stability and favorable orientation to the ssDNA probe molecules. Using confocal fluorescence microscopy, the influence of buffer type for the removal of excess 10-UDA and ssDNA, the probe ssDNA length, the probe ssDNA concentration, and the presence of the COOH-linker on the density and functionality of the ssDNA probe layer were investigated. It was determined that the most homogeneously dense and functional DNA layer was obtained when 300 pmol of short ssDNA was applied to COOH-modified NCD samples, while H-terminated NCD was resistant for DNA attachment. Exploiting this surface functionality dependence of the DNA attachment efficiency, a shadow mask was applied during the photochemical introduction of the COOH-functionalities, leaving certain regions on the NCD H-terminated. The subsequent DNA attachment resulted in a fluorescence pattern corresponding to the negative of the shadow mask. Finally, NCD surfaces covered with mixtures of the 10-UDA linker molecule and a similar molecule lacking the COOH functionality, functioning as a lateral spacer, were examined for their suitability in preventing nonspecific adsorption to the surface and in decreasing steric hindrance. However, purely COOH-modified NCD samples, patterned with H-terminated regions and treated with a controlled amount of probe DNA, proved the most efficient in fulfilling these tasks.
Notes: Hasselt Univ & Transnatl Univ Limburg, Biomed Res Inst, Sch Life Sci, B-3590 Diepenbeek, Belgium. Hasselt Univ & Transnatl Univ Limburg, Inst Mat Res, Sch Life Sci, B-3590 Diepenbeek, Belgium. IMEC VZW, Div IMOMEC, B-3590 Diepenbeek, Belgium.
URI: http://hdl.handle.net/1942/8438
DOI: 10.1021/la800946v
ISI #: 000258377900110
ISSN: 0743-7463
Category: A1
Type: Journal Contribution
Validation: ecoom, 2009
Appears in Collections: Research publications

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