title: Comparison of methods for generating planar DNA-modified surfaces for hybridization studies.
abstract: The surface conformation and accessibility of oligonucleotides within arrays are two key parameters that affect the utility of immobilized nucleic acids in sensor technologies. In this work, a novel combination of analytical techniques was used to compare two methods for DNA immobilization on glass. The aim of the study was to identify a method that generated a high surface density of hybridization-accessible oligonucleotides in a true planar monolayer. The first method based on direct coupling of silanized DNA to the glass surface showed a high immobilization density of 0.013 molecules/nm2 but low surface accessibility, as shown by the hybridization measurements (< or =15%). The second method, based on the biotin-streptavidin interaction, generated a high immobilization density (0.02 molecules/nm2) and high surface accessibility (90%). Atomic force microscopy and X-ray photoelectron spectroscopy indicated that both methods achieved uniform surfaces. Using the biotin-streptavidin system, the intermolecular distance between the hybridized molecules could be tightly controlled by titrating biotinylated complementary and noncomplementary oligonucleotides.
authors.affiliation: Cardiff School of Biosciences, Biomedical Science Building, Museum Avenue, Cardiff CF10 3AX, UK. kasrya@cf.ac.uk
authors.forename: Amal
authors.initials: A
authors.lastname: Kasry
authors.forename: Paola
authors.initials: P
authors.lastname: Borri
authors.forename: Philip R
authors.initials: PR
authors.lastname: Davies
authors.forename: Adrian
authors.initials: A
authors.lastname: Harwood
authors.forename: Nick
authors.initials: N
authors.lastname: Thomas
authors.forename: Stefan
authors.initials: S
authors.lastname: Lofas
authors.forename: Trevor
authors.initials: T
authors.lastname: Dale
mesh_terms: D015374:Biosensing Techniques / Q000379:methods; D001710:Biotin / Q000737:chemistry; D019921:Biotinylation; D004247:DNA / Q000737:chemistry* / Q000235:genetics; D005898:Glass; D018625:Microscopy, Atomic Force; D008967:Molecular Biology / Q000379:methods; D009693:Nucleic Acid Hybridization; D009841:Oligonucleotides / Q000737:chemistry; D056951:Photoelectron Spectroscopy / Q000379:methods; D019809:Streptavidin / Q000737:chemistry; D013499:Surface Properties
chemical_list: D009841:Oligonucleotides; D001710:Biotin; D004247:DNA; D019809:Streptavidin