Surfaces for tuning of oligonucleotide biosensing selectivity based on surface-initiated atom transfer radical polymerization on glass and silicon substrates.
Covalently immobilized mixed films of oligonucleotide and oligomer components on glass and silicon surfaces are reported. This work has investigated how such films can improve selectivity for the detection of multiple base-pair mismatches. The intention was to introduce a "matrix isolation" effect on oligonucleotide probe molecules by surrounding the probes with oligomers, thereby reducing oligonucleotide-to-oligonucleotide and/or oligonucleotide-to-surface interactions. Thiol-functionalized oligonucleotide was coupled onto (3-aminopropyl)trimethoxysilane (APTMS) via a heterobifunctional linker, succinimidyl 4-[N-maleimidomethyl]cyclohexane-1-carboxylate (sulfo-SMCC). Using a variety of monomers such as 2-hydroxyethyl methacrylate (HEMA), oligomers were grown by surface-initiated atom transfer radical polymerization (ATRP) from a bromoisobutyryl NHS ester initiator which was immobilized onto APTMS sites that coated glass and oxidized silicon substrates. Various surface modification steps on silicon substrates were characterized by ellipsometry, wettability, atomic force microscopy, X-ray photoelectron spectroscopy, and time-of-flight secondary ion mass spectrometry. Polymerized HEMA (PHEMA) in mixture with oligonucleotide probes was evaluated for fluorescence transduction of hybridization. The presence of PHEMA was found to provide a sharper melt curve for hybrids containing both fully complementary and three base-pair mismatched targets, and this surface derivatization also minimized non-selective adsorption. The maximum increase in slope was improvement by a factor of 3-fold. An increase of up to 30% in difference of melting temperatures between fully complementary and 3 base-pair mismatched targets was achieved using PHEMA. The results suggest that the presence of oligomers dispersed among DNA hybrids can improve selectivity through what is believed to be a reduction of dispersity of interactions of probes with targets, and probes within their local environment at a surface.
Chemical Sensors Group, University of Toronto Mississauga, 3359 Mississauga Rd. N., Mississauga, ON, Canada L5L 1C6.
April K Y
AK
Wong
Ulrich J
UJ
Krull
D000327:Adsorption; D001547:Benzaldehydes / Q000737:chemistry; D015374:Biosensing Techniques / Q000379:methods*; D003432:Cross-Linking Reagents / Q000737:chemistry; D005453:Fluorescence; D005898:Glass / Q000737:chemistry*; D007700:Kinetics; D008689:Methacrylates / Q000737:chemistry; D009691:Nucleic Acid Denaturation; D009841:Oligonucleotides / Q000032:analysis* / Q000737:chemistry* / Q000378:metabolism; D010084:Oxidation-Reduction; D011108:Polymers / Q000737:chemistry*; D011437:Propylamines / Q000737:chemistry; D012680:Sensitivity and Specificity; D012821:Silanes / Q000737:chemistry; D012825:Silicon / Q000737:chemistry*; D013499:Surface Properties; D044366:Transition Temperature
D001547:Benzaldehydes; D003432:Cross-Linking Reagents; D008689:Methacrylates; D009841:Oligonucleotides; D011108:Polymers; D011437:Propylamines; D012821:Silanes; C088294:3-aminopropyltrimethoxysilane; C005044:hydroxyethyl methacrylate; C032175:benzaldehyde; D012825:Silicon