title: A sol-gel-integrated protein array system for affinity analysis of aptamer-target protein interaction.
abstract: A sol-gel microarray system was developed for a protein interaction assay with high activity. Comparing to 2-dimensional microarray surfaces, sol-gel can offer a more dynamic and broad range for proteins. In the present study, this sol-gel-integrated protein array was used in binding affinity analysis for aptamers. Six RNA aptamers and their target protein, yeast TBP (TATA-binding protein), were used to evaluate this method. A TBP-containing sol-gel mixture was spotted using a dispensing workstation under high-humidity conditions and each Cy-3-labeled aptamer was incubated. The dissociation constants (K(d)) were calculated by plotting the fluorescent intensity of the bound aptamers as a function of the TBP concentrations. The K(d) value of the control aptamer was found to be 8 nM, which agrees well with the values obtained using the conventional method, electric mobility shift assay. The sol-gel-based binding affinity measurements fit well with conventional binding affinity measurements, suggesting their possible use as an alternative to the conventional method. In addition, aptamer affinity measurements by the sol-gel-integrated protein chip make it possible to develop a simple high-throughput affinity method for screening high-affinity aptamers.
authors.affiliation: Department of Biomedical Engineering, Dongguk University, Joong-gu, Seoul, Korea.
authors.forename: Ji-Young
authors.initials: JY
authors.lastname: Ahn
authors.forename: Eunkyung
authors.initials: E
authors.lastname: Kim
authors.forename: Jeehye
authors.initials: J
authors.lastname: Kang
authors.forename: Soyoun
authors.initials: S
authors.lastname: Kim
mesh_terms: D000345:Affinity Labels; D052157:Aptamers, Nucleotide / Q000737:chemistry*; D001483:Base Sequence; D005782:Gels; D008969:Molecular Sequence Data; D049329:Nanostructures; D044367:Phase Transition; D040081:Protein Array Analysis / Q000379:methods*; D035181:TATA-Box Binding Protein / Q000737:chemistry*; D015003:Yeasts / Q000737:chemistry
chemical_list: D000345:Affinity Labels; D052157:Aptamers, Nucleotide; D005782:Gels; D035181:TATA-Box Binding Protein