title: Label-free detection of DNA-binding proteins based on microfluidic solid-state molecular beacon sensor.
abstract: A solid-state molecular beacon using a gold support as a fluorescence quencher is combined with a polydimethylsiloxane (PDMS) microfluidic channel to construct an optical sensor for detecting single-stranded DNA binding protein (SSBP) and histone protein. The single-stranded DNA-Cy3 probe or double-stranded DNA-Cy3 probe immobilized on the gold surface is prepared for the detection of SSBP or histone, respectively. Due to the different quenching ability of gold to the immobilized single-stranded DNA-Cy3 probe and the immobilized double-stranded DNA-Cy3 probe, low fluorescence intensity of the attached single-stranded DNA-Cy3 is obtained in SSBP detection, whereas high fluorescence intensity of the attached double-stranded DNA-Cy3 is obtained in histone detection. The amounts of SSBP in sample solutions are determined from the degree of fluorescence recovery of the immobilized single-stranded DNA-Cy3 probe, whereas that of histone in sample solutions is determined from the degree of fluorescence quenching of the immobilized double-stranded DNA-Cy3 probe. Using this approach, label-free detection of target proteins at nanomolar concentrations is achieved in a convenient, general, continuous flow format. Our approach has high potential for the highly sensitive label-free detection of various proteins based on binding-induced conformation changes of immobilized DNA probes.
authors.affiliation: Department of Applied Chemistry, Graduate School of Engineering, Nagoya University, Japan. wjzyy988@gmail.com
authors.forename: Jun
authors.initials: J
authors.lastname: Wang
authors.forename: Daisuke
authors.initials: D
authors.lastname: Onoshima
authors.forename: Michihiko
authors.initials: M
authors.lastname: Aki
authors.forename: Yukihiro
authors.initials: Y
authors.lastname: Okamoto
authors.forename: Noritada
authors.initials: N
authors.lastname: Kaji
authors.forename: Manabu
authors.initials: M
authors.lastname: Tokeshi
authors.forename: Yoshinobu
authors.initials: Y
authors.lastname: Baba
mesh_terms: D001483:Base Sequence; D015374:Biosensing Techniques / Q000295:instrumentation*; D015342:DNA Probes / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D004268:DNA-Binding Proteins / Q000378:metabolism*; D006657:Histones / Q000378:metabolism; D046210:Microfluidic Analytical Techniques / Q000379:methods*; D015345:Oligonucleotide Probes / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D013050:Spectrometry, Fluorescence; D013997:Time Factors
chemical_list: D015342:DNA Probes; D004268:DNA-Binding Proteins; D006657:Histones; D015345:Oligonucleotide Probes