title: Preparation of scFv-immobilized quartz crystal microbalance sensor by PS-tag-mediated solid-phase refolding.
abstract: The activation of a single-chain Fv antibody on the surface of a quartz crystal microbalance (QCM) sensor chip was investigated in order to develop an economical and sensitive immuno-QCM sensor system for use in clinical diagnosis. On the bare gold surface of a QCM sensor chip, approximately 60nm of hydrophilic polystyrene (phi-PS) thin film that specifically binds the polystyrene-binding peptide (PS-tag) was prepared by spin-coating and O(2)-plasma irradiation. When the adsorption of PS-tag-fused anti-ED-B scFv (scFv-PS) onto the phi-PS surface was directly monitored, the maximum density of scFv-PS attained was 1.56μg/cm(2), 1.6-times higher than that of scFv. The specific antigen-binding activity of scFv-PS after solid-phase refolding increased with the density of immobilized scFv-PS, and, consequently, activity 1.7 times higher than that of scFv was retained. The scFv-PS-immobilized QCM sensor chip rapidly allowed the detection of clear signals for antigen at the range of 0.1-10μg/ml, while no signal was detectable for 10μg/ml BSA as a negative control. The scFv-PS-immobilized QCM sensor developed in the present study will therefore be very useful for the rapid and highly sensitive detection of biomarkers, and should be applicable to clinical diagnosis.
authors.affiliation: Department of Chemistry and Materials Technology, Kyoto Institute of Technology, Hashigami-cho, Kyoto 606-8585, Japan. kumada@chem.kit.ac.jp
authors.forename: Yoichi
authors.initials: Y
authors.lastname: Kumada
authors.forename: Eiju
authors.initials: E
authors.lastname: Sasaki
authors.forename: Michimasa
authors.initials: M
authors.lastname: Kishimoto
mesh_terms: D055910:Antibodies, Immobilized / Q000737:chemistry* / Q000276:immunology; D000937:Antigen-Antibody Reactions; D000941:Antigens / Q000032:analysis; D015374:Biosensing Techniques / Q000379:methods*; D011137:Polystyrenes / Q000737:chemistry*; D017510:Protein Folding; D058525:Quartz Crystal Microbalance Techniques; D057127:Single-Chain Antibodies / Q000737:chemistry* / Q000276:immunology
chemical_list: D055910:Antibodies, Immobilized; D000941:Antigens; D011137:Polystyrenes; D057127:Single-Chain Antibodies