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A Reusable and Specific Protein A‐Coated Piezoelectric Biosensor for Flow Injection Immunoassay
Author(s) -
Lu HsinChun,
Chen HsiuMei,
Lin YuSheng,
Lin JengWei
Publication year - 2000
Publication title -
biotechnology progress
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.572
H-Index - 129
eISSN - 1520-6033
pISSN - 8756-7938
DOI - 10.1021/bp9901320
Subject(s) - chemistry , chromatography , biosensor , lysis , covalent bond , immunoassay , polyclonal antibodies , protein a , matrix (chemical analysis) , adsorption , biochemistry , antibody , organic chemistry , biology , immunology
A hydrophilic matrix of periodate‐oxidized dextran was used as a double‐sided linker to covalently immobilize Staphylococcus aureus protein A (SpA) molecules onto a poly‐ L ‐lysine‐modified piezoelectric crystal surface to improve their stability, activity, and binding specificity with human immunoglobulin G (IgG) in flow injection assays. The prepared sensing crystals displayed best sensitivity and reusability at a flow rate of 140 μL/min. A human IgG concentration as low as 0.3 nM can be detected by this system. Up to 19 successive assay repetitions were achieved without significant loss of sensitivity using the same crystal. The analysis of adsorption kinetics indicates that such a preparation can greatly increase the amount of available active human IgG binding sites on immobilized SpA. Hardly any response arising from unspecific binding was detected. In addition, the sensing crystal prepared by this method was found to retain activity better than one prepared via direct deposition when stored in either wet or dry states. Finally, the prepared SpA‐coated crystals were applied to the affinity immobilization of polyclonal goat anti‐ Schistosoma japonicum glutathione‐S‐transferase (GST) and were able to subsequently detect GST and its genetically engineered mutant either in a purified form or in the crude cell lysate.