title: Multiresidue Method for Analysis of β Agonists in Swine Urine by Enzyme Linked Receptor Assay Based on β2 Adrenergic Receptor Expressed in HEK293 Cells.
abstract: A novel enzyme-linked receptor assay (ELRA) based on β2-adrenergic receptor (β2-AR) has been developed for rapid and high-throughput detection of β-adrenergic agonists (β-agonists) in urine. Human embryonic kidney cells (HEK293) were introduced as the expression system to enhance the functionality of the recombinant β2-AR, and the attempt to detect β-agonists in swine urine using such approaches was accomplished unprecedentedly. In this article, a recombinant porcine β2-AR was produced in the inner membrane of HEK293 cells and purified from crude membrane protein by nickel-nitrilotriacetic acid affinity chromatography. After activity identification, the recombinant receptor was used in the development of direct competitive ELRA. Several parameters such as blocking buffer and blocking process were optimized and the performance of the system was determined. The IC50 concentrations of clenbuterol, salbutamol, and ractopamine were 34, 53 and 63 μg/L, and the average recovery rates were 68.2%, 60.3% and 65.5%, respectively. ELRA based on β2-AR shows a series of advantages such as safety, easy operation, and high efficiency, making it promising for the rapid screening of β-agonists in animal urine.
authors.affiliation: Institute of Quality Standards and Testing Technology for Agro-products of CAAS, Key Laboratory of Agro-Product Quality and Safety, Ministry of Agriculture, Beijing, 100081, P. R. China; Department of Food Science, Hebei North University, Zhangjiakou, 075000, P. R. China.
authors.forename: Jian
authors.initials: J
authors.lastname: Wang
authors.affiliation: Institute of Quality Standards and Testing Technology for Agro-products of CAAS, Key Laboratory of Agro-Product Quality and Safety, Ministry of Agriculture, Beijing, 100081, P. R. China.
authors.forename: Yongxin
authors.initials: Y
authors.lastname: She
authors.affiliation: Institute of Quality Standards and Testing Technology for Agro-products of CAAS, Key Laboratory of Agro-Product Quality and Safety, Ministry of Agriculture, Beijing, 100081, P. R. China.
authors.forename: Miao
authors.initials: M
authors.lastname: Wang
authors.affiliation: Institute of Quality Standards and Testing Technology for Agro-products of CAAS, Key Laboratory of Agro-Product Quality and Safety, Ministry of Agriculture, Beijing, 100081, P. R. China.
authors.forename: Maojun
authors.initials: M
authors.lastname: Jin
authors.affiliation: Institute of Quality Standards and Testing Technology for Agro-products of CAAS, Key Laboratory of Agro-Product Quality and Safety, Ministry of Agriculture, Beijing, 100081, P. R. China.
authors.forename: Yongfei
authors.initials: Y
authors.lastname: Li
authors.affiliation: Institute of Quality Standards and Testing Technology for Agro-products of CAAS, Key Laboratory of Agro-Product Quality and Safety, Ministry of Agriculture, Beijing, 100081, P. R. China.
authors.forename: Jing
authors.initials: J
authors.lastname: Wang
authors.affiliation: Department of Food Science, Hebei North University, Zhangjiakou, 075000, P. R. China.
authors.forename: Yuan
authors.initials: Y
authors.lastname: Liu
mesh_terms: D058666:Adrenergic beta-2 Receptor Agonists / Q000378:metabolism / Q000494:pharmacology* / Q000652:urine*; D000818:Animals; D004798:Enzymes / Q000378:metabolism*; D015870:Gene Expression; D057809:HEK293 Cells; D057166:High-Throughput Screening Assays; D006801:Humans; D057230:Limit of Detection; D018343:Receptors, Adrenergic, beta-2 / Q000235:genetics* / Q000378:metabolism*; D011994:Recombinant Proteins / Q000378:metabolism; D013552:Swine; D016482:Urinalysis / Q000379:methods*
chemical_list: D058666:Adrenergic beta-2 Receptor Agonists; D004798:Enzymes; D018343:Receptors, Adrenergic, beta-2; D011994:Recombinant Proteins