title: Coupling purification and in situ immobilization process of monoclonal antibodies to clenbuterol for immunosensor application.
abstract: Clenbuterol (CL), which promotes the growth of muscular tissue and the reduction of body fat in pigs and cattle, has been confirmed to be a potential hazard to human health. In this study, a monoclonal antibody to clenbuterol (CL mAb) from a hybridoma culture supernatant was purified by an aqueous two-phase system (ATPS) at different polyethylene glycol (PEG) concentrations, PEG molecular weights, pH values, and NaCl concentrations. Then the CL mAb was immobilized in situ by directly adding polystyrene microspheres (PSMSs) into a PEG phase containing CL mAb. Using the immobilized antibody, an immunosensor was constructed to detect the CL residues in pork samples. The results showed that using an ATPS composed of 15% (w/w) PEG6000, 15% (w/w) phosphate, and 15% (w/w) NaCl at pH 8.0, the partition coefficient was 7.24, the activity recovery was 87.86%, and the purification fold was 2.88. The PEG-CL mAb-PSMS retained approximately 98% of its initial activity after 30-ml phosphate buffer (PBS) washings. After 30days of storage, the CL mAb-PSMS lost nearly 75% of its activity, whereas the PEG-CL mAb-PSMS retained as much as 95% of its initial activity. Furthermore, the constructed immunosensor obtained recoveries of 90.5 to 102.6% when applied to pork samples spiked with CL.
authors: School of Medical Instruments and Food Engineering, University of Shanghai for Science and Technology, Shanghai 200093, People's Republic of China.
authors: Hui
authors: H
authors: Cao
authors: School of Medical Instruments and Food Engineering, University of Shanghai for Science and Technology, Shanghai 200093, People's Republic of China.
authors: Min
authors: M
authors: Yuan
authors: School of Medical Instruments and Food Engineering, University of Shanghai for Science and Technology, Shanghai 200093, People's Republic of China.
authors: Lili
authors: L
authors: Wang
authors: School of Medical Instruments and Food Engineering, University of Shanghai for Science and Technology, Shanghai 200093, People's Republic of China.
authors: Jingsong
authors: J
authors: Yu
authors: School of Medical Instruments and Food Engineering, University of Shanghai for Science and Technology, Shanghai 200093, People's Republic of China. Electronic address: xufei8135@126.com.
authors: Fei
authors: F
authors: Xu
mesh_terms: D000818:Animals; D000911:Antibodies, Monoclonal / Q000302:isolation & purification*; D002417:Cattle; D002976:Clenbuterol / Q000032:analysis* / Q000276:immunology*; D006801:Humans; D011092:Polyethylene Glycols / Q000737:chemistry
chemical_list: D000911:Antibodies, Monoclonal; D011092:Polyethylene Glycols; D002976:Clenbuterol
keywords: Aqueous two-phase system; Clenbuterol detection; Immunosensor; In situ immobilization; Monoclonal antibody to clenbuterol