Preparation of Novel Monoclonal Antibodies Against Chelated Cadmium Ions
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Preparation of Novel Monoclonal Antibodies Against Chelated Cadmium Ions Tao Kong & Xue-Qin Hao & Xiao-Bing Li & Guo-Wen Liu & Zhi-Gang Zhang & Zhi-Jun Yang & Zhe Wang & Jia Tang & Wei Yang & Jia Sun
Received: 18 September 2012 / Accepted: 1 January 2013 / Published online: 2 February 2013 # Springer Science+Business Media New York 2013
Abstract The detection of cadmium ions using enzymelinked immunosorbent assays (ELISA) has been reported by several research groups. Because cadmium ions are too small to stimulate the immune system, high molecular weight immunogens of cadmium are constructed using bifunctional chelators. At present, the most commonly used bifunctional chelator for the preparation of antigens for heavy metal ions is 1-(4-isothiocyanobenzyl) ethylenediamine N,N,N′,N′-tetraacetic acid (ITCBE). However, the price of ITCBE is high. So we are interested in a cheaper bifunctional chelator, 1-(4-aminobenzyl) ethylenediamine N,N,N′,N′-tetraacetic acid (aminobenzyl-EDTA). Here, cadmium ions were conjugated to carrier proteins using aminobenzyl-EDTA to make artificial antigens. Then, sev-
eral mice were immunized with the antigen. And monoclonal antibodies (MAbs) against cadmium were produced. Spleen cells of immunized mice were fused with myeloma cells. The resulting hybridomas were screened using protein conjugates which were covalently bound to metal-free EDTA or cadmium. Three hybridoma cell lines (A3, E4 and B5) that produced MAbs with high selectivity and sensitivity were expanded for further study. Crossreactivities with other metals were below 1 %. These antibodies were used to construct competitive ELISAs. The IC50 for A3 was 8.4 μg/l. The detection range and the lowest detection limit using the antibody A3 was 0.394–64.39 and 0.051 μg/l, respectively. Spike–recovery studies in tap water showed that the antibody A3 could be used for cadmium detection in drinking water.
Authors Tao Kong, Xue-Qin Hao, and Xiao-Bing Li of Jilin University and Henan University of Science and Technology contributed equally to this work.
Keywords Cadmium . Bifunctional chelator . Antigen . Monoclonal antibody
T. Kong : X.-Q. Hao : Z.-J. Yang College of Animal Science and Veterinary Medicine, Henan University of Science and Technology, 70 Tianjin Road, Luoyang, Henan 471003, People’s Republic of China X.-B. Li (*) : G.-W. Liu (*) : Z. Wang : J. Tang : W. Yang Laboratory of Animal Nutritional and Metabolic Diseases & Toxicopathy, College of Animal Science and Veterinary Medicine, Jilin University, 5333 Xian Road, Changchun, Jilin 130062, People’s Republic of China e-mail: [email protected] e-mail: [email protected] Z.-G. Zhang College of Veterinary Medicine, Northeast Agriculture University, Harbin 150030, People’s Republic of China J. Sun Karolinska Institutet Huddinge, Novum, plan 8 Baxter Novum, 141 86 Stockholm, Sweden
Abbreviations AminobenzylEDTA EDTA MAb BSA KLH HAT HT FBS OPD ELISA PEG Ka CR
1-(4-Aminobenzyl) ethylenediamine N,N,N′,N′-tetraacetic acid N,N,N′,N′-Ethylenediamine-tetraaceticacid Monoclonal a
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