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| 莱克多巴胺单克隆抗体间接竞争ELISA检测方法的建立 | |
| 引用本文： | 高以明,吴艳涛,王寿利,张小荣,薛峰,陆承平.莱克多巴胺单克隆抗体间接竞争ELISA检测方法的建立[J].畜牧与兽医,2010,42(5). |
| 作者姓名： | 高以明 吴艳涛 王寿利 张小荣 薛峰 陆承平 |
| 作者单位： | 1. 南京农业大学动物医学院,江苏,南京,210095;南京出入境检验检疫局,江苏,南京,210001 2. 扬州大学兽医学院,江苏,扬州,225009 3. 江苏出入境检验检疫局,江苏,南京,210001 4. 南京农业大学动物医学院,江苏,南京,210095 |
| 摘 要： | 用混合酸酐法将莱克多巴胺(Rac)与匙孔蓝蛋白(KLH)、牛血清白蛋白(BSA)偶联,经紫外光谱扫描确定偶联成功,测得偶联物Rac-KLH浓度为3.6 mg/mL,Rac-BSA浓度为6.2 mg/mL。以偶联物Rac-KLH作为免疫原,免疫6～8周龄雌性BALB/c小鼠,取小鼠脾细胞和骨髓瘤细胞SP2/O-Ag-14进行融合。以Rac-BSA作为包被抗原,经间接ELISA筛选出阳性细胞株,再用有限稀释法进行多次亚克隆,获得稳定分泌单克隆抗体的杂交瘤细胞株1D9、1F3、2C11、5C3、3A10、4A8、6B7、6D5、7C11、7D3。其中单克隆抗体5C3和3A10间接ELISA效价1∶500 000,对莱克多巴胺的半数阻断浓度(IC50)均为3.98 ng/mL,对盐酸克伦特罗和沙丁醇胺的IC50均大于2 000 ng/mL;对盐酸克伦特罗和沙丁醇胺的交叉反应率均小于0.2%。间接竞争ELISA检测莱克多巴胺在0.5～100 ng/mL范围内为线性分布,确定的最低检测限为0.5 ng/mL。 |
| 关 键 词： | 莱克多巴胺 单克隆抗体 竞争ELISA |
Development of an indirect competitive ELISA based on monoclonal antibodies for detection of ractopamine |
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| GAO Yi-Ming,WU Yan-tao,WANG Shou-i,ZHANG Xiao-rong,XUE Feng,LU Cheng-ping.Development of an indirect competitive ELISA based on monoclonal antibodies for detection of ractopamine[J].Animal Husbandry & Veterinary Medicine,2010,42(5). | |
| Authors: | GAO Yi-Ming WU Yan-tao WANG Shou-i ZHANG Xiao-rong XUE Feng LU Cheng-ping |
| Institution: | GAO Yi-Ming1,2,WU Yan-tao3,WANG Shou-li3,ZHANG Xiao-rong3,XUE Feng4,LU Cheng-ping1(1.College of Veterinary Medicine,Nanjing Agricultural University,Nanjing 210095,China,2.Nanjing Entry Exit Inspection and Quarantine Bureau,Nanjing 210001,3.College of Veterinary Medicine,Yangzhou University,Yangzhou 225009,4.Jiangsu Entry-Exit Inspection and Quarantine Bureau,China) |
| Abstract: | Ractopamine was coupled with keyhole limpet hemocyanin(KLH) and bovine serum albumin(BSA) to prepare complete antigens Rac-KLH and Rac-BSA,respectively.The two antigens were identified by ultraviolet radiation,with the concentrations of 3.6 mg/mL and 6.2 mg/mL.The spleen cells of BALB/c mice immunized by Rac-KLH were fused with SP2 /0-Ag-14 myeloma cells using PEG4000.The Rac-BSA was used as the coating antigen for indirect enzyme linked immunosorbent assay(ELISA) to screen hybridoma cells,and limited dilut... |
| Keywords: | ractopamine monoclonal antibody competitive indirect ELISA |
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