摘要
琥珀酸酐法将雌二醇（E2）衍生化,再用碳二亚胺法（EDC）将半抗原与BSA和OVA偶联,制备免疫原和检测抗原,紫外扫描和红外光谱鉴定表明人工抗原合成成功,E2与BSA偶联比为12.3∶1。将E2-BSA免疫BALB/c小鼠,细胞融合技术筛选抗E2杂交瘤细胞株,制备单克隆抗体。结果表明,筛选的3株杂交瘤细胞E2D5、E3F7和E4A6抗体效价均在8×104以上。以E3F7细胞株建立间接竞争ELISA（icELISA）标准曲线,其线性范围为0.06~66.2μg/L,检测限和IC50值分别为0.03和0.76μg/L,除与去氢甲睾酮22.9%的交叉反应率外,与其他化合物无交叉反应。本研究为开发ELISA试剂盒,检测食源性动物产品中E2残留奠定基础。
Succinic anhydride was employed to synthesis estradiol （E2） derivative,which was con- jugated to BSA as immunogen and to OVA as coating antigen,by the EDC method. The results of IR and UV-visible spectrum indicated that the artificial antigen was synthesized successfully and the conjugation ratio of E2-BSA was 12.3：1. E2-BSA was introduced to immunize BALB/c mice, and cell fusion technology was employed to screen hybridoma cell lines and produce anti-E2 mono- clonal antibodies. Finally,Three hybridoma cells named E2D5,E3F7 and E4A6 were screened out, which antibody titers were more than 8 × 10^4. Based on the E3F7 hybridoma,an icELISA standard curve was developed,and the dynamic range was from 0.06 to 66.2 μg/L,with a detection limit and IC50 values of 0.03 and 0.76 μg/L,respectively. No cross-reaction with other compounds was observed except 22.9% for dehydrogenation. Therefore, this research put a potential for the development of ELISA kits and provide an alternative method for the detection of E2 residues in food producing animals.
出处
《中国兽医学报》
CAS
CSCD
北大核心
2014年第5期772-775,780,共5页
Chinese Journal of Veterinary Science
基金
国家自然科学基金资助项目(U1204310)
河南省教育厅科技攻关项目(2011A230003)
