| “新吉富”罗非鱼免疫球蛋白单克隆抗体的制备及其在免疫效果评价上的应用 |
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| 引用本文： | 吴斌.“新吉富”罗非鱼免疫球蛋白单克隆抗体的制备及其在免疫效果评价上的应用[J].福建农林大学学报(自然科学版),2017,46(2). |
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| 作者姓名： | 吴斌 |
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| 作者单位： | 福建省水产技术推广总站/福建省淡水水产研究所,福建 福州,350002 |
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| 基金项目： | 现代农业产业技术体系建设专项，福建省公益类科研院所专项，福建省海洋与渔业厅重点项目 |
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| 摘 要： | 制备了"新吉富"罗非鱼血清免疫球蛋白IgM单克隆抗体(McAb)并进行了特性分析,以血清IgM McAb为基础,建立了IgM捕获ELISA检测方法.采用亲和层析法纯化健康"新吉富"罗非鱼IgM,纯化蛋白经SDS-PAGE检测,重链、轻链的相对分子质量分别为75~78、23~25 ku;以纯化的IgM为抗原制备IgM McAb,制备McAb杂交瘤细胞株系3株,分别命名为2H5、2D4、2B11,抗体亚型均为IgM,小鼠腹水效价分别为1.0×10~(-5)、1.0×10~(-4)、1.0×10~(-5),对IgM敏感度测定表明,2H5、2D4、2B11检测灵敏度分别为31.25、125.00、62.50 ng.以抗罗非鱼IgM McAb包被酶标板,建立IgM捕获ELISA检测方法.以无乳链球菌灭活疫苗免疫罗非鱼,受免血清按建立的IgM捕获ELISA方法检测特异性抗体,结果表明,建立的IgM捕获ELISA检测方法可应用于免疫应答抗体水平分析.
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| 关 键 词： | “新吉富”罗非鱼 免疫球蛋白IgM 单克隆抗体 ELISA检测方法 |
Preparation of McAbs against immunoglobulinin from Oreochromis niloticus and its application in immune effect analysis |
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| WU Bin.Preparation of McAbs against immunoglobulinin from Oreochromis niloticus and its application in immune effect analysis[J].Journal of Fujian Agricultural and Forestry University,2017,46(2). |
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| Authors: | WU Bin |
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| Abstract: | To prepare effective monoclonal antibody ( McAb) against serum immunoglobulin ( Ig) of tilapia, antibody capture en-zyme-linked immunosorbent assay ( ELISA) was conducted, and Ig of healthy Oreochromis niloticus was purified by protein A affinity chromatography to characterize its immunological feature. Then purified IgM was used as antigen to immunize Balb/c mice. SDS-PAGE analysis showed that molecular weight of tilapia Ig heavy chain and light chain was 75-78 and 23-25 ku, respectively. Three hybridomas strains secreting monoclonal antibodies ( McAb) were obtained and named 2H5, 2D4, and 2B11, with the common iso-type being IgM. And titers in the ascites of Balb/c mice were 1.0×10-5, 1.0×10-4, 1.0×10-5, respectively. Sensitivities of McAb against tilapia IgM was high, with detection limits being 31. 25, 125. 00, and 62. 50 ng, respectively. Furthermore, MAC-ELISA showed that specific antibody was detected within tilapia immuned by Streptococcus agalaciate inactivated vaccine. |
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| Keywords: | Oreochromis niloticus immunoglobulin monoclonal antibody ELISA |
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