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| 糖化酶单抗的研制及其抗原识别位点的分析 | |
| 引用本文： | 张丹,姜威,李双喜,汪惠泽,冯云飞,刘云迎,李东明,何祥来,王捍东.糖化酶单抗的研制及其抗原识别位点的分析[J].中国畜牧兽医,2012,39(11):39-42. |
| 作者姓名： | 张丹 姜威 李双喜 汪惠泽 冯云飞 刘云迎 李东明 何祥来 王捍东 |
| 作者单位： | 1. 扬州大学兽医学院,江苏省普通高校重点学科内科实验室,江苏扬州 225009 2. 江苏省常州市出入境检验检疫局,江苏常州,213022 3. 江苏畜牧兽医职业技术学院,江苏泰州,225300 |
| 基金项目： | 江苏高校优势学科建设工程和江苏省高校重点实验室开放课题资助 |
| 摘 要： | 为建立掺糖造假蜂蜜中残留糖化酶的检测方法,用从黑曲霉中提取的糖化酶作为免疫原,免疫BALB/c小鼠,利用杂交瘤技术获得了12株稳定分泌针对糖化酶抗体的杂交瘤细胞株。单克隆抗体亚型鉴定结果显示,10株为IgG1,2株为IgG2b,轻链均为κ轻链。Western blotting分析结果表明,12株抗体均可特异性结合糖化酶。其中6株单抗(McAb-2H4F9、6H9D8、8F2F11、8F2E9、1A8G6、1C4D5)细胞株采用体内诱生法制备的腹水效价均1∶1×104以上。采用抗体叠加试验对这6株抗糖化酶单抗的抗原识别位点进行检测,反应增殖结果表明,6株单抗分别针对4类不同抗原位点,McAb-6H9D8和McAb-8F2F11针对第Ⅰ种抗原决定簇;McAb-1A8G6和McAb-1C4D5针对第Ⅱ种抗原决定簇;McAb-8F2E9针对第Ⅲ种抗原决定簇;McAb-2H4F9针对第Ⅳ种抗原决定簇。制备的抗体针对不同的抗原表位,为双抗夹心ELISA方法的建立提供前提。 |
| 关 键 词： | 淀粉糖化酶 单克隆抗体 抗原识别位点 |
| 收稿时间： | 2012-03-30 |
The Development and Antigenic Analysis of Monoclonal Antibodies to Glucoamylase |
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| ZHANG Dan , JIANG Wei , LI Shuang-xi , WANG Hui-ze , FENG Yun-fei , LIU Yun-ying , LI Dong-ming , HE Xiang-lai , WANG Han-dong.The Development and Antigenic Analysis of Monoclonal Antibodies to Glucoamylase[J].China Animal Husbandry & Veterinary Medicine,2012,39(11):39-42. | |
| Authors: | ZHANG Dan JIANG Wei LI Shuang-xi WANG Hui-ze FENG Yun-fei LIU Yun-ying LI Dong-ming HE Xiang-lai WANG Han-dong |
| Institution: | 1. Clinical Veterinary Medicine of Jiangsu Provincial Key Discipline,College of Veterinary Medicine,Yangzhou University, Yangzhou 225009,China;2. Changzhou Entry-exit Inspection and Quarantine Bureau of Jiangsu Province,Changzhou 213022,China;3. Jiangsu Animal Husbandry and Veterinary College, Taizhou 225300,China |
| Abstract: | To develop a detection method for glucoamylase in honey, BALB/c mice were immunized with the glucoamylase from Aspergillus niger. Splenocytes of the immunized mice were fused with SP2/0 cells with PEG. 12 monoclonal antibodies(McAbs) against the glucoamylase were obtained and identified. 10 hybridoma cell lines represented subtypes IgG1 and the other were IgG2b, and their light chains were κ chain. Western blotting analysis showed that the 12 McAbs specifically recognized glucoamylase. Hybridoma 2H4F9,6H9D8,8F2F11,8F2E9,1A8G6,1C4D5 excreting anti-glucoamylase McAbs were used to be injected into abdominal cavity of mice(ICR). The titers of the McAbs were all above 1∶1×104. Six strains of monoclonal antibodies were confirmed to be specific for four different kinds of epitopes on glucoamylase competitive binding assay. Among them, McAb-6H9D8 and McAb-8F2F11 might direct to the same antigenic determinant on the GA, McAb-1A8G6 and McAb-1C4D5 to the second antigenic site, McAb-8F2E9 to the third antigenic site, McAb-2H4F9 to the fourth antigenic site. |
| Keywords: | glucoamylase monoclonal antibodies epitopes |
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