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[1]周 勇 张佳琪 廖浩祥 马煜宁** 程金平.抗DEP单克隆抗体杂交瘤细胞株的构建*[J].应用与环境生物学报,2019,25(03):1-11.[doi:10.19675/j.cnki.1006-687x.2018.08025]
 ZHOU YongZHANG JiaqiLIAO Haoxiang MA Yuning** & CHENG Jinping.CLC X592 Development of hybridoma cell lines secreting monoclonal antibodies against diethyl phthalate*[J].Chinese Journal of Applied & Environmental Biology,2019,25(03):1-11.[doi:10.19675/j.cnki.1006-687x.2018.08025]
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抗DEP单克隆抗体杂交瘤细胞株的构建*()
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《应用与环境生物学报》[ISSN:1006-687X/CN:51-1482/Q]

卷:
25卷
期数:
2019年03期
页码:
1-11
栏目:
研究论文
出版日期:
2019-06-25

文章信息/Info

Title:
CLC X592 Development of hybridoma cell lines secreting monoclonal antibodies against diethyl phthalate*
文章编号:
201808025
作者:
周 勇 张佳琪 廖浩祥 马煜宁** 程金平
(上海交通大学环境科学与工程学院,上海200240)
Author(s):
ZHOU YongZHANG JiaqiLIAO Haoxiang MA Yuning** & CHENG Jinping
(School of Environmental Science and Engineering, Shanghai Jiaotong University, Shanghai 200240, China )
关键词:
邻苯二甲酸二乙酯杂交瘤细胞株单克隆抗体HAT筛选胶体金免疫层析
Keywords:
DEP hybridoma cell line McAb HAT screening GICA
DOI:
10.19675/j.cnki.1006-687x.2018.08025
摘要:
为了构建针对水环境中邻苯二甲酸二乙酯(Diethyl phthalate, DEP)等邻苯二甲酸酯类(Phthalates, PAEs)化合物的免疫学快速检测方法,需获得高效稳定的分泌抗DEP单克隆抗体的杂交瘤细胞株。本文以邻苯二甲酸单乙酯(Monoethylphthalate, MEP)-BSA为免疫抗原免疫BALb/c小鼠,间接竞争ELISA测定小鼠血清效价,用细胞融合筛选技术筛选抗DEP单克隆抗体杂交瘤细胞株。结果显示,实验小鼠免疫血清效价最高能达到1:5000,经过HAT筛选、复检和亚克隆后得到9个能分泌抗DEP单克隆抗体的细胞株,细胞株上清液效价最高能达到1:10000,对DEP的抑制率最高能达到86.5%,且其对11种结构类似物的交叉反应率均低于0.1%,特异性较好。研究表明,本方法成功构建的杂交瘤细胞株能稳定高效分泌抗DEP单克隆抗体,为DEP的ELISA检测方法建立及胶体金免疫层析(Colloidal g old immunochromatography assay, GICA)试纸条的研制提供了稳定高效的单克隆抗体源。(图8表2参23)
Abstract:
In order to establish a rapid immunological detection method of diethyl phthalate (DEP) and other phthalic acid ester(PAEs), highly efficient and stable hybridoma cell line secreting anti-DEP McAb is urgently needed. In this study, mass spectrometry was used to determine whether the MEP-BSA is built, MEP-BSA was used as immune antigen to immunize BALb/c mice in order to detect serum titer in mice coupled with indirect competitive ELISA, and the technique of cell hybridoma screening was used to screen hybridoma cell line resistant to anti-DEP monoclonal antibody. And the results showed that the MEP-BSA was successfully synthetic; the mice serum titer can reach the maximum value of 1:5000 ; the 17 cell lines which can secrete anti-DEP monoclonal antibody after selection by HAT, and subclones whose cell supernatant titer can reach a maximum value of 1:10000. The successfully constructed hybridoma cell lines can stably secrete anti-DEP monoclonal antibody, and the inhibition ratio can reach 86.5% , and the cross-reaction rate of 11 structural analogs is less than 0.1%. Our method provides a stable and efficient source of monoclonal antibody for establishing the detecting method of ELISA as well as a reference for developing the colloidal gold immunochromatographic (GICA) method for ELISA detection of DEP GICA strip.

备注/Memo

备注/Memo:
收稿日期:2018-08-25 接受日期 Accepted: 2019-01-04
*科技部海洋重大专项(2016YFC1402403) 资助?
**通讯作者(E-mail:yuningma@sjtu.edu.cn)
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更新日期/Last Update: 2019-01-15