转基因外源蛋白Bt Cry1Ab/Ac时间分辨荧光试纸条的制备与应用

引用本文: 谭桂玉, 祝一帆, 张漫, 何青青, 陈晓娇, 张坤, 王保民. 转基因外源蛋白Bt Cry1Ab/Ac时间分辨荧光试纸条的制备与应用[J]. 分析化学, 2021, 49(5): 752-758. doi: 10.19756/j.issn.0253-3820.191515 shu

Citation: TAN Gui-Yu, ZHU Yi-Fan, ZHANG Man, HE Qing-Qing, CHEN Xiao-Jiao, ZHANG Kun, WANG Bao-Min. Development and Application of Time-Resolved Fluorescence Lateral Flow Immunoassay for Bt Cry1Ab/Ac Protein Detection in Transgenic Plant[J]. Chinese Journal of Analytical Chemistry, 2021, 49(5): 752-758. doi: 10.19756/j.issn.0253-3820.191515 shu

转基因外源蛋白Bt Cry1Ab/Ac时间分辨荧光试纸条的制备与应用

谭桂玉 ^1,2, ,
祝一帆 ^1, ,
张漫 ^1, ,
何青青 ^1, ,
陈晓娇 ^{1,
,} ,
张坤 ^2,3, ,
王保民 ^{1,
,}

1.
中国农业大学农学院, 北京 100193;
2.
广西壮族自治区药用植物园, 南宁 530023;
3.
福建安欣睿捷生物科技有限公司, 福州 350026

通讯作者: 陈晓娇,E-mail:chenxj913@126.com; 王保民,E-mail:wbaomin@263.net

基金项目:
国家重点研发计划项目（No.2018YFC1602904）和福州市仓山区科技计划项目（No.2017C03）资助。

摘要: 随着转基因作物种植面积的不断扩大，不仅需要对转基因作物进行定性检测，还需对其所表达的外源蛋白进行快速定量分析，用于判断转基因作物的抗虫或抗除草剂强度。本研究利用前期筛选的两株识别Bt Cry1Ab/Ac不同抗原表位的小鼠单克隆抗体4E5和1E6，以羧基荧光微球作为标记物与抗体1E6偶联，4E5单抗和羊抗鼠抗体作为试纸条检测线（T线）和质控线（C线），制备了检测Bt Cry1Ab/Ac的时间分辨荧光试纸条。结合便携式荧光免疫分析仪，建立了定量检测转基因作物中Bt Cry1Ab/Ac外源蛋白的侧向层流免疫检测方法。通过筛选抗体量、反应时间等对试纸条进行优化，方法的检测时间为20 min，标准曲线线性方程为y=0.5804x-1.9123，相关系数R²=0.998，定量检出限为1.5 ng/mL。采用所制备的时间分辨荧光试纸条测定植物样品中Bt Cry1Ab/Ac外源蛋白的含量，与商品化试剂盒检测结果（检测时间为1.5 h）比较，两种方法检测结果相关性高（R²=0.999），一致性良好。本研究建立的时间分辨荧光试纸条方法可作为快速、准确检测转基因作物Bt Cry1Ab/Ac外源蛋白的定量方法。

关键词:

English

Development and Application of Time-Resolved Fluorescence Lateral Flow Immunoassay for Bt Cry1Ab/Ac Protein Detection in Transgenic Plant

1.
College of Agronomy and Biotechnology, China Agricultural University, Beijing 100193, China;
2.
Guangxi Botanical Garden of Medicine Plant, Nanning 530023, China;
3.
Fujian Antiorigin Biotechnology Co, Ltd., Fuzhou 350026, China

Corresponding author: CHEN Xiao-Jiao, ; WANG Bao-Min,

Received Date: 25 August 2019
Revised Date: 01 April 2021
Fund Project:
Supported by the National Key Research and Development Program of China (No. 2018YFC1602900) and the Fuzhou Cangshan District Science and Technology Plan Project (No. 2017C03).

Abstract: By now, genetically modified crops have been widely planted in the world. It is an urgent need not only to conduct qualitative testing of transgenes crop, but also to conduct rapid quantitative analysis of exogenous protein content expressed in transgenic crops, so as to judge the resistance to insects or herbicides. Two mouse monoclonal antibodies that recognize different epitopes of Bt Cry1Ab/Ac have been prepared in the previous work. On this basis, time-resolved fluorescent strips of Bt Cry1Ab/Ac were prepared using fluorescent microspheres as markers in this work. Combined with portable fluorescence immunoanalyzer, a lateral flow immunoassay was developed for the quantitative analysis of exogenous protein Bt Cry1Ac in transgenic crops. The detection time was 20 min, the linear equation of the standard curve was y=0.5804x-1.9123, with correlation coefficient (R²) of 0.998. The limit of quantitation was 1.5 ng/mL. With the established method, the content of Bt Cry1Ab/Ac protein in samples was determined, and the results were compared with those of commercial kit with detection time of 1.5 h. The results of these two methods correlated well (R²=0.999). The time-resolved fluorescent strip established here could be used as a rapid and accurate quantitative method for determination of Bt cry1Ab/Ac in transgenic crops.

Key words:

Transgenic crop
/ Bt Cry1Ab/Ac
/ Time-resolved fluorescent lateral flow immunoassay

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沈阳化工大学材料科学与工程学院沈阳 110142

转基因外源蛋白Bt Cry1Ab/Ac时间分辨荧光试纸条的制备与应用

通讯作者: 陈晓娇,E-mail:chenxj913@126.com; 王保民,E-mail:wbaomin@263.net

English

Development and Application of Time-Resolved Fluorescence Lateral Flow Immunoassay for Bt Cry1Ab/Ac Protein Detection in Transgenic Plant

Corresponding author: CHEN Xiao-Jiao, ; WANG Bao-Min,

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通讯作者: 陈斌, bchen63@163.com

中国化学会秘书处

转基因外源蛋白Bt Cry1Ab/Ac时间分辨荧光试纸条的制备与应用

通讯作者: 陈晓娇,E-mail:chenxj913@126.com; 王保民,E-mail:wbaomin@263.net

English

Development and Application of Time-Resolved Fluorescence Lateral Flow Immunoassay for Bt Cry1Ab/Ac Protein Detection in Transgenic Plant

Corresponding author: CHEN Xiao-Jiao, ; WANG Bao-Min,

CCS Chemistry：嵌段共聚物自组装成 “三明治”二维有机材料，实现纳米级压力传感功能

CCS Chemistry：颜徐州、鲍哲南： 你的皮肤可能是一片SPMs

CCS Chemistry：工作高效不疲劳，只须让催化剂动起来

CCS Chemistry：静电组装导向聚合- 聚电解质纳米凝胶量化制备新策略

CCS Chemistry：金黄色葡萄球菌醛基脱氢酶是如何识别特异性底物的？

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