数字农科院2.0

A novel sustainable immunoassay for sensitive detection of atrazine based on the anti-idiotypic nanobody and recombinant full-length antibody

文献类型： 外文期刊

作者： Jing Zhao;Peipei Li;A. M. Abd El-Aty;Lingyuan Xu;Xingmei Lei;Song Gao;Jia Li;Yun Zhao;Yongxin She;Fen Jin;Jing Wang;Bruce D. Hammock;Maojun Jin

作者机构：

关键词： Anti-idiotypic nanobody;Atrazine;Recombinant full-length antibody;Specific molecular mechanism;Sustainable immunoassay

期刊名称： Chemical Engineering Journal

ISSN： 1385-8947

年卷期： 2024 年 491 卷

页码：

收录情况： SCIE（2024版） ; ; EI（2024版）

摘要： Immunoassays have been widely used to determine small-molecule compounds in food and the environment, meeting the challenge of obtaining false positive or negative results because of the variance in the batches of antibodies and antigens. To resolve this problem, atrazine (ATR) was used as a target, and anti-idiotypic nanobodies for ATR (AI-Nbs) and a recombinant full-length antibody against ATR (ATR-rAb) were prepared for the development of a sustainable enzyme-linked immunosorbent assay (ELISA). AI-Nb-7, AI-Nb-58, and AI-Nb-66 were selected from an immune phage display library. ATR-rAb was produced in mammalian HEK293 (F) cells. Among the four detection methods explored, the assay using AI-Nb-66 as a coating antigen and ATR-rAb as a detection reagent yielded a half maximal inhibitory concentration (IC50) of 1.66 ng mL−1 for ATR and a linear range of 0.35 − 8.73 ng mL−1. The cross-reactivity of the assay to ametryn was 64.24 %, whereas that to terbutylazine was 38.20 %. Surface plasmon resonance (SPR) analysis illustrated that these cross-reactive triazine compounds can bind to ATR-rAb to varying degrees at high concentrations; however, the binding/dissociation kinetic curves and the response values at the same concentration are different, which results in differences in cross-reactivity. Homology modeling and molecular docking revealed that the triazine ring is vital in recognizing triazine compounds. The proposed immunoassay exhibited acceptable recoveries of 84.40 − 105.36 % for detecting fruit, vegetables, and black tea. In conclusion, this study highlights a new strategy for developing sustainable immunoassays for detecting trace pesticide contaminants.

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