特异性纳米抗体的制备及其在水样中节球藻毒素-R快速检测中的应用

Preparation of Specific Nanobodies and Their Application in the Rapid Detection of Nodularin-R in Water Samples.

作者信息

Yang Jinyi, Si Rui, Wu Guangpei, Wang Yu, Fang Ruyu, Liu Fei, Wang Feng, Lei Hongtao, Shen Yudong, Zhang Qi, Wang Hong

机构信息

Guangdong Provincial Key Laboratory of Food Quality and Safety, National-Local Joint Engineering Research Center for Processing and Safety Control of Livestock and Poultry Products, College of Food Science, South China Agricultural University, Guangzhou 510642, China.

Guangzhou Institute of Food Inspection, Guangzhou 510080, China.

出版信息

Foods. 2021 Nov 10;10(11):2758. doi: 10.3390/foods10112758.

DOI:10.3390/foods10112758

PMID:34829042

原文链接:https://pmc.ncbi.nlm.nih.gov/articles/PMC8622565/

Abstract

Nanobodies have several advantages, including great stability, sensibility, and ease of production; therefore, they have become important tools in immunoassays for chemical contaminants. In this manuscript, nanobodies for the detection of the toxin Nodularin-r (NOD-R), a secondary metabolite of cyanobacteria that could cause a safety risk for drinks and food for its strong hepatotoxicity, were for the first time selected from an immunized Bactrian camel VHH phage display library. Then, a sensitive indirect competitive enzyme-linked immunosorbent assay (ic-ELISA) for NOD-R, based on the nanobody N56 with great thermostability and organic solvent tolerance, was established under optimized conditions. The results showed that the limit of detection for NOD-R was 0.67 µg/L, and the average spike recovery rate was between 84.0 and 118.3%. Moreover, the ic-ELISA method was validated with spiked water sample and confirmed by UPLC-MS/MS, which indicated that the ic-ELISA established in this work is a reproducible detection assay for nodularin residues in water samples.

摘要

纳米抗体具有多种优势，包括高度稳定性、敏感性以及易于生产；因此，它们已成为化学污染物免疫分析中的重要工具。在本论文中，首次从免疫双峰驼VHH噬菌体展示文库中筛选出用于检测微囊藻毒素-R（NOD-R）的纳米抗体，微囊藻毒素-R是蓝藻的一种次生代谢产物，因其强烈的肝毒性可能对饮料和食品造成安全风险。然后，基于具有高热稳定性和有机溶剂耐受性的纳米抗体N56，在优化条件下建立了一种灵敏的NOD-R间接竞争酶联免疫吸附测定法（ic-ELISA）。结果表明，NOD-R的检测限为0.67μg/L，平均加标回收率在84.0%至118.3%之间。此外，ic-ELISA方法通过加标水样进行了验证，并经超高效液相色谱-串联质谱法（UPLC-MS/MS）确认，这表明本研究建立的ic-ELISA是一种可重复的水样中微囊藻毒素残留检测方法。