Mimotope peptides for nanobodies: A nontoxic alternative to ochratoxin a and its application in chemiluminescence immunoassays for analysis of pepper samples

IF 8.5 1区农林科学 Q1 CHEMISTRY, APPLIED Food Chemistry Pub Date : 2024-11-16 DOI:10.1016/j.foodchem.2024.142061

Fujing Mao, Xun Yang, Zhenyun He, Zhichang Sun, Sihang Zhang, Xing Liu

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Abstract

Ochratoxin A (OTA) is a common food contaminant and poses a significant threat to human health, which requires rigorous monitoring. Mimotope peptides (MPs) are commonly used as non-toxic alternatives to toxic small molecules in eco-friendly immunoassays. Herein, with an anti-OTA nanobody as the target protein, cyclic 7-mer MPs of OTA were screened using phage display and immunomagnetic separation. The phage MPs (PMP) with the highest sensitivity and its alkaline phosphatase-tagged MP fusion (ALP-MP) were used to develop a PMP-based chemiluminescent immunoassay (PMP-CLIA) and an ALP-MP-based CLIA (AMP-CLIA). After optimization, PMP-CLIA and AMP-CLIA exhibited a limit of detection of 0.128 ng/mL and 0.232 ng/mL. Good accuracy and selectivity were confirmed for both CLIAs by recovery experiments and cross-reactions. Moreover, they were validated by high performance liquid chromatography in detecting real pepper samples. Thus, two CLIAs based on the nanobody and MPs were demonstrated as reliable tools for monitoring OTA in pepper.

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纳米抗体的拟态肽：赭曲霉毒素 a 的无毒替代品及其在化学发光免疫分析法中的应用，用于分析辣椒样品

赭曲霉毒素 A（OTA）是一种常见的食品污染物，对人类健康构成严重威胁，需要进行严格监测。在环保型免疫测定中，拟态肽（MPs）通常被用作有毒小分子的无毒替代品。本文以抗 OTA 纳米抗体为靶蛋白，利用噬菌体展示和免疫磁分离技术筛选了 OTA 的环状 7-mer肽。灵敏度最高的噬菌体 MPs（PMP）及其碱性磷酸酶标记的 MP 融合体（ALP-MP）被用于开发基于 PMP 的化学发光免疫分析法（PMP-CLIA）和基于 ALP-MP 的 CLIA（AMP-CLIA）。经过优化，PMP-CLIA 和 AMP-CLIA 的检测限分别为 0.128 纳克/毫升和 0.232 纳克/毫升。回收实验和交叉反应证实了这两种 CLIA 具有良好的准确性和选择性。此外，它们还通过高效液相色谱法在检测真实辣椒样品时得到了验证。因此，基于纳米抗体和 MPs 的两种 CLIA 被证明是监测辣椒中 OTA 的可靠工具。

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来源期刊

Food Chemistry 工程技术-食品科技

CiteScore

16.30

自引率

10.20%

发文量

3130

审稿时长

122 days

期刊介绍： Food Chemistry publishes original research papers dealing with the advancement of the chemistry and biochemistry of foods or the analytical methods/ approach used. All papers should focus on the novelty of the research carried out.