Mao Fujing, Yang Xun, He Zhenyun, Sun Zhichang, Zhang Sihang, Liu Xing
School of Food Science and Engineering, Hainan University, Haikou 570228, China.
School of Food Science and Engineering, Hainan University, Haikou 570228, China.
Food Chem. 2025 Feb 15;465(Pt 2):142061. doi: 10.1016/j.foodchem.2024.142061. Epub 2024 Nov 16.
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.
赭曲霉毒素A（OTA）是一种常见的食品污染物，对人类健康构成重大威胁，需要进行严格监测。模拟表位肽（MPs）在环保免疫分析中通常用作有毒小分子的无毒替代品。在此，以抗OTA纳米抗体为靶蛋白，利用噬菌体展示和免疫磁分离筛选OTA的环状七聚体MPs。使用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）开发了基于PMP的化学发光免疫分析（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 ng/mL和0.232 ng/mL。通过回收率实验和交叉反应证实了两种CLIA均具有良好的准确性和选择性。此外，它们在检测实际辣椒样品中通过高效液相色谱法进行了验证。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。
