摘要
玉米赤霉烯酮(zeralenone,ZEN)具有雌激素活性,主要污染谷物和饲料,大量聚积可导致流产和死胎,给动物和人类健康带来严重威胁。本研究通过将ZEN偶联抗原ZEN-BSA包被于纳米磁珠(magnetic nanoparticles,MNPs),制备纳米磁珠-偶联抗原复合物(MNPs-BSA-ZEN),同时使用金颗粒(Au nanoparticles,AuNPs)和辣根过氧化物酶(horseradish peroxidase,HRP)双标记的ZEN单克隆抗体,建立新型酶联免疫检测方法(MNPs-HRP-AuNPsIC-ELISA)。检测下限(IC10)达到0.03ng/mL,检测区间(IC20–IC80)为0.05–0.89ng/mL,半数抑制率(IC50)为0.22ng/mL,与ZEN类似物(α-zearalanol、zearalanone、α-zearalenol、β-zearalenol和β-zearalanol)的交叉反应性依次为19.2%、11.7%、8.3%、1.2%和4.3%,与黄曲霉毒素B1、赭曲霉毒素A、伏马毒素B1、桔青霉素和展青霉毒素几乎不存在交叉反应。在玉米、面粉和大豆样本中的加标回收率可达81.6%–113.5%,与LC-MS/MS同时对天然样本中ZEN含量的检测结果表明,两种方法相关性良好。本研究建立的MNPs-HRP-AuNPs IC-ELISA具备快速和高灵敏的双重优势,也可为其他霉菌毒素精准检测技术的开发提供参考。
Zearalenone(ZEN), a mycotoxin, is produced by Fusarium graminearum and Fusarium roseum growing primarily on corn or hay exposed to high moisture and threatens to food safety and public health.Children are more vulnerable to ZEN than adults. Sensitive, accurate and rapid analytical methods with new strategies for signal enhancement are needed for detection of ZEN. Therefore, a novel enzyme linked immunosorbent assay(MNPs-HRP-AuNPs IC-ELISA) using magnetic nanoparticles(MNPs) and double labeled monoclonal antibody complex(Anti-ZEN-HRP-AuNPs) was prototyped and used for detecting ZEN in cereal samples. The detection signal was enhanced which in turn improved the sensitivity of the assay.The lower limit of detection using MNPs-HRP-AuNPs IC-ELISA was 0.03 ng/mL while the IC50 was0.22 ng/mL. The linear working range was 0.05-0.89 ng/mL. The cross-reactivities with zeralenone analogues(α-zearalanol, zearalanone,α-zearalenol,β-zearalenol and β-zearalanol) were 19.2%, 11.7%,8.3%, 1.2% and 4.3%, respectively. No cross-reactivity(<0.01%) was observed with other co-occurring mycotoxins(AFB1, OTA, FB1, CIT and PAT). The recovery rates in spiked corn, wheat and soybean samples were 81.6%-113.5%, and the intra-day and inter-day relative standard deviations were <10%.Simultaneous analysis of commercial samples(corn, wheat, and feedstuff) showed a good correlation between MNPs-HRP-AuNPs IC-ELISA and liquid chromatography tandem mass spectrometry(LC/MS-MS).Hence, this new method can be used as a rapid, sensitive, and less time-consuming method to determine levels of zeralenone in cereal and feedstuff samples.
作者
肖佳丽
叶青
方云
方维焕
宋厚辉
章先
XIAO Jia-Li;YE Qing;FANG Yun;FANG Wei-Huan;SONG Hou-Hui;ZHANG Xian(Cllege of Animal Science and Technology, Zhejiang A&F University, Lin'an, Zhejiang 311300, China;Key Laboratory of Applied Technology on Green-Eco-Healthy Animal Husbandry of Zhejiang Provinee, Lin'an, Zhejiang 311300, China;Zhejia ng Provincial Engin eeri ng Laboratory for Animal Health In specti on and In ter net Tech nology, Lin 'an, Zhejia ng 311300, China;College of Animal Sciences, Zhejiang University, Hangzhou, Zhejiang 310058, China;Journal of Zhejiang University-SCIENCE B (Biomedicine & Biotechnology), Hangzhou, Zhejiang 310027, China;Zhejiang Academy of Scienee and Technology for Inspect!on and Quarantine, Hangzhou, Zhejiang 310012, China)
出处
《菌物学报》
CAS
CSCD
北大核心
2019年第3期440-450,共11页
Mycosystema
基金
浙江省自然科学基金(LQ17C170002)
浙江农林大学人才项目(2016FR025)
浙江省重点研发计划(2018C02041)
国家高新技术研究发展计划(863计划)(2012AA101602)~~
关键词
玉米赤霉烯酮
纳米磁珠
金颗粒
辣根过氧化物酶
单克隆抗体
zerale none
mag netic nan oparticles
gold particles
horseradish peroxidase
monoclonal an tibody
