Peltomaa Riikka, Farka Zdeněk, Mickert Matthias J, Brandmeier Julian C, Pastucha Matěj, Hlaváček Antonín, Martínez-Orts Mónica, Canales Ángeles, Skládal Petr, Benito-Peña Elena, Moreno-Bondi María C, Gorris Hans H
Institute of Analytical Chemistry, Chemo- and Biosensors, University of Regensburg, Universitätsstraße 31, 93040, Regensburg, Germany; Department of Analytical Chemistry, Faculty of Chemistry, Universidad Complutense de Madrid, Plaza de las Ciencias, Ciudad Universitaria, 28040, Madrid, Spain.
Institute of Analytical Chemistry, Chemo- and Biosensors, University of Regensburg, Universitätsstraße 31, 93040, Regensburg, Germany; Department of Biochemistry, Faculty of Science, Masaryk University, Kamenice 5, 625 00, Brno, Czech Republic.
Biosens Bioelectron. 2020 Dec 15;170:112683. doi: 10.1016/j.bios.2020.112683. Epub 2020 Oct 5.
Due to increasing food safety standards, the analysis of mycotoxins has become essential in the food industry. In this work, we have developed a competitive upconversion-linked immunosorbent assay (ULISA) for the analysis of zearalenone (ZEA), one of the most frequently encountered mycotoxins in food worldwide. Instead of a toxin-conjugate conventionally used in competitive immunoassays, we designed a ZEA mimicking peptide extended by a biotin-linker and confirmed its excellent suitability to mimic ZEA by nuclear magnetic resonance (NMR) and surface plasmon resonance (SPR) analysis. Upconversion nanoparticles (UCNP, type NaYF:Yb,Tm) served as background-free optical label for the detection of the peptide mimetic in the competitive ULISA. Streptavidin-conjugated UCNPs were prepared by click reaction using an alkyne-PEG-neridronate linker. The UCNP conjugate clearly outperformed conventional labels such as enzymes or fluorescent dyes. With a limit of detection of 20 pg mL (63 pM), the competitive ULISA is well applicable to the detection of ZEA at the levels set by the European legislation. Moreover, the ULISA is specific for ZEA and its metabolites (α- and β-zearalenol) without significant cross-reactivity with other related mycotoxins. We detected ZEA in spiked and naturally contaminated maize samples using liquid chromatography-tandem mass spectrometry (UPLC-MS/MS) as a reference method to demonstrate food analysis in real samples.
由于食品安全标准日益提高，霉菌毒素分析在食品工业中变得至关重要。在这项工作中，我们开发了一种竞争性上转换免疫吸附测定法（ULISA）用于分析玉米赤霉烯酮（ZEA），它是全球食品中最常见的霉菌毒素之一。我们设计了一种通过生物素连接子延伸的ZEA模拟肽，以替代竞争性免疫测定中传统使用的毒素缀合物，并通过核磁共振（NMR）和表面等离子体共振（SPR）分析证实了其模拟ZEA的优异适用性。上转换纳米颗粒（UCNP，NaYF:Yb,Tm型）用作竞争性ULISA中肽模拟物检测的无背景光学标记。通过使用炔基-聚乙二醇-奈替膦酸连接子的点击反应制备了链霉亲和素缀合的UCNP。UCNP缀合物明显优于传统标记物，如酶或荧光染料。该竞争性ULISA的检测限为20 pg/mL（63 pM），非常适用于检测欧盟法规规定水平的ZEA。此外，ULISA对ZEA及其代谢物（α-和β-玉米赤霉醇）具有特异性，与其他相关霉菌毒素无明显交叉反应。我们使用液相色谱-串联质谱法（UPLC-MS/MS）作为参考方法检测了加标和天然污染的玉米样品中的ZEA，以证明在实际样品中的食品分析。
