Su Zixian, Zhao Guangying, Dou Wenchao
Food Safety Key Laboratory of Zhejiang Province, College of Food Science and Biotechnology, Zhejiang Gongshang University, Hangzhou, 310018, China.
School of Engineering, Westlake University, 18 Shilongshan Road, Zhejiang, 310024, Hangzhou, China.
Mikrochim Acta. 2020 Nov 17;187(12):658. doi: 10.1007/s00604-020-04636-6.
Aflatoxin M1 (AFM1) residues in milk pose a major threat to human health, so there is an urgent need for a simple, rapid, and sensitive method for the determination of trace AFM1 in milk. In this study, a competitive immunochromatographic assay (ICA), using visual (PEI/PSS) red silica nanoparticles (SiNPs) as signal amplification probes, was used for the highly sensitive detection of AFM1. The (PEI/PSS) red SiNPs were used to label AFM1 monoclonal antibody (mAb) to prepare ICA for the detection of AFM1. After exploring the optimal conditions of mAb and immunoprobe dosage conditions, the lowest visual detection limit (VDL) of AFM1 in phosphate-buffered saline with Tween 20 (PBST, 10 mM, pH 7.4, containing 1% BSA, 3% sucrose, 1% trehalose, and 0.5% Tween 20) can reach 0.1 pg/mL. The intuitive visually visible value of AFM1 in both PBST and milk was 10 pg/mL. The results showed that the immunochromatographic system based on high chroma color (PEI/PSS) red SiNPs has high sensitivity and broad application prospects for the detection of trace AFM1 residues in milk. The high chroma (PEI/PSS) red SiNPs are expected to be a convenient biomarker for improving the sensitivity of immune chromatography bands. Graphical abstract The schematic diagram shows the detection principle. In this work, in the competitive experiment, (PEI/PSS) red SiNPs were selected as visual labeling materials, and the specific antibody combined with the labeled material was selected as an immune probe. The AFM1-BSA antigen coupled with the macromolecular BSA was fixed on the T line of the nitrocellulose (NC) membrane. The AFM1 in sample solution competes with AFM1-BSA for the specific binding site of immune probe. The detection sensitivity of this method for AFM1 is obtained by judging the change of the red signal intensity produced by the positive sample, compared with the color at the T line of the negative sample.
牛奶中的黄曲霉毒素M1（AFM1）残留对人类健康构成重大威胁，因此迫切需要一种简单、快速且灵敏的方法来测定牛奶中的痕量AFM1。在本研究中，一种竞争性免疫层析测定法（ICA），使用可视化（PEI/PSS）红色二氧化硅纳米颗粒（SiNPs）作为信号放大探针，用于高灵敏检测AFM1。（PEI/PSS）红色SiNPs用于标记AFM1单克隆抗体（mAb）以制备用于检测AFM1的ICA。在探索mAb和免疫探针剂量条件的最佳条件后，在含吐温20的磷酸盐缓冲盐水（PBST，10 mM，pH 7.4，含有1%牛血清白蛋白、3%蔗糖、1%海藻糖和0.5%吐温20）中AFM1的最低可视化检测限（VDL）可达到0.1 pg/mL。在PBST和牛奶中AFM1直观的可视化可见值均为10 pg/mL。结果表明，基于高色度颜色（PEI/PSS）红色SiNPs的免疫层析系统对检测牛奶中痕量AFM1残留具有高灵敏度和广阔的应用前景。高色度（PEI/PSS）红色SiNPs有望成为提高免疫层析条带灵敏度的便捷生物标志物。图形摘要示意图显示了检测原理。在本工作中，在竞争性实验中，选择（PEI/PSS）红色SiNPs作为可视化标记材料，并选择与标记材料结合的特异性抗体作为免疫探针。与大分子牛血清白蛋白偶联的AFM1 - BSA抗原固定在硝酸纤维素（NC）膜的T线上。样品溶液中的AFM1与AFM1 - BSA竞争免疫探针的特异性结合位点。通过判断阳性样品产生的红色信号强度的变化，与阴性样品T线处的颜色相比，获得该方法对AFM1的检测灵敏度。
