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基于多拷贝基因等温扩增的环介导等温扩增-侧向流动检测法:在食品中高灵敏度低技术筛选致敏大豆()的适用性

LAMP-LFD Based on Isothermal Amplification of Multicopy Gene : Applicability for Highly Sensitive Low-Tech Screening of Allergenic Soybean () in Food.

作者信息

Allgöwer Stefanie M, Hartmann Chris A, Lipinski Clarissa, Mahler Vera, Randow Stefanie, Völker Elke, Holzhauser Thomas

机构信息

Division of Allergology, Paul-Ehrlich-Institut (PEI), D-63225 Langen, Germany.

出版信息

Foods. 2020 Nov 26;9(12):1741. doi: 10.3390/foods9121741.

Abstract

Soybean () allergy can be life threatening. A lack of causative immunotherapy of soybean allergy makes soybean avoidance indispensable. Detection methods are essential to verify allergen labeling and unintentional allergen cross contact during food manufacture. Here, we aimed at evaluating our previously described primers for loop-mediated isothermal amplification (LAMP) of multicopy gene , combined with a lateral flow dipstick (LFD)-like detection, for their performance of soybean detection in complex food matrices. The results were compared with those obtained using quantitative real-time Polymerase Chain Reaction (qPCR) as the current standard of DNA-based allergen detection, and antibody-based commercial lateral flow device (LFD) as the current reference of protein-based rapid allergen detection. LAMP-LFD allowed unequivocal and reproducible detection of 10 mg/kg soybean incurred in three representative matrices (boiled sausage, chocolate, instant tomato soup), while clear visibility of positive test lines of two commercial LFD tests was between 10 and 10 mg/kg and depending on the matrix. Sensitivity of soybean detection in incurred food matrices, commercial retail samples, as well as various processed soybean products was comparable between LAMP-LFD and qPCR. The DNA-based LAMP-LFD proved to be a simple and low-technology soybean detection tool, showing sensitivity and specificity that is comparable or superior to the investigated commercial protein-based LFD.

摘要

大豆()过敏可能危及生命。由于缺乏针对大豆过敏的病因性免疫疗法,避免食用大豆必不可少。检测方法对于验证食品生产过程中的过敏原标签以及意外的过敏原交叉接触至关重要。在此,我们旨在评估我们先前描述的用于多拷贝基因环介导等温扩增(LAMP)的引物,并结合类似侧向流动试纸条(LFD)的检测方法,以检测其在复杂食品基质中检测大豆的性能。将结果与使用定量实时聚合酶链反应(qPCR)作为基于DNA的过敏原检测的当前标准以及基于抗体的商业侧向流动装置(LFD)作为基于蛋白质的快速过敏原检测的当前参考所获得的结果进行比较。LAMP-LFD能够明确且可重复地检测出三种代表性基质(煮香肠、巧克力、速溶番茄汤)中添加的10 mg/kg大豆,而两种商业LFD测试阳性测试线的清晰可见度在10至10 mg/kg之间,且取决于基质。在添加了大豆的食品基质、商业零售样品以及各种加工大豆产品中,LAMP-LFD和qPCR对大豆的检测灵敏度相当。基于DNA的LAMP-LFD被证明是一种简单且技术要求低的大豆检测工具,其灵敏度和特异性与所研究的基于蛋白质的商业LFD相当或更优。

https://cdn.ncbi.nlm.nih.gov/pmc/blobs/67ce/7760099/bbeca8685ba4/foods-09-01741-g001.jpg

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