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首页> 美国卫生研究院文献>ACS Omega >Enzyme-Free Immunoassay Using Silver Nanoparticlesfor Detection of Gliadin at Ultralow Concentrations
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Enzyme-Free Immunoassay Using Silver Nanoparticlesfor Detection of Gliadin at Ultralow Concentrations

机译：使用银纳米粒子的无酶免疫测定超低浓度麦醇溶蛋白的检测

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摘要

Determination of biomarkers in clinical or food samples is of crucial importance for monitoring, prevention, and care of public health. The standard procedure used for this purpose is the enzyme-linked immunosorbent assay (ELISA), which makes use of the specific antibody–antigen biorecognition and the catalytic effect of the enzymes. One of the main shortcomings of this technique is the use of enzymes that often present low chemical and thermal stabilities compared to other chemicals. Other drawbacks include the nonspecific binding process that could lead to false-positive results, the use of relatively large amounts of the sample, and the number of time-consuming steps involved. Recently, an enzyme-free and ultrasensitive analytical method for antigen detection denoted as intensity depletion immunolinked assay (IDILA) has been proposed by our laboratory. The assay is based on the inhibition to form Ag nanosphere dimers linked by a specific antibody in the presence of the corresponding antigen. In this work, we go a step further demonstrating how the performance of this method could be improved by using silvernanoparticles (Ag NPs) of different diameters (58 and 78 nm). Theexperiments are performed for detecting gliadin, an antigen of utmostimportance in celiac disease, and the results are compared with ELISA,the standard technique homologated by the Food Codex Alimentarius.It is found that the IDILA assay could be around 1000 or 10 000times more sensitive than ELISA, also having lower limits of detection,depending on the conditions explored (fraction of dimers and Ag NPdiameter). Using the appropriate conditions, the IDILA assay is shownto be able to detect femtomolar concentrations of the antigen, besidesbeing robust, reliable, cheap, rapid (around 2 h), and of easy implementationusing the standard equipment and biomolecular reagents used for theELISA assay.
机译：临床或食品样品中生物标志物的测定对于监测，预防和护理公共卫生至关重要。用于此目的的标准方法是酶联免疫吸附测定（ELISA），该方法利用特异性抗体-抗原的生物识别和酶的催化作用。该技术的主要缺点之一是使用的酶与其他化学药品相比通常具有较低的化学和热稳定性。其他缺点包括可能导致假阳性结果的非特异性结合过程，使用相对大量的样品以及涉及的耗时步骤。最近，我们的实验室提出了一种无酶超灵敏的抗原检测分析方法，称为强度消耗免疫分析法（IDILA）。该测定基于在相应抗原存在下通过特异性抗体连接形成Ag纳米球二聚体的抑制作用。在这项工作中，我们进一步说明了如何通过使用白银来改善此方法的性能。不同直径（58和78 nm）的纳米颗粒（Ag NPs）。的为检测麦醇溶蛋白（最大抗原）进行了实验在腹腔疾病中的重要性，并将结果与​​ELISA进行比较，食品法典认可的标准技术。发现IDILA分析可能在1000或10 000左右是ELISA的两倍，检测限较低，取决于探索的条件（二聚体和Ag NP的分数直径）。在适当的条件下，显示了IDILA分析能够检测抗原的飞摩尔浓度坚固，可靠，便宜，快速（大约2小时），并且易于实施使用标准设备和生物分子试剂ELISA测定。

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