A.N. Bach Institute of Biochemistry, Research Center of Biotechnology of the Russian Academy of Sciences, Leninsky Prospect 33, 119071 Moscow, Russia.
Biosensors (Basel). 2023 May 7;13(5):525. doi: 10.3390/bios13050525.
Finding optimal conditions for competitive lateral flow immunoassay is a controversial task. The content of specific antibodies labeled by nanoparticles should be simultaneously high to reach intense signals and low to register an influence on the signals for minimal concentrations of the target analyte. We propose to use two kinds of complexes of gold nanoparticles in the assay, with antigen-protein conjugates and with specific antibodies. The first complex interacts both with immobilized antibodies in the test zone and with antibodies on the surface of the second complex. In this assay, the coloration is enhanced by the binding of two-colored preparations in the test zone, whereas the antigen in the sample inhibits both the binding of the first conjugate with the immobilized antibodies and with the second conjugate. This approach is realized for the detection of insecticide imidacloprid (IMD), an important toxic contaminant connected with the recent global death of bees. The proposed technique expands the working range of the assay, that is, in accordance with its theoretical analysis. The reliable change of coloration intensity is achieved for a 2.3-times-lower concentration of the analyte. The limit of IMD detection is 0.13 ng/mL for tested solutions and 1.2 µg/kg for initial honey samples. The combination of two conjugates doubles the coloration in the absence of the analyte. The developed lateral flow immunoassay is applicable for five-fold-diluted honey samples without extraction, does not require additional stages (all reagents are pre-applied to the test strip), and is implemented in 10 min.
找到竞争性侧向流动免疫分析的最佳条件是一项有争议的任务。纳米粒子标记的特异性抗体的含量既要高到达到强烈的信号，又要低到能够记录出目标分析物的最小浓度对信号的影响。我们建议在测定中使用两种金纳米粒子复合物，一种与抗原-蛋白缀合物结合，另一种与特异性抗体结合。第一个复合物与测试区中的固定化抗体以及第二个复合物表面上的抗体都相互作用。在该测定中，通过测试区中双色制剂的结合增强了显色，而样品中的抗原抑制了第一种缀合物与固定化抗体的结合以及与第二种缀合物的结合。这种方法用于检测杀虫剂吡虫啉（IMD），这是一种与最近全球蜜蜂死亡有关的重要有毒污染物。所提出的技术扩展了测定的工作范围，即根据其理论分析。可靠地改变了显色强度，分析物的浓度降低了 2.3 倍。对于测试溶液，IMD 的检测限为 0.13 ng/mL，对于初始蜂蜜样品，检测限为 1.2 µg/kg。在没有分析物的情况下，两种缀合物的结合使显色增加一倍。开发的侧向流动免疫分析可用于未经提取的五倍稀释蜂蜜样品，不需要额外的步骤（所有试剂都预先应用于测试条），并且在 10 分钟内完成。
