A.N. Bach Institute of Biochemistry, Research Centre of Biotechnology of the Russian Academy of Sciences, Leninsky Prospect 33, 119071 Moscow, Russia.
Agricultural-Technological Institute, RUDN University, Miklukho-Maklaya Street 8/2, 117198 Moscow, Russia.
Sensors (Basel). 2018 Nov 15;18(11):3975. doi: 10.3390/s18113975.
A simple approach was proposed to decrease the detection limit of sandwich lateral flow immunoassay (LFIA) by changing the conditions for binding between a polyvalent antigen and a conjugate of gold nanoparticles (GNPs) with antibodies. In this study, the potato virus Y (PVY) was used as the polyvalent antigen, which affects economically important plants in the family. The obtained polyclonal antibodies that are specific to PVY were characterized using a sandwich enzyme-linked immunosorbent assay (ELISA) and surface plasmon resonance (SPR). For LFIA, the antibodies were conjugated with GNPs with a diameter of 17.4 ± 1.0 nm. We conducted LFIAs using GNP conjugates in a dried state on the test strip and after pre-incubation with a sample. Pre-incubating the GNP conjugates and sample for 30 s was found to decrease the detection limit by 60-fold from 330 ng∙mL to 5.4 ng∙mL in comparison with conventional LFIA. The developed method was successfully tested for its ability to detect PVY in infected and uninfected potato leaves. The quantitative results of the proposed LFIA with pre-incubation were confirmed by ELISA, and resulted in a correlation coefficient of 0.891. The proposed approach is rapid, simple, and preserves the main advantages of LFIA as a non-laboratory diagnostic method.
提出了一种简单的方法，通过改变多价抗原与金纳米粒子（GNP）与抗体缀合物之间结合的条件，降低夹心侧向流动免疫分析（LFIA）的检测限。在本研究中，马铃薯 Y 病毒（PVY）被用作多价抗原，它影响茄科中具有经济重要性的植物。使用夹心酶联免疫吸附测定（ELISA）和表面等离子体共振（SPR）对获得的特异性针对 PVY 的多克隆抗体进行了表征。对于 LFIA，将抗体与直径为 17.4 ± 1.0nm 的 GNP 缀合。我们在测试条上以干燥状态和在与样品预孵育后进行 GNP 缀合物的 LFIA。与传统 LFIA 相比，发现将 GNP 缀合物和样品预孵育 30s 可将检测限降低 60 倍，从 330ng·mL降至 5.4ng·mL。该方法已成功用于检测受感染和未感染马铃薯叶片中的 PVY。通过 ELISA 验证了具有预孵育的拟议 LFIA 的定量结果，相关系数为 0.891。所提出的方法快速、简单，并保留了 LFIA 作为非实验室诊断方法的主要优势。
