A.N. Bach Institute of Biochemistry, Federal Research Centre "Fundamentals of Biotechnology" of the Russian Academy of Sciences, Leninsky prospect 33, 119071, Moscow, Russia.
Mikrochim Acta. 2020 Apr 13;187(5):268. doi: 10.1007/s00604-020-04253-3.
The influence of Au@Pt nanoparticles' composition, morphology, and peroxidase-mimicking activity on the limit of detection (LOD) of lateral flow immunoassay (LFIA) has been investigated. Fourteen types of nanoparticles were synthesized by varying the concentration of Pt (20-2000 μM), using gold nanoparticles (GNP, diameter 20.0 ± 2.6 nm) as the seeds and ascorbic acid as a reducing agent. Au@Pt nanoparticles and GNPs were conjugated with antibodies specific to the target analyte, a widespread and dangerous phytopathogenic bacteria species (Clavibacter michiganensis). We found that the 100-fold growth of the Pt concentration was accompanied by an increase of the Au@Pt nanoparticle diameter (24-55 nm) and surface area with the formation of urchin-shaped morphology. These changes led to a 70-fold increase in peroxidase-mimicking activity in the solution (specific activity 0.06-4.4 U mg) and a 30-fold decrease in LOD using the catalytic activity of Au@Pt. The Au@Pt nanoparticles synthesized at 1000-2000 μM of Pt demonstrated statistically indistinguishable catalytic activity. The highest sensitivity of LFIA was reached for Au@Pt nanoparticles synthesized at Pt concentration equal to 1000 μM. Au@Pt nanoparticles saved most of their peroxidase-mimicking activity, whereas endogenous plant peroxidases were completely inhibited by sodium azide. The LOD of LFIA with Au@Pt nanoparticles synthesized at 1200 μM of Pt was 300 colony-forming units (CFU) per mL of buffer and 500 CFU per mL of potato tuber extract, which provides 330- and 200-fold improvement compared to the conventional LFIA with GNPs. The assay consists of three rapid 5-min stages, namely, extraction, lateral flow, and color enhancement (oxidation of diaminobenzidine by Au@Pt nanoparticles). LFIA with the urchin Au@Pt nanoparticles allows the detection of latent bacterial infections rapidly without equipment or special skills. Graphical abstract.
研究了 Au@Pt 纳米粒子的组成、形态和过氧化物酶模拟活性对侧向流动免疫分析（LFIA）检测限（LOD）的影响。通过改变 Pt 的浓度（20-2000 μM），使用金纳米粒子（GNP，直径 20.0±2.6nm）作为种子，抗坏血酸作为还原剂，合成了 14 种纳米粒子。Au@Pt 纳米粒子和 GNP 与针对靶分析物（一种广泛存在且危险的植物病原菌（野油菜黄单胞菌））的抗体结合。我们发现，Pt 浓度增加 100 倍伴随着 Au@Pt 纳米粒子直径（24-55nm）和表面积的增加，并形成了刺猬状形态。这些变化导致溶液中过氧化物酶模拟活性增加了 70 倍（比活性 0.06-4.4Umg），并且使用 Au@Pt 的催化活性使 LOD 降低了 30 倍。在 1000-2000μM 的 Pt 下合成的 Au@Pt 纳米粒子表现出统计学上无明显差异的催化活性。当 Pt 浓度等于 1000μM 时，LFIA 达到最高灵敏度。在 1200μM 的 Pt 下合成的 Au@Pt 纳米粒子的 LOD 为缓冲液中的 300 个集落形成单位（CFU）/mL 和马铃薯块茎提取物中的 500CFU/mL，与传统的使用 GNP 的 LFIA 相比，提高了 330 倍和 200 倍。该测定由三个快速的 5 分钟阶段组成，即提取、侧向流动和颜色增强（Au@Pt 纳米粒子氧化二氨基联苯胺）。具有刺猬状 Au@Pt 纳米粒子的 LFIA 无需设备或特殊技能即可快速检测潜伏性细菌感染。
