State Key Laboratory of Analytical Chemistry for Life Science, School of Chemistry and Chemical Engineering, Nanjing University, Nanjing 210093, China.
Department of Chemistry, University of California, Riverside, CA 92521, United States.
Biosens Bioelectron. 2018 Jun 30;109:206-213. doi: 10.1016/j.bios.2018.02.029. Epub 2018 Feb 23.
A highly efficient method for aptamer screening with real-time monitoring of the SELEX process was described by silver decahedra nanoparticles (Ag-NPs) enhanced surface plasmon resonance imaging (SPRI). A microarray chip was developed by immobilization of target protein (Lactoferrin (Lac)) and control proteins (α-lactalbumin (α), β-lactoglobulin (β), casein, and bovine serum albumin (BSA)) on the biochip surface. Ag-NPs were conjugated with an ssDNA library (lib) (Ag-NPs-library) that consisted of a central 40 nt randomized sequence and a 20 nt fixed primer sequence. Introduction of the Ag-NPs-library to the SPRI flow channels drastically increased the sensitivity of SPRI signal for real-time monitoring of SELEX. The work allows rapid screening of potential targets, and yields nine aptamers with high affinity (nanomolar range) for Lac after only six-rounds of selection. The aptamer Lac 13-26 was then further tested by SPRI, and the results demonstrated that the aptamer had the capacity to be ultra-sensitive for specific detection of Lac. The novel SPRI-SELEX method demonstrated here showed many advantages of real-time evaluation, high throughput, and high efficiency.
本文描述了一种利用银纳米十面体（Ag-NPs）增强表面等离子体共振成像（SPRI）实时监测 SELEX 过程的高效适体筛选方法。通过将靶蛋白（乳铁蛋白（Lac））和对照蛋白（α-乳白蛋白（α）、β-乳球蛋白（β）、酪蛋白和牛血清白蛋白（BSA））固定在生物芯片表面，开发了一种微阵列芯片。Ag-NPs 与包含中心 40nt 随机序列和 20nt 固定引物序列的 ssDNA 文库（Ag-NPs-library）偶联。将 Ag-NPs-library 引入 SPRI 流道中，大大提高了 SPRI 信号的灵敏度，实现了 SELEX 的实时监测。该工作允许快速筛选潜在的靶标，经过仅六轮筛选，得到了九个对 Lac 具有高亲和力（纳摩尔范围）的适体。随后，通过 SPRI 进一步测试了 Lac 13-26 适体，结果表明该适体具有超灵敏特异性检测 Lac 的能力。这里展示的新型 SPRI-SELEX 方法具有实时评估、高通量和高效率等诸多优点。
