Non-SELEX isolation of DNA aptamers for the homogeneous-phase fluorescence anisotropy sensing of tau Proteins.
Non-SELEX isolation of DNA aptamers for the homogeneous-phase fluorescence anisotropy sensing of tau Proteins.
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DOI：
10.1016/j.aca.2018.07.029
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发表时间：
2018-12
影响因子：
6.2
通讯作者：
S. Lisi;Emmanuelle Fiore;S. Scarano;E. Pascale;Yannik Boehman;F. Ducongé;S. Chierici;M. Minunni;E. Peyrin;C. Ravelet
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文献类型：
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作者：
S. Lisi;Emmanuelle Fiore;S. Scarano;E. Pascale;Yannik Boehman;F. Ducongé;S. Chierici;M. Minunni;E. Peyrin;C. Ravelet
在此，我们首次报道了针对整个tau蛋白（一种重要的阿尔茨海默病（AD）生物标志物）的DNA适体的分离。采用基于毛细管电泳分离技术的非SELEX方法，仅需三轮和一个工作日即可分离出对靶具有高亲和力的DNA序列池。接下来进行高通量测序，并使用固定在芯片上的蛋白质靶标通过表面等离子体共振初步评估五种选择的适体的识别能力。最后，最亲和的适体的分析潜力，证明通过设计的双相荧光各向异性测定。该DNA适体不仅能识别τ-441，而且能识别τ-381、τ-352、τ-383亚型。传感平台允许测定这四种靶标，检测限分别为28 nM、3.2 nM、6.3 nM和22 nM。
Herein, we report for the first time the isolation of DNA aptamers directed against the whole tau protein, an important Alzheimer's disease (AD) biomarker. Non-SELEX approach based on the capillary electrophoresis partitioning technique was employed to isolate a high-affinity DNA sequence pool towards the target in only three rounds and one working day. High-throughput sequencing was next performed and the recognition ability of five selected aptamers was preliminary evaluated by surface plasmon resonance using the protein target immobilized on the chip. Finally, the analytical potential of the most affine aptamer was demonstrated through the design of a homogeneous-phase fluorescence anisotropy assay. This DNA aptamer was found to be able to recognize not only the whole τ-441 but also the τ-381, τ-352, τ-383 isoforms. The sensing platform allowed the determination of these four targets with a detection limit of 28 nM, 3.2 nM, 6.3 nM and 22 nM, respectively.