Detection of Non-Nucleic Acid Targets with an Unmodified Aptamer and a Fluorogenic Competitor.
Detection of Non-Nucleic Acid Targets with an Unmodified Aptamer and a Fluorogenic Competitor.
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DOI：
10.1016/j.jala.2010.02.002
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发表时间：
2010-06-01
期刊：
影响因子：
--
通讯作者：
Li, Na
中科院分区：
文献类型：
--
作者：
Li, Na
适体是可以结合各种非核酸靶标的寡核苷酸，其范围从蛋白质到小分子，具有与抗体相当的特异性和亲和力。大多数基于适体的检测策略需要对适体进行修饰，这可能导致其对靶标的亲和力和特异性的显著损失。在这里，我们报告了一个通用的策略来设计基于适体的光学探针。对靶特异性的未修饰的适体和与适体互补的荧光竞争物分别用于靶识别和信号产生。竞争剂是一种发夹寡核苷酸，其一端连接有荧光团，另一端连接有猝灭剂。当不存在靶标时，竞争剂与适体结合。然而，当引入靶时，竞争物将被靶从适体中置换，从而导致荧光信号的靶特异性降低。已经证明了这种策略对不同类型的靶标（小分子和蛋白质）以及不同类型的适体（DNA和RNA）的成功应用。此外，建立了一个基于几何学的预测模型，以进一步合理化的优化过程。由于其快速性和简单性，这种基于适体的检测策略在高通量应用中具有很大的前景。
Aptamers are oligonucleotides that can bind to various non-nucleic acid targets, ranging from proteins to small molecules, with a specificity and affinity comparable to that of antibodies. Most aptamer-based detection strategies require modification on the aptamer, which could lead to a significant loss in its affinity and specificity to the target. Here we reported a generic strategy to design aptamer-based optical probes. An unmodified aptamer specific to the target and a fluorogenic competitor complementary to the aptamer are utilized for target recognition and signal generation, respectively. The competitor is a hairpin oligonucleotide with a fluorophore attached on one end and a quencher attached on the other. When no target is present, the competitor binds to the aptamer. However, when the target is introduced, the competitor will be displaced from the aptamer by the target, thus resulting in a target-specific decrease in fluorescence signal. Successful application of this strategy to different types of targets (small molecules and proteins) as well as different types of aptamers (DNA and RNA) has been demonstrated. Furthermore, a thermodynamics-based prediction model was established to further rationalize the optimization process. Due to its rapidness and simplicity, this aptamer-based detection strategy holds great promise in high throughput applications.
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