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Fluorescence Polarization Based Displacement Assay for the Determination of Small Molecules with Aptamers

机译:基于荧光偏振的置换法测定适体的小分子

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The conversion of an aptamer-target binding event into a detectable signal is an important step in the development of aptamer-based sensors. In this work, we show that the displacement of a fluorescently labeled oligo from the aptamer by the target can be detected by fluorescence polarization (FP). We used Ochratoxin A (OTA), a small organic molecule (MW velence 403) as a case study. A detection limit of 5 nM OTAwas achieved. The method presented here provides an advantage over fluorophore-quenching systems and other steady-state fluorescence approaches in that no modification of the aptamer or the target is required. Additionally, the signal is produced by the displacement event itself, so no further agregation or conformational events have to be considered. This analytical method is particularly useful for small targets, as for large targets a direct measurement of the FP change of a labeled aptamer upon binding can be used to determine the concentration of the target. The results presented here demonstrate that aptamers and inexpensive labeled oligos can be used for rapid, sensitive, and specific determination of small molecules by means of FP.
机译:将适体-靶标结合事件转化为可检测信号是开发基于适体的传感器的重要步骤。在这项工作中,我们表明可以通过荧光偏振(FP)来检测荧光标记的寡核苷酸从适体到目标的位移。我们以O曲霉毒素A(OTA)(一种有机小分子(MW velence 403))作为案例研究。达到了5 nM OTA的检测限。本文介绍的方法提供了优于荧光团猝灭系统和其他稳态荧光方法的优势,因为不需要修饰适体或靶标。另外,信号是由位移事件本身产生的,因此不必考虑其他聚集或构象事件。该分析方法对小目标特别有用,因为对于大目标,可以使用结合后直接测量标记适体的FP变化来确定目标浓度。此处显示的结果表明,适体和廉价的标记寡核苷酸可用于通过FP快速,灵敏和特异性地测定小分子。

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