Electrochemical arrays for the detection of small molecule drugs

描述（由申请人提供）：用于检测小分子药物的电化学阵列概述：用于检测DNA和RNA的微阵列已经建立，蛋白质微阵列正在迅速发展。相比之下，用于小分子定量检测的阵列的开发则明显滞后。在这里，我们提出了一种无试剂的电化学方法，用于在血液和其他复杂介质中并行、真实的监测一组小分子，这一进步应该证明在从诊断到药物发现到基础生物科学（例如，代谢组学）。我们的方法是基于电化学，基于适体（E-AB）的平台，是敏感的，快速的，无试剂的，和选择性足以直接在血清和其他大部分未加工的生物材料。虽然我们的长期目标是制造E-AB设备，以有效地实时监测任何小分子，但拟议的研究重点是滥用药物和氨基糖苷类抗生素的具体，高价值的例子。无试剂E-AB平台利用电化学来监测靶诱导的电极结合适体的折叠。到目前为止，我们已经制造了针对蛋白质，小分子和无机离子目标的E-AB传感器，其中每一个都是敏感的（微摩尔到皮摩尔的检测限）和选择性足以直接部署在血清，唾液和土壤提取物。此外，我们的传感器稳定且快速平衡，允许连续实时检测分析物流中的小分子，例如流动的血清。为了支持这种新生的方法，拟议的研究计划侧重于自上而下的开发E-AB阵列，旨在同时实时定量小分子药物套件。实现这一点将需要开发改进的方案，用于选择针对小分子的高亲和力、高特异性DNA适体，选择新的适体及其在高增益、高灵敏度E-AB传感器中的适应，进一步改进E-AB平台本身，以及最后，制造和测试微米级E-AB阵列，这些进展将坚定地确立这种潜在的有前途的和一般的传感技术的效用。用于同时检测多个RNA的微阵列已经彻底改变了我们对转录和基因调控的理解，蛋白质阵列正在迅速提高我们对翻译调控和蛋白质-蛋白质相互作用的理解。监测小分子的定量的、可并行的手段可能在包括基础生物科学（例如，代谢组学）、诊断学和药物发现。然而，目前用于检测血液中的药物、代谢物和其他小分子的方法通常限于色谱或竞争测定，这些技术不适合于快速、平行的分析物检测。在这里，我们提出了一种电化学，适体为基础的（E-AB）技术的发展，用于定量的多个小分子分析物平行，在真实的时间，在血液和其他复杂的媒体。虽然我们的长期目标是开发适用于有效地连续实时监测任何小分子的E-AB阵列，但这里提出的努力将集中在滥用药物和氨基糖苷类抗生素作为高价值的临床相关目标，以开发这种潜在的有前途的通用传感技术。

DESCRIPTION (provided by applicant): Electrochemical arrays for the detection of small molecule drugs Summary: Microarrays for the detection of DNA and RNA are well established, and protein microarrays are advancing rapidly. The development of arrays for the quantitative detection of small molecules, in contrast, has lagged significantly. Here we propose a reagentless, electrochemical method for monitoring a panel of small molecules in parallel, in real time, in blood and other complex media, an advance that should prove of significant utility in applications ranging from diagnostics through drug discovery to the basic biological sciences (e.g., metabolomics). Our approach is based on an electrochemical, aptamer-based (E-AB) platform that is sensitive, rapid, reagentless, and selective enough to employ directly in serum and other largely unprocessed biological materials. And while our long-term objective is the fabrication of E-AB devices for the real-time monitoring of effectively any small molecule, the proposed research focuses on specific, high-value examples of the drugs of abuse and aminoglycosidic antibiotics. The reagentless E-AB platform utilizes electrochemistry to monitor the target-induced folding of an electrode-bound aptamer. To date we have fabricated E-AB sensors against protein, small molecule and inorganic ion targets, each of which is sensitive (micromolar to picomolar detection limits) and selective enough to deploy directly in blood serum, saliva, and soil extracts. Moreover, our sensors are stable and equilibrate rapidly, allowing for the continuous, real-time detection of small molecules in an analyte stream, such as flowing blood serum. In support of this nascent approach, the proposed research program focuses on the top-to-bottom development E-AB arrays directed at the simultaneous, real-time quantification of suites of small-molecule drugs. Achieving this will require the development of improved schemes for the selection of high affinity, high-specificity DNA aptamers against small molecules, the selection of new aptamers and their adaptation in high gain, high- sensitivity E-AB sensors, further improvements in the E-AB platform itself and, finally, the fabrication and testing of micron-scale E-AB arrays, advances that will firmly establish the utility of this potentially promising and general sensing technology.Relevance. Microarrays for the simultaneous detection of multiple RNAs have revolutionized our understanding of transcription and gene regulation, and protein arrays are rapidly improving our understanding of translational regulation and protein-protein interactions. A quantitative, parallelizable means of monitoring small molecules would likely prove similarly transformative in applications including the basic biological sciences (e.g., metabolomics), diagnostics and drug discovery. Current methods for the detection of drugs, metabolites and other small molecules in blood, however, are generally limited to chromatographic or competition assays, techniques that are ill suited for rapid, parallel analyte detection. Here we propose the development of an electrochemical, aptamer-based (E-AB) technology for the quantification of multiple small molecule analytes in parallel, in real time, in blood and other complex media. And while our long-term objective is the development of E-AB arrays suitable for the continuous, real-time monitoring of effectively any small molecule, the efforts proposed here will focus on the drugs of abuse and the aminoglycosidic antibiotics as high-value, clinically relevant targets with which to develop this potentially promising and general sensing technology.