Probes and cell arrays for detection of bacterial toxins

项目概述/摘要：微阵列技术为同时监测哺乳动物细胞内多个标记物的行为提供了机会，最终可用于检测和阐明不同生物活性剂的作用机制，包括那些在生物防御领域被认为是威胁的生物活性剂。用于诊断阵列的探针的“预测能力”及其应用成本将是将阵列技术引入常规诊断实践的关键参数。我们建议开发一种创新的强大系统，该系统将允许鉴定和分离特定的人类DNA序列，这些序列在真核细胞与特定生物活性物质或生物活性物质群之间的相互作用中被上调或下调。这些序列可作为构建基于细胞阵列的检测系统的基础。在第一阶段，我们将演示应用程序所有基本方面的技术可行性，以便在随后的阶段进行细化。我们的方法将是使用流式细胞仪细胞分选过程来鉴定和分离遗传元件，这些遗传元件先前与报告基因序列融合，这些基因序列对特定的毒物挑战作出反应。在第二阶段，我们的主要工作将是扩展我们的反应元件库，这些元件将用于检测主要有毒物质，并构建诊断细胞阵列的原型。这项研究的最终目标是为检测生物活性药物提供一种通用的、可靠的方法，特别是当这些药物被设计为逃避目前可用的免疫测定时。项目描述：细胞阵列为灵敏和精确监测生物活性物质提供了新的机会。本申请的目标是开发一种用于识别调节元件的系统，该系统将允许细胞阵列的鉴别能力的实质性扩展，并创建能够检测和识别潜在生物战剂的细胞阵列。

DESCRIPTION (provided by applicant): Project Summary/Abstract: Microarray technology offers an opportunity for simultaneous monitoring of the behavior of multiple markers within a mammalian cell and ultimately could be used for detection and elucidation of mechanisms of action of different biologically active agents, including those that are considered a threat in the biodefense area. The "predictive power" of probes used in diagnostic arrays and the cost of their application will be crucial parameters for introduction of array technology into routine diagnostic practices. We propose to develop an innovative robust system that will allow for the identification and isolation of the specific human DNA sequences that are up or down regulated in response to interaction between a eukaryotic cell with specific biologically active agents or groups of biologically active agents. Such sequences could be used as the basis for the construction of a cell array based detection system. In Phase I, we will demonstrate the technical feasibility of all fundamental aspects of the application for elaboration during subsequent phases. Our approach will be to use a flow cytometer cell sorting process to identify and isolate genetic elements, previously fused with the reporter sequences, which respond to the specific toxic agent challenge. In Phase II, our principal effort will be to extend our library of responsive elements that will be used for detection of major toxic agents and construct a prototype of the diagnostic cell array. The ultimate goal of this research will be to provide a general and robust approach for the detection of biologically active agents, especially when these agents have been engineered to elude immunoassays which are currently available. Project Narrative: Cell arrays offer a new opportunity for sensitive and precise monitoring of biologically active substances. The goal of this application is to develop a system for identification of regulatory elements that will allow a substantial extension of discriminative abilities of cell arrays and creation of cell arrays capable of detection and identification of potential biowarfare agents.