Rapid Detection of Agents that Cause Respiratory Disease

快速检测引起呼吸道疾病的病原体

• 批准号: 6789688
• 负责人: James R. Prudent
• 金额: $ 32.57万
• 依托单位: ERAGEN BIOSCIENCES, INC.
• 依托单位国家: 美国
• 财政年份: 2004
• 资助国家: 美国
• 起止时间: 2004-03-15 至 2005-02-28
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6789688
• 关键词: biotechnology; clinical research; communicable disease diagnosis; diagnosis design /evaluation; feces; human tissue; nucleic acid chemical synthesis; polymerase chain reaction; rapid diagnosis; respiratory infections; sputum

DESCRIPTION (provided by applicant): The ultimate goal of this proposed project is to develop a clinical diagnostic in three years that targets 15 agents that cause infections in the respiratory tract that may be misdiagnosed for the SARS virus. Our preliminary targets have genomes constructed from RNA or DNA and therefore the test will be required to analyze both, possibly in a single tube. The MULTI-CODE platform will be used to for the diagnostic proposed. This platform is extremely simple to perform, is complete in less than three hours, does not require washing, filtration of expensive equipment and is uniquely suited for the clinical lab. Specifically: i. The platform has been shown in a mock experiment using synthetic targets to simultaneously detect RNA and DNA even when the targets differ by a single nucleotide. ii. The platform is fast and robust. Detecting low copy number in less than three hours with minimal "hands-on" requirements. iii. Previously, PCR based tests in both singleplex and low number multiplexes have been able to diagnose these targets in throat swabs as suggested in this application. iv. EraGen's collective expertise in diagnostic platform development (Chemistry, molecular biology, manufacturing) and market implementation makes it uniquely qualified to explore capabilities of this highly innovative technology. The Aims of Phase I are to: 1. Design multiple MULTI-CODE PCR primers and ERA-CODE primer extenders for each target. 2. Design multiple MULTI-CODE PCR primers and ERA-CODE primer extenders for at least two internal positive control RNA and DNA targets. 3. Synthesize all primers and targets from Aim 1. 4. Determine the correct mixture of primers and extenders that include one of the DNA and RNA internal positive control targets that allow for the highest multiplexing possible for both sputum and stool samples that continues to allow 1-100 copy sensitivity. 5. Using a number of sample preparation systems, validate sample preparation methods in conjunction with multiplexed assays using blinded controlled samples spiked with synthetic targets in multiple combinations.

描述（由申请人提供）：该拟议项目的最终目标是在三年内开发出一种临床诊断方法，针对 15 种导致呼吸道感染的病原体，这些病原体可能被误诊为 SARS 病毒。我们的初步目标是由 RNA 或 DNA 构建的基因组，因此测试需要对两者进行分析，可能在单个试管中进行。多代码平台将用于建议的诊断。该平台执行起来极其简单，不到三个小时即可完成，不需要清洗、过滤昂贵的设备，特别适合临床实验室。 具体来说： 我。该平台已在模拟实验中展示，使用合成靶标可以同时检测 RNA 和 DNA，即使靶标只有一个核苷酸不同。 二.该平台快速且强大。在不到三个小时的时间内检测低拷贝数，并且“动手”要求极低。 三.此前，基于 PCR 的单重和低数量多重测试已经能够诊断咽拭子中的这些目标，如本申请中所建议的。 四. EraGen 在诊断平台开发（化学、分子生物学、制造）和市场实施方面的集体专业知识使其具有独特的资格来探索这种高度创新技术的能力。 第一阶段的目标是： 1. 为每个靶标设计多个 MULTI-CODE PCR 引物和 ERA-CODE 引物延伸剂。 2. 为至少两个内部阳性对照 RNA 和 DNA 靶标设计多个 MULTI-CODE PCR 引物和 ERA-CODE 引物延伸剂。 3. 合成目标 1 中的所有引物和靶标。 4. 确定正确的引物和延伸剂混合物，其中包括 DNA 和 RNA 内部阳性对照靶标之一，使痰液和粪便样本能够实现最高的多重检测，并继续保持 1-100 拷贝的灵敏度。 5. 使用多种样品制备系统，结合多重测定验证样品制备方法，使用盲法对照样品掺入多种组合的合成靶标。