Multipled POC device for antigen/molecular detection of influenza & other viruses

描述(由申请人提供)： 在接下来的十年中，很有可能出现流感(流感)大流行、新型呼吸道病毒(RV)[例如SARS]的大流行，或者导致呼吸道症状的生物恐怖主义袭击。此外，每年有多达50万人死于流感，临床上没有方法将流感与许多其他常见的RV感染区分开来。我们迫切需要灵敏、高特异性、廉价、便携、低复杂性的设备/检测方法，能够对流感和其他常见的轮状病毒进行多重检测和鉴定。长期以来，我们一直在开发针对流感和其他轮状病毒的多重商业分析方法，并开发了世界上第一个针对流感A、B、RSV A、B和副流感1、2、3的商业多重分子检测方法(Hexaplex，1996)和第一个针对SARS的检测方法(2003年)。我们现在已经开发了基于独特的样品采集设备(SCD)、稳定的Eu荧光微粒(EFM)和基于PRNA的结合化学的新型全集成技术，允许对患者进行灵敏的(比FDA批准的检测方法高400x&gt；)POC抗原检测。这项技术使用电池运行，需要15分钟，可以同时检测多个病毒。更重要的是，SCD收集并稳定核酸(RNA和DNA)样本，以根据需要用于分子分析(允许在保持生物遏制的同时测试阴性样本或进一步分析阳性样本)。在本申请中，我们提出了以下具体目标：！选择和优化化学成分、底物和硬件，建立一种SCD-EFM-PRNA检测方法，用于检测13种引起儿童和成人LRI或重大疾病的常见轮状病毒(流感A、B、RSV、[Paraflu 1，2，3，hMPV]、冠状病毒[NL63、HKU1、229E、OC43]、腺病毒和肠道病毒)。2.建立针对这13种病毒的多重RT-PCR电子微阵列(EMR)检测方法，作为确证分子检测方法(也作为产品)，与SA-1及其他已建立的分子检测方法测定并比较分析的LOD、敏感性和特异性。3.确定FDA要求的最佳参数。(例如，确保所有组件的设计、易用性指南和非专业实验室人员的结果解释符合CLIA在SA#1中放弃的分析标准)；4.过程开发(确保诊断组件的可制造性和以合理成本实现商业化的方法)。5.临床样本测试(冰冻样本)；以确定SA#1和SA#2中开发的针对大多数产生“流感样”症状的RV和LRI的多重POC分析是否可以证明与目前可用的标准诊断分析相比，临床敏感性、特异性和成本效益都有所提高。在这些SA完成后，临床医生和公共卫生官员将拥有两个重要的新工具来对抗流感和病毒性呼吸道感染，无论是从他们的医院、办公室还是地球上最偏远的地方。

DESCRIPTION (provided by applicant): There is a significant probability that in the next decade there will be either an influenza (flu) pandemic, a major epidemic of a novel respiratory virus (RV) [e.g., SARS], or a bioterrorism attack that results in respiratory symptoms. In addition, as many as 500,000 people die each year of flu and there is no method to clinically distinguish flu from the many other common RV infections. There is an urgent need for sensitive, highly specific, inexpensive, portable, low complexity devices/assays that allow for the multiplex detection and identification of flu and other common RV's. We have a long record of developing multiplex commercial assays for flu and other RVs and developed the first commercial multiplex molecular assay for flu A, B, RSV A, B, and parainflu 1,2,3 in the world (Hexaplex, 1996) and the first assay for SARS in the US (2003). We have now developed novel fully integrated technology based on a unique sample collection device (SCD), stable europium fluorescent microparticles (EFM), and binding chemistry based on pRNA that allows for sensitive (400x > than FDA approved assays) bedside or in-the-field multiplex POC antigen testing of patients. This technology runs on batteries, takes 15 minutes, and can detect multiple viruses simultaneously. Even more importantly the SCD collects and stabilizes a sample of nucleic acid (RNA and DNA) for use in molecular assays as needed (allowing for testing of negative samples or further analysis of positives while maintaining biocontainment). In this application we propose the following specific aims:! Select and optimize the chemistry, substrate, and hardware to produce a SCD-EFM-pRNA assay to detect 13 common RV's causing LRI or significant disease in children and adults (flu A, B, RSV, [paraflu 1,2,3, hMPV], corona- viruses [NL63, HKU1, 229E, OC43], adenovirus, and enterovirus). 2. Develop a multiplex-RT-PCR electronic microarray (eMR) assay to these same 13 viruses as a confirmatory molecular assay (also as a product), determine and compare analytical LOD, sensitivity and specificity with multiplex immunoassay developed in SA 1 and other established molecular assays. 3. Determine optimal parameters required by FDA. (e.g., ensure design of all components, ease of use guidelines, and results interpretation by non-professional laboratory personnel meets CLIA waived standards for assay in SA#1); 4. Process Development (methods to ensure manufacturability of the diagnostic components and commercialization at a reasonable cost). 5. Clinical sample testing (frozen samples); To determine if the multiplex POC assays developed in SA#1 & 2 to the majority of RV that produce "Flu-like" symptoms and LRI can demonstrate improved clinical sensitivity, specificity, and cost effectiveness compared to currently available standard diagnostic assays. After completion of these SAs, clinicians and public health officials will have two important new tools to fight flu and viral respiratory infections, whether from their hospital, office or the most remote locations on the planet.