An Assay for Rapid and Sensitive Flu Virus Detection

描述（由申请方提供）：本I期SBIR提案的具体目的是确定用于快速和灵敏检测流感病毒的新型检测技术的可行性。具体而言，我们将确定手动检测是否快速（约15分钟），灵敏度（至少是FDA批准的抗原检测的100倍，或更好，就TCID 50单位/mL而言），可重复性（CV<15%）和特异性（在浓度高达106 cfu/mL时，无潜在干扰细菌物种的假阳性结果）。建议的研究有五个具体目标，包括： 1.黑曲霉素-4，7-二甲基-O-神经氨酸底物的合成、纯化和确认-第D1节。我们的目标是获得至少1克高度纯化的底物（HPLC纯度>98%）。 2.检测混合物的配制-第D2节。我们将准备一个检测混合物，其中包含检测流感病毒神经氨酸酶的所有必要试剂。检测混合物将在一次性检测管中冻干。 3.试验优化-第D3节 4.分析性能评价-第D4节。将评价试验的分析性能。我们将确定超过10种人甲型流感和人B流感病毒株的分析灵敏度。 5.使用人体样本的比较研究和初步评价-第D5节。我们将使用FDA批准的基于抗原的检测和RT-PCR检测进行比较研究。此外，我们将检测从人类受试者中采集的一些样本。 公共卫生相关建议的第一阶段SBIR项目将评估一种新的检测技术的可行性，以快速和灵敏地检测流感病毒。该技术通过使用主混合物简化了检测过程，该主混合物能够实现一步检测，即，向主混合物中加入样品，然后进行光信号测量，所有这些都在同一检测管中进行。整个过程需要10-15分钟。成功发展拟议的流感病毒测试，将为快速诊断流感提供更好的选择，从而大大提高我们应对流感大流行期间和潜在流感大流行的医疗能力。

DESCRIPTION (provided by applicant): The specific aims of this Phase I SBIR proposal are to determine the feasibility of a novel detection technology for rapid and sensitive detection of influenza virus. Specifically, we will determine if the manual assay is rapid (about 15 minutes), sensitive (as least 100 times as good as, or better than, an FDA approved antigen test in terms of TCID50 units/mL), reproducible (with CV<15%) and specific (without false positive results from potentially interfering bacterial species at concentrations of upto106 cfu/mL). There are five specific aims in the proposed studies, including: 1. Synthesis, purification and qualification of luciferin - 4,7-di-methyl-O-neuraminic acid substrate - Section D1. Our goal is to obtain at least 1 gram of highly purified substrate (>98% purity by HPLC). 2. Formulation of the detection mix - Section D2. We will prepare a detection mix that contains all necessary reagents for detection of flu viral neuraminidase. The detection mix will be lyophilized in a single-use detection tube. 3. Assay optimization - Section D3 4. Evaluation of analytical performance - Section D4. Analytical performance of the assay will be evaluated. We will determine the analytical sensitivity for more than 10 strains of human influenza A and human influenza B. 5. Comparison studies and preliminary evaluation using human samples - Section D5. We will perform comparison studies using FDA-approved antigen-based test and RT-PCR assay. In addition, we will test some samples collected from human subjects. PUBLIC HEALTH RELVANCE The proposed Phase I SBIR project will evaluate the feasibility of a novel detection technology for rapid and sensitive detection of influenza virus. The technology simplifies the detection process by using a master mix which enables one-step detection, i.e., addition of a sample to the master mix followed by light signal measurement, all performed in the same detection tube. The entire process takes 10-15 minutes. Successful development of the proposed influenza virus test will provide a better alternative for rapid flu diagnosis, thereby significantly improving our medical capability for interpandemic influenza and potential pandemic influenza.