Developing specific, rapid and cost-effective immunoassays for Zika detection

开发用于寨卡病毒检测的特异性、快速且具有成本效益的免疫测定方法

• 批准号: 9345836
• 负责人: Ling Tian
• 金额: $ 30万
• 依托单位: MEDIOMICS, LLC
• 依托单位国家: 美国
• 财政年份: 2017
• 资助国家: 美国
• 起止时间: 2017-03-10 至 2019-02-28
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9345836
• 关键词: Amino Acid Sequence; Antibodies; Antigens; Biological Assay; Blood Transfusion; Blood specimen; Brazil; Centers for Disease Control and Prevention (U.S.); Collaborations; Country; Cross Reactions; Culicidae; Dengue Virus; Detection; Diagnosis; Enzyme-Linked Immunosorbent Assay; Epitopes; European; Evaluation; Family; Flavivirus; Foundations; Goals; Hepatitis C; Human; Immunity; Immunoassay; Immunoglobulin M; Incidence; Individual; Infection; Japanese encephalitis virus; Laboratories; Libraries; Ligands; Methods; Molecular; Molecular Conformation; Patient Care; Patients; Peptide Library; Peptides; Performance; Phase; Phylogenetic Analysis; Pregnancy; Proteins; RNA Viruses; Reproducibility; Reverse Transcriptase Polymerase Chain Reaction; Ribosomes; Risk; Saliva; Sampling; Serological; Serum; Solid; Specificity; Symptoms; Techniques; Technology; Testing; Urine; Vaccination; Vaccines; Vascular blood supply; Viral Antigens; Virus Diseases; West Nile virus; Yellow Fever Virus Infection; Yellow fever virus; Zika Virus; base; cost; cost effective; cross reactivity; env Gene Products; high throughput screening; improved; next generation sequencing; novel; performance tests; prototype; screening; viral RNA; viral detection

This proposal aims to develop highly specific, rapid and cost-effective immunoassays to detect Zika virus (ZIKV) infection. ZIKV is a single-stranded RNA virus in the Flaviviridae family that is transmitted to humans through infected mosquitos, blood transfusions, and sexual contact. As of June 2016, around 500,000 Zika Virus disease cases have been estimated in Brazil alone. Recently, ZIKV has been detected in many other countries including the USA and most European countries. The FDA has recently mandated screening the entire US blood supply for ZIKV. Key challenges in ZIKV surveillance include the proportion of cases that remain asymptomatic and the nonspecificity of ZIKV symptoms. Current methods available for detection of ZIKV are not rapid, and furthermore, suffer from problems of unreliability and risks of cross-reactivity. A highly specific, easily performed antibody test is urgently needed for both surveillance and patient care. Mediomics has developed and commercialized a new assay platform (PINCER assays) that allows simple homogenous mix-and-read detection of a variety of targets. These rapid-result assays can reach high specificity while remaining low cost and easy to use. In a recent collaboration with the CDC, Mediomics developed a PINCER-based homogeneous assay prototype for hepatitis C (HCV) that achieved 97.3% specificity, 98.7% sensitivity, and inter and intra assay CV’s better than 10%. These promising analytical parameters, combined with the simplicity of homogenous detection, indicate a great potential for this assay platform. In addition, our long term collaborator, Dr. Tomasz Heyduk’s laboratory, developed a novel screening platform by combining both ribosome display and Next Generation Sequencing (NGS) technologies to quickly identify linear and conformational epitopes of antibodies from patient samples. We hypothesize that by combining our PINCER platform and the peptide epitopes unique to the ZIKV infection, we will be able to develop a specific, rapid, and cost effective immunoassay for ZIKV detection. Toward this goal, we have three specific aims in our Phase I project. Aim 1. Develop a PINCER-based homogeneous immunoassay for ZIKV detection. Aim 2. Explore entire protein sequence space of ZIKV to identify peptide ligands specifically recognized by ZIKV antibodies in patient blood samples. Aim 3. Optimize and evaluate the performance of both peptide and improved antigen-based PINCER assays. A third party will perform the evaluation of the prototype assay. If this project is successful, we should have established a solid foundation for a Phase II project to fully develop and validate a novel immunoassay that is simple (mix-and-read), rapid (10-30 minutes), cost-effective (<5-10% of ELISA), and suitable for low to high throughput screening of ZIKV infection.

该提案旨在开发高度特异性、快速且具有成本效益的免疫测定法来检测寨卡病毒（ZIKV） ZIKV 是黄病毒科的一种单链 RNA 病毒，通过传播给人类。 截至 2016 年 6 月，约 50 万例寨卡病毒感染者为蚊子、输血和性接触。 仅在巴西就估计有病例，最近在包括巴西在内的许多其他国家也发现了寨卡病毒。 美国和大多数欧洲国家最近强制要求对整个美国的血液供应进行筛查。 ZIKV 监测的主要挑战包括无症状病例的比例和 ZIKV 症状的非特异性。目前可用于检测 ZIKV 的方法并不快速，而且， 高度特异性、易于执行的抗体测试存在不可靠性和交叉反应风险的问题。 监测和患者护理都迫切需要 Mediomics 开发并商业化了一种新的方法。 检测平台（PINCER 检测）允许对各种靶标进行简单的同质混合和读取检测。 这些快速结果测定可以达到高特异性，同时保持低成本和易于使用。 Mediomics 与 CDC 合作开发了基于 PINCER 的肝炎均质检测原型 C (HCV) 的特异性为 97.3%，敏感性为 98.7%，检测间和检测内 CV 优于 10%。 这些有前途的分析参数，加上同质检测的简单性，表明了一个伟大的 此外，我们的长期合作者 Tomasz Heyduk 博士的实验室， 通过结合核糖体展示和下一代测序开发了一种新颖的筛选平台 (NGS) 技术可快速识别患者样本中抗体的线性和构象表位。 我们通过结合我们的 PINCER 平台和 ZIKV 感染特有的肽表位来发展这一点， 为了实现这一目标，我们将能够开发一种特异性、快速且具有成本效益的免疫分析方法来检测 ZIKV。 我们的第一阶段项目有三个具体目标：开发基于 PINCER 的同质化。 用于 ZIKV 检测的免疫分析 目标 2. 探索 ZIKV 的整个蛋白质序列空间以识别肽。 目标 3. 优化和评估患者血液样本中的 ZIKV 抗体特异性识别的配体。 肽和改进的基于抗原的 PINCER 检测的性能将由第三方进行。 如果这个项目成功的话，我们应该已经为原型测定的评估奠定了坚实的基础。 II 期项目，旨在全面开发和验证一种简单（混合读取）、快速（10-30 分钟）、成本效益高（< ELISA 的 5-10%），并且适用于低到高通量的 ZIKV 感染筛查。