Integrated Cost-Effective Point of Care Nucleic Acid Testing for TB diagnosis

用于结核病诊断的经济高效的综合护理点核酸检测

• 批准号: 9114458
• 负责人: Gerard A Cangelosi
• 金额: $ 54.14万
• 依托单位: KECK GRADUATE INST OF APPLIED LIFE SCIS
• 依托单位国家: 美国
• 财政年份: 2014
• 资助国家: 美国
• 起止时间: 2014-08-08 至 2018-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9114458
• 关键词: Address; Affect; Award; Biological Assay; Clinical; Collaborations; Complex; Country; Coupled; Coupling; Cross-Priming; Cytolysis; DNA; DNA amplification; Data; Detection; Developing Countries; Devices; Diagnosis; Diagnostic; Diagnostic Procedure; Disease; Disinfection; Docking; Early Diagnosis; Electronics; Engineering; Ensure; Europe; Goals; Health; Human; Human Resources; Incidence; Institution; Intervention; Laboratories; Lateral; Life; Marketing; Methods; Microscopy; Monitor; Mycobacterium tuberculosis; Nucleic Acid Amplification Tests; Patients; Performance; Population; Preparation; Process; Production; Pulmonary Tuberculosis; Quality Control; Reader; Reagent; Recovery; Reproducibility; Research Infrastructure; Research Personnel; Resources; Sampling; Sensitivity and Specificity; Sputum; System; Testing; Training; Tuberculosis; United States National Institutes of Health; Validation; Wireless Technology; amplification detection; base; biodefense; biothreat; communicable disease diagnosis; cost; cost effective; cross reactivity; design; effective therapy; improved; instrument; instrumentation; manufacturing scale-up; meetings; novel; pathogen; point of care; prototype; quality assurance; rapid diagnosis; scale up; transmission process; usability; validation studies

DESCRIPTION (provided by applicant): The goal of this project is to enable diagnosis of pulmonary tuberculosis at the point of care in low resource high burden countries by developing a portable, easy to use, integrated nucleic acid testing device that executes sample preparation, isothermal DNA amplification and lateral flow based detection without user intervention, providing a result in < 1.5 h sample-in-to-answer-out, at a significantly lower cost than currently available fully integrated bench-top systems. The project builds upon ongoing efforts that involve the same team of investigators and institutions, and is strengthened by new partners. To date, we have developed a novel, rapid sample preparation process for sputum liquefaction and disinfection, pathogen lysis, and DNA extraction, and have established suitable performance of this process using clinical sputum samples. We have demonstrated that our system can accommodate different isothermal amplification methods coupled to lateral flow detection, and have developed a novel process for master-mix reagent stabilization. Going forward we will utilize an isothermal cross-priming amplification (CPA), clinically validated for TB diagnosis, which is coupled to lateral flow, and includes an internal amplification control. We have designed and built a prototype integrated cartridge and instrument, and we have demonstrated that the sample preparation and amplification/detection subunits are functional. By the beginning of the proposed project, we aim to demonstrate fully automated process execution in the integrated alpha prototype system. The proposed project will enable us to further develop and validate the system, and to obtain regulatory approval for market release as an IVD. In Aim 1, we will optimize key system components related to sample pre-processing, DNA extraction, CPA, and lateral flow detection. We will improve the thermal stability of master-mix reagents, refine on-board reagent storage, incorporate a lateral flow strip reader, and ensure overall system robustness. In Aim 2, we will refine the design for manufacturability, and perform industrial design and human factors engineering. After finalizing the design of the cartridge and instrument, we will scale up manufacturing under ISO 13485 to produce a sufficient number of cartridges and instruments to execute Aim 3, which entails analytical and clinical validation of the system. These three aims will culminate in application for CE-IVD regulatory approval, to enable market entry. In Aim 4, we will develop a docking station to which four of the portable instrument units can be attached, as an upgrade to the basic system. This docking station will provide enhanced quality control, a better user interface, plus wireless connectivity, enabling external quality assurance and electronic result transmission, two important operational aspects of disseminated TB diagnostics in low resource near patient settings. This project presents a unique opportunity to enable early diagnosis of all TB cases, which was identified as a key objective in the Global Plan to Stop TB. In the long term, the system can be applied to other pathogens relevant to bio-threat detection and infectious disease diagnosis in resource-limited settings.

描述（由申请人提供）：该项目的目标是通过开发一种便携式、易于使用的集成核酸检测设备，在无需用户干预的情况下执行样品制备、等温 DNA 扩增和基于侧向层析的检测，在 < 1.5 小时的样品输入到应答输出时间内提供结果，从而以比目前显着降低的成本，在资源匮乏、高负担国家实现肺结核诊断。 可用的完全集成的台式系统。该项目建立在同一研究人员和机构团队持续努力的基础上，并通过新的合作伙伴得到加强。迄今为止，我们已经开发出一种用于痰液化和消毒、病原体裂解和 DNA 提取的新型快速样品制备工艺，并使用临床痰样品建立了该工艺的合适性能。我们已经证明，我们的系统可以适应与侧流检测相结合的不同等温扩增方法，并开发了一种用于稳定预混试剂的新工艺。展望未来，我们将利用经过临床验证可用于结核病诊断的等温交叉引发扩增 (CPA)，该扩增与侧向流动相结合，并包括内部扩增控制。我们设计并制造了集成盒和仪器的原型，并且证明了样品制备和扩增/检测子单元的功能。在拟议项目开始时，我们的目标是在集成 alpha 原型系统中演示完全自动化的流程执行。拟议的项目将使我们能够进一步开发和验证该系统，并获得监管部门批准作为 IVD 投放市场。在目标 1 中，我们将优化与样本预处理、DNA 提取、CPA 和层析检测相关的关键系统组件。我们将提高预混试剂的热稳定性，改进机载试剂存储，采用侧流试纸条读取器，并确保整体系统的稳健性。在目标2中，我们将细化可制造性设计，并进行工业设计和人因工程。在完成卡盒和仪器的设计后，我们将根据 ISO 13485 扩大生产规模，以生产足够数量的卡盒和仪器来执行目标 3，这需要对系统进行分析和临床验证。这三个目标最终将申请 CE-IVD 监管批准，以实现市场准入。在目标 4 中，我们将开发一个可以连接四个便携式仪器单元的扩展坞，作为基本系统的升级。该扩展坞将提供增强的质量控制、更好的用户界面以及无线连接，从而实现外部质量保证和电子结果传输，这是在患者附近资源匮乏的情况下进行播散性结核病诊断的两个重要操作方面。该项目为实现所有结核病病例的早期诊断提供了独特的机会，这被确定为全球遏制结核病计划的关键目标。从长远来看，该系统可以应用于资源有限环境中与生物威胁检测和传染病诊断相关的其他病原体。