An integrated platform for blood collection, HIV viral RNA purification, and stab

血液采集、HIV 病毒 RNA 纯化和刺杀的集成平台

• 批准号: 8740553
• 负责人: Erwin Berthier
• 金额: $ 20万
• 依托单位: TASSO, INC.
• 依托单位国家: 美国
• 财政年份: 2013
• 资助国家: 美国
• 起止时间: 2013-09-24 至 2015-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8740553
• 关键词: Acute; Address; Antibodies; Biological Markers; Biological Models; Biological Preservation; Blood; Blood specimen; Clinic; Clinical Research; Clinical Trials; Cold Chains; Collaborations; Collection; DNA; Development; Device Designs; Devices; Diagnostic; Diagnostic Procedure; Economics; Elements; Encapsulated; Environment; Equipment; Error Sources; Gold; HIV; Hour; Human immunodeficiency virus test; Infection; Joints; Laboratories; Light; Liquid substance; Methods; Microfluidic Microchips; Microfluidics; Motion; Nature; Needles; Nucleic Acids; One-Step dentin bonding system; Pain-Free; Patients; Phase; Population; Positioning Attribute; Preparation; Process; Puncture procedure; RNA; RNA purification; Radial; Research Institute; Resources; Reverse Transcription; Sampling; Serum; Shipping; Ships; Site; Skin; Small Business Innovation Research Grant; Solutions; Specialist; Spottings; Staging; Techniques; Technology; Testing; Time; Training; Uganda; Venipunctures; Viral; Viral Load result; Waxes; Whole Blood; base; cost; cost effective; experience; improved; interest; novel; nuclease; operation; particle; point of care; professor; prototype; public health relevance; sample collection; seal; viral RNA

DESCRIPTION (provided by applicant): Economic, logistical, and physical barriers limit HIV viral load quantification, the gold standard for HIV diagnostics. The cost of the test is high (approximately $100 per test) and the intricate equipment required limits testing to laboratories staffed by highly trained technicians. Also, the labile nature of the biomarker, viral RNA, require testing to be performed within 6 hours of blood draw unless the RNA can be stabilized, thus limiting the geographical reach of this test. Current point-of-care (POC) HIV tests rely on antibody titer, but this biomarker is limited in early, acute-stage infections, resulting in many false negatives. Additionally, strategies to preserve blood sample (e.g., dried blood spots) add additional steps to the workflow (e.g., spotting, drying, elution), resulting in added labor and increased potential for error. We propose a device that merges a novel blood collection method in commercial development by Tasso Inc., with a technology for sample preparation and stabilization developed at UW - Madison. Through a simple-to-use method integrating blood collection and sample preparation, the device will alleviate many of the barriers (time, cost, trained specialist) to the applications of HIV diagnostics. Further, a novel method for sample stabilization enables this method to be used and transported from any number of low-resource settings, expanding the reach of blood diagnostic methods. To achieve this project, Tasso is uniquely positioned through three key collaborations: 1) Professor David Beebe as an expert consultant for simple microfluidic devices for sample preparation with startup experience in 4 different companies, 2) Dr. Frank Graziano as an expert consultant for diagnostics for low-resource settings with a special affiliation with the Joint Clinical Research Centre (JCRC) in Uganda, and 3) the Morgridge Institutes for Research, enabling Tasso to have direct access to established prototyping facilities, along with access to experts in device design for manufacturability (including Thomas Mackie, founder of TomoTherapy). The proposal consists of three Aims. First, the Tasso device for blood acquisition will be modified for economically-limited deployment by making the non-blood contacting portion of the device reusable. Next, a one-step sample preparation cartridge, the SLIDE, will be optimized to extract viral RNA from blood samples collected with the Tasso device. Lastly, a wax-based sealing mechanism will be integrated into the SLIDE to enable on-chip reverse transcription, which stabilizes viral RNA into DNA, and sealing for transport. At the conclusion of Phase I, an integrated, optimized proof-of-concept platform will be capable of blood acquisition, viral RNA extraction, and RNA stabilization and storage. In Phase II, this prototype will be tested on patients with the assistance of consultant Dr. Frank Graziano, an expert in HIV- related clinical trials. Additionall, a nucleic acid amplification / quantitation component will be developed, enabling complete sample-to-answer operation and further mitigating barriers to HIV viral load testing access.

描述（由申请人提供）：经济、后勤和物理障碍限制了HIV病毒载量定量，这是HIV诊断的金标准。测试的成本很高（每次测试大约100美元），所需的复杂设备限制了由训练有素的技术人员组成的实验室进行测试。此外，生物标志物病毒RNA的不稳定性要求在抽血后6小时内进行检测，除非RNA能够稳定，从而限制了该检测的地理范围。目前的即时艾滋病毒检测依赖于抗体滴度，但这种生物标志物在早期急性期感染中受到限制，导致许多假阴性。此外，保存血液样本的策略（例如，干燥的血点）增加了工作流程的额外步骤（例如，斑点，干燥，洗脱），导致增加了劳动力和增加了出错的可能性。我们提出了一种设备，它融合了Tasso公司在商业开发中的一种新的血液采集方法，以及威斯康星大学麦迪逊分校开发的一种样品制备和稳定技术。通过一种集血液采集和样品制备于一体的简单易用的方法，该设备将减轻艾滋病诊断应用的许多障碍（时间、成本、训练有素的专家）。此外，一种新的样品稳定方法使该方法能够在任何数量的低资源环境中使用和运输，扩大了血液诊断方法的范围。为了实现这个项目，Tasso通过三个关键合作处于独特的地位：1) David Beebe教授是样品制备用简单微流体装置的专家顾问，在4家不同的公司有创业经验；2)Frank Graziano博士是资源匮乏地区的诊断专家顾问，与乌干达联合临床研究中心（JCRC）有特殊的联系；3)Morgridge研究所，使Tasso能够直接使用已建立的原型设施。同时还可以接触到设备可制造性设计方面的专家（包括TomoTherapy的创始人托马斯·麦基）。该建议包括三个目标。首先，将对用于血液采集的Tasso装置进行修改，使该装置的非血液接触部分可重复使用，从而在经济上限制部署。接下来，一步制样盒SLIDE将被优化，用于从Tasso装置收集的血液样本中提取病毒RNA。最后，将蜡基密封机制集成到SLIDE中，以实现芯片上的逆转录，从而将病毒RNA稳定为DNA，并密封运输。在I期结束时，一个集成的，优化的概念验证平台将能够采集血液，病毒RNA提取，RNA稳定和储存。在第二阶段，这个原型将在顾问Frank Graziano博士的帮助下在患者身上进行测试，Frank Graziano博士是HIV相关临床试验的专家。此外，将开发核酸扩增/定量组件，实现完整的样本到答案操作，并进一步减轻艾滋病毒载量检测的障碍。