Self-test HIV diagnostics utilizing structurally novel, shark-derived binding domains

利用结构新颖的鲨鱼衍生结合域进行 HIV 自检诊断

• 批准号: 10539323
• 负责人: Helen M. Dooley
• 金额: $ 46.94万
• 依托单位: UNIVERSITY OF MARYLAND BALTIMORE
• 依托单位国家: 美国
• 财政年份: 2022
• 资助国家: 美国
• 起止时间: 2022-01-01 至 2024-12-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10539323
• 关键词: 3-Dimensional; Acquired Immunodeficiency Syndrome; Acute; Address; Affinity; Animals; Antibodies; Antigen-Antibody Complex; Antigens; Area; Binding; Biological; Biological Assay; Blood; Blood specimen; Buffers; Capsid Proteins; Cellular Phone; Clinic Visits; Coupling; Data; Databases; Detection; Diagnosis; Diagnostic; Diagnostic tests; Dissociation; Drops; Drug resistance; Early Diagnosis; Environment; Epidemiology; Epitopes; Equipment; Feedback; Fingers; Generations; HIV; HIV Core Protein p24; HIV Infections; HIV Seropositivity; HIV-1; Harvest; Health Personnel; Healthcare; Home; Human; Human immunodeficiency virus test; Immunize; Individual; Interruption; Laboratories; Libraries; Link; Microfluidics; Microspheres; Monitor; Monoclonal Antibodies; Nurses; Outcome; Patients; Performance; Peripheral Blood Lymphocyte; Phase; Porosity; Privatization; Proteins; Reader; Reagent; Recombinants; Reproducibility; Resources; Risk; Sampling; Secure; Self Administration; Sepharose; Serology test; Shark; Specialist; Specificity; Surface; System; Tablets; Target Populations; Technology; Testing; Time; Training; Validation; Viral; Viral Load result; Virus; Weight; accurate diagnostics; antiretroviral therapy; assay development; cloud based; detection platform; diagnostic platform; diagnostic technologies; diagnostic tool; digital; drug resistant virus; falls; home test; imaging system; improved; microchip; minimally invasive; novel; point of care; programs; progression risk; prototype; repository; response; self testing; sensor; thermostability; virology

PROJECT SUMMARY Since the first serologic tests were introduced at the peak of the AIDS crisis in the 1980s, continuous improvements in diagnostic technology have enabled the earlier detection and routine monitoring of HIV positive individuals. However, there are two areas where current HIV diagnostics fall short of ideal, and which we will attempt to directly address in this proposal; the first is the need for a simple diagnostic tool that allows individuals to self‐test for acute phase HIV infections. The second challenge is the provision of a diagnostic tool that will allow patients previously diagnosed as HIV positive to monitor their viral load following ART interruption or check for loss of viral control due to the emergence of drug‐ resistance. Given these diagnostics are to be used in a patients own home (or other non‐clinical, setting), they need to be cheap, robust, and suitable for use without prior training or specialist equipment. Further, they should use a biological sample that can be obtained in a minimally invasive manner (e.g. a finger‐stick blood drop) to encourage frequent retesting in target populations. To address the above problems, we will integrate two existing technologies to generate a sensitive home‐test diagnostic for HIV. The first technology is a structurally novel binding domain, so called VNARs, that we isolate from immunized sharks. Despite their diminutive size (12kDa), the VNAR domains raised thus far have binding affinities equal to classical antibodies but are intrinsically much more chemo‐ and thermostable. VNARs interact with antigen in unique ways and can be raised against epitopes that are inaccessible to conventional antibodies; we will exploit this fact to target the HIV proteins p24 and Env, allowing us to capture and accurately quantify free proteins or whole virus in unmanipulated blood. We will integrate these VNARs with our second technology, the programmable Bio‐Nano‐Chip (p‐BNC), a microchip‐ based detection system which utilizes porous agarose microbeads as 3D diagnostic surfaces. Immunometric assays can be performed by loading the beads with biomolecules such as antibodies, or in this case VNARs, allowing the capture and quantitation of desired target(s) in biological samples. To enable use of this technology in a non‐clinical setting, we will develop credit card‐sized diagnostic cartridges that have a microbead‐based sensor array at their core, and microfluidic system for the delivery of sample, wash buffer, and detection reagent. This will be deployed and read by a battery‐ powered handheld reader system that will be developed and fabricated during our project. This system will use an imaging system derived from smartphones and enable the safe and secure uploading of data to a cloud‐based repository. Together these technologies will allow us to deliver an inexpensive, robust, sensitive, and accurate diagnostic test that can be used by an individual to assess their HIV status or closely monitor their viral load, without a clinic visit.

项目摘要 由于第一个血清学测试是在1980年代艾滋病危机的高峰期引入的，因此持续改进 在诊断技术中，已经使艾滋病毒阳性个体的较早检测和常规监测。然而， 在两个领域，当前的艾滋病毒诊断不理想，我们将尝试直接解决这一问题 提议;首先是需要一种简单的诊断工具，该工具允许个人自我测试以患急性期HIV感染。 第二个挑战是提供诊断工具，该工具将允许先前被诊断为HIV阳性的患者 由于药物出现而导致的ART中断或检查病毒控制丧失后，监测其病毒载荷 反抗。鉴于这些诊断要在患者自己的家（或其他非临床设置）中使用，它们需要是 便宜，健壮且适合不用事先培训或专业设备使用。此外，他们应该使用生物学 可以以微创方式获得的样本（例如，手指粘液下降），以鼓励经常重新测试 在目标人群中。 为了解决上述问题，我们将集成两种现有技术以产生敏感的房屋测试 艾滋病毒的诊断。第一种技术是一个结构新颖的绑定域，所谓的VNAR，我们从 免疫鲨鱼。尽管它们的大小较小（12KDA），但迄今为止提出的VNAR域具有相等的结合亲和力 对于经典抗体，但本质上更加化学和热稳定性。 vnars与抗原互动独特 可以与传统抗体无法访问的表位来提出。我们将利用这一事实为目标 HIV蛋白p24和Env，使我们能够在未经操纵的 血。我们将将这些VNAR与我们的第二种技术，可编程的Bio-Nano-Chip（P-BNC），一种微芯片 - 基于基于多孔琼脂糖微粒作为3D诊断表面的检测系统。免疫测定可能是 通过用抗体等生物分子加载珠子或在这种情况下，在这种情况下，可以捕获和 生物样品中所需靶标的定量。为了在非临床环境中使用该技术，我们将 开发信用卡大小的诊断墨盒，其核心具有基于微粒的传感器阵列，而微流体 用于输送样品，洗涤缓冲液和检测试剂的系统。这将由电池部署和阅读 将在我们项目期间开发和制造的电力手持式阅读器系统。该系统将使用成像 从智能手机派生的系统，可以将数据安全上传到基于云的存储库中。一起 这些技术将使我们能够提供廉价，健壮，敏感和准确的诊断测试 由个人评估其艾滋病毒状况或密切监测其病毒负荷的情况，没有诊所就诊。