A sensitive, low-cost, fluorescence assay for determination of HIV p24 in infants

一种灵敏、低成本的荧光测定法，用于测定婴儿中的 HIV p24

• 批准号: 8790356
• 负责人: Linda G Lee
• 金额: $ 15万
• 依托单位: SONG DIAGNOSTIC RESEARCH, LLC
• 依托单位国家: 美国
• 财政年份: 2014
• 资助国家: 美国
• 起止时间: 2014-08-06 至 2015-12-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8790356
• 关键词: Acute; Anti-Retroviral Agents; Antibodies; Antigen-Antibody Complex; Area; Bedside Testings; Binding; Biological Assay; Biological Models; Blood; Child; Complex; Computer software; Core Protein; Country; Coupled; Data; Data Analyses; Detection; Devices; Diagnostic Services; Dissociation; Dyes; Enzyme-Linked Immunosorbent Assay; Eye; Fluorescence; Fluorescent Dyes; Gold; HIV; HIV Antibodies; HIV Core Protein p24; Healthcare; Heating; Human; Human Chorionic Gonadotropin; Human immunodeficiency virus test; Image; Image Analysis; Immunoassay; Improve Access; Infant; Infection; Investigation; Laboratories; Lateral; Life; Lighting; Modeling; Monitor; Mothers; Newborn Infant; Optics; Outcome; Patients; Performance; Phase; Phycoerythrin; Process; Property; Proteins; Publishing; Reagent; Recombinant Proteins; Recombinants; Reporter; Reporting; Resources; Sampling; Serum; Stream; System; Techniques; Telephone; Testing; Vertical Disease Transmission; Viral Load result; Work; antiretroviral therapy; base; clinically significant; cost; design; detector; improved; innovation; internal control; mortality; point of care; prototype; public health relevance; reagent testing; transmission process

DESCRIPTION (provided by applicant): Each year, 1.5 million children are born to mothers living with HIV resulting in vertical transmission of HIV to newborns in 350,000 cases. Early testing followed by prompt anti-retroviral therapy (ART) is known to save lives, yet lack of point-of-care (POC) tests for infants results in delays or absence of therapy. Our approach will provide a low-cost yet quantitative and sensitive lateral flow immunoassay (LFIA) test to analyze and quantitate the presence of HIV p24. Fluorescent dyes will be used to improve the limit of detection compared to existing rapid tests. A smart phone is utilized for imaging, analysis and automated reporting. A long Stokes shift dye, R-phycoerythrin (PE) will be used as the fluorescent reporter, either singly or bundled into a cluster. The optical properties of PE allow the use of an inexpensive illumination device; the minimal non-specific binding of PE results in sensitive detection with a wide dynamic range. Additional gains in sensitivity will be achieved through an integrated test feature designed to dissociate target protein immune-complexes for improved access by the test reagents. In Phase I we will: 1) Optimize the PE reporter (cluster) on anti-HIV p24 antibody; 2) Develop a smartphone application to allow automated analysis; 3) Develop an on-LFIA strip "zone of dissociation" to aid in analysis of complexed p24; 4) Develop the use of the control line as a quantification standard on the lateral flow strip to allow absolut quantitation of p24; 5) Design, fabricate and test a prototype of a low cost fluorescence system designed with a materials cost less than $35. 6) Validate the p24 LFIAs by comparing results with commercially available ELISA and LFIA tests using heat inactivated serum spiked with recombinant HIV p24. These investigations will demonstrate the performance advantage of our system for detection and analysis of HIV p24. For phase II we hope to test authentic patient samples.

描述（由申请人提供）：每年有150万名儿童的母亲感染了艾滋病毒，导致35万例艾滋病毒垂直传播给新生儿。众所周知，早期检测之后及时进行抗逆转录病毒治疗（ART）可以挽救生命，但缺乏对婴儿的即时检测会导致治疗延误或缺乏治疗。我们的方法将提供一种低成本、定量和敏感的横向流动免疫测定（LFIA）测试来分析和定量HIV p24的存在。与现有的快速检测相比，荧光染料将用于提高检测极限。智能手机用于成像、分析和自动报告。一个长斯托克斯移位染料，r -植红蛋白（PE）将被用作荧光报告者，单独或捆绑成一个簇。PE的光学特性允许使用廉价的照明装置；极小的PE非特异性结合使得检测灵敏度高，动态范围宽。灵敏度的额外增益将通过一个集成的测试功能来实现，该测试功能设计用于解离靶蛋白免疫复合物，以改善测试试剂的访问。在第一阶段，我们将：1)优化抗hiv p24抗体PE报告（簇）；2)开发一个智能手机应用程序，允许自动分析；3)建立一个lfia上的条带“解离区”，以帮助分析络合的p24；4)开发使用控制线作为横向流带的定量标准，以实现p24的绝对定量；5)设计、制造和测试一个低成本荧光系统的原型，使用成本低于35美元的材料设计。6)将重组HIV p24热灭活血清与市售ELISA和LFIA检测结果进行比较，验证p24 LFIA的有效性。这些研究将证明我们的系统在检测和分析HIV p24方面的性能优势。对于第二阶段，我们希望测试真实的患者样本。