Proof-of-Concept for Card-based CD4 Cell Counting

描述（由申请人提供）：这项拟议研究的长期目标是开发一种简单、廉价的即时检测设备，该设备将对全血中的CD 4 + T细胞进行绝对计数和百分比计数。这将通过调整和测试一种新的“混合”方法来实现微流体样品制备和分析。一次性使用的薄膜层压微流体试剂卡将收集、计数和储存血液样本，同时将储存所需的混合试剂并从台式装置注入试剂卡中。集成系统将使用一次性微流体试剂卡上包含的新体积组合电阻抗/荧光传感器进行总白色血细胞计数、绝对CD 4 + T细胞计数和CD 4计数百分比。为了实现这一目标，第二阶段将扩大我们的第一阶段的努力，并专注于系统优化和验证。目的之一是优化微流控盒设计和CD 4血液标记方案，用于自动化、卡上CD 4 + T细胞计数。第二个目标将集中在制造和测试的综合台式系统。目标三将使用改良的市售正常人全血和来自HIV+患者的临床样本测试和确认完整的CD 4计数系统，并开始获得FDA批准的II类医疗器械所需的实验室研究。当在拟议的使用场景中进行充分验证和评估时，所产生的设备将是简单的，强大的和独特的，因为它提供了一个完全定量的CD 4 + T细胞分析，仪器成本低于1000美元。这比已建立的流式细胞术装置的成本低许多倍。试剂和一次性用品的成本也将低于任何目前可用的设备，每次测试的总成本低于1.50美元。该工具将立即应用于各级公共保健系统，特别是在资源贫乏地区和发展中国家，以监测感染人体免疫缺陷病毒（艾滋病毒）的人，作出鉴别诊断，并就抗逆转录病毒药物治疗制度，包括高效抗逆转录病毒疗法（HAART）办法作出治疗决定。公共卫生相关性廉价便携式CD 4计数器的成功完成将为医疗保健提供者提供一个更便宜的移动的平台，以进行目前世界上最常见的CD 4诊断测试，从而在艾滋病毒监测和治疗方面取得重大突破。这一突破将大大提高其一次性测试盒的测试效率，并使其能够在床边，偏远地区或医疗资源有限的地区快速测试患者。最后，这种新的即时系统可以显著降低当前的CD 4设备和每次测试的成本，从而为美国医疗保健和保险行业节省数百万美元，并在发展中国家产生更大的全球医疗保健影响。

DESCRIPTION (provided by applicant): The long-term objective of this proposed research is to develop a simple, inexpensive point-of-care device that will perform absolute and percent CD4+ T cell counts from whole blood. This will be accomplished by adapting and testing a novel "hybrid" approach to microfluidic sample preparation and analysis. One-time use thin-film laminated microfluidic cassettes will collect, count, and store the blood sample while the required cocktail reagents will be stored and injected into the cassettes from a bench-top unit. The integrated system will perform a total white blood cell count, absolute CD4+ T cell count, and percent CD4 count using a new volumetric combination electric impedance/fluorescence sensor contained on a disposable microfluidic cassette. Toward this objective, Phase II will expand on our Phase I effort and focus on system optimization and validation. Aim one will optimize the microfluidic cassette design and CD4 blood labeling protocol for automated, on-card CD4+ T cell counting. Aim two will focus on fabrication and testing of the integrated bench-top system. Aim three will test and validate the completed CD4 enumeration system, using both modified commercially available normal human whole blood and clinical samples from HIV+ patients, and begin the bench-top studies required to obtain FDA clearance for a Class II medical device. The resulting device, when fully validated and evaluated in the proposed scenarios of use, will be simple, robust, and unique in that it provides a fully quantitative CD4+ T cell analysis with an instrument costing less than $1000. This is many times below the cost of established flow cytometry devices. Reagent and disposable costs will also be lower than that of any currently available device with per-test total costs of less than $1.50. The tool will find immediate application in all levels of the public health care system, particularly in resource-poor areas and developing countries, for monitoring persons with human immunodeficiency virus (HIV), formulating differential diagnoses, and making therapeutic decisions regarding anti-retroviral drug treatment regimes, including the highly active anti-retroviral therapy (HAART) approach. PUBLIC HEALTH RELEVANCE The successful completion of an inexpensive, portable CD4 counter would constitute a significant breakthrough in HIV monitoring and treatment by offering healthcare providers a less expensive, mobile platform to conduct the most common CD4 diagnostic test currently performed in the world. Such a breakthrough will dramatically improve testing efficiencies with its single-use, disposable test cartridges and will make it possible to rapidly test patients at bedside, in remote locations, or in areas with limited healthcare resources. Finally, this new point-of-care system could significantly reduce current CD4 equipment and per-test costs, thereby resulting in millions of dollars saved in the US healthcare and insurance industries, with even greater worldwide healthcare impacts in developing countries.