Rapid point of-care test for Lyme serodiagnosis based on novel ultra-sensitive de

描述(申请人提供)：建议的项目旨在开发和实施一种用于快速医学诊断测试的高灵敏度比色检测技术，并将其应用于基于多肽的莱姆病快速测试作为第一个例子。新技术的核心是一种新的多色底物系统，用于广泛使用的酶标记辣根过氧化物酶，该系统利用固定化的显色试剂。这项技术为应用酶联免疫分析技术开发高灵敏度的快速诊断试验开辟了新的机会。新的检测技术将提供目前传统技术无法达到的灵敏度水平，但保留了传统POC测试的简单性、低成本、可制造性和稳定性。这项新技术在POC测试中的应用将使开发快速测试成为可能，测试时间非常短，但分析灵敏度超过使用传统比色标记的测试，如胶体金或染色乳胶颗粒，以及微孔板ELISA。此外，灵敏度有望与目前正在开发的依赖于快速测试的新的复杂仪器技术的灵敏度相媲美。该技术将适用于目前所有主要形式的快速诊断、侧流、流过或浸渍-条带以及珠子过滤分析。新的稳定的衬底系统具有独特的多路复用功能，将在诊断膜上产生明亮的、高对比度的彩色区域，非常适合于视觉读取或密度测量。为了验证新检测技术在体外诊断试验中的适用性，将开发一种新的莱姆病血清侧向流动试验。目前，市场上还没有临床上有用的灵敏度和特异度的莱姆病护理点检测。新的快速检测将利用VlsE的C6多肽和OspC的C10多肽两种多肽抗原，并将显著提高对疾病早期诊断的敏感性，目前的方法包括微板法。快速检测的周转时间将为3-5分钟，全血作为样本类型将允许现场使用。新快速检测的性能特征将使用培养确认的莱姆病患者或医生记录的早期莱姆病患者和对照人群的特征良好的血清样本进行评估。 公共卫生相关性：该项目旨在开发一种新的比色检测技术，适用于就诊诊断测试，它比目前的方法具有更高的灵敏度。这项技术将为使用多色视觉标签的护理点测试提供快速周转、经济高效的解决方案。作为第一个应用，将开发一种基于多肽抗原的莱姆病快速检测方法，这将是第一个能够对早期莱姆病进行准确的医疗点血清诊断的方法。

DESCRIPTION (provided by applicant): The proposed project is aimed at the development and implementation of a highly sensitive colorimetric detection technology for use in rapid medical diagnostic tests, and its application to a peptide-based Lyme disease rapid test as a first example. At the core of the new technology is a novel multi-color chromogenic substrate system for the widely used enzyme label horseradish peroxidase utilizing immobilized forms of color-generation reagents. This technology opens new opportunities for the application of enzyme -linked immunoassays to development of highly sensitive rapid diagnostic tests. The new detection technology will provide a level of sensitivity which is currently unattainable with conventional technologies, but preserving the simplicity, low cost, manufacturability and stability of conventional POC tests. Implementation of the new technology in POC tests will make it possible to develop rapid tests with very short assay time but with analytical sensitivity exceeding that of tests utilizing conventional colorimetric labels, such as colloidal gold or dyed latex particles, as well as microplate ELISA's. Moreover, the sensitivity is expected to compare favorably with the sensitivities of new sophisticated instrument-dependant technologies currently in development for rapid tests. The technology will be applicable to all major formats of current rapid diagnostic, lateral flow, flow through or dip-strip as well as bead filtration assays. The new stable substrate system with its unique multiplexing capabilities will produce bright, high-contrast, colored zones on diagnostic membranes ideally suited for either visual reading or densitometry. To validate applicability of the new detection technology to in vitro diagnostic tests, a new serological lateral flow test for Lyme disease will be developed. At present, there is no point-of-care test for Lyme disease on the market with clinically useful sensitivity and specificity. The new rapid test will utilize two peptide antigens, the C6 peptide of VLsE and the C10 peptide of OspC, and will provide a significant improvement in the diagnostic sensitivity for early phases of the disease over current methods including microplate ELISA's. Turnaround time for the rapid test will be 3-5 minutes, and whole blood as sample type will allow point-of-care use. Performance characteristics of the new rapid test will be evaluated using well-characterized serum samples from Lyme patients with culture-confirmed or physician documented early stage Lyme disease, and on control populations. PUBLIC HEALTH RELEVANCE: This project is aimed at development of a new colorimetric detection technology suitable for point-of-care diagnostic tests, which offers significantly greater sensitivity than current methods. This technology will provide a rapid turnaround, cost-effective solution for point-of-care testing using multicolor visual labels. As a first application, a rapid test for Lyme disease will be developed based on peptide antigens, which will be the first to enable accurate point-of-care serodiagnosis for early Lyme disease.