A Multiplex Platform for Lyme Disease Diagnosis and Treatment Response

莱姆病诊断和治疗反应的多重平台

• 批准号: 8841437
• 负责人: MONICA E EMBERS
• 金额: $ 33.24万
• 依托单位: TULANE UNIVERSITY OF LOUISIANA
• 依托单位国家: 美国
• 财政年份: 2012
• 资助国家: 美国
• 起止时间: 2012-06-01 至 2017-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8841437
• 关键词: Aftercare; Antibiotics; Antibodies; Antibody Formation; Antigens; Binding; Biological Assay; Bite; Blood Circulation; Borrelia burgdorferi; Case Study; Centers for Disease Control and Prevention (U.S.); Clinical; Coupling; Detection; Diagnosis; Diagnostic; Diagnostic Procedure; Diagnostic tests; Disease; Effectiveness; Enzyme-Linked Immunosorbent Assay; Epitopes; Goals; Human; Immune Sera; Immunoblotting; Indium; Individual; Infection; Ixodes; Knowledge; Laboratories; Life; Lyme Disease; Measures; Monoclonal Antibodies; Nature; North America; OspA protein; OspC protein; Patients; Pattern; Peptides; Phase; Physicians; Process; Production; Proteins; Reporting; Sampling; Sensitivity and Specificity; Serologic tests; Serum; Signs and Symptoms; Source; Specificity; Staging; Systemic disease; Technology; Testing; Ticks; Time; Treatment Efficacy; Variant; Vector-transmitted infectious disease; Work; base; clinical Diagnosis; design; disease diagnosis; disorder prevention; immunogenic; improved; nonhuman primate; response; success; treatment response

ABSTRACT Lyme disease is the most commonly reported vector-borne disease in North America, with nearly 30,000 confirmed cases and > 8,500 probable cases reported to the CDC in 2009. The etiologic agent, B. burgdorferi is transmitted to humans via Ixodes ticks and spreads hematogenously to cause a systemic disease. The protean nature of this disease lends to difficult clinical diagnosis, so laboratory testing needs to be reliable. An improved diagnostic test for Lyme disease must detect infection: a) early (within 2 weeks of tick bite); b) across multiple B. burgdorferi genospecies, and c) in the different phases of disease. The ability of a diagnostic test to distinguish persistent infection from clinical cure would be an asset as well. New Luminex(R) technology incorporating monospecific antibodies bound to fluorometric beads has allowed for the quantification of multiple proteins within a single sample. We aim to reverse the process by coupling multiple to the beads for use in the detection of specific antibodies for the diagnosis of Lyme disease. Along with the C6 peptide, the most successful standalone diagnostic antigen for Lyme disease, we have carefully selected four additional antigens (DbpA, OspA, OspC and OppA-2) to incorporate into the initial testing of this platform. Each is broadly immunogenic, but elicits different responses over the course of infection. The overall goal is to produce a single diagnostic test for Lyme disease that is sensitive, specific and detects infection with multiple variants at all stages of disease. The objective of the proposed work is to create a platform test, based on Luminex(R) technology and our knowledge of antibody responses to B. burgdorferi throughout infection, which will overcome the deficiencies of current tests. The central hypothesis is that the combination of select antigens that elicit distinctive host antibody response patterns into a fluorometric bead-based quantitative assay will provide assay sensitivity and specificity exceeding that of current Lyme diagnostics. We propose the following aims (Phase I): (1) Proof of concept: to combine antigens into bead arrays for testing and optimization of sensitivity using monoclonal antibodies and immune serum. The working hypothesis for this aim is that monoclonal antibodies can be used to produce standard curves to determine levels of specific antibody in a clinical sample and to define the dynamic range of detection for each antigen; (2) Proof of feasibility for human use: to test assay specificity compared to standard ELISA using serum from multiple sources, including Lyme disease patients at different phases of disease (early, disseminated, or late- disseminated) versus healthy control patient serum. The working hypothesis for this aim is that by coupling to beads only 5 carefully-selected B. burgdorferi-specific purified antigens, the sensitivity of specific antibody detection will be enhanced without compromising specificity. Following the completion of these aims, in Phase II the diagnostic assay will be optimized, streamlined and tested against standard diagnostic methods currently employed for the diagnosis of Lyme disease.

摘要 莱姆病是北美最常见的媒介传播疾病， 2009年向CDC报告了30，000例确诊病例和> 8，500例可能病例。病原体，B。 伯氏疏螺旋体通过硬蜱传播给人类，并通过血源性传播引起全身性疾病。 这种疾病的多变性导致临床诊断困难，因此实验室检测需要可靠。 一种改进的莱姆病诊断测试必须检测感染：a）早期（蜱叮咬后2周内）; B） 在多个B之间。burgdorferi基因种，和c）在疾病的不同阶段。诊断的能力 区分持续性感染和临床治愈的测试也是一种资产。新的Luminex（R）技术 掺入与荧光珠结合的单特异性抗体允许定量 在一个单一的样品中的多种蛋白质。我们的目标是通过将多个耦合到珠子来逆转这个过程， 用于检测特异性抗体以诊断莱姆病。沿着C6肽， 最成功的莱姆病独立诊断抗原，我们精心挑选了四个额外的 抗原（DbpA、OspA、OspC和OppA-2）纳入该平台的初始检测。每一个都是 广泛的免疫原性，但在感染过程中产生不同的反应。总体目标是 针对莱姆病进行单一诊断测试，该测试敏感、特异并可检测多种感染 在疾病的各个阶段。建议工作的目标是创建一个平台测试，基于 Luminex（R）技术和我们对B抗体反应的知识。在整个感染过程中， 这将克服当前测试的不足。核心假设是， 选择引发独特的宿主抗体应答模式的抗原， 定量分析将提供超过当前莱姆诊断的分析灵敏度和特异性。我们 提出以下目标（I期）：（1）概念验证：将联合收割机抗原结合到珠阵列中进行检测 以及使用单克隆抗体和免疫血清优化灵敏度。工作假设 这一目的是单克隆抗体可用于产生标准曲线， （2）证明临床样品中的抗体，并确定每种抗原的检测动态范围; 人用途：与标准ELISA相比，使用来自多个 来源，包括疾病不同阶段的莱姆病患者（早期、播散性或晚期）， 播散的）与健康对照患者血清。这一目标的工作假设是，通过耦合到 珠子只有5个精心挑选的B。特异性纯化抗原，特异性抗体的敏感性 在不损害特异性的情况下，检测将得到加强。在完成这些目标后， II诊断分析将被优化，简化，并根据目前的标准诊断方法进行测试。 用于莱姆病的诊断。