Recombinant antigen assays for Lassa and other arenaviru

拉沙病毒和其他沙粒病毒的重组抗原测定

• 批准号: 7009491
• 负责人: Robert F Garry
• 金额: $ 386.06万
• 依托单位: TULANE UNIVERSITY OF LOUISIANA
• 依托单位国家: 美国
• 财政年份: 2005
• 资助国家: 美国
• 起止时间: 2005-09-30 至 2009-09-29
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7009491
• 关键词: Arenaviridae; Lassa virus; Macaca mulatta; bioterrorism /chemical warfare; cell line; clinical research; clone cells; communicable disease diagnosis; cooperative study; diagnosis design /evaluation; diagnostic tests; enzyme linked immunosorbent assay; glycoproteins; guinea pigs; hemorrhagic fever; human subject; humoral immunity; immunoglobulin G; immunoglobulin M; laboratory mouse; monoclonal antibody; nucleoproteins; patient oriented research; recombinant proteins; serology /serodiagnosis; virus antigen; virus genetics; virus protein

DESCRIPTION (provided by applicant): This project will develop Diagnostics for Biodefense against Lassa virus (LASV), Junin virus and several other arenaviruses that induce severe, often fatal, hemorrhagic diseases, and are classified as Biosafety Level 4 and NIAID Biodefense Category A agents. The goal of our proposed study is to develop and validate multi-agent diagnostic immunoassays for arenaviruses. Specific Aim 1 is to clone cell lines producing murine and rhesus macaque monoclonal antibodies (MAb) specific for LASV glycoproteins and nucleoprotein. LASV glycoproteins GPC, GP1 and GP2, and the nucleoprotein (NP) will be expressed in bacteria or eukaryotic cell lines, and the recombinant proteins will be used to immunize mice and rhesus macaques. Specific Aim 2 is to develop LASV antigen-capture and immunoglobulin M (IgM) and immunoglobulin G (IgG) antibody-capture enzyme-linked immunosorbent assays (ELISA). These assays will be directly compared for sensitivity and specificity to current assays based on BSL4 grown virus, and optimized for the ability to detect various strains of LASV, including Josiah, Macenta, Z132 and LP. Specific Aim 3 is to characterize the humoral immune response to LASV virus proteins in adult rhesus macaques and validate the antigen-capture and antibody-capture ELISA for rhesus serological samples. Cross-reactive epitopes of arenavirus glycoproteins and nucleoprotein based on murine and rhesus MAb and human antibodies from LASV infected patients will be identified, including early IgM specific epitopes that appear to be the most important diagnostically. Specific Aim 4 is to broaden the specificity of the antigen-capture and antibody-capture ELISA to include both Old World arenaviruses and New World arenaviruses with potential for use as bioweapons. Specific Aim 5 is to validate the antigen-capture and antibody-capture ELISA using confirmed clinical samples and control samples field-collected from Guinea, Sierra Leone and South America. The potential use of arenaviruses as biological weapons directed against civilian or military targets necessitates development of effective diagnostics. The multi-agent diagnostic immunoassays for arenaviruses would be used to determine the attack agent following a deliberate release, and allow the virus used to be distinguished from other hemorrhagic fever viruses such as dengue virus or Ebola virus that may have similar case presentations. Development of rapid immunodiagnostic assays will also improve treatment of arenaviral diseases, facilitate studies to understand their prevalence and natural history, and lead to vaccines for preventing these major causes of morbidity and mortality.

描述（由申请人提供）：该项目将开发针对拉沙病毒（LASV）、胡宁病毒和其他几种沙粒病毒的生物防御诊断，这些沙粒病毒会诱发严重的、往往致命的出血性疾病，并被归类为生物安全 4 级和 NIAID 生物防御 A 类药剂。我们提出的研究的目标是开发和验证沙粒病毒的多试剂诊断免疫测定法。具体目标 1 是克隆产生 LASV 糖蛋白和核蛋白特异性鼠和恒河猴单克隆抗体 (MAb) 的细胞系。 LASV糖蛋白GPC、GP1和GP2以及核蛋白（NP）将在细菌或真核细胞系中表达，重组蛋白将用于免疫小鼠和恒河猴。具体目标 2 是开发 LASV 抗原捕获以及免疫球蛋白 M (IgM) 和免疫球蛋白 G (IgG) 抗体捕获酶联免疫吸附测定 (ELISA)。这些检测方法将直接与基于 BSL4 生长病毒的当前检测方法进行敏感性和特异性比较，并针对检测各种 LASV 毒株（包括 Josiah、Macenta、Z132 和 LP）的能力进行优化。具体目标 3 是表征成年恒河猴对 LASV 病毒蛋白的体液免疫反应，并验证恒河猴血清学样品的抗原捕获和抗体捕获 ELISA。将鉴定基于小鼠和恒河猴 MAb 以及来自 LASV 感染患者的人类抗体的沙粒病毒糖蛋白和核蛋白的交叉反应表位，包括似乎在诊断上最重要的早期 IgM 特异性表位。具体目标 4 是扩大抗原捕获和抗体捕获 ELISA 的特异性，以包括具有用作生物武器潜力的旧世界沙粒病毒和新世界沙粒病毒。具体目标 5 是使用从几内亚、塞拉利昂和南美洲现场收集的经确认的临床样本和对照样本来验证抗原捕获和抗体捕获 ELISA。沙粒病毒作为针对民用或军事目标的生物武器的潜在用途需要开发有效的诊断方法。沙粒病毒的多试剂诊断免疫分析将用于确定故意释放后的攻击剂，并允许将病毒与可能具有类似病例表现的其他出血热病毒（例如登革热病毒或埃博拉病毒）区分开来。快速免疫诊断检测的发展还将改善沙粒病毒疾病的治疗，促进研究以了解其患病率和自然史，并开发出预防这些发病和死亡主要原因的疫苗。

项目成果

Bacterial-based systems for expression and purification of recombinant Lassa virus proteins of immunological relevance.

• DOI: 10.1186/1743-422x-5-74
• 发表时间: 2008-06-06
• 期刊: Virology journal
• 影响因子: 4.8
• 作者: Branco LM;Matschiner A;Fair JN;Goba A;Sampey DB;Ferro PJ;Cashman KA;Schoepp RJ;Tesh RB;Bausch DG;Garry RF;Guttieri MC
• 通讯作者: Guttieri MC