LAMPoles for Dengue Diagnosis

用于登革热诊断的 LAMPoles

• 批准号: 8969846
• 负责人: Dennis John Grab
• 金额: $ 26万
• 依托单位: JOHNS HOPKINS UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2015
• 资助国家: 美国
• 起止时间: 2015-07-01 至 2016-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8969846
• 关键词: Acids; Acute; Affinity; African; Americas; Antibodies; Antibody Response; Antigens; Archives; Benign; Binding; Biological Assay; Blood; Blood Circulation; Body Fluids; Cells; Centers for Disease Control and Prevention (U.S.); Cessation of life; Chalcone synthase; Chimera organism; Clinical; Clinical Management; Clinical Trials; Country; DNA; DNA-Directed DNA Polymerase; Dengue; Dengue Shock Syndrome; Dengue Virus; Detection; Diagnosis; Diagnostic; Diagnostic tests; Disease; Disease Outbreaks; Early Diagnosis; Enrollment; Enzyme Gene; Enzyme-Linked Immunosorbent Assay; Epidemiology; Evolution; Excision; Fever; Genetic; Hemorrhage; Human; Human body; IgG2; IgG3; Immune; Immunoglobulin Fragments; Immunoglobulin G; Immunoglobulin M; Infection; Intervention; Ligand Binding; Light; Link; Malaria; Malaysia; Measurement; Measures; Mediating; Memory; Methods; Morbidity - disease rate; Mutation; Neutralization Tests; Nicaragua; Oligonucleotides; PTPN11 gene; Pathogenesis; Patients; Peptides; Phase; Physiological; Plants; Polymerase Chain Reaction; Positioning Attribute; Prevention Protocols; Procedures; Protein Dynamics; Proteins; Public Health; Reaction; Reading; Reagent; Recombinants; Sampling; Sensitivity and Specificity; Serologic tests; Serotyping; Serum; Sri Lanka; Staging; Supportive care; Tadpoles; Tail; Technology; Testing; Time; Trypanosoma; Vaccination; Vaccines; Vector-transmitted infectious disease; Viral Antigens; Virus; Visual; accurate diagnosis; anti-IgM; antibody engineering; antigen binding; antimicrobial; base; care seeking; design; disorder prevention; improved; innovation; mortality; nanobodies; point of care; population based; prevent; prototype; public health relevance; small molecule; tool; trend

 DESCRIPTION (provided by applicant): Dengue is a major public health threat throughout the tropics and subtropics will considerable morbidity and mortality It is the most widespread vector-borne disease after malaria and cannot be prevented by vaccination now Dengue can progress rapidly from benign febrile illness to dengue hemorrhage fever (DHF) and/or dengue shock syndrome (DSS) and death; supportive care is important to limit morbidity and mortality from complicated dengue. Hence, the ability to diagnose dengue early and accurately is important for clinical management of the disease. This proposal is innovative in that links 3 powerful technologies - LAMP, `tadpoles' and Nanobodies (Nbs) - to create a dynamic protein-DNA chimera-based diagnostic for DF/DHF with high specificity and sensitivity compared with existing technologies. We successfully created protein-DNA chimeras, termed `LAMPoles', by fusing DNA oligonucleotide tails based on a sequence of a plant enzyme gene with a previously described protein L/G-DNA chimera whose protein moiety binds most classes of mammalian antibodies to create a protein L/G LAMPole for ultrasensitive detection of essentially any mammalian IgG. For proof-of-concept we show that protein LG-LAMPoles enhanced the sensitivity of ELISA to detect host anti-African trypanosome antibodies in blood several hundred-fold leading to our hypothesis that improved detection of acute dengue is also possible. The discovery of single chain camelid antibody fragments called Nanobodies (Nb) is a major breakthrough in antibody engineering that will enable measurement of DV Ags even in the presence of host Abs. By creating LAMPoles from Nbs targeting both human IgM and DV antigens, we will create an ultrasensitive and specific assay for the diagnosis of acute dengue. Our team has archived serum and blood from patients in Malaysia, Sri Lanka, and Nicaragua with confirmed and excluded dengue. We will generate LAMPoles from camelid nanobodies (Nbs) specific for DV antigens (Ags) and evaluate the sensitivity and specificity of LAMPoles versus state-of-the-art diagnosis of acute dengue (vs. past and no dengue) using archived clinical samples.

 描述（由申请人提供）： 登革热是整个热带和亚热带地区的主要公共卫生威胁，具有相当大的发病率和死亡率。 它是继疟疾之后最广泛的媒介传播疾病，目前无法通过疫苗接种来预防。 登革热可以从良性发热性疾病迅速进展登革热出血热（DHF）和/或登革热休克综合征（DSS）和死亡；支持治疗对于限制复杂登革热的发病率和死亡率非常重要。早期准确诊断登革热的能力对于该疾病的临床管理非常重要，该提案的创新之处在于将 LAMP、“蝌蚪”和纳米抗体 (Nbs) 等 3 种强大技术联系起来，以创建基于蛋白质-DNA 嵌合体的动态诊断。与现有技术相比，DF/DHF 具有高特异性和灵敏度，通过将基于植物酶基因序列的 DNA 寡核苷酸尾部与先前描述的蛋白质 L/G-DNA 嵌合体，其蛋白质部分结合大多数类别的哺乳动物抗体，产生蛋白质 L/G LAMPole，用于基本上任何哺乳动物 IgG 的超灵敏检测。 ELISA 检测血液中宿主抗非洲锥虫抗体的灵敏度提高了数百倍，这使得我们假设改进急性登革热的检测也是可能的，发现了称为单链骆驼抗体片段。纳米抗体 (Nb) 是抗体工程领域的一项重大突破，即使在存在宿主抗体的情况下也能测量 DV Ag。通过从针对人类 IgM 和 DV 抗原的 Nb 中创建 LAMPole，我们将创建一种超灵敏且特异的诊断检测方法。我们的团队已经存档了马来西亚、斯里兰卡和尼加拉瓜确诊和排除的登革热患者的血清和血液，我们将从骆驼纳米抗体 (Nbs) 中产生 LAMPole。对 DV 抗原 (Ag) 具有特异性，并使用存档的临床样本评估 LAMPole 与最先进的急性登革热诊断（与过去和无登革热）的敏感性和特异性。