Ultrasensitive Bispecific Antibody Assay for SARS Virus

SARS 病毒超灵敏双特异性抗体测定

• 批准号: 7237955
• 负责人: MAVANUR R SURESH
• 金额: $ 18.12万
• 依托单位: UNIVERSITY OF ALBERTA
• 依托单位国家: 美国
• 财政年份: 2005
• 资助国家: 美国
• 起止时间: 2005-03-15 至 2010-02-28
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7237955
• 关键词: Affinity; Alberta province; Aluminum; Antibodies; Antibody Binding Sites; Antigen Targeting; Antigens; Architecture; Bacteria; Binding; Biological Assay; Bispecific Antibodies; Bispecific Monoclonal Antibodies; Body Fluids; Canada; Cells; Community Health; Detection; Diagnostic; Diagnostic tests; Enzyme-Linked Immunosorbent Assay; Epitopes; Evaluation; Glycoproteins; Gold; Health; Health Personnel; Hybridomas; Immunoassay; Individual; Injection of therapeutic agent; Label; Laboratories; Membrane Glycoproteins; Microbiology; Molecular; Multiple Myeloma; Mus; Nose; Nucleocapsid Proteins; Patients; Peroxidase; Peroxidases; Pharyngeal structure; Proteins; Protocols documentation; Recombinants; Reproduction spores; Research Design; Risk; SARS coronavirus; Safety; Saliva; Sampling; Screening procedure; Serum; Severe Acute Respiratory Syndrome; Specificity; Splenocyte; Sputum; Swab; Testing; Therapeutic; Universities; Virus; Work; base; design; env Gene Products; point of care; prototype

DESCRIPTION (provided by applicant): I. BROAD,LONG-TERM OBJECTIVES: The current objective is to develop inexpensive point of care diagnostics using new monoclonals against SARS antigens. The same antibodies may have therapeutic utility as well. II. SPECIFIC AIMS: (a)To develop high affinity MAbs against SARS associated envelope (E), membrane glycoprotein (M), the nucleocapsid protein (NP) or spike proteins (SP) useful in homosandwich(repeat epitopes as in whole virus) or heterosandwich (as in shed monomeric antigens) assay; (b) To develop bispecific antibodies with specificity to NP (or SG) and HRPO; (c) To optimize an ultrasensitive molecular velcro homosandwich assay of the whole SARS virus on microplates(d) To develop an immunoswab or dipstick assay for samples collected from throat or sputum for screening applications. III. RESEARCH DESIGN: The various SARS antigens will be used to immunize mice by short-term and long-term protocols with s.c., i.p.,and i.v. injections. Immunized splenocytes and myeloma cells will be fused to obtain antigen specific hybridomas. The hybridoma supernatants from different clones will be tested for their ability to detect antigens in homosandwich and heterosandwich formats with recombinant antigen. We will also develop quadromas to generate bispecific monoclonal antibodies (bsMAb) with one paratope capable of binding to SARS antigen and the other to an enzymatic marker like peroxidase. We will explore different ELISA formats for detection of whole virus, for NP or other shed antigens. The immunoswab assay will be optimized using the commercially available nasopharyngeal swab with a flexible aluminum shaft for direct detection of SARS virus or its shed antigen using bsMAbs. Alternatively, we will also test the utility of our Mabs labeled with collidal gold to develop a prototype immunoswab assay that could potentially be used for rapid screening of suspected individuals. The simplicity of the proposed immunoswab assay is its utility in screening at airports and high risk communities and health care workers.

描述（申请人提供）：I.广泛、长期的治疗：目前的目标是使用新的抗SARS抗原的单克隆抗体开发廉价的护理点诊断。相同的抗体也可以具有治疗效用。二.具体目标：（a）开发抗SARS相关包膜（E）、膜糖蛋白（M）、核衣壳蛋白（NP）或刺突蛋白（SP）的高亲和力单克隆抗体，（重复表位，如在整个病毒中）或异源夹心（B）开发对NP（或SG）和HRPO具有特异性的双特异性抗体;（c）优化在微孔板上对整个SARS病毒进行的超灵敏分子Velcro homosandwich分析。（d）开发用于筛查应用的从喉咙或痰中收集的样品的免疫拭子或试纸分析。 三.研究设计：各种SARS抗原将被用于通过短期和长期的皮下注射方案免疫小鼠，腹腔注射，和静脉注射。 将免疫的脾细胞和骨髓瘤细胞融合以获得抗原特异性杂交瘤。将检测来自不同克隆的杂交瘤上清液检测具有重组抗原的同夹心和异夹心形式的抗原的能力。我们还将开发quadromas以产生双特异性单克隆抗体（bsMAb），其中一个互补位能够结合SARS抗原，另一个能够结合酶标记物如过氧化物酶。我们将探索用于检测全病毒、NP或其他脱落抗原的不同ELISA形式。将使用市售鼻咽拭子（带有柔性铝杆）优化免疫拭子检测方法，以便使用bsMAbs直接检测SARS病毒或其脱落抗原。 或者，我们也将测试我们的单克隆抗体的实用性与collidal金标记，以开发一个原型免疫分析，可能用于快速筛选可疑的个人。所提出的immunoswab测定的简单性在于其在机场和高风险社区和卫生保健工作者中的筛查中的实用性。