Synthetic Peptide SARS Coronavirus Diagnostic Kit

描述(由申请人提供)： 其长期目标是开发一种基于合成肽抗原的严重急性呼吸综合征(SARS)的酶联免疫分析(ELISA)。该检测将是稳定的，没有生物危险的制造操作和成分，旨在作为一种安全和快速的血清学监测方法。该测试将适用于大规模生产，并与全自动诊断筛查兼容。具体目标是1)鉴定和优化多肽抗原，包括来自非结构SARS冠状病毒(SCOV)蛋白的抗原，以及已鉴定的结构蛋白S、M和N的多肽；2)研制一种针对敏感性和特异性进行优化的SARS EL ISA检测试剂盒；3)用一组SARS患者的特征血清和未接触SCOV的受试者的多个试剂盒进行验证；4)在美国、台湾和中国的多个地点进行大规模临床试验；4)完成试剂盒的商业化生产；以及5)提交PMA或PDP申请，以获得美国和外国监管部门的批准。以CoVOV S、M和N结构蛋白的多肽抗原为抗原的SARS ELISA原型将通过添加非结构多肽进行改进，这些非结构多肽的序列来自CoVOV基因组的Pol区。重叠的多肽将被合成以覆盖整个pol1a和lb开放阅读框架。多肽的长度为20-60个氨基酸。个别多肽将通过酶联免疫吸附试验方法进行筛选，以检测其对SARS确诊患者血清的反应性。多肽上的血清反应性表位将被提炼，以获得最佳的序列和构象。新发现的非结构多肽抗原将被添加到SARS ELISA的固相抗原配方中。这些多肽和试剂盒将按照cGMP标准进行制造和表征。将在亚洲的设施中对大量收集的SARS阳性血清进行大规模临床试验。

DESCRIPTION (provided by applicant): The long-term objectives are to develop an enzyme-linked immunoassay (ELISA) for Severe Acute Respiratory Syndrome (SARS), based on synthetic peptide antigens. The test will be stable and free of biohazardous manufacturing operations and components, and is intended as a safe and rapid method for serological surveillance. The test will be amenable to large scale manufacture and compatible with fully automated diagnostic screening. Specific aims are 1) to identify and optimize the peptide antigens, including antigens from non-structural SARS coronavirus (SCoV) proteins, in addition to peptides from structural proteins S, M, and N that have already been identified; 2) formulate a SARS ELISA test kit optimized for sensitivity and specificity; 3) validate the kit with a panel of characterized sera from SARS patients and various panels from the sera of subjects that have not been exposed to the SCoV; 4) perform a large-scale clinical trial at multiple sites in the US, Taiwan and China; 4) accomplish the commercial scale production of kits; and 5) submit a PMA or PDP application for US and foreign regulatory approvals. A prototype SARS ELISA having peptide antigens from the S, M, and N structural proteins of SCoV is to be improved by the addition of non-structural peptides whose sequences are deduced from the putative pol region of the SCoV genome. Overlapping peptides will be synthesized to cover the entire pol la and lb open reading frames. Peptides will be of lengths from 20-60 amino acids. Individual peptides will be screened by the ELISA method for reactivity to a collection of sera from confirmed SARS patients. Serologically reactive epitopes on the peptides will be refined for optimum sequences and conformation. The newly identified non-structural peptide antigens will be added to the solid-phase antigen formulation of the SARS ELISA. The peptides and kits will be manufactured and characterized with cGMP standards. Large-scale clinical trials will be done at facilities in Asia on extensive collections of SARS positive sera.