Synthetic Peptide SARS Coronavirus Diagnostic Kit

描述（由申请人提供）： 长期目标是开发基于合成肽抗原的严重急性呼吸道综合征（SARS）酶联免疫测定（ELISA）。该检测将保持稳定，不含生物危害性生产操作和组分，预期作为一种安全、快速的血清学监测方法。该测试将适合大规模生产，并与全自动诊断筛查兼容。具体目标是1）鉴定和优化肽抗原，包括来自非结构SARS冠状病毒（SCoV）蛋白的抗原，以及已经鉴定的来自结构蛋白S、M和N的肽; 2）配制针对灵敏度和特异性优化的SARS ELISA检测试剂盒; 3）用一组来自SARS患者的表征血清和来自未暴露于SCoV的受试者的血清的各种组验证试剂盒; 4）在美国、中国台湾和中国大陆的多个地点进行大规模临床试验; 4）完成试剂盒的商业规模生产;以及5）提交PMA或PDP申请以获得美国和外国监管机构的批准。具有来自SCoV的S、M和N结构蛋白的肽抗原的原型SARS ELISA将通过添加非结构肽来改进，所述非结构肽的序列从SCoV基因组的推定pol区推导。将合成重叠肽以覆盖整个polla和lb开放阅读框。肽的长度为20-60个氨基酸。将通过ELISA方法筛选单个肽对确诊SARS患者血清的反应性。将对肽上的血清学反应性表位进行优化以获得最佳序列和构象。新鉴定的非结构肽抗原将被添加到SARS ELISA的固相抗原制剂中。将使用cGMP标准品生产和表征肽和试剂盒。大规模的临床试验将在亚洲的设施中进行，以广泛收集SARS阳性血清。

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.