Point-of-care lateral flow immunoassay for diagnosis of pertussis

百日咳（百日咳）是一种高度传染性的空气传播疾病， 美国的公共卫生威胁。尽管疫苗覆盖率很高，但美国每年报告的病例 最近一直在增加，2010 - 2019年平均每年25，000人。这是一个前所未有的水平， 60年代初百日咳的诊断是具有挑战性的，因为它的早期症状（在传染性卡他性 阶段，阵发性咳嗽之前）与其他呼吸道疾病重叠。这类似于诊断 COVID-19的挑战持续的COVID-19大流行表明了公共卫生的重要性， 可获得的快速诊断，以控制高度传播性呼吸道疾病的传播， 包括百日咳因此，与COVID-19一样，迫切需要开发改进的百日咳 诊断，以使及时，适当的病人治疗，以及限制不必要的社区传播。 该项目的目标是一种快速、即时（POC）、侧流免疫测定（LFIA）检测百日咳杆菌 鼻咽标本的抗原。目标人群将是出现以下症状的患者： 儿科医生办公室、紧急护理诊所或小型医院急诊室。该产品将是一个敏感和具体的测试 能够立即采取行动，类似于其他高度使用和接受的LFIAs（例如COVID-19，RSV）。 本提案以第一阶段和第二阶段的数据和结果为基础。首先是B。百日咳蛋白气管定植 因子A（TcfA）被鉴定为细菌存在的特异性生物标志物。第二，表位文库- 开发了对TcfA具有高亲和力和特异性的定位单克隆抗体（mAbs）。第三，超过784 评估mAb组合的排列以构建LFIA。第四，LFIA的原型 为了检测纯化的TcfA和临床应用的高特异性和灵敏度， B的相关浓度。百日咳细胞第五，原型LFIA显示出高灵敏度和特异性， 在狒狒模型中诊断百日咳。最后，用患者标本证明了概念验证。 根据最初的计划，IIb期建立在I期和II期的基础上，以提供FDA批准的诊断。三个目标是 提出了目标1包括FDA预提交和从研究级原型LFIA过渡到 根据GMP生产的商业级产品。目的2是精密度和重现性的临床前评价， 检测限、分析专属性和包容性、稳定性、耐用性、干扰物质和微生物 干扰目标3是一项临床评价，将以FDA 510（k）提交结束。如果成功，这项研究 将提供一种百日咳POC测试，所有诊所都可以轻松获得，并提供可操作的结果 时间框架-从而降低患者发病率并防止不必要的社区传播。

Pertussis (Whooping cough) is a highly contagious, airborne disease that is rapidly re-emerging as a serious public health threat in the United States. Despite high vaccine coverage, annual reported cases in the U.S. have been increasing recently, with an average of 25,000 per year from 2010-2019. This is a level unseen since the early 1960s. Diagnosis of pertussis is challenging because its early symptoms (during the contagious catarrhal stage, prior to paroxysmal cough) overlap with other respiratory diseases. This is analogous to the diagnostic challenge of COVID-19. The ongoing COVID-19 pandemic has illustrated the critical public health importance of accessible, rapid diagnostics for controlling the spread of highly transmissible respiratory diseases, a category that includes pertussis. Consequently, as with COVID-19, there is a critical need to develop improved pertussis diagnostics to enable prompt, appropriate patient treatment as well as limit needless community transmission. The project goal is a rapid, point-of-care (POC), lateral flow immunoassay (LFIA) to detect Bordetella pertussis antigen from nasopharyngeal specimens. The target population will be symptomatic patients who present in pediatrician offices, urgent care clinics, or small hospital ERs. The product will be a sensitive and specific test that enables immediate action, akin to other highly used and accepted LFIAs (e.g. COVID-19, RSV). This proposal builds on data and results from Phases I and II. First, the B. pertussis protein tracheal colonization factor A (TcfA) was identified as a specific biomarker for presence of the bacterium. Second, a library of epitope- mapped monoclonal antibodies (mAbs) with high affinity and specificity to TcfA was developed. Third, over 784 permutations of mAb combinations were evaluated for construction of a LFIA. Fourth, the prototype LFIA’s configuration was optimized for high specificity and sensitivity for detection of both purified TcfA and clinically relevant concentrations of B. pertussis cells. Fifth, the prototype LFIA showed high sensitivity and specificity for diagnosis of pertussis in a baboon model. Finally, proof-of-concept was demonstrated with patient specimens. Per the original plan, Phase IIb builds on Phases I and II to deliver an FDA-cleared diagnostic. Three aims are proposed. Aim 1 includes an FDA Pre-Submission and transitions from a research-grade, prototype LFIA to a commercial-grade product made under GMP. Aim 2 is a pre-clinical evaluation of precision and reproducibility, limit of detection, analytical specificity and inclusivity, stability, ruggedness, interfering substances, and microbial interference. Aim 3 is a clinical evaluation that will end with a FDA 510(k) Submission. If successful, this study will deliver a pertussis POC test that is easily accessible to all clinics and that provides results in an actionable timeframe – thereby reducing patient morbidity and preventing unnecessary community transmission.