Circulating Cell-free Antigens for Monitoring of Machining Fluids

用于监测加工液的循环无细胞抗原

• 批准号: 9979088
• 负责人: Jagjit S Yadav
• 金额: $ 21.5万
• 依托单位: UNIVERSITY OF CINCINNATI
• 依托单位国家: 美国
• 财政年份: 2020
• 资助国家: 美国
• 起止时间: 2020-09-30 至 2022-09-29
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9979088
• 关键词: Circulating; Cell; free; Antigens; Monitoring

Project summary (Abstract) Mycobacteria colonizing industrial metalworking fluids (MWFs) are recognized causal antigens of the occupational lung disease hypersensitivity pneumonitis (HP) and related respiratory symptoms in machinists. Currently there are no immunoassays for routine monitoring of these industrial process fluids for the offending antigens for exposure assessment. This is primarily due to the lack of knowledge of specific mycobacterial antigens prevalent in these fluids. Considering the fact that intact cell count may underestimate the total antigen load for mycobacteria (due to the accumulating cell-free antigens) especially during prolonged circulation of these fluids, monitoring for the antigens may be more informative. Therefore, the overall goal of this study is to identify major circulating cell-free (cf) mycobacterial antigens prevalent in these fluids and target them for development of antigen monitoring assay for exposure assessment. In this context, assays targeting whole antigens may lack specificity because of presence of multiple and cross-reactive epitopes in individual antigens, necessitating identification of specific key epitopes as detection antibody targets. This led us to hypothesize that identification of mycobacterial antigenic proteins selectively expressed and accumulated in machining fluids as dominant cell-free antigens and their key epitopes could form the basis for an informative antigen monitoring in these industrial fluids by serving as assay targets in a multiplex immunoassay, offering an early and reliable exposure assessment strategy in the machining industry. The specific aims are to: (1). Identify circulating cell-free mycobacterial antigens (and their key epitopes) predominant in field MWF; (2). Designing of a multiplex immunoassay platform for routine fluid monitoring targeting the cf mycobacterial antigens (epitopes) in industrial in-use MWF. Candidate antigens will be identified using immunoproteomic approach based on shotgun proteomics and their key epitopes identified. In parallel efforts, an ELISA-based immunoassay will be optimized for MWF matrix and expanded to a multiplex immunoassay using key identified epitopes and antibodies; the assay will be validated for rapid and sensitive detection of circulating cf antigens in industrial MWF samples for mycobacterial exposure assessment. The resulting information will further the NORA’s objectives and NIOSH’s r2p initiative by providing a set of tools for HP-causative antigen exposure assessment in machining fluids. These outcomes will facilitate future epidemiological studies and development of intervention strategies for exposures to these industrial process fluids in long-term.

项目摘要（摘要） 分枝杆菌定植工业金属加工液（MWF）是公认的因果抗原 机械师中占据肺部疾病过敏性肺炎（HP）和相关呼吸道症状。 目前尚无免疫测定法以常规监控这些工业流程的烟气以进行犯罪 暴露评估的抗原。这主要是由于缺乏特定分枝杆菌的知识 这些液体中普遍存在的抗原。考虑完整的细胞计数可能低估总数的事实 分枝杆菌的抗原负荷（由于累积的细胞抗原），尤其是在长时间 这些烟道的循环，对抗原的监测可能会更有益。因此，总体目标 这项研究是为了确定这些液体中普遍存在的主要无细胞（CF）分枝杆菌抗原 它们用于开发抗原监测测定法进行暴露评估。在这种情况下，针对目标的测定 整个抗原可能缺乏特异性，因为个体中存在多重和交叉反应表位 抗原，必要的特定关键表位为检测抗体靶标。这导致了我们 假设鉴定分枝杆菌抗原蛋白有选择地表达和累积 在将液体加工为主要细胞的抗原及其关键表位时，可以构成 这些工业烟道中有益的抗原监测，通过作为多重测定目标 免疫测定法，在加工行业提供早期可靠的暴露评估策略。 具体目标是：（1）。识别无细胞的分枝杆菌抗原（及其关键表位） MWF领域主要是； （2）。设计用于常规流体监测的多重免疫测定平台 靶向工业内使用MWF中的CF分枝杆菌抗原（Eppitopes）。候选抗原将是 使用基于shot弹枪蛋白质组学及其关键表位的免疫蛋白质组学方法鉴定。在 平行努力，基于ELISA的免疫测定法将针对MWF矩阵进行优化，并扩展到多重 使用密钥确定的表位和抗体的免疫测定；该测定将得到验证，以快速和敏感 在工业MWF样品中检测循环CF抗原用于分枝杆菌暴露评估。这 最终的信息将通过提供一组工具来进一步进一步进一步 加工液体中的HP抗原暴露评估。这些结果将促进未来 流行病学研究和开发干预策略以接触这些工业过程 长期流体。