Circulating Cell-free Antigens for Monitoring of Machining Fluids

项目摘要(摘要) 工业金属加工液(MWF)中的分枝杆菌是公认的致病原 机械工职业性肺病、过敏性肺炎及相关呼吸道症状。 目前还没有免疫分析方法来对这些违规的工业过程流体进行常规监测 用于暴露评估的抗原。这主要是由于对特定的分枝杆菌缺乏了解。 这些液体中普遍存在的抗原。考虑到完整的细胞计数可能低估了总数 分枝杆菌的抗原负荷(由于积累无细胞抗原)，特别是在长时间 这些液体的循环，对抗原的监测可能会提供更多信息。因此，总的目标是 本研究旨在确定在这些液体中普遍存在的主要循环无细胞(Cf)分枝杆菌抗原和靶标。 用于建立用于暴露评估的抗原监测试验。在这种情况下，检测靶向 由于个体中存在多个交叉反应表位，整个抗原可能缺乏特异性。 抗原，有必要确定特定的关键表位作为检测抗体的目标。这让我们找到了 假设分枝杆菌抗原蛋白的鉴定是选择性表达和积累的 在机械加工中，流体作为主要的无细胞抗原及其关键表位可能构成 以多重分析为靶点监测这些工业流体中的信息抗原 免疫分析，为机械加工行业提供了一种早期可靠的暴露评估策略。这个 具体目标是：(1)。鉴定循环中的无细胞分枝杆菌抗原(及其关键表位) 田间MWF占优势；(2)一种多功能常规体液监测免疫分析平台的设计 针对工业中使用的MWF中的分枝杆菌抗原(表位)。候选抗原将是 采用基于鸟枪式蛋白质组学的免疫蛋白质组学方法进行鉴定，并对其关键表位进行鉴定。在……里面 同时，一种基于ELISA法的免疫分析将针对MWF矩阵进行优化，并扩展为多路 使用关键识别表位和抗体的免疫分析；该分析将得到快速和灵敏的验证 用于分枝杆菌暴露评估的工业mwf样本中循环cf抗原的检测。这个 由此产生的信息将通过提供一套工具来促进Nora的目标和NIOSH的R2P倡议 机械加工液中幽门螺杆菌致病抗原暴露评估。这些结果将促进未来 暴露于这些工业过程的流行病学研究和干预策略的发展 体液长期存在。

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.