Proteomics of Epithelial Lining Fluid Obtained by Bronchoscopic Microprobe Sampling
Proteomics of Epithelial Lining Fluid Obtained by Bronchoscopic Microprobe Sampling
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DOI：
10.1007/978-1-61779-319-6_2
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发表时间：
2011-01-01
期刊：
影响因子：
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通讯作者：
Bischoff, Rainer
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文献类型：
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作者：
Franciosi, Lorenza;Govorukhina, Natalia;Bischoff, Rainer
上皮内衬液 (ELF) 形成一层薄薄的液体层，覆盖肺泡、小气道和大气道的粘膜。由于它构成了肺部与外界之间的第一道屏障，因此它是专注于肺部疾病的蛋白质组学研究的一个有趣的目标。 ELF 的支气管镜微探针 (BMP) 采样使用带有吸收尖端的小探针，通过支气管镜引入。与迄今为止用于从肺部收集生物流体的其他方法（例如支气管肺泡灌洗液、诱导痰）相比，该技术的优点是 ELF 不被稀释并且含有高浓度的生物分子。此外，气管支气管树中的研究位置明确，并且没有口咽细菌或唾液污染。尽管探针偶尔会因刮擦气道粘膜而受到血液污染，但对微探针采样的 ELF 进行蛋白质组学分析为肺部疾病的研究开辟了新的可能性。我们的工作特别关注香烟烟雾诱发的慢性阻塞性肺疾病 (COPD) 的诱导和进展。在本章中，我们描述了 ELF 采样的实际方面，然后详细描述了通过 SDS-PAGE 和凝胶内消化分离蛋白质后通过 LC-MS/MS 进行的蛋白质组学分析。例如，我们应用这个蛋白质组平台来鉴定和定量来自 COPD 患者和健康受试者的 ELF 中的蛋白质。
Epithelial lining fluid (ELF) forms a thin fluid layer that covers the mucosa of the alveoli, the small airways, and the large airways. Since it constitutes the first barrier between the lung and the outer world, it is an interesting target for proteomics studies that focus on lung disease. Bronchoscopic microprobe (BMP) sampling of ELF uses small probes with an absorptive tip that are introduced bronchoscopically.In contrast to other methods used so far for the collection of biofluids from the lung (e.g., bronchoalveolar lavage fluid, induced sputum), this technique has the advantage that ELF is not diluted and contains high concentrations of biomolecules. In addition, the investigated location in the tracheobronchial tree is well defined, and there is no contamination with oropharyngeal bacteria or saliva. Despite occasional blood contamination of the probes by scratching the mucosa of the airways, the proteomic analysis of microprobe-sampled ELF opens new possibilities for research in lung diseases. Our work focuses particularly on the induction and progression of cigarette smoke-induced Chronic Obstructive Pulmonary Disease (COPD).In this chapter, we describe the practical aspects of sampling ELF followed by a detailed description of proteomics analysis by LC-MS/MS after protein separation by SDS-PAGE and in-gel digestion. As an example, we apply this proteomic platform to the identification and quantification of proteins in ELF from COPD patients and healthy subjects.