| Popis: |
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. |
