Shotgun proteomic analysis of human induced sputum

Sputum is increasingly recognised as a biologically relevant sample of the environment of the airways when collected under controlled conditions. As such, it has been used for the analysis of bacterial load, tumour and inflammatory cell content, and of biomarkers of airways diseases such as cystic fibrosis, asthma and chronic obstructive pulmonary disease. While related biofluids such as broncho-alveolar lavage1 fluid, nasal lavage fluid,2 and in particular saliva3 have been well-characterised using standard 2-D gel based proteomics, sputum has not. This is probably due to the presence of mucin glycoproteins that compromise the separation of proteins during the IEF stage of 2-D gel analysis, preventing good resolution.We have used a combination of 2-D gel electrophoresis (fig 1BGo) and a multidimensional mass-spectrometric technique (GeLC-MS/MS) (fig 1AGo) to examine the proteome of induced sputum. 191 individual human proteins were confidently assigned using these techniques. In addition to expected components, several hitherto unreported proteins were discovered, including three members of the annexin family, kallikreins 1 and 11, and peroxiredoxins 1, 2, and 5. The resultant data represent the first detailed survey of the human induced sputum proteome and allow a comparison of the induced sputum proteome with other biologically related fluids such as broncho-alveolar lavage fluid and saliva. They also provide a platform for the future identification of biomarkers of airways disease