Cancer is a quite heterogeneous disease and each cancer type can be divided in different subentities. Normally this is done by pathologist using classical dye-staining protocols or considering specific biomarkers. To identify new biomarkers, allowing a more specific diagnosis clinical tissue specimen is the material of choice. But the amount of clinical material obtained by resection or biopsy is often limited. In order to perform analytical studies with such scarce sample material, a sensitive analysis method is required. Using two-dimensional electrophoresis (2DE) for the analysis of small protein amounts, protein saturation labeling using fluorescence dyes has been successfully applied. Here, we describe the application of saturation labeling in combination with microdissection for the analysis of lung tumor cells and bronchial epithelium cells. The presented study demonstrates all relevant steps of differential proteome analysis with scarce protein amount: experimental design, manual microdissection, optimization of saturation labeling, 2DE, protein identification and validation. As a result, 32 non-redundant proteins could be identified to be differentially regulated between the respective tissue types and are candidate biomarkers for describing lung cancer in more detail.
DIGE Saturation labeling Cancer proteomics Microdissection Reference proteome
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The authors would like to thank Anna Lendzian and Sabine Roggenbrodt for excellent technical assistance. This work was supported by a grant from the European Commission (LCVAC, COOP-CT-2004-512855).
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