Application of saturation labeling in lung cancer proteomics.
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.
Molecular Proteomics Laboratory, Heinrich-Heine-Universität, Düsseldorf, Germany. gereon.poschmann@rub.de
Gereon
G
Poschmann
Barbara
B
Sitek
Bence
B
Sipos
Kai
K
Stühler
D001980:Bronchi / Q000473:pathology; D002294:Carcinoma, Squamous Cell / Q000473:pathology; D002452:Cell Count; D015180:Electrophoresis, Gel, Two-Dimensional; D004847:Epithelial Cells / Q000378:metabolism / Q000473:pathology; D005456:Fluorescent Dyes / Q000378:metabolism; D006801:Humans; D007091:Image Processing, Computer-Assisted; D008175:Lung Neoplasms / Q000378:metabolism* / Q000473:pathology*; D042282:Microdissection; D020543:Proteome / Q000032:analysis / Q000302:isolation & purification / Q000378:metabolism; D040901:Proteomics / Q000379:methods* / Q000592:standards; D012015:Reference Standards; D013194:Staining and Labeling / Q000379:methods*
D005456:Fluorescent Dyes; D020543:Proteome