title: Application of highly sensitive fluorescent dyes (CyDye DIGE Fluor saturation dyes) to laser microdissection and two-dimensional difference gel electrophoresis (2D-DIGE) for cancer proteomics.
abstract: Proteome data combined with histopathological information provides important, novel clues for understanding cancer biology and reveals candidates for tumor markers and therapeutic targets. We have established an application of a highly sensitive fluorescent dye (CyDye DIGE Fluor saturation dye), developed for two-dimensional difference gel electrophoresis (2D-DIGE), to the labeling of proteins extracted from laser microdissected tissues. The use of the dye dramatically decreases the protein amount and, in turn, the number of cells required for 2D-DIGE; the cells obtained from a 1 mm2 area of an 8-12 microm thick tissue section generate up to 5,000 protein spots in a large-format 2D gel. This protocol allows the execution of large-scale proteomics in a more efficient, accurate and reproducible way. The protocol can be used to examine a single sample in 5 d or to examine hundreds of samples in large-scale proteomics.
authors.affiliation: Proteome Bioinformatics Project, National Cancer Center Research Institute, 5-1-1 Tsukiji, Chuo-ku, Tokyo 104-0045, Japan. takondo@gan2.res.ncc.go.jp
authors.forename: Tadashi
authors.initials: T
authors.lastname: Kondo
authors.forename: Setsuo
authors.initials: S
authors.lastname: Hirohashi
mesh_terms: D015180:Electrophoresis, Gel, Two-Dimensional / Q000379:methods*; D005456:Fluorescent Dyes; D006801:Humans; D007834:Lasers; D013058:Mass Spectrometry; D042282:Microdissection / Q000379:methods*; D009369:Neoplasms / Q000473:pathology*; D040901:Proteomics / Q000379:methods*; D013194:Staining and Labeling / Q000379:methods*; D013223:Statistics as Topic
chemical_list: D005456:Fluorescent Dyes