Protocol abstract
Nature Protocols 1, - 2940 - 2956 (2007)
Published online: 25 January 2007 | Corrected online: 22 February 2007 | doi:10.1038/nprot.2006.421
Subject Categories: Genomics and proteomics | Isolation, purification and separation | Spectroscopy and structural analysis
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
Tadashi Kondo1 & Setsuo Hirohashi1
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 
m 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.
NOTE: In the PDF version of this article initially published online, the publication date was shown as 25 January 2006 instead of 25 January 2007. The error has been corrected in the PDF version of the article.
- Proteome Bioinformatics Project, National Cancer Center Research Institute, 5-1-1 Tsukiji, Chuo-ku, Tokyo 104-0045, Japan.
Correspondence to: Tadashi Kondo1 e-mail: takondo@gan2.res.ncc.go.jp
nature-products
A-Z product listing
- 0.5% eosin alcohol solution(Muto Pure Chemicals)
- 87% glycerol(GE Healthcare Bio-sciences)
- 96-well thin-wall plate(Corning)
- Acetic acid(Wako)
- Acetonitrile(Sigma-Aldrich)
- acrylamide(Wako)
