CyDye DIGE Fluor saturation
dye (saturation
dye, GE Healthcare Amersham Biosciences) enables highly sensitive 2-D PAGE. As the
dye reacts with all reduced
cysteine thiols, 2-D PAGE can be performed with a lower amount of
protein, compared with CyDye DIGE Fluor minimal
dye (GE Healthcare Amersham Biosciences), the sensitivity of which is equivalent to that of
silver staining. We constructed a 2-D map of the saturation
dye-labeled
proteins of a
liver cancer cell line (HepG2) and identified by MS 92
proteins corresponding to 123
protein spots. Functional classification revealed that the identified
proteins had chaperone, protein binding,
nucleotide binding,
metal ion binding,
isomerase activity, and motor activity. The functional distribution and the
cysteine contents of the
proteins were similar to those in the most comprehensive 2-D database of
hepatoma cells (Seow et al.., Electrophoresis 2000, 21, 1787-1813), where
silver staining was used for
protein visualization. Hierarchical clustering on the basis of the quantitative expression profiles of the 123 characterized spots labeled with two charge- and mass-matched saturation
dyes (Cy3 and
Cy5) discriminated between nine
hepatocellular carcinoma cell lines and primary cultured hepatocytes from five individuals, suggesting the utility of saturation
dye and our database for proteomic studies of
liver cancer.