Identification of phosphoproteins in breast tumors. (A) Phosphoprotein staining by Pro-Q Diamond and SYPRO-Ruby dyes. Total proteins from both HER-2/neu positive and -negative breast tumors were separated by 2-DE gels and stained with fluorescent Pro-Q Diamond for phosphoproteins, followed by SYPRO-Ruby for total proteins. Stained gels were scanned with a Typhoon 9600 fluorescence scanner. Images were captured and the relative phosphorylation levels of differentially phosphorylated spots were analyzed using the ImageMaster 2D Elite software. Proteins were in-gel digested with trypsin, analyzed using 4800 MALDI-TOF/TOF™ analyzer and identified by NCBInr database search. Four phosphoproteins were unambiguously identified (A: in table, right). (B) Peptide sequences and Mowse scores of Hsp27.