The combination of
laser microdissection and two-dimensional gel electrophoresis (2-D PAGE) has been developed to perform proteomic analysis on specific populations of cells in
cancer tissues. However, as conventional low sensitivity
silver staining was used for spot detection, the microdissection required to obtain an adequate amount of
protein for 2-D PAGE is laborious and only a restricted number of
protein spots could be visualized. As a consequence, this technology was impractical for direct clinical applications and had a limited impact on
cancer studies. To solve these problems, we developed an application in which
fluorescent dyes label the
proteins extracted from microdissected tissues prior to 2-D PAGE separation. In this application, a small amount of
protein, less than 6.6 microg, was enough to generate a 2-D profile with approximately 1500
protein spots. This technique was applied to compare the
proteome of normal intestinal epithelium with that of
adenoma in Min mice. Thirty-seven
protein spots reproducibly showed significant differences in intensities. Mass spectrometric analysis and Western blotting identified eight of them, including
prohibitin, 14-3-3zeta,
tropomyosin 3 and Hsp84. These results indicate that fluorescence labeling of
proteins from microdissected tissues prior to 2-D PAGE is a powerful
cancer proteomic study tool.