Synthesis and validation of cyanine-based dyes for DIGE.

The application of difference gel electrophoresis (DIGE), in particular its most common "minimal labeling" variety, utilizes N-hydroxysuccinimide esters of Cy2, Cy3, and Cy5 dyes, which are commercially available. We describe methods for the efficient synthesis of all three dyes from relatively inexpensive and commercially available precursors in only a few steps and with relatively high yields. In model DIGE experiments, the newly synthesized dyes proved to be indistinguishable from commercially available ones and have been shown to be stable for years while stored under argon as dry solids or after being dissolved in N,N-dimethylformamide.