Why can’t I pick directly from a gel containing CyDye™ DIGE Fluor labelled proteins?
When a CyDye DIGE Fluor is coupled to a protein it will add approximately Mr 500 to the proteins mass. This causes a shift between labelled and unlabelled proteins and is more marked for lower molecular weight proteins. Because of this shift, the most fluorescent point on a spot represents the highest concentration of the labelled protein, but does not correspond to the highest concentration of unlabelled protein. Thus, to maximize the amount of protein extracted from each spot, preparative gels are run using higher protein loadings and post-staining e.g., with SYPRO™ Ruby. These gels are matched to the quantitative data obtained from the analytical gels so that spots of interest can be accurately picked from the preparative gel(s).
Related Questions
- When imaging the SYPRO™ Ruby stained gel, can I still image for the original CyDye™ DIGE Fluor minimal dyes and get three (or four) related images?
- When imaging a SYPRO™ Ruby stained gel, can I still image for the original CyDye™ DIGE Fluor minimal dyes and get three (or four) related images?
- Why can’t I pick directly from a gel containing CyDye™ DIGE Fluor labelled proteins?
