CyDye™ DIGE Fluor Cy™2, Cy3 and Cy5 minimal dyes have different quantum yields. Does this effect the accuracy with which protein abundance changes can be quantified?
No. The quantum yield of Cy5 is greater than that of Cy2 and Cy3 so the signal from a sample labelled with CyDye™ DIGE Fluor Cy5 minimal dye would be expected to be the most intense. Selecting the correct PMT when scanning will ensure that the signal for each dye falls within the linear dynamic range of the Typhoon Variable Mode Imager. For example, using a higher PMT for Cy3 images should result in similar signal intensities in both channels. Images are then normalized in DeCyder Differential Analysis Software by setting the total signal for each image at the same level. This normalization process negates differences in signal intensity between the dyes.
Related Questions
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