(152d) Development Of Flamingo™, A Novel Fluorescent Dye For Non-Specific Detection And Quantitation Of Proteins In Gels

Gel-separated proteins are most commonly detected and quantitated by dye binding, utilizing the property of some dyes to bind to proteins non-specifically, rendering the proteins optically detectable and quantifiable. Fluorescent dyes are widely used for this due to the high sensitivity and dynamic range of fluorescent detection. Flamingo is a novel dye that was developed from a class of dyes that are minimally fluorescent at low pH in the absence of protein, but acquire strong fluorescence in the presence of denatured protein. This property was reasoned to be potentially useful for staining proteins separated in polyacrylamide gels, which are typically denatured and fixed in acid solution. Modifications of the dye structure were undertaken to enhance protein binding, resulting in development of a dye with particularly useful gel staining properties. Fixed protein gels can be stained by immersion in an acid solution of Flamingo. High sensitivity protein detection is possible without the need for destaining, since unbound dye is relatively non-fluorescent. Investigations of Flamingo Fluorescent Gel Stain were undertaken to characterize the product and compare it to other fluorescent stains with respect to gel staining sensitivity, linearity of response to protein, protein-to-protein variability and compatibility with mass spectrometric analysis. Flamingo was found to allow protein detection down to 30 pg and allow linear quantitation over four orders of magnitude. Flamingo staining was found to be fully compatible with peptide mass fingerprinting by MALDI-MS and to result in a lower incidence of oxidative protein modification than SYPRO? Ruby.