Fluorescence polarisation of green fluorescent protein (GFP). A strategy for improved wavelength discrimination for GFP determinations

The fluorescence of green fluorescent protein (GFP), present both within wh ole yeast cells and in protein extracts from yeast cells, has been observed to be significantly polarised. Fluorescence polarisation is proposed as a useful technique to allow some discrimination between GFP fluorescence and that of other interfering species in cell or media matrices, which have flu orescence bands that overlap those of GFP, which should lead to improved re solution and limits of detection. The method has been tested by discriminat ing between the fluorescence of GFP in cell extracts and added fluorescein, both of which fluoresce brightly at the same wavelength, The flow-through instrumentation incorporating an argon-ion laser developed for this work is also described.