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The strategy of our fluorescence quenching-based membrane binding assay. To the solution of an EGFP-fusion protein (e.g., PDK1-PH-EGFP) is added vesicles containing their favorite lipids (e.g., PtdIns(3,4,5)P3 shown as a shaded hexagon) and a dark quencher, dabsyl-PE (shown in black). As the protein binds the membrane surface, the fluorescence emission intensity of EGFP is greatly reduced due to quenching by dabsyl-PE, allowing quantitative analysis of membrane-protein interaction.
