Quantification of Genetically Encoded Lipid Biosensors.

Lipids, like phosphoinositides, can be visualized in living cells in real time using genetically encoded biosensors and fluorescence microscopy. Sensor localization can be quantified by determining the fluorescence intensity of each fluorophore. Enrichment of lipids at membranes can be determined by generating and applying an organelle-specific binary mask. In this chapter, we provide a detailed list of reagents and methods to visualize and quantify relative lipid levels. Applying this approach, changes in lipid levels can be assessed in cases when lipid metabolizing enzymes are mutated or otherwise altered.