Spatiotemporal stimulation of single cells using flow photolysis

Quantitative studies of chemotactic signaling require experimental techniques that can expose single cells to chemical stimuli with high resolution in both space and time. Recently, we have introduced the method of flow photolysis (Anal. Chem. 79:3940-3944, 2007), which combines microfluidic techniques with the photochemical release of caged compounds. This method allows us to tailor chemical stimuli on the length scale of individual cells with subsecond temporal resolution. In this chapter, we provide a detailed protocol for the setup of flow photolysis experiments and exemplify this versatile approach by initiating membrane translocation of fluorescent fusion proteins in chemotactic Dictyostelium discoideum cells.