Quantitative studies of cellular systems require experimental techniques that can expose single cells to well-controlled chemical stimuli with high spatiotemporal resolution. Here, we combine microfluidic techniques with the photochemical release of caged signaling molecules to generate tailored stimuli on the length scale of individual cells with subsecond switching times. We exemplify this flexible approach by initiating membrane translocation of fluorescent fusion proteins in chemotactic Dictyostelium discoideum cells.