When cells are exposed to hormones that act on cell surface receptors, information is processed through the plasma membrane into the cell interior via second messengers generated in the inner leaflet of the plasma membrane. Individual biochemical steps along this cascade have been characterized from ligand binding to receptors through to activation of guanine nucleotide binding proteins and their downstream effectors such as adenylate cyclase or phospholipase C. However, the complexity of temporal and spatial integration of these molecular events requires that they are studied in intact cells. The great expansion of fluorescent techniques and improved imaging technologies such as confocal and TIRF microscopy combined with genetically-engineered protein modules has provided a completely new approach to signal transduction research. Spatial definition of biochemical events followed with real-time temporal resolution has become a standard goal, and several new techniques are now breaking the resolution barrier of light microscopy.