Glucose metabolism has traditionally been assayed via biochemical means. Fluorescence monitoring of NAD(P)H levels has provided a noninvasive method to assay glucose metabolism in cells and tissues. However, these measurements have traditionally been of low resolution (no subcellular information) because of limitations imposed by optical and cellular photodamage problems. The recent advent of two-photon excitation microscopy as a dependable tool for biological research has opened the possibility of real-time, high- resolution analysis of glucose metabolism in living cells. Such measurements have the potential to provide subcellular information from intact tissue that cannot be obtained by other techniques.