Quantitation of the oxygen pressure distribution in the heart provides an accurate definition of metabolically viable cells adjacent to ischemic injury. By injection of a phosphorescent oxygen probe into the blood of newborn piglets and illumination of the heart with monochromatic light, a series of images of the heart were collected with use of a gated intensified CCD camera. These sets of images were used to calculate two-dimensional maps of the oxygen pressure in the epicardial microvasculature. Occlusion of a distal arterial vessel resulted in a hypoxic area containing a central focus of near-zero oxygen pressure bordered by tissue with diminished but nonzero oxygen pressures. These border zones extended for several millimeters from the hypoxic core, with the oxygen pressures progressively increasing from the focus to the normoxic region. The maps of oxygen distribution obtained by phosphorescence imaging provide rapid and quantitative measure of the oxygen pressures in the beating heart in vivo. They accurately show the degree of oxygen deprivation and the extent of an ischemic border zone that may respond to treatment.