A new, phosphorescence‐based optical instrument for making oxygen tension (PO2) measurements in individual vessels of microvascular preparations is described. Palladium–porphyrin compounds injected into the systemic vasculature are excited by light and the measured phosphorescence emission lifetime used to calculate intravascular PO2. Because this technique is relatively new, we have undertaken a rigorous examination of its usefulness and inherent limitations for microvascular studies. ©1995 American Institute of Physics.