A relative deficiency in kidney oxygenation, i.e., renal hypoxia, may contribute to the initiation and progression of acute and chronic kidney disease. A critical barrier to investigate this is the lack of methods allowing measurement of the partial pressure of oxygen in kidney tissue for long periods in vivo. We have developed, validated, and tested a novel telemetric method that can do this. Here we provide details on the calibration, implantation, implementation for data recording, and reuse of this telemetry-based technology for measurement of medullary tissue oxygen tension in conscious, unrestrained rats. This technique provides an important additional tool for investigating the impact of renal hypoxia in biology and pathophysiology.