The oxygen consumption rate in tumors affects tumor oxygenation and response to therapies. A new EPR method was developed to measure tissue oxygen consumption non-invasively. The protocol was based on the measurement of pO₂ during a carbogen challenge. The following sequence was used: (1) basal value during air breathing; (2) saturation of tissue with oxygen by carbogen breathing; (3) switch back to air breathing. The assumption was that the kinetics of the return to the basal value after oxygen saturation would be governed mainly by tissue oxygen consumption. This challenge was applied in hyperthyroid mice (generated by chronic treatment with l-thyroxine) and control mice because hyperthyroidism is known to dramatically affect the oxygen consumption rate of tumor and muscle cells. Muscle and tumor cells from the hyperthyroid mice consumed oxygen faster than muscle and tumor cells from the control mice, which is consistent with the results of in vitro studies. Tumor perfusion was not affected by the treatment with l-thyroxine. This method gives an index that may reasonably be ascribed to the local oxygen consumption and has the unique advantage of being adaptable to in vivo studies.