An EPR method for the measurement of the oxidative stress status in biological systems is described. The method is based on the X-band EPR detection of a persistent nitroxide generated under physiological or pseudo-physiological conditions by oxidation of a highly lipophylic hydroxylamine probe. The probe employed is bis(1-hydroxy-2,2,6,6-tetramethyl-4-piperidinyl)-decandioate which is administrated as hydrochloride salt. This probe is able to give a fast reaction with the majority of radical species involved in the oxidative stress. Furthermore, it crosses cell membranes and distributes in a biological environment without the need to alter or destroy compartmentation. The method is therefore suitable for quantitative measurements of ROS and can be applied to human tissues in real clinical settings. It has been successfully employed in systems of growing complexity and interest, ranging from subcellular fractions to whole animals and human liver.