Studies devoted to the detection and measurement of free radicals in biological systems generally generated accepted methods of reactive oxygen species (ROS) level analysis. When out of control, ROS induces tissue damage, chronic inflammatory processes and cellular functional disturbances. Aerobic organisms have adapted to defense against ROS aggression by developing potent antioxidant mechanisms. Recent advances in ROS measurement methodology allow the study of ROS biology at a previously unachievable level of precision. However, their high activity, very short life span and extremely low concentration, make ROS measurement a challenging subject for researchers.