Quantification of superoxide and hydrogen peroxide in leaves

Reactive oxygen species (ROS) are produced in plants under both non-stressful and stressful conditions. Various histochemical staining methods have been developed and are widely used to visualize ROS accumulation sites. In contrast to qualitative analysis, quantification of ROS has been time- and labor consuming. As a consequence, the number of samples, which could be analyzed in parallel, has been limited. To overcome this problem, we introduce an improved semiquantitative method, in which ROS levels are quantified after histochemical staining in plant organs with the digital image analysis package ImageJ.