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Plant pathogens such as fungi, oomycetes, viruses and bacteria infect important crops and account for significant economic losses worldwide. Therefore, it is critical to gain insights into plant-pathogen interactions at the cellular and molecular level. The outcome of the interaction between plants and pathogens greatly differs depending on the species, strains and cultivars involved as well as environmental factors, yet typically results in stress for the plant, the pathogen or both. These biotic-induced stresses can be monitored using a wide range of techniques, of which some of the most commonly used techniques are outlined in this chapter. One widely observed feature of biotic stress in plants is the generation of reactive oxygen species (ROS) such as hydrogen peroxide (H2O2) and superoxide (O2-). We describe the quantification of hydrogen peroxide by 3,3'-diaminobenzidine (DAB) staining and luminol-based assays, and of superoxide by nitroblue tetrazolium (NBT) staining. Other techniques detailed here include measurement of callose deposition by aniline blue staining, evaluation of cell death by trypan blue staining and analysis of the loss of membrane integrity by monitoring electrolyte leakage.

Original publication




Journal article


Methods Mol Biol

Publication Date





241 - 255


Aniline blue, Biotic stress, Callose deposition, Cell death, DAB, Electrolyte leakage, Luminol, NBT, Nicotiana benthamiana, Plant pathogen, Pseudomonas syringae, ROS, Trypan blue