Polymorphonuclear neutrophils (PMNs) generate reactive oxygen species (ROS) during phagocytosis and in response to soluble agonists. This functional response, termed oxidative burst, contributes to host defense, but it can also result in collateral damage of host tissues. To study this important PMN response, different methods have been developed that are based on the assessment of oxidative burst by measuring intracellular ROS production or formation of ROS in the extracellular space. Among the different methods that were developed, the following two are particularly widely used because of their convenience and accuracy. The first method depends on the reduction of cytochrome c, which can be assessed by photometry, while the second method relies on changes in the fluorescence properties of dihydrorhodamine 123, which can be assessed by flow cytometry.