NADPH oxidase, cytoskeleton, oxygen radicals, killing/digestion of microbes. Phagocytes and in particular neutrophils constitute the first line of defence of the innate immune system against invading pathogens. These white blood cells are able to chase, bind and finally engulf bacteria in a sealed phagocytic vacuole formed by invagination of the plasma membrane. Within this specialised organelle, pathogens are exposed to proteolytic enzymes that are downloaded from cytoplasmic granules. Setting ionic conditions within the vacuole that are optimal for the activity of these enzymes is critical for an efficient killing and digestion of the pathogen. To this contribute the NADPH oxidase, as well as a range of ion channels, ion exchangers and ion pumps. As it generates some reactive oxygen species in the vacuole, the NADPH oxidase has also been considered as bactericidal by itself. We use a range of electrophysiological, imaging and spectroscopy techniques to delineate the respective role of proteases and reactive oxygen species in the killing process.