Not all phosphorus removed in activated sludge systems can be accounted for by polyphosphate accumulating organisms (PAO). A method for the qualitative and quantitative in situ characterization of PAO cell clusters and closely associated extracellular polysaccharides (EPS) is described. X-ray microanalysis was performed on samples from four activated sludge plants situated in Pretoria, South Africa. Analyses were done by means of Scanning Electron Microscopy (SEM) combined with Energy Dispersive Spectrometry (EDS). Cell clusters with associated EPS on average contained between 57 and 59% phosphorus, while EPS alone contained on average between 23 and 30% phosphorus. Results suggest that phosphorus removal in activated sludge might be due not only to PAO, but also by EPS acting as a phosphorus reservoir. Extraction of EPS from two different activated sludge plants yielded different amounts of EPS, which, in combination with SEM-EDS, may shed light on different phosphate uptake abilities of different activated sludges.