Microbial bioaggregates and biofilms are hydrated three-dimensional structures of cells and extracellular polymeric substances (EPS). Microbial communities associated with interfaces and the samples thereof may come from natural, technical, and medical habitats. For imaging such complex microbial communities confocal laser scanning microscopy (CLSM) is the method of choice. CLSM allows flexible mounting and noninvasive three-dimensional sectioning of hydrated, living, as well as fixed samples. For this purpose a broad range of objective lenses is available having different working distance and resolution. By means of CLSM the signals detected may originate from reflection, autofluorescence, reporter genes/fluorescence proteins, fluorochromes binding to specific targets, or other probes conjugated with fluorochromes. Recorded datasets can be used not only for visualization but also for semiquantitative analysis. As a result CLSM represents a very useful tool for imaging of microbiological samples in combination with other analytical techniques.