The involvement of microorganisms in infections is often inferred from their presence during disease and their absence in healthy humans or animals, which is an oversimplification. Microorganisms often form stable, structured polymicrobial consortia, such as biofilms and cohesive sludges, which exhibit properties that differ from those of the organisms involved. The detection of such biofilms and proof of their direct involvement is crucial. Fluorescence in situ hybridization (FISH) combines the specific identification of microorganisms with the morphological aspect of the host tissue and is therefore particularly useful for these purposes. This paper describes FISH methods that we use in the ambulance for Polymicrobial Infections and Bacterial Biofilms at Charité Hospital to visualize pathogens (pathogenic consortia) in clinical samples.