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One of the most groundbreaking discoveries over the last 40 years has been the recognition of microbial biofilms as the predominant lifestyle of microorganisms, outside laboratory conditions. Bacterial biofilms are defined as aggregated, often sessile bacteria, which differ from free-floating cells by their slow growth and tolerance to antibiotics and immune cells.1 It is estimated that over 65% of microbial infections are biofilm-mediated, conferring biofilms a high clinical relevance.2 Moreover, in industrial settings, biofilms cause biofouling, corrosion, machinery damage and product contamination that cause delays, add cost and give rise to energy losses.3 In a biofilm, living cells integrate into a structured multicellular community that is enclosed by a self-produced polymeric matrix, composed of polysaccharides, extracellular DNA and proteins. As biofilm cells multiply, they dynamically produce matrix components, which make the 3D-community more stable and tolerant to external threats. Thus, the study of the biofilm matrix provides essential clues into the community’s functional behavior, which are critical for the understanding of biofilms from an industrial, environmental, biomedical, pharmaceutical and even purely microbiological perspective. Exploration of Microbial Communities Using the Thermo Scientific Varioskan LUX Multimode Reader and the Invitrogen EVOS FL Cell Imaging System