Exploiting advances in imaging technology to study biofilms by applying multiphoton laser scanning microscopy as an imaging and manipulation tool.

Biofilms are an important element of the natural ecosystems but can be detrimental in health care and industrial settings. To improve our ability to combat biofilms, we need to understand the processes that facilitate their formation and dispersal. One approach that has proven to be invaluable is to image biofilms as they grow. Here we describe tools and protocols to visualize biofilms with multiphoton laser scanning microscopy, compare this with single photon laser scanning confocal microscopy and highlight best working procedures. Furthermore, we describe how with multiphoton laser scanning microscopy the laser can be used to manipulate the biofilm, specifically to achieve localized bleaching, killing or ablation within the biofilm biomass. These applications open novel ways to study the dynamics of biofilm formation, regeneration and dispersal.