Confocal laser scanning microscopy (CLSM) is one of the most relevant technologies for studying biofilms in situ. Several tools have been developed to investigate and quantify the architecture of biofilms. However, an approach to accurately quantify the intensity of a fluorescent signal over biofilm depth is still lacking. Here we present a tool developed in the ImageJ open-source software that can be used to extract both structure and fluorescence intensity from CLSM data: BIAM (Biofilm Intensity and Architecture Measurement). This is of utmost significance when studying the fundamental mechanisms of biofilm development, differentiation, and in situ gene expression or when aiming to understand the effect of external molecules on biofilm phenotypes.
Keywords: Biofilms; Bioimage analysis; Confocal microscopy; Fluorescence intensity; Structure.