The analysis of bacteria at the single cell level is essential to characterize processes in which cellular heterogeneity plays an important role. BACMMAN (BACteria Mother Machine ANalysis) is a software allowing fast and reliable automated image analysis of high-throughput 2D or 3D time-series images from experiments using the "mother machine", a very popular microfluidic device allowing biological processes in bacteria to be investigated at the single-cell level. Here we describe how to use some of the BACMMAN features including i) segmentation and tracking of bacteria and intracellular fluorescent spots, ii) visualization and editing of the results iii) configuration of the image processing pipeline for different datasets, and iv) BACMMAN coupling to data analysis software for visualization and analysis of data subsets with specific properties. Among software specifically dedicated to the analysis of mother machine data, only BACMMAN allows segmentation and tracking of both bacteria and intracellular spots. For a single position, single channel with 1000 frames (2GB dataset) image processing takes about 6 min on a regular computer. Numerous implemented algorithms, easy configuration and high modularity ensure wide applicability of the BACMMAN software.