Array-based mRNA profiling offers a variety of opportunities to address different issues relevant to anti-bacterial research. The technique can be used to investigate the mode of action of antibiotics, bacterial resistance mechanisms and virulence factors, and to identify novel targets. This review discusses recent developments of this highly innovative field of technology with respect to technical requirements and experimental design. Several applications are described in which bacterial mRNA profiling has already been successfully performed, illustrating how the generation of large numbers of diverse datasets can be used as a powerful tool for evaluating anti-bacterial compounds and consequent counteracting mechanisms in the cell.