With readily applicable hybridization assays, mainly based on rRNA-targeted nucleic acid probes, and direct, cultivation-independent sequence retrieval, microbiologists can for the first time determine the true composition of microbial communities. Phylogenetic identification and exact spatiotemporal quantification of microorganisms will in the future become prerequisites for high quality studies in microbial ecology just as good taxonomy and solid quantification have always been for macroecology. This review is intended to give a short history of the development of rRNA-targeted nucleic acid probes and probe technologies, as well as of their application in microbial ecology. The current state of the art is described, and we will try to look into the future. Over the last decade, rRNA-targeted probes have become a handy tool for microbial ecologists. In order to speed up the transformation of microbial ecology from a mostly descriptive to a hypothesis-driven, experimental science more intense use must be made of the taxonomic precision and quantitativeness of rRNA-targeted probes.