Immunofluorescence colony‐staining (IFC), as a concept in bacterial detection, has developed during the past decade, providing several new options for the study of culturable plant‐associated bacteria: (1) in situ detection of target colony‐forming units on or in plant parts, such as on roots or in stem sections; (2) selective isolation of cross‐reacting bacteria for research on the improvement of the reliability of serological detection systems; (3) distinction between culturable and non‐culturable cells; (4) simultaneous detection of different target bacteria using different fluorescent markers and a dual fluorochrome‐filter system. IFC‐positive colonies can be directly identified by isolation or PCR, which makes it easier to recognize false positive results. The use of IFC for the selective isolation of cross‐reacting bacteria forms a major tool for the further improvement of the specificity of IFC and serological assays.