In cardiac muscle variations of the cell cycle (endoreduplication) make it technically challenging to provide evidence for cell division. The only definitive proof for proliferative activity followed by cytokinesis in cardiac muscle is the observation of a contractile ring or a midbody prior to abscission. To visualize these cell cycle specific events we developed a new in vivo proliferation marker that indicates M-phase in great detail. This was achieved by fusing eGFP to the scaffolding protein Anillin. A transgenic G4 embryonic stem (ES) cell line was generated stably expressing the eGFP-Anillin fusion protein under the control of the ubiquitous CAG promoter. Undifferentiated ES cells of this line displayed a high overlap of eGFP-Anillin expression with the mitotic marker Ki-67 in immunofluorescent stainings (97%) and flow cytometry analysis (94%) respectively. During differentiation in embryoid bodies the rate of eGFP-Anillin positive cells declined with the appearance of postmitotic cells and cells of all three germ layers showed expression of the fusion-protein. During embryonic development of eGFP-Anillin mice proliferative cells could be monitored in all organs based on the eGFP-Anillin expression. At postnatal day 3 we detected the highest expression of the eGFP-Anillin fusion protein in cardiac muscle due to the strong expression of the CAG promoter but eGFP-Anillin positive cells were abundant also in other tissues i.e. testis and skin. We probed adult eGFP-Anillin mice for the identification of proliferating cells after myocardial injury. Four days post infarction a high amount of eGFP-Anillin positive cells with typical localization indicating cytokinesis was detectable in cryosections. This new system for visualization of proliferation in vivo will allow to accurately monitor cell division, especially in tissues where cells undergo endoreduplication or acytokinetic mitosis. It will be possible to address e.g. the open question of the regeneration of the heart muscle upon injury.