Intracellular Fe3+ amount is one of the critical determinants of human health. The development of simple and effective probes for the quantitative detection of Fe3+in vivo is of great significance for the early diagnosis of disease or disorder associated with iron deficiency or overload. In this study, remarkable carbon dots, which can serve as a biosensor for efficient intracellular Fe3+ detection, were synthesized by hydrothermal carbonization of Fusobacterium nucleatum, an anaerobic bacterium. The achieved F. nucleatum-carbon dots (Fn-CDs) possessed the features of strong fluorescence, high stability and excellent biocompatibility. The obtained Fn-CDs could easily internalize into both plant cells and human cells with excellent ability for cell tracking and biomedical labeling. The fluorescence of Fn-CDs could still remain for another 24 hours after penetrating into cells. Furthermore, the fluorescent Fn-CDs were very sensitive to the presence of Fe3+ ions even in cells, exhibiting great promising applications in in vivo detection of Fe3+ ions. In addition, the Fn-CDs posed no harm to the mice, being circulated and excreted within a short time, making the Fn-CDs an excellent candidate for bioimaging and biosensing in vivo.