Doxorubicin entrapped carbon dots (DOX-CDs) were prepared for bioimaging and enhanced intracellular drug delivery. The CDs were synthesized via the hydrothermal method using citrate and urea under 200 °C for 1 h. Then, DOX was successfully conjugated on the CDs via physicochemical interactions. The DOX-CDs exhibited good crystal structure, remarkable aqueous stability, excellent photoluminescence property, and a high quantum yield of 93%. The fluorescent images revealed that the DOX-CDs could be readily taken up by the cancer cells for cell labeling. Furthermore, endo-lysosomal pH-assisted DOX release behavior was observed from DOX-CDs, and the cytotoxicity of DOX-CDs was confirmed by the MTS assay against H0-8910 ovarian cancer cells. In addition, the CDs indicated bright fluorescent signal in the animal imaging test and demonstrated low toxicity after administration for 7 and 21 days. Therefore, the prepared CDs could be a promising imaging probe for biomedical imaging and intracellular drug delivery.