Carbon dots (CDs) hold great promise as a class of fluorophores for bioimaging. Here, we report a simple solvothermal approach to prepare photoluminescent CDs derived from the antineoplastic drug etoposide. The CDs show photoluminescence ranging from 365 to 650 nm when excited at 360 nm. When excited at 405, 488 and 543 nm, strong blue, green and red fluorescence can be detected. Our experiments reveal that the CDs can penetrate into the cells in a short time and remain emissive, even at low temperature. Also, optical signals can be detected from mice after hypodermic or intravenous injection. We demonstrated that the CDs can serve as a low-cytotoxicity biological imaging agent for bioimaging in vitro and in vivo.