A novel electrochemical sensor based on a carbon paste electrode modified with electrospun carbon nanofibers and TiO2 nanoparticles was prepared. It was designed to electrochemical determination of Idarubicin (IDA) in biological samples. To this end, carbon nanofibers were prepared by electrospinning process from polyacrylonitrile (PAN) and then modified with TiO2 nanoparticles. Scanning electron microscopy (SEM) and electrochemical impedance spectroscopy (EIS) techniques were employed for characterization of the various nanocomposite modified electrode. The results revealed that the modified electrodes showed an obvious electrocatalytic activity toward the oxidation of IDA by a remarkable enhancement in the current response compared with bare electrode. The modified electrode was applied to determine of IDA with square wave voltammetry (SWV) technique. Under optimized conditions, IDA gave a linear response in the concentration range of 0.012 to 10.0 mu M with the detection limit of 3 nM. Interfering species of other anticancer drug such as doxorubicin, epirubicin and daunorubicin showed no interference with the selective determination of IDA. The finding suggests that the mentioned method was completely validated and successfully applied for the determination of IDA in real samples such as serum.