Molecularly imprinted polymer based working electrode was prepared for the determination of ciprofloxacin (CF). Electrochemical behavior of CF was investigated by differential pulse voltammetric (DPV) method in the range of 10-60% (v/v) acetonitrile (MeCN)-H2O binary mixture at the pH between 3-7 prepared Britton-Robinson (BR) buffers. Voltammetric analyzes were performed with molecularly imprinted (MIP) and non-imprinted (NIP) modified electrodes. The influence of the electropolymerization cycles, solution pH, and MeCN-H2O ratio, on the performance of both electrodes were evaluated and optimized. Detection limit was obtained as 3.34×10-5 M (S/N=3). Recommended method was successfully employed for determination of CF in pharmaceutical, human serum and urine.