A novel potentiometric sensor with high selectivity and sensitivity has been developed for the determination of heparin, based on the modification of heparin-imprinted polymer film onto a glassy carbon. The performance of the developed heparin sensor was evaluated, and the results indicated that a sensitive potentiometric sensor could be fabricated. The obtained heparin sensor shows high-selectivity monitoring of heparin, shorter response time (2 months). The potentiometric sensor was successfully applied to the determination of heparin in heparin sodium injection with recoveries between 97.1% and 110.0%.