Herein, impedimetric detection of THR could be achieved using an aptasensing platform based on carbon nano-fibers enriched screen-printed electrodes (CNF-SPE). The resistance to charge transfer (R-ct) was recorded using electrochemical impedance spectroscopy (EIS) technique before/after the immobilization of amino-modified DNA aptamer (APT) selective to thrombin (THR) onto the surface of CNF-SPEs and the specific interaction between APT and THR. The selectivity of the aptasensor was also tested in the presence of a random DNA oligonucleotide and a DNA aptamer that were different from THR specific APT. The impedimetric aptasensing of target protein was also explored in the fetal bovine serum (FBS) medium at different concentration levels of THR. Additionally, the selectivity of the aptasensor was tested against bovine serum albumin (BSA) and protein C (PC) in FBS medium. This CNF-SPE based aptasensor platform allows a reliable, sensitive and selective impedimetric monitoring of THR. (C) 2015 Elsevier B.V. All rights reserved.