In the paper，adenosine complementary aptamer strand（S1） and carboxylic acid terminated DNA（S2） were firstly modified on the gold nanoparticles（GNPs） surface through Au-S bonds.With the help of the amidation reaction of toluidine blue（TB） and S2，TB was labeled on the surface of gold nanoparticles for the preparation of TB labeled DNA probe TB-S2-GNPs-S1.Then a layer of gold nanoparticles was electro-deposited on the glass carbon electrode surface，which was used as the immobilization matrix for adenosine aptamer with terminal mercapto group.Subsequently，bovine serum albumin（BSA） as a blocking agent was used to eliminate non-specific adsorption.TB-S2-GNPs-S1 was immobilized on the electrode surface through the hybridization of S1 and the adenosine aptamer.An aptamer biosensor was constructed with toluidine blue as the electrochemical redox probe.The synthesized gold nanoparticles and TB-S2-GNPs-S1 composites were characterized by UV-Vis spectroscopy and scanning electron microscopy.The electrode assembly process was monitored by cyclic voltammetry and electrochemical impedance spectroscopy.The sensor’s performance was studied by differential pulse voltammetry and electrochemical impedance spectroscopy.The sensor showed a linear relationship to adenosine from 1.0×10-4 ng/mL to 100.0 ng/mL，with a correlation coefficient（r） of 0.994 and a detection limit（S/N=3） of 64.7 fg/mL.