A biosensor for glucose determination was fabricated by the immobilization of glucose oxidase (GOx) on carbon nanotubes (CNTs) and/or 1,10-phenanthroline-5,6-dione (PD) modified graphite rod electrodes (GOx/PD/CNTs/GR) and its amperometric response toward glucose was investigated under aerobic and anaerobic conditions. The sensitivity of the GOx/PD/CNTs/GR electrode was found to be higher compared to that of a PD-modified GR electrode without CNTs (GOx/PD/GR), implying that CNTs play an important role in the facilitation of electron transfer between the redox active site of GOx and the electrode surface. The GOx/PD/CNTs/GR biosensor exhibited a linear dependency on substrate concentration in a range from 0.0 until 50.0 mM of glucose with oxygen present and from 0.0 until 62.5 mM of glucose in the absence of oxygen. With oxygen present, the limit of detection (LOD) values were determined to be 5.4 and 8.0 mM, and the limit of quantitation values (LOQ) were calculated as 16.2 and 24 mM for GOx/PD/GR and GOx/PD/CNTs/GR, respectively. In the absence of oxygen, the LOD values were calculated as 4.2 and 10.7 mM, and the LOQ values were calculated as 12.6 and 32.1 mM for GOx/PD/GR and GOx/PD/CNTs/GR, respectively. When examining the interference effect of uric acid for GOx/PD/GR and GOx/PD/CNTs/GR electrodes, no significant changes in the amperometric response of the modified electrodes were observed up to 100.0 mM of uric acid.