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Abstract The correlation between the anatomical cardiac and electrocardiographic axes has been discussed for several years. Using upright computed tomography, this study aimed to reveal the relationship between the anatomical cardiac and electrocardiographic axes in both the supine and upright positions. Upright CT and standard supine CT were performed for healthy volunteers, following electrocardiography in both upright and supine position. On CT images, the coordinates of apex, the center of aortic valve (AV) and mitral valve (MV) were recorded, and the vectors and angles were calculated. Subcutaneous and visceral fat volume were semi-automatically calculated in a workstation. From a total 190 volunteers, 41 males were performed electrocardiography and included for this study. The QRS and anatomical axes (AV-apex and MV-apex axis) were significantly correlated in both supine and upright positions, while the angle of the AV-apex to Z axis was the most correlated (supine: r = − 0.54, p = 0.0002, upright: r = − 0.47, p = 0.0020). The anatomical axis moved in the dorsal and caudal directions from the supine to upright position. Multiple regression analysis revealed that the anatomical axis from the AV-apex to the Z-axis was determined according to age, body height, subcutaneous and visceral fat volumes.