Heart volume may provide important information on the status of fetal hemodynamics. However, traditionally fetal heart volume has been assessed with the erroneous assumption that the fetal heart is spherical or elliptical. With the advent of three-dimensional ultrasound, accurate assessment of organ volume has become feasible. The objectives of this study were to compare the reproducibility of two-dimensional ultrasound and three-dimensional ultrasound in the assessment of heart volume, and to test whether heart volume assessed by the traditional method of two-dimensional ultrasound equates to that assessed by three-dimensional ultrasound. If it proved to be significantly different, we aimed to find a new constant which, if incorporated into the traditional formula used to determine heart volume, would enable us to achieve more accurate volumes with two-dimensional ultrasound. In total, 50 normal singleton fetuses ranging from 20 to 30 weeks' gestation were included in the study. Both the traditional two-dimensional and the new three-dimensional volume measurements were compared. The results showed that three-dimensional ultrasound has a better reproducibility than two-dimensional ultrasound in heart volume assessment and that heart volume assessed by the traditional formula of two-dimensional ultrasound is significantly larger than that measured by three-dimensional ultrasound (p < 0.001). We therefore propose that, if the traditional two-dimensional equation is to be used, the constant for heart volume could be modified to 0.4563 (SE = 0.0153, n = 50) to achieve more accurate results. With this new constant, the heart volume derived by two-dimensional ultrasound was not found to differ from that measured by three-dimensional ultrasound. From our series, we conclude that three-dimensional ultrasound is theoretically the best method for the assessment of heart volume. However, because of the limitations of three-dimensional ultrasound (i.e. it is not routinely available, it is more expensive and more time-consuming) two-dimensional ultrasound in practical terms should be the method of choice. Although we were able to improve the accuracy of the heart volume measurements using a new constant in the traditional two-dimensional formula, the new constant will not improve variability, which can only be reduced by three-dimensional ultrasound.