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Introduction: The development of real-time three-dimensional echocardiography (RT3DE) can circumvent the limitations of two-dimensional echocardiographic (2DE), avoiding the geometric assumption limitation by permitting direct planimetry of the cross-sectional left ventricular outflow tract (LVOT) area (a) and stroke volume (SV) assessment.Purpose: To study whether measurement of LVOT-diameter (d) by using color Doppler in order to more accurately define LVOTd might be more robust for determination of SV than gray scale determination of LVOTd. Furthermore, we wanted to elucidate whether direct measurement of LVOTa by RT3DE could improve SV calculation. Methods and Results: 2DE and RT3DE were acquired in 21 volunteers for SV assessment. LVOTa was calculated by 2DE using: π * (d/2)2, where d being the LVOT diameter measured with gray scale and color Doppler. Using RT3DE in parasternal long axis (PLAX) view, direct planimetry of LVOTa was performed 5mm above the aortic valve. Eccentricity Index (EI) was calculated using the lateral and antero-posterior LVOTd. SV was obtained by 4 different methods 1) 2D gray scale SV = π×(LVOTd/2)2×LVOT velocity time integral (VTI), 2) 2D color SV = π×(LVOTd/2)2×LVOTVTI, 3) LVOTa×LVOTVTI, 4) SV assessed by Simpson's biplane method. LVOTd measured with grey scale was significantly smaller in comparison to LVOTd obtained with color Doppler (20,4±2,1mm vs. 21,5±1,9mm, p<0,05). Significant differences were observed between LVOTa grey scale and color Doppler (3,29±0,74cm2 vs. 3,67±0,70cm2, p<0,05) and also between LVOTa assessed by grey scale in comparison to RT3DE planimetry; (3,29±0,74cm2 vs. 3,61±0,89cm2, p=0,011). No significant differences were observed with color Doppler and RT3DE planimetri regardning LVOTa assessment (3,67±0,70cm2 vs. 3,61±0,89cm2). EI was 0,21±0,13mm and its median 0,17mm, which means that half of the subjects had at least 17% difference between the lateral and the antero-posterior LVOTd. Regarding SV, there were significant differences between 2D grey scale and 2D color Doppler (82,8±17,1mL vs. 92,4±16,8mL, p<0,05) and between 2D grey scale and RT3DE LVOTa planimetry (82,8±17,1mL vs. 90,7±19,8mL, p=0,025). Inter-observer variability for LVOTa with color Doppler and RT3DE LVOTa planimetry was <5%.Conclusion: This study showed that LVOT shape is usually not round. We also found similar results between 2DE color Doppler and RT3DE planimetry in LVOTa and SV assessment, suggesting 2DE color Doppler as an alternative method for accurate SV determinnation in daily clinical practice.