Previous studies assessing endothelial function as flow-mediated changes in the brachial artery diameter have not been able to measure the true inner luminal diameter. This is due to the lack of image quality, which has hampered the visualisation of the lumen-intimal interface. Because increases in resolution and scanning frequency have recently led to improved ultrasound (US) image quality, we assessed the feasibility of measuring the true brachial artery diameter using digital US and 13-MHz scanning frequency. Satisfactory true inner diameter measurements were obtained in all subjects (n = 148, middle-aged men, mean age 54 +/- 7 y) participating in a risk factor study. At baseline flow, the intima to intima diameter was 4.03 +/- 0.49 and 4.67 +/- 0.52 mm measured conventionally from the anterior to the posterior media-adventitia interface (difference 0.64 +/- 0.10 mm). After hyperaemia, the intima to intima diameter was 4.23 +/- 0.46 mm and the adventitia to adventitia diameter 4.86 +/- 0.50 mm. Flow-mediated dilation (FMD) expressed as the percentage change from the baseline diameter measured 5.3 +/- 4.3% using the true inner diameters and 4.3 +/- 3.7% using the conventional outer diameters. The difference in FMD values was systematic, and there was a good linear correlation between them (r = 0.93, p < 0.0001). If FMD is presented as the percentage change from baseline to hyperaemia, this new method gives values that are approximately 1% unit higher, compared with values when brachial luminal diameter is measured in the conventional way between the adventitia-media interfaces.