Doppler tissue imaging (DTI) is a new echocardiographic imaging technique from which quantitative data regarding myocardial velocity can be extracted. The purpose of this study was to compare endocardial velocities determined from DTI with those determined from M-mode echocardiography and to assess the range of myocardial velocities in normal subjects. Nineteen subjects were evaluated by M-mode echocardiography and quantitative DTI for maximal systolic velocities of the anteroseptal and inferoposterior walls. Mid-myocardial and epicardial velocities were also measured by DTI for each wall. Maximal systolic velocities of the anteroseptal and inferoposterior endocardium determined by DTI correlated significantly with those derived from M-mode echocardiography (r = 0.87). The velocity of the inferoposterior wall by DTI (27.3 ± 4.8 mm/sec) was greater than that of the anteroseptum (20.8 ± 4.1 mm/sec) by a mean difference of 7.0 ± 5.7 mm/sec (p < 0.001). A peak velocity gradient of 5.9 ± 3.5 mm/sec (p < 0.001) between the epicardium to endocardium was detected. These data confirm that (1) regional myocardial velocities may be quantified with DTI in human beings; (2) the high resolution of DTI allows velocities to be determined at different levels within the myocardium; and (3) heterogeneity of myocardial velocities may be demonstrated in normal subjects.