Tissue Doppler derived longitudinal strain and strain rate were studied in 48 healthy term neonates by measuring peak systolic strain (PSS) and peak systolic strain rate (PSSR) in 18 heart segments on day 1-3 of life. The mean PSS for each examination was -21.8% (-22.1, -21.4) [mean (95%CI)], and the mean PSSR was -1.78/s (-1.81, -1.74). Age (d), fetal shunts, and heart rate had negligible impact on measurements for any segment. The fractional shortening had negligible impact on septum and left heart segments and the pulmonary artery pressure had no impact on the right heart segments. Values varied significantly between segments and individuals, and were lower (closer to zero) in images of low quality than in images of high quality (p < 0.05). Both apical and basal segments values were lower in the septum and higher in the right ventricle than in the left ventricle (p < 0.05), except for no difference between the basal left and right segments PSSR. Apical segments values were higher than basal segments values in the right ventricle (p < 0.05) but not in the septum or the left ventricle. At present, PSS and PSSR are more feasible for quantifying differences between segment groups and patient groups than between individuals and segments within individuals.