Post-systolic shortening (PSS) does not contribute to the ejection of blood and may inhibit diastolic filling. We determined normal values of PSS in healthy subjects and investigated associations with echocardiographic and invasive measures of systolic and diastolic function. We prospectively analyzed participants from the general population (n = 620, mean age 47 ± 14 years) with no cardiovascular disease. Participants underwent echocardiography, including speckle tracking assessment of the post-systolic index (PSI), strain and time. We defined the PSI as: 100 × [(peak global longitudinal strain - peak systolic longitudinal strain)/(peak global longitudinal strain)]. We also included stable patients (n = 44) referred for left ventricle (LV) catheterization and echocardiography. Normal values: median PSI 2.0% (IQR 0.7, 4.8), post-systolic strain 0.4% (IQR 0.2, 0.8) and post-systolic time 22.6 ms (IQR 10.7, 40.8). Sex modified the relationship between PSI and age (P interaction = 0.037), such that PSI increased with age in women but not in men. PSI was associated with diastolic function (e', E/e' and E/A) (P < 0.05 for all), but not with LV ejection fraction (P = 0.08). PSI was associated with invasively measured LV pressure decline in early diastole, dP/dt min ([Formula: see text] = 0.12, P = 0.010), but not with LV pressure rise in early systole, dP/dt max ([Formula: see text]= - 0.05, P = 0.30). A PSI > 5% had 82% specificity and 99% sensitivity for identifying impaired LV systolic and/or diastolic function. Normal values of PSS are modified by sex. The PSI is associated with most validated echocardiographic and invasive measures of cardiac systolic and diastolic function.