To assess ventricular function and dyssynchrony using three-dimensional (3D) computed tomography (CT) strain in adult congenital heart disease (ACHD). We prospectively analyzed a multiphase cardiac CT data set for 22 adult patients with CHD, including 8 patients with congenital systemic right ventricle (RV) and 14 patients with repaired Tetralogy of Fallot (TOF). Eight patients had a cardiac pacemaker. Volume of Interest was drawn on a multiplanar reconstruction of the ventricle with strain overlay using a 3D-strain algorithm. Ventricular strain, inter- and intraventricular dyssynchrony, and right ventricle outflow tract (RVOT)-apex dyssynchrony were calculated. RVOT-apex dyssynchrony by ventriculography was also compared in 15 patients. Pulmonary ventricular strain, systemic ventricular strain, and septal wall strain were lower in ACHD patients than in the controls, and lower in the ACHD with pacing group than without pacing group as well. Maximum interventricular time difference and intraventricular time difference were longer than in ACHD than in the controls, and longer in the ACHD with pacing group than without pacing group as well. RVOT-apex delay was significantly longer in patients with a pacemaker than in those without a pacemaker (118.1 ± 31.9 ms vs. 76.1 ± 36.2 ms, p = 0.03). RVOT delay determined by 3D CT strain significantly correlated with that determined by ventriculography (Pearson r = 0.55, p = 0.03). 3D CT strain can detect reduced biventricular contraction and inter- and intraventricular and RVOT-apex mechanical dyssynchrony can be assessed in patients with ACHD.