In order to investigate the usefulness and accuracy of two-dimensional echocardiography (2-D echo) for the diagnosis of bovine congenital heart disease, a series of 40 cases, mostly calves, with various types of cardiac malformations were studied prospectively. All but one case was confirmed postmortem. The patients were divided into 2 groups according to the 2-D echo methods used and the results for both groups were compared. The group studied initially (25 cases) using a linear scanner showed a substantially lower degree of diagnostic accuracy than the group studied subsequently (15 cases) using our new systematic 2-D echo approach with a sector scanner. With the latter approach, complex forms of cardiac defect such as transposition of the great arteries and double-outlet right ventricle were correctly diagnosed. However, difficulty was encountered in the definitive diagnosis of such extracardiac defects as patent ductus arteriosus and total anomalous pulmonary venous connection, in which the additional use of contrast or pulsed Doppler echocardiography was a great aid to confirmation. The 2-D echo technique is non-invasive and easily applicable. Therefore, it appears to be a very promising clinical procedure for the diagnosis of bovine congenital heart disease.