The objective of this study was to determine intra- and inter-observer variability of echocardiographic measurements in awake cats. Four observers with different levels of experience in echocardiography performed 96 echocardiographic examinations in four cats on four different days over a 3-week period. The examinations were randomized and blinded. The maximum within-day and between-day CV values were 17.4 and 18.5% for inter-ventricular septal thickness in diastole, 18.7 and 22.6% for left ventricular free-wall thickness in diastole, 9.8 and 14.9% for left ventricular end-diastolic diameter, 20.8 and 15.2% for left ventricular end-systolic diameter, and 21.2 and 18.4% for left ventricular shortening fraction. The maximum within-day CV values were most often associated with the least competent observer (i.e. the graduate student) and, the minimum CV values with the most competent observer (i.e. the associate professor in cardiology). A significant interaction between cat and observer was also evidenced. Thus, the most competent observer could not be replaced by any of the other observers.