We recently speculated that mitral-aortic separation (MAS) might be increased in discrete subaortic stenosis (DSS). We have examined this hypothesis in 22 heart specimens in which the subaortic obstruction originated on the muscular ventricular septum below the right aortic sinus, either as a discrete band, an accumulation of several bands or a diffuse ridge, and extended posteriorly into the MAS or anterior leaflet of the mitral valve or both, with a variable relationship to the aortic cusps and sinuses. No specimen had ventricular septal defect, supravalvular aortic stenosis or other features of Shone syndrome. The mean MAS was nearly twice that of 80 normal specimens (4.9 vs 2.5 mm), the range of MAS was increased from normal (0.11 vs 0.7 mm) and the mean diameter of the aortic annulus was decreased compared with the normal specimens, data that will be of interest to echo- and angiocardiographers in the clinical description of DSS, and to the surgeon who must resect these lesions.