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Objectives This study sought to describe, in detail, the angiographic, gross macroscopy, and histological dissection of the conduction axis in humans. Background The recent upsurge of interest in specific pacing of the atrioventricular conduction axis has emphasized the need for precise knowledge of the location of the atrial and penetrating components of the atrioventricular conduction axis. Methods A total of 41 human hearts were studied by serial histological sectioning and an additional 3 hearts by gross dissection. One of the hearts studied histologically was also dissected to show the location of the conduction axis prior to serial sectioning. The anatomical findings were then compared with the results of angiography undertaken in the catheter laboratory in 60 patients undergoing electrophysiological studies. Results Marked variation of the location of the transition from atrioventricular conduction axis to the penetrating atrioventricular bundle, or the bundle of His, relative to the landmarks of the triangle of Koch was observed. In just over one-half of both the specimens and the patients, the site of penetration was on the atrial aspect of the hinge of the septal leaflet of the tricuspid valve, with further variation noted relative to the apex of the triangle of Koch. Based on measurements of the histological sections, marked variation in the dimensions of the axis and its adjacency to the right-sided endocardium were also found. In almost three-fifths of hearts, an interventricular component of the fibrous membranous septum was not able to be identified. The significance of these findings to those who seek to perform selective pacing of the atrioventricular conduction axis are discussed. Conclusions Marked variability of the location of the conduction axis within the triangle of Koch is reported. In three-fifths of hearts, the interventricular component of the fibrous membranous septum is nonexistent.