The interatrial septum (IAS) is thought to be involved in the mechanism of persistent atrial fibrillation (PeAF). Simultaneous contact mapping of both sides of the IAS has not been performed previously.The purpose of this study was to describe wave front (WF) activation patterns and extent of left and right atrial septal electrical dissociation in patients with PeAF.Simultaneous mapping of both atrial septal surfaces using 2 high-density grid catheters was performed. Filtered electrograms of continuous atrial fibrillation, sinus rhythm (SR), and atrial pacing recordings were exported to MATLAB for off-line phase/activation analysis, and activation patterns on paired surfaces were analyzed. WF activation patterns between the 2 grids were evaluated to determine whether activation WFs were associated or dissociated.Eight patients with PeAF undergoing catheter ablation were included. Complete dissociation of WF activation patterns between the 2 sides of the septum existed throughout the mapping period with no 2 consecutive WF activation patterns matching. Single linear WFs were the most prevalent activation pattern on both septal grids. No focal breakthroughs were seen. Transient rotational activity was seen in 10% of phase activations. During SR and atrial pacing, both grids appeared to be activated independent of each other with no evidence of contralateral conduction across the 2 grids.Simultaneous biatrial septal mapping of human PeAF, SR, and atrial pacing shows complete WF dissociation between the left and right IAS with no evidence of trans-septal conduction, indicating that the 2 sides function as electrically discrete structures. No stable septal drivers were observed. These findings may have implications for mapping and ablation of PeAF.