This study aims to assess the efficacy of various irrigation protocols on the dentinal tubule penetration of a bioceramic-based endodontic sealer. Sixty-four single-rooted extracted human mandibular incisors were used. After instrumentation, teeth were randomly divided into the following four groups (n = 16 each) according to the final irrigation technique: group 1, conventional endodontic needle (CEN); group 2, EndoActivator (EA); group 3, Er,Cr: YSGG laser; group 4, XP-endo Finisher (XPF). The root canals were finally irrigated with 17% EDTA and 5.25% NaOCl for 1 min (min) respectively. The teeth were then obturated with Endosequence BC Points and rhodamine B dye-labeled BC Sealer. After 2 weeks, 1-mm-thick transverse sections were cut 2 and 5 mm from the apex, and examined by confocal laser scanning microscopy at 5 × magnification. The total percentage of sealer penetration (TPSP), sealer penetration area (SPA), and maximum sealer penetration depth (MSPD) was measured. Data were analyzed by Kruskal–Wallis, Dunn’s multiple comparison, and Wilcoxon tests, with significance set at P < 0.05. At 2 mm level, no significant differences were detected among the groups (P > 0.05). At the 5 mm level, the XPF group showed significantly higher values for both TPSP and SPA in comparison with the Er,Cr: YSGG laser and CEN groups (P < 0.05), but no significant difference was observed with the EA group. The choice of different final irrigation techniques can affect dentinal tubule penetration.