This study compared several irrigation protocols and application systems for sealer penetration into dentinal tubules. Single-rooted-human teeth were divided into 5 experimental groups (n = 15) and a control group (n = 5), according to final irrigation protocols: standard needle irrigation (SNI); Vibringe; Vibringe + NaviTip FX (Vibringe NFX); Endo Spray (ES); and passive-ultrasonic-irrigation (PUI). Following obturation of the root canals, the percentage of the sealer penetration was measured at different depths using stepwise CLSM analysis. The sealer penetration in the experimental groups was significantly higher than the control group at all levels (p<.05). No significant differences were observed between Vibringe and SNI or Vibringe NFX, ES, and PUI at all depths (p>.05). The Vibringe NFX, ES, and PUI groups allowed deeper sealer penetration than SNI at 100, 250, and 500 mu m levels (p<.05). The irrigant activation, the needle design, and the application form (syringe or spray) may impact the quality of the seal that is achieved with root canal filling.