Comparison of Three Final Irrigation Activation Techniques: Effects on Canal Cleanness,Smear Layer Removal,and Dentinal Tubule Penetration of Two Root Canal Sealers

Abstract

Objective: The aim of this study was to compare three final irrigation activation techniques with respect to their effects on debridement efficacy, smear layer removal, and dentinal tubule penetration of two different root canal sealers. Background data: Different applications to improve the delivery of irrigating solutions within the root canal system are currently being investigated, as not all of the mechanisms and effects of these techniques have been clearly identified. Materials and methods: One hundred forty-two single-rooted teeth were randomly divided into a control group and three experimental groups based on the irrigant activation technique used: EndoVac (EV) system, photon-induced photoacoustic streaming (PIPS), and conventional syringe irrigation (CSI). Thirteen specimens from each experimental group were evaluated for debris and smear layer removal using scanning electron microscopy. The remaining 30 specimens per group were divided into two subgroups according to the root canal sealer used: AH Plus and TotalFill BC. The maximum depth and total percentage of sealer penetration were measured using confocal laser scanning microscopy. Results: PIPS resulted in significantly less debris in the middle third of the root canal compared with CSI (p < 0.01). There were no significant differences among CSI, EV, and PIPS concerning debris removal at coronal and apical levels or smear layer removal at all levels (p > 0.05). TotalFill BC use after final irrigation with EV and CSI at 2 mm or PIPS at 5 mm exhibited a significantly higher percentage of sealer penetration than that with AH Plus (p < 0.05). When AH Plus was used, PIPS allowed deeper sealer penetration than CSI at 2 mm (p < 0.05). Conclusions: The effects of EV, PIPS, and CSI on debridement efficacy, smear layer removal, and dentinal tubule penetration were almost comparable. TotalFill BC showed superior tubular penetration than AH Plus.

Introduction

During endodontic instrumentation, debris and smear layer form on the root canal wall.¹ Debris is defined as dentinal particles and vital and necrotic pulp tissue remnants that adhere weakly to the root canal walls after chemomechanical preparation.² The smear layer is an amorphous, irregular, tenacious structure consisting of dentin, pulp tissue remnants, and microbial elements that occlude dentinal tubule openings.³ Both can harbor microorganisms and their by-products.^2,4 Also, the smear layer can act as a barrier to the penetration of intracanal disinfectants and root canal sealers into dentinal tubules, and this can lead to a subsequent microleakage between root canal walls and filling materials.⁵ For these reasons, it is desirable to completely remove debris and the smear layer by alternate use of sodium hypochlorite (NaOCl), a deproteinizing agent,⁶ and the calcium-chelating agent ethylenediaminetetraacetic acid (EDTA).⁷

Different equipment and concepts have been introduced in effort to achieve sufficient delivery of irrigating solutions within the root canal system.^8
–10 The EndoVac system (EV) (SybronEndo, Orange, CA), based on method of apical negative pressure irrigation system, was designed to distribute irrigating solutions to working length and evacuate the root canal of debris.¹¹ Recently, a new laser activation technique, referred to as photon-induced photoacoustic streaming (PIPS), has become commercially available. In this concept, by using subablative energy and restricting tip placement to within the coronal part of the tooth, impulses interact with water molecules within the irrigation solution, generating photon-induced shock waves that lead to rapid fluid streaming and enhance the action of the irrigation solutions.¹²

In addition, preparing the root canal chemomechanically, hermetic three-dimensional filling of the root canal is also considered a key factor in successful endodontic therapy.¹³ During root canal filling, penetration of root canal sealer into the dentinal tubule is necessary because it increases the connection between the core material and dentin, thereby helping to optimize the adaptability and sealing capability of the root canal filling.⁷ Further, sealer penetration can promote an antimicrobial effect in dentinal tubules, which increases when the sealer comes in close contact with microorganisms.¹⁴

Different physical properties and compositions of root canal sealers can also influence penetration depth.¹⁵ TotalFill BC sealer (FKG Dentaire SA, La Chaux-de-Fonds, Switzerland) is a new calcium silicate-based bioceramic root canal sealer containing zirconium oxide, tricalcium silicate, calcium hydroxide, dicalcium silicate, calcium phosphate monobasic, thickening agents, and filler.¹⁶ AH Plus sealer (Dentsply DeTrey, Konstanz, Germany) is a widely used epoxy resin-based sealer demonstrated to have excellent tubular penetration and adaptation.¹⁷

Both the diameter and density of dentinal tubules vary among root regions. Dentinal tubule orifices are larger and denser in the coronal and middle thirds compared with the apical third.¹⁸ Therefore, it is important to compare debris and smear layer removal and tubular penetration of sealers among canal thirds.

The aim of this study was to compare EV, PIPS, and conventional syringe irrigation (CSI) with respect to their effects on debridement efficacy; smear layer removal in the coronal, middle, and apical thirds of the root canal; and dentinal tubule penetration of root canal sealers AH Plus and TotalFill BC at three separate root regions.

Materials and Methods

This study was approved by the Baskent University Institutional Review Board (Project No. D-KA15/04). One hundred forty-two straight and mature roots from extracted human maxillary central incisor and mandibular canine teeth with single canals were selected for the study. The teeth were decoronated with a 0.3-mm diamond disc to standardize the root length to 17 mm from the anatomic apex. The roots were prepared with ProTaper rotary instruments (Dentsply Maillefer, Ballaigues, Switzerland) up to F4 (#40). The root canals were irrigated with 2 mL of 5% NaOCl between instruments. The root apices were covered with sticky wax to simulate clinical conditions and create a closed root canal system to obtain a vapor lock effect.⁹ A size 40 gutta-percha point was placed into the root canal to prevent melted wax from entering it.

One hundred forty-two specimens were randomly assigned into a control group (n = 13) and three experimental groups (n = 43) based on the irrigation activation technique used (Table 1). In the PIPS group, an erbium:yttrium–aluminum–garnet (Er:YAG) laser with a wavelength of 2940 nm was used. A 14-mm long, 300 μm laser fiber tip was operated at 0.3 W, 15 Hz, and 20 mJ/pulse. The pulse duration was 50 μsec (Table 2). In all the experimental groups, root canals were subsequently irrigated with 5 mL distilled water using a 27-gauge notched-tip needle for 30 sec to avoid prolonged effects of EDTA solution, and were dried with sterile paper points. For all groups tested, the irrigation and activation methods involved 15 mL total irrigation solution volume and 90 sec total irrigation solution delivery time.

Table 1.

Experimental Groups and Protocols

Group name	n	Activation	Protocol
Control	13	CSI	Root canals were irrigated with 15 mL distilled water using a 27-gauge notched-tip needle (Ultradent, South Jordan, UT) for 90 sec. During irrigation, the needle was inserted deeply at 1 mm from the WL and moved ∼1–2 mm in a coronoapical direction.
CSI	43	CSI	Final rinse with 5 mL 5% NaOCl followed by 5 mL 17% EDTA using a 27-gauge notched-tip needle (Ultradent) for 30 sec each. During irrigation, the needle was inserted deeply at 1 mm from the WL and moved ∼1–2 mm in a coronoapical direction.
EV	43	EV	The master delivery tip was located in the coronal part of the root canal to deliver solutions into the root canal and then two cycles of microirrigation were performed. Each cycle of microcannular irrigation involved placing the tip at the full WL for 6 sec, withdrawing 2 mm from the full WL for 6 sec and then returning back to the full WL for the next 6 sec. This up and down motion continued for 30 sec. The first microirrigation cycle was completed using 5 mL 5% NaOCl, and the second cycle was completed using 5 mL 17% EDTA. At the end of the second cycle, the microcannula was left at the WL to remove excess irrigant.
PIPS	43	PIPS	The air and water spray in the laser system were switched off. Then, 0.5 mL 5% NaOCl was applied into the root canal and activated with the laser fiber tip for 30 sec. The laser fiber tip remained stationary in the coronal part of the root canal (no further than 4 mm in the root canal) during the irrigation activation protocol. When the irrigating solution in the root canal decreased, a fresh NaOCl solution was added. Subsequently, 0.5 mL 17% EDTA was placed in the root canal and activation was performed as before for 30 sec. When the irrigating solution in the root canal decreased, a fresh EDTA solution was added. The total activation time was 60 sec, and irrigation resulted in 5 mL total irrigant volume for NaOCl and 5 mL total irrigant volume for EDTA.

CSI, conventional syringe irrigation; EDTA, ethylenediaminetetraacetic acid; EV, EndoVac system; NaOCl, sodium hypochlorite; PIPS, photon-induced photoacoustic streaming; WL, working length.

Table 2.

Laser Parameters

Manufacturer	Fotona, Ljubljana, Slovenia
Model	Fidelis AT
Laser system	Er:YAG
Probe	PIPS™ tip (300 μm diameter, 14-mm long fiber tip design)
Wavelength	2940 nm
Power	0.3 W
Energy per pulse	20 mJ
Repetition rate	15 Hz
Pulse duration	SSP, 50 μsec
Placement of the tip	Coronal part of the root canal
Application type	Stationary
Duration of each activation protocol	30 sec

Scanning electron microscopy evaluation

Thirteen specimens from each experimental group were evaluated for debris and smear layer removal using scanning electron microscopy (SEM; Fig. 1). To facilitate fracturing, shallow longitudinal grooves were created on the buccal and lingual root surfaces with a diamond disc. Roots were then split in half using a chisel with a ProTaper F4 gutta-percha cone in the root canal to limit tooth fragments covering canal walls. For each specimen, the half containing the most visible part of the endodontic wall was selected and labeled. Labeled specimens were then dehydrated through graded ethanol baths up to 100%. Each specimen was mounted on an aluminum stub, sputter-coated with ∼10 nm gold, and then examined under SEM (Quanta 200 FEG, Hillsboro, OR) at 5 kV. For each specimen, the coronal, middle, and apical thirds of the root canal were examined under 1000 × magnification. Three representative areas were identified randomly and a mean score was calculated. Two researchers performed a blind evaluation separately. The intraclass correlation coefficients method was used to confirm the intraobserver and interobserver agreement for SEM evaluation. The efficacy of debris² and smear layer¹⁹ removal was assessed using criteria proposed by Hulsmann et al.

FIG. 1.

Experimental design. CLSM, confocal laser scanning microscopy; CSI, conventional syringe irrigation; EV, EndoVac system; PIPS, photon-induced photoacoustic streaming; SEM, scanning electron microscopy.

Confocal laser scanning microscopic analysis of sealer penetration in dentinal tubules

The remaining 30 specimens from each experimental group were assigned into two subgroups (each n = 15) according to the root canal sealer used: AH Plus or TotalFill BC (Fig. 1). The fluorescent rhodamine B isothiocyanate dye (Merck Chemistry, Dannstadt, Germany) was incorporated to each sealer at an approximate concentration of 0.1 wt% to be able to visualize the specimens under confocal laser scanning microscopy (CLSM). The sealer was delivered to the canal using a #40 paper point (SureDent Corporation, Seongnam, Korea). Root canals were then filled using the cold lateral compaction technique. Excess gutta-percha was removed using a heated instrument and vertical compaction was applied with a plugger at the coronal level for 10 sec. Access cavities were sealed with Cavit G (3M; ESPE, St. Paul, MN) and specimens were stored in 100% humidity at room temperature for 1 week to allow the sealers to set completely.

The roots were embedded in self-cure acrylic repair material (Meliodent; Heraeus Kulzer, Hanau, Germany). Each root was sliced 90° to its long axis in 1-mm-thick sections using a 0.3-mm microtome saw (Metkon, Bursa, Turkey) under water cooling. The cuts were made at 2, 5, and 8 mm from the root apex. All specimens were mounted onto glass slides and examined under CLSM (Carl Zeiss LSM 510; Carl Zeiss Microscopy, Jena, Germany) at a wavelength of 575 nm and 4 × magnification with a zoom oil lens. Each section was evaluated using Zeiss LSM Image Browser Version 4.2.0.121 (Carl Zeiss MicroImaging GmbH 1997–2006) with a calibrated measuring tool. The maximum depth and total percentage of sealer penetration were measured as previously described.²⁰

Statistical analysis

All values are presented as mean ± standard deviation and mean (maximum − minimum) percentage and frequencies. Normally distributed groups with homogeneous variances were compared by Student's t-test or analysis of variance as appropriate. Regarding variables for which parametric test assumptions were not available, comparisons were made using the Mann–Whitney U test or Kruskal–Wallis test. For comparisons of means of repeated measures, repeated measures analysis of variance was used. If preconditions were not met for parametric tests, the Friedman test was used. Multiple comparisons were adjusted for using Bonferroni's correction. Categorical data were analyzed using Fisher's exact test or the chi-square test. p Values <0.05 were considered statistically significant.

Results

Evaluation of debris and smear layer removal

Intergroup comparison (irrigation activation technique comparison)

In the middle third of the root, PIPS removed debris more effectively than CSI (p < 0.01), whereas there was no significant difference between EV and PIPS (p > 0.05). In the coronal and apical thirds (Fig. 2A), there were no significant differences among CSI, EV, and PIPS regarding debris removal. There were no significant differences among the methods with respect to smear layer removal at any of the three levels (p > 0.05; Fig. 2B).

FIG. 2.

Intergroup and intragroup analysis for debris (A) and smear layer (B) removal in apical, middle, and coronal thirds. *Significance at p < 0.01.

All experimental groups except CSI had better debris and smear layer removal than the control group at all levels of the radicular dentin (p < 0.01). There were no significant differences between the CSI and control groups regarding debris removal at the middle and apical thirds (p > 0.05).

Intragroup comparison (comparison of canal thirds)

CSI, EV, and PIPS removed debris and smear layer more effectively in the coronal and middle thirds than at the apical third (p < 0.01). CSI and EV removed debris more effectively (Fig. 2A) and CSI, EV, and PIPS removed smear layer more effectively (Fig. 2B) at the coronal third compared with the middle third (p < 0.01). The PIPS group had a significantly lower mean debris score in the middle third than in the coronal third (p < 0.01; Fig. 2A). In the control group, there were no significant differences regarding debris or smear layer removal among the three levels of radicular dentin (p > 0.05).

None of the irrigation activation systems was capable of removing debris and smear layer completely. Characteristic SEM images in the coronal, middle, and apical thirds of all groups are presented in Fig. 3.

FIG. 3.

Representative scanning electron microscopic images showing coronal, middle, and apical thirds of the root canal after irrigation activation with CSI, EV, and PIPS. The most intense debris and amorphous smear layer with no open dentinal tubules were observed in the control group. Experimental groups demonstrated minimal debris with a few occluded tubules in the coronal third, fewer open dentinal tubules with more smear plugs in the middle third, and a thick smear layer and more debris with few patent tubules in the apical third.

Evaluation of sealer penetration

One specimen from the CSI + AH Plus group and one from the EV + TotalFill BC group were excluded from the study because of technical issues during preparation for confocal laser scanning microscopic analysis. Characteristic CLSM images from all groups at 2, 5, and 8 mm are shown in Fig. 4.

FIG. 4.

Representative confocal laser scanning microscopic images from each group at 2, 5, and 8 mm.

Irrigation activation technique comparison

Maximum depth of sealer penetration

Among the groups in which AH Plus was used, there was a significant difference between CSI + AH Plus and PIPS + AH Plus regarding the maximum depth of sealer penetration at 2 mm (p < 0.05; Fig. 5A). There were no other significant differences at 2, 5, or 8 mm (p > 0.05). Among the groups in which TotalFill BC was used, there were no significant differences in the maximum depth of sealer penetration at any of the levels (p > 0.05).

FIG. 5.

Comparison of different irrigation activation techniques and sealers relative to maximum depth (A) and percentage (B) of sealer penetration at 2, 5, and 8 mm. *Significance at p < 0.05. A, AH Plus; T, TotalFill BC.

Percentage of sealer penetration

For both sealer types, no significant differences in the percentage of penetration were observed among the irrigation activation techniques at any of the levels (p > 0.05; Fig. 5B).

Sealer comparison

Maximum depth of sealer penetration

TotalFill BC showed a significantly greater maximum depth of penetration than AH Plus for CSI at 2 mm (p < 0.05), however, no significant differences were observed for EV or PIPS (p > 0.05). In addition, no significant differences were observed between AH Plus and TotalFill BC at 5 and 8 mm (p > 0.05; Fig. 5A).

Percentage of sealer penetration

TotalFill BC had greater percentage of penetration than AH Plus in CSI and EV at 2 mm and in PIPS at 5 mm (p < 0.05). However, no other significant differences were observed between the sealers at 2, 5, and 8 mm (p > 0.05; Fig. 5B).

Sectional comparisons

For each experimental group, the maximum depth and highest total percentage of sealer penetration were observed at 8 mm, followed by 5 and 2 mm. Statistically significant differences among sections are shown in Table 3.

Table 3.

Comparisons of Maximum Depth and Total Percentage of Sealer Penetration by Section

		AH Plus		TotalFill BC
Group name	Sectional comparison (mm)	Max. depth, mean (±SD), (μ)	Percentage, mean (±SD)	Max. depth, mean (±SD), (μ)	Percentage, mean (±SD)
CSI	2–5	391.9 (570.7)–1212.7 (626.3)^*	29.3 (25.1)–60.6 (28.7)^*	834.1 (105.7)–1558.2 (470.4)	58 (32.6)–78.9 (19.2)
	2–8	391.9 (570.7)–1915.3 (213.7)^†	29.3 (25.1)–83.1 (18.3)^†	834.1 (105.7–2034.6 (445.4)^†	58 (32.6)–86.5 (92)^†
	5–8	1212.7 (626.3)–1915.3 (213.7)^*	60.6 (28.7)–83.1 (18.3)	1558.2 (470.4)–2034.6 (445.4)	78.9 (19.2)–86.5 (92)
EV	2–5	616.1 (472.4)–1368.5 (286.1)^†	51.9 (26.8)–81.9 (16.8)^*	932.3 (370.9)–1470.4 (491.9)^†	72.8 (23.9)–84.9 (16.2)
	2–8	616.1 (472.4)–1586.8 (385.3)^†	51.9 (26.8)–87.3 (17.2)^†	932.3 (370.9)–1947.1 (585.7)^†	72.8 (23.9)–84.9 (16.2)
	5–8	1368.5 (286.1)–1586.8 (385.3)	81.9 (16.8)–87.3 (17.2)	1470.4 (491.9)–1947.1 (585.7)	84.9 (16.2)–84.9 (16.2)
PIPS	2–5	805.2 (636.7)–1547.5 (509.1)^†	42 (33.9)–78.9 (13.6)^*	813.5 (532.9)–1458 (430.8)^†	54.6 (29.6)–89.8 (8.1)
	2–8	805.2 (636.7)–1930 (496.9)^†	42 (33.9)–91.6 (9.4)^†	813.5 (532.9)–1713.8 (361.5)^†	54.6 (29.6)–93.1 (11.3)^†
	5–8	1547.5 (509.1)–1930 (496.9)	78.9 (13.6)–91.6 (9.4)	1458 (430.8)–1713.8 (361.5)	89.8 (8.1)–93.1 (11.3)

Significant differences are indicated by asterisks (^* p < 0.05) and number sign (^† p < 0.01).

Max., maximum; SD, standard deviation.

Discussion

Smear layer removal from root canal walls during instrumentation allows endodontic irrigants and filling materials to enter the dentinal tubules. In previous studies, activation of NaOCl and EDTA using the EndoVac (EV) system and PIPS appeared to improve smear layer removal and the cleaning capability of irrigants compared with CSI.^21
–23 However, in this study, no significant differences among these groups were noted regarding smear layer removal; the most important step in removing the smear layer appeared to be introduction of EDTA into the canal.²⁴ In addition, no significant differences among experimental groups were observed in debris removal at apical and coronal levels. These findings are similar to the study by Howard et al.,²⁵ who presented no significant differences in canal and isthmus cleanliness between EV and side-ported needle irrigation. They reported that irrigation volume was a greater factor in debris and bacteria removal than the irrigation method. In this study, standardized irrigation/activation procedures (15 mL total irrigation solution volume and 90 sec total irrigation solution delivery time) were tested for all groups. The similar debridement efficacy of CSI observed may be attributed to the high volume and long action time of the irrigation solutions. On the other hand, a study by Versiani et al.²⁶ using the micro-computed tomography has shown significantly greater percentage of aggregated debris in the mesial canals (mainly in the isthmus area) of mandibular molars with CSI than EV. The methodology of this study might have some limitations, making it difficult to compare the results with the clinical environment. Therefore, further studies on molar teeth with different assessment techniques are needed.

In this study, none of the irrigation activation systems was able to remove debris and smear layer completely in the apical third of the canal, even for single-canal teeth. The placement of the PIPS endodontic fiber tip in the coronal part of the root may be too far away to activate fluid flow toward the apical part of the canal, affecting its debridement efficacy.^27,28 In a previous study,¹² PIPS was activated for 20 sec, whereas in this study, the activation time was extended to 60 sec. Despite the longer activation time used in our study, compared with CSI, significant improvement of debridement efficacy was observed only in the middle third of the canal using PIPS.

Penetration of root canal sealers into chemomechanically exposed dentinal tubules is considered advantageous with respect to filling materials' sealing and bonding capability,⁷ as well as microbial activity.¹⁴ Although recent studies demonstrated no relationship between sealer penetration and sealability²⁹ or bonding ability³⁰ of root filling materials, presence of sealer within the dentinal tubules can be considered an indicator of efficient smear layer removal.³¹ Therefore, we evaluated the maximum depth of sealer penetration along with the percentage of penetration in this study.

When subgroups with the same sealer were compared, PIPS allowed greater sealer penetration of AH Plus than CSI at 2 mm. This finding is in accord with the result of a previous study in which PIPS demonstrated greater dentinal penetration than CSI.³² Our findings for EV and CSI were partially supported by the findings of Kara Tuncer and Unal.³³ They reported that the maximum depth and percentage of sealer penetration using EV were significantly greater than those for CSI at 1 and 3 mm. In contrast, they reported that there was no difference between EV and CSI at 5 mm. Given that the percentage of sealer penetration is considered to be more clinically related than the maximum depth of sealer penetration,³¹ it is reasonable to consider that the smear layer removal efficacy and dentinal tubule penetration were consistent among all three irrigation activation methods tested in this study.

When subgroups using the same irrigation activation technique were compared, TotalFill BC use after final irrigation procedures with EV and CSI at 2 mm and with PIPS at 5 mm showed a significantly greater percentage of penetration than AH Plus use. In addition, TotalFill BC use after final irrigation procedures with CSI at 2 mm exhibited a significantly greater maximum depth of penetration than AH Plus use. To date, no studies have yet analyzed the effectiveness of TotalFill BC for dentinal tubule penetration. iRoot SP sealer (Innovative BioCeramix, Inc., Vancouver, Canada), which has a similar composition to TotalFill BC, was shown to have a greater penetration area than AH Plus.³² The greater dentinal tubule penetration area of iRoot SP was found to be related to sealer's small particle content and high viscosity.³² Further, after drying the root canals with paper points, the amount of remaining irrigation solution in the root canal system is unpredictable because canal anatomy varies.³⁴ As shown previously, residual moisture in the root canal is critical for AH Plus, which may decrease dentinal tubule penetration of the sealer.³⁴ Recently, an epoxy-amine resin-based sealer has been advocated for use with an ultrasonic activation method that allowed an enhanced sealer penetration into lateral canals.³⁵

Regarding the overall comparison of canal thirds, previous reports indicated that debris and smear layer removal^22,23 and tubular penetration of the sealers^17,32,36 were greater in the coronal and middle thirds of root canals than in the apical third. This may be caused by a decreased flow and irrigant distribution,³⁷ a decrease in the diameter and number of the dentinal tubules, and greater formation of tubular sclerosis in the apical third compared with those in the coronal and middle thirds.^18,38

The literature contains some controversy regarding the relationship between the smear layer and dentinal tubule penetration of sealers. Some studies have demonstrated no sealer penetration when a smear layer was present,^15,39 while others have shown that the presence of this layer does not completely block the dentinal tubule; a trace of sealer penetration can be observed.⁴⁰ One potential limitation of our study's methodology might be that no investigation of dentinal tubule penetration in the presence of the smear layer was performed. Therefore, further studies are needed to investigate the effect of the smear layer on sealer penetration.

Conclusions

EV, PIPS, and CSI produced almost similar debridement efficacy, smear layer removal, and dentinal tubule penetration. TotalFill BC sealer achieved superior tubular penetration compared with AH Plus.

Footnotes

Acknowledgments

This study was supported by the Baskent University Research Fund (Project No. D-KA15/04).

Author Disclosure Statement

No competing financial interests exist.

References

McComb

, Smith

. A preliminary scanning electron microscopic study of root canals after endodontic procedures. J Endod, 1975; 1:238–242.

Hulsmann

, Rümmelin

, Schäfers

. Root canal cleanliness after preparing with different endodontic hand pieces and hand instruments: a comparative SEM investigation. J Endod, 1997; 23:301–306.

Mader

, Baumgartner

, Peters

. Scanning electron microscopic investigation of the smeared layer on root canal. J Endod, 1984; 10:477–483.

Yang

, Cha

, Kim

, Kum

, Lee

, Jung

. Effect of smear layer and chlorhexidine treatment on the adhesion of Enterococcus faecalis to bovine dentin. J Endod, 2006; 32:663–667.

Depraet

, De Bruyne

, De Moor

. The sealing ability of an epoxy resin root canal sealer after Nd:YAG laser irradiation of the root canal. Int Endod J, 2005; 38:302–309.

Carson

, Goodell

, McClanahan

. Comparison of the antimicrobial activity of six irrigants on primary endodontic pathogens. J Endod, 2005; 31:471–473.

Mamootil

, Messer

. Penetration of dentinal tubules by endodontic sealer cements in extracted teeth and in vivo. Int Endod J, 2007; 40:873–881.

Munoz

, Camacho-Cuandra

. In vitro efficacy of three different endodontic irrigation systems for irrigant delivery to working length of mesial canals of mandibular molars. J Endod, 2012; 38:445–448.

Tay

, Gu

, Schoeffel

, et al. Effect of vapor lock on root canal debridement by using a side-vented needle for positive-pressure irrigant delivery. J Endod, 2010; 36:745–750.

10.

, Kim

, Ling

, Choi

, Pashley

, Tay

. Review of contemporary irrigant agitation techniques and devices. J Endod, 2009; 35:791–804.

11.

Schoeffel

. The EndoVac method of endodontic irrigation: part 2-efficacy. Dent Today, 2008; 27:82, 84, 86–87.

12.

DiVito

, Peters

, Olivi

. Effectiveness of the erbium:YAG laser and new design radial and stripped tips in removing the smear layer after root canal instrumentation. Lasers Med Sci, 2012; 27:273–280.

13.

Schilder

. Filling root canals in three dimensions. Dent Clin North Am, 1967; 11:723–744.

14.

Heling

, Chandler

. The antimicrobial effect within dentinal tubules of four canal sealers. J Endod, 1996; 22:257–259.

15.

Okşan

, Aktener

, Sen

, Tezel

. The penetration of root canal sealers into dentinal tubules. A scanning electron microscopic study. Int Endod J, 1993; 26:301–305.

16.

TotalFill BC Sealer-Instructions for use. [PDF on Internet]. La Chaux-de-Fonds: FKG Dentaire SA, Switzerland [cited 7 June 2016]. Available at: www.fkg.ch/sites/default/files/201511_Totalfill_BC_Sealer_ifu_xx.pdf (Last accessed January 14, 2017 ).

17.

Balguerie

, van der Sluis

, Vallaeys

, Gurgel-Georgelin

, Diemer

. Sealer penetration and adaptation in the dentinal tubules: a scanning electron microscopic study. J Endod, 2011; 37:1576–1579.

18.

Carrigan

, Morse

JDR

, Furst

, Sinai

. A scanning electron microscope evaluation of human dentinal tubules according to age and location. J Endod, 1984; 10:359–363.

19.

Hulsmann

, Heckendorff

, Schafers

. Comparative in vitro evaluation of three chelator pastes. Int Endod J, 2002; 35:668–679.

20.

Gharib

, Tordik

, Imamura

. A confocal laser scanning microscope investigation of the epiphany obturation system. J Endod, 2007; 33:957–961.

21.

Abarajithan

, Dham

, Velmurugan

, Valerian-Albuguergue

, Ballal

, Senthilkumar

. Comparison of Endovac irrigation system with conventional irrigation for removal of intracanal smear layer: an in vitro study. Oral Surg Oral Med Oral Pathol Oral Radiol Endod, 2011; 112:407–411.

22.

Mancini

, Cerroni

, Iorio

, Armellin

, Conte

, Cianconi

. Smear layer removal and canal cleanliness using different irrigation systems (EndoActivator, EndoVac, and passive ultrasonic irrigation): field emission scanning electron microscopic evaluation in an in vitro study. J Endod, 2013; 39:1456–1460.

23.

Akyuz Ekim

, Erdemir

. Comparison of different irrigation activation techniques on smear layer removal: an in vitro study. Microsc Res Techn, 2015; 78:230–239.

24.

Nasher

, Franzen

, Gutknecht

. The effectiveness of the Erbium:Yttrium aluminum garnet PIPS technique in comparison to different chemical solutions in removing the endodontic smear layer-an in vitro profilometric study. Lasers Med Sci, 2016; 31:1871–1882.

25.

Howard

, Kirkpatrick

, Rutledge

, Yaccino

. Comparison of debris removal with three different irrigation techniques. J Endod, 2011; 37:1301–1305.

26.

Versiani

, Alves

FRF

, Andrade-Junior

, et al. Micro-CT evaluation of the efficacy of hard-tissue removal from the root canal and isthmus area by positive and negative pressure irrigation systems. Int Endod J, 2016; 49:1079–1087.

27.

Zhu

, Yin

, Chang

JWW

, Wang

, Cheung

, Zhang

. Comparison of the antibacterial effect and smear layer removal using photon-initiated photoacoustic streaming aided irrigation versus a conventional irrigation in single-rooted canals: an in vitro study. Photomed Laser Surg, 2013; 31:371–377.

28.

Arslan

, Guneser

, Dincer

, Kustarci

, Er

, Siso

. Comparison of smear layer removal ability of QMix with different action techniques. J Endod, 2016; 42:1279–1285.

29.

De-Deus

, Brandão

, Leal

, et al. Lack of correlation between sealer penetration into dentinal tubules and sealability in nonbonded root fillings. Int Endod J, 2012; 45:642–651.

30.

Kara Tuncer

, Tuncer

, Gokyay

. Correlation between sealer penetration into dentinal tubules and bond strength of two new calcium silicate-based and an epoxy resin-based, endodontic sealer. J Adhes Sci Technol, 2014; 28:702–710.

31.

Bolles

, He

, Svoboda

, Schneiderman

, Glickman

. Comparison of Vibringe, EndoActivator, and needle irrigation on sealer penetration in extracted human teeth. J Endod, 2013; 39:708–711.

32.

Akcay

, Arslan

, Durmus

, Mese

, Capar

. Dentinal tubule penetration of AH Plus, iRoot SP, MTA Fillapex, and GuttaFlow Bioseal root canal sealers after different final irrigation procedures: a confocal microscopic study. Laser Surg Med, 2016; 48:70–76.

33.

Kara Tuncer

, Unal

. Comparison of sealer penetration using the EndoVac irrigation system and conventional needle root canal irrigation. J Endod, 2014; 40:613–617.

34.

Gibby

, Wong

, Kulild

, Williams

, Yao

, Walker

. Novel methodology to evaluate the effect of residual moisture on epoxy resin sealer/dentine interface: a pilot study. Int Endod J, 2011; 44:236–244.

35.

Arslan

, Abbas

, Karatas

. Influence of ultrasonic and sonic activation of epoxy-amine resin-based sealer on penetration of sealer into lateral canals. Clin Oral Invest, 2016; 20:2161–2164.

36.

McMichael

, Primus

, Opperman

. Dentinal tubule penetration of tricalcium silicate sealers. J Endod, 2016; 42:632–636.

37.

Teixeira

, Felippe

. The effect of application time of EDTA and NaOCl on intracanal smear layer removal: an SEM analysis. Int Endod J, 2005; 38:285–290.

38.

Paque

, Luder

, Sener

, Zehnder

. Tubular sclerosis rather than the smear layer impedes dye penetration into the dentine of endodontically instrumented root canals. Int Endod J, 2006; 39:18–25.

39.

Kokkas

, Boutsioukis

Ach

, Vassiliadis

, Stavrianos

. The influence of the smear layer on dentinal tubule penetration depth by three different root canal sealers: an in vitro study. J Endod, 2004; 30:100–102.

40.

Kuçi

, Alaçam

, Yavaş

, Ergul-Ulker

, Kayaoglu

. Sealer penetration into dentinal tubules in the presence or absence of smear layer: a confocal laser scanning microscopic study. J Endod, 2014; 40:1627–1631.