Isomet 2000 precision saw

Prior to specimen sectioning, 10 bonded specimens in each adhesive group (CEA, HAA-5%, and HAA-10%) were thermo cycled (TC) in water baths at 5 °C and 55 °C for 30 s each and 5 s dwelling time (THE-1100, SD Mechatronik GmbH, Germany). However, the other 10 specimens remained non-thermocycled (NTC) in each adhesive group and were stored in distilled water for 1 week. The sectioning of all bonded specimens in the study was performed with a slow speed diamond saw (Buehler Isomet 2000 Precision saw, Lake Bluff, IL, USA) to form 1 mm × 1 mm, composite–dentin bonded beams. Six beams from each tooth were used for μTBS assessment (sixty) in each adhesive subgroup (CEA-TC, CEA-NTC, HAA-5%-TC; HAA-5%-NTC, HAA-10%-TC, HAA-10%-NTC). Bonded beams were secured to the micro-tensile tester (Bisco Inc., Richmond, VA, USA) jaws with cyanoacrylate (Superglue, Minneapolis, MN, USA) and were assessed under tension at a crosshead speed of 0.5 mm/minute until failure. Failure modes were evaluated among the groups, classified as adhesive, cohesive, and mixed types using a digital microscope (Hirox KH 7700, Tokyo, Japan). Means and standard deviations of μTBS among the groups were analyzed using ANOVA and a post hoc multiple comparisons test.

Thirty-six extracted mandibular molars were collected, sterilized in 10% buffered formalin. All teeth were decoronated, and the lengths of roots were standardized to 16 mm. The roots were split at the furcation area by using ISOMET 2000 PRECISION SAW (Buehler, USA). Straight and angulated conventional radiographs were taken for all mesial roots to verify the inclusion criteria. The inclusion criteria were as follows: mesial roots with two separate canals (Vertucci Type IV), free from calcifications and pulp stones, and a root curvature between 10° and 30° as verified by measurement of Schneider [22 (link)]. Finally, nineteen mesial roots with thirty-eight root canals were selected and included in this study (N = 38). Each root was mounted in clear acrylic block with a mark on the buccal side. This mark will assist the positioning of each sample in the same orientation for pre- and postinstrumentation micro-CT scan.

Eighty teeth (twenty from each group) were used for µTBS analysis. From the twenty bonded teeth, ten teeth from each group were aged using thermocycling at 5 and 55 °C in distilled water baths (THE-1100, SD Mechatronik GmbH, Feldkirchen-Westerham, Germany). A total of 10,000 cycles for 30 s with a dwell time of 5 s were used for aging. The other ten bonded teeth remained non-aged and were kept safe in distilled water for one day, pre-sectioning. The sectioning of specimens belonging to each adhesive group was carried out to form beams of 1 mm × 1 mm of composite resin-adhesive with the help of a water-cooled diamond saw (Buehler Isomet 2000 Precision saw, IL, USA). In every tooth, six beams were formed (sixty beams in total), and for each group, five beams were analyzed for μTBS analysis.

The remaining five bonded teeth from every group were partitioned using a slow speed saw (Buehler Isomet 2000 Precision saw, Lake Bluff, IL, USA) in order to create beams of 1 mm × 1 mm. The bonded adhesive–resin interface was observed using SEM-EDX spectroscopy. Employing a polisher (Beuhler Polisher, Lake Bluff, IL, USA), beams were wet polished, and placed in an ultrasonic bath containing distilled water (Bandelin Digital-Sigma-Aldrich Darmstadt, Germany) for 5 min. For the samples’ conditioning, 36% phosphoric acid (DeTrey conditioner, Dentsply, PA, USA) was applied. This was followed by washing and immersion in 5.25% sodium hypochlorite solution for 15 min. The samples were cleaned with distilled water, and then their dehydration was carried out with 80–100% concentrations of ethanol solution. The specimens were sputter coated with gold (as mentioned previously). An SEM (JEOL, JSM-6513, SEM, Tokyo, Japan) was used to observe samples’ adhesive–dentin interfaces with 30 kV voltage using different magnifications. EDX spectroscopy was also implemented to analyze the elemental distribution and presence of Si nanoparticles in the adhesives.

For microtensile bond testing (μTBS), sixty samples were used (twenty from each group). Out of 20 samples in each group, 10 bonded specimens in all adhesive groups were thermocycled (TC) in distilled water baths at 5 °C and 55 °C for 30 s with 5 s dwelling time (THE-1100, SD Mechatronik GmbH, Germany) and 10,000 cycles were used. The remaining 10 samples remained non-thermocycled (NTC) and were kept in distilled water for 24 h prior to sectioning. The bonded samples in each adhesive group were partitioned in order to construct 1 × 1 mm beams of composite–adhesive-dentin using a diamond slow speed saw (Buehler Isomet 2000 Precision saw, Rapid, IL, USA). Every group involved 20 teeth and each tooth formed six beams. In every adhesive group, at least five bonded beams were assessed for μTBS analysis. The beams were attached to the jaws of micro tensile tester (Bisco Inc., VA, USA) with cyanoacrylate (Zapit, Dental ventures Inc., Corona, CA, USA) and loaded in tension at 0.5 mm/min crosshead speed up to fracture. The failure modes were assessed in each group, and were classified as adhesive, cohesive, and mixed types by means of a digital microscope (Hirox KH 7700, Tokyo, Japan). The observed μTBS values in every group were matched using ANOVA and multiple comparisons test.

Pre-sample sectioning, ten samples from each group were thermo-cycled (TC) with 10,000 cycles while the remaining ten remained non-TC (NTC, stored in DW for 1 week). The TC was carried out inside a water bath (THE-1100, SD Mechatronik GmbH, Berlin, Germany) containing separate water chambers at 5 °C and 55 °C. The samples were immersed for 30 s, whereas the dwell time was 5 s. A slow-speed diamond saw (Buehler Isomet 2000 Precision saw, Lake Bluff, IL, USA) was utilized to section the bonded samples to form composite-dentin bonded beams (1 mm × 1 mm). Individually, every tooth produced seven beams (seventy in each adhesive group) for μTBS evaluation (β-TCPA-5%-TC, β-TCPA-5%-NTC, EA-TC, and EA-NTC). These beams were fixed in the micro-tensile tester (Bisco Inc., Richmond, VA, USA) jaws using cyanoacrylate (Superglue, MN, USA). The assessment was carried out in tension at a crosshead speed of 0.5 mm/minute until failure. The nature of the failure modes in our study were also appraised and regarded as adhesive, cohesive, and mixed kinds employing a digital microscope (Hirox KH 7700, Tokyo, Japan).