Table 1.

Description of the experimental groups with applied laser parameters and final irrigation procedure.

Group	Description	Irrigation procedure	Final methylene blue application
POS	Positive control	Rinsing with ultrapure water during instrumentation (10 mL), drying the canal with paper points (Large, WaveOne Gold, Dentsply Sirona, Baden, Switzerland), etching the canal walls with 35% H3PO4 (Ultra-Etch, Ultradent, South Jordan, UT, USA) for 60 s under constant movement with a paper-point and verifying that excess etching gel extruded the apex. Rinsing with ultra-pure water (5 mL)	100 s
NEG	Negative control (smear layer)	2 mL ultra-pure water between each file and a final rinse with 5 mL ultra-pure water	100 s
CONV	Conventional syringe irrigation	2 mL NaOCl (3%, Hänseler Swiss Pharma, Herisau, Switzerland) between each file followed by 5 mL EDTA (17%, Pharma24 SA, Geneva, Switzerland) and 5 mL ultra-pure water	100 s
Er:YAG	ER:YAG laser ((AS7066(X), 1.3×14 mm; 50 mJ, 15 Hz	2 mL ultra-pure water between each file	5× 20 s of laser activation
CO2	9.3 µm CO2 laser (Ultra-guide hand piece with endo-tip Ø 1.25 mm; 40%)	2 mL ultra-pure water between each file	5× 20 s of laser activation
D455	SIROLASE 455 nm (black coated tip Ø 0.4 mm; 0.8 W, 40% duty cycle, 15 Hz)	2 mL ultra-pure water between each file	5× 20 s of laser activation
D970	SIROLASE 970 nm (black coated tip Ø 0.4 mm; 0.8 W, 40% duty cycle, 15 Hz)	2 mL ultra-pure water between each file	5× 20 s of laser activation
D808	WISER 808 nm (black coated tip Ø 0.2 mm; 1 W, 26,666 µs on, 40,000 off)	2 mL ultra-pure water between each file	5× 20 s of laser activation
D980	WISER 980 nm (black coated tip Ø 0.2 mm; 1 W, 26,666 µs on, 40,000 off)	2 mL ultra-pure water between each file	5× 20 s of laser activation

EDTA: ethylenediaminetetraacetic acid; H3PO4: Phosphoric acid; NaOCl: Sodium hypochlorite