The effect of the mouthwash on the viability of human gingival fibroblasts (HGFs) was evaluated by a CCK-8 cell counting kit (CCK-8; Biosharp, China). HGFs were seeded into 96-well plates with high-glucose DMEM medium (BasalMedia Technologies Co., Shanghai, China), and transferred to a CO2 incubator for 24 h at 37 °C. Cells were then stimulated with the different concentrations of test mouthwash for 30 s, and a mouthwash containing 0.12% chlorhexidine and 0.02% metronidazoleand as the positive control, only medium as the negative control. Afterwards, the cells were incubated in a medium containing 10 µL test reagents for 1 h at 37 ℃. The optimal absorbance at 450 nm was determined using a microplate reader.
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