Related Experiment Video
Updated: May 5, 2026

Visualizing the Effects of Sputum on Biofilm Development Using a Chambered Coverglass Model
Published on: December 14, 2016
Effects of Sodium Hypochlorite Concentration and Application Time on Bacteria in an Ex Vivo Polymicrobial Biofilm
Hazuki Maezono1, Kittipit Klanliang2, Tsuyoshi Shimaoka1
1Department of Restorative Dentistry and Endodontology, Osaka University Graduate School of Dentistry, Suita, Osaka, Japan.
Introduction:
In endodontic treatment, it is important to remove or inactivate biofilms in the root canal system. We investigated the effects of different concentrations and application times of sodium hypochlorite (NaOCl) on the viability of bacteria in ex vivo polymicrobial biofilms of different maturation levels.
Methods:
Polymicrobial biofilms were prepared from dental plaque samples and grown for 1, 2, and 3 weeks under anaerobic conditions on collagen-coated hydroxyapatite discs as an ex vivo biofilm model. The biofilms were then exposed to NaOCl at concentrations ranging from 0.1% to 2% for 1 or 3 minutes. The control group was exposed to sterile distilled water. Viability staining was performed and examined by confocal laser scanning microscopy to determine the percentage of biofilm bacteria killed by NaOCl. Scanning electron microscopy was also performed to visually examine the biofilms.
Results:
Application of NaOCl at 0.5%-2% for both 1 and 3 min killed significantly more bacteria when compared to the controls (P < .05). Cell viability tended to be lower after the application of NaOCl for 3 minutes than that for 1 minute.
Conclusions:
Our experiments using an ex vivo model showed that within the range of 0.1%-2% of NaOCl, higher NaOCl concentrations and longer application times were more effective in killing biofilm bacteria, and that mature biofilms were more resistant to NaOCl than younger biofilms.

