Objective   To explore the bactericidal effect of silver nanoparticle on P. aeruginosa in an in vitro model of biofilms.    Methods   The ethylene-vinyl acetate (EVA) plastics with 0.01% or 0.05% silver nanoparticle in it were used as carriers for the bacteria adhesion

on which bacterial biofilm models in vitro were built by using shaking method to mimic the micro-environment in vivo. The samples not containing silver nanoparticle which had been cultured for 3 d were scanned by scanning electron microscopy (SEM) and were observed by fluorescence microscope after stained with FITC-ConA to observe biofilm formation. The model specimens which had been cultured for 0.5

1

2

3 or 5 d with shaking method were stained by the fluorescent antibody SYTO9/PI

and then detected under confocal laser scanning microscopy (CLSM) to take the images at different levels. The image were were then analyzed by the software of Image-Pro Plus 6.0. The bacterial mortalities under the different conditions (concentration of silver nanoparticles and culture times) were calculated.    Results   SEM and fluorescence microscope displayed the linear flow of bacterial biofilm formation in the 3-day-cultured EVA plastic carriers

indicating the establishment of in vitro model of biofilm. Mortality of bacteria was obviously affected by the different concentrations of silver ion

duration of action (F=84.62

85.67

P0.0001). The mortality of bacterial of silver nanoparticle was significantly higher than control group at different time points (P0.05). However

there was no significant difference in the mortality between the concentration of 0.05% and 0.1% silver nanoparticle. The mortality of bacteria in the specimens cultured for 1

2 and 3 d was significantly higher than those cultured for 0.5 and 5 d. The mortality rate of bacteria on 0.1% silver nanoparticle EVA plastic cultured for 2 d was the highest among all groups [(88.53±1.88)％].    Conclusion   In vitro model of bacterial biofilm is successfully established by using the shaking method. The bacteria in biofilm can be killed by silver nanoparticle.