Background: Common and persistent isolate ina the teeth following failed therapy of the root canal is the gram-positive facultative bacterium Enterococcus faecalis and Escherichia coli, which develop biofilm through a complicated process that results in the formation of a biofilm. Enterococcus faecalis and Escherichia coli are significant factors that cause chronic periradicular lesions after root canal therapy. Aim: This study aimed to treat the root canal tooth infected with Escherichia coli and Enterococcus faecalis Methods: In this study biofilm formation was done for Escherichia coli in growth phase cultured in a brain heart broth Enterococcus faecalis and Escherichia coli cultured in Luria-Bertani (LB) infusion medium for 18 hrs. Then we studied the effect of plasma jet that works with argon gas, and it is generated by a power supply that operates at alternating high voltages in the form of a sinusoidal wave with peak-to-peak value of about 12 kV at a frequency of 30 KHz and its power is about 200 watts. Results: Enterococcus faecalis biofilms were treated for 0.5, 1, 1.5, 2, 2.5 minute with a constant gas flow rate 2.5 . The killing rate decreased from (0.979 to 0.361). In the Enterococcus faecalis biofilms was treated for a longer period of time (5, 10, 15, 20, 25, 30) min, as the killing rates decreased from (0.739 to 0.179). Conclusions: The result of this study indicates that plasma jet is useful in disinfecting root canal tooth and reducing the formation of biofilm.