Aim: The purpose of this study was to evaluate in-vitro the efficacy of prepared dressing gel materials from conventional used root canal irrigate at a special concentration namely in a standard volume (0.1 ml) was applied. ECHXT: 0.01% EDTA, 0.01% CHX and 6% Triss buffer, ENaOCl (used separate dressing gel 2.5% NaOCl / 18.6% EDTA gel) and ECHX (used separate dressing gel 18.6% EDTA and 17% CHX gel). Materials and methods: The dressing gel materials were prepared and rheological determination was done. Neutralizers were prepared for each to be used in the biofilm group. One hundred and five distal roots of extracted lower molars were standardized to 12± 1 mm length and up to #20 K- file apex’s width. A cervical seat was created of 1x1 mm dimensions at the coronal third of the root. The root surfaces were coated with varnish and root apices were sealed with cyanoacrylate. Each root was placed separately inside Eppendorf tube. Initially prepared roots were divided into two equal groups (N=45) according to the use of dressing gel for removal of smear layer or disruption and/ or removal of biofilm. In the smear layer group after mechanical preparation, randomly selected samples were used to confirm the formation of smear layer by ESEM. Dressing of the canals were done and the roots were incubated for 1, 24 hour and 1-week. Debris and smear layer removal evaluation were done using the stereomicroscope and ESEM, respectively. For biofilm groups; the canals were contaminated with 1 McFarland suspensions of E. faecalis and incubated for 14 days at 37 ºC and the biofilm formation was confirmed by SEM. For biofilm group all the steps were done under aseptic conditions inside ClⅡlaminar flow cabinet with HEPA filter. Biofilm disruption evaluation was done through the presence of viable microorganisms using the fluorescent microscope. Results: The statistically significant lowest mean smear layer percentage at the apical third showed after 1 hour in ECHX group while it was after 1 and 24 hour in ENaOCl group. The bacterial viability results showed no statistically significant difference between the three used dressings after recommended time interval in fluid samples. While in dentin shavings samples; ENaOCl and ECHX groups showed the statistically significant lowest mean viability after 24 hour. The comparison between the fluid and dentin chip shavings revealed the statistically significant highest mean viability percentage with the fluid samples. Conclusion: Under the limitations of the present study, we conclude that: Variation in the physical nature of irrigate did improve its role and one week could not be recommended. The combined use of conventional irrigate with chelating agent as intracanal medicament may be promising in removal of smear layer and biofilm disruption and their effect could be improved by increase concentration and minimizing the time of application.