Background:The microbiological ecosystem is considered the main obstacle in endodontics. Therefore, canal disinfection became a great priority. Aim of the study:This study was established to assess cysteamine's antibacterial effect against E. Faecalis and compare the improvement effect to its combination with various intracanal medicaments. Methods and Materials:Cleaning and shaping were performed on human single-rooted teeth then inoculated by E. faecalis biofilm. Samples were divided randomly into three groups: Group I to verify the maturation of E. faecalis biofilm inside the root canal using SEM. Group II were divided into four subgroups according to the intracanal medication used. Subgroup A was treated with Cysteamine. Subgroup B was treated with Chlorhexidine Cysteamine combination. Subgroup C was treated with Calcium hydroxide (CaOH) Cysteamine combination. Subgroup D was treated with Triple antibiotic paste Cysteamine combination. Group III were divided into two subgroups treated with plain gel: positive control and negative control. Samples collected from root canals were used for viable count (CFU/ml). Log transformation of the data was carried out. The statistical significance level was set at P < 0.05. Results:There was a significant difference between different groups (p<0.001). Post hoc pairwise comparisons showed statistically significant difference between subgroups except for the CaOH Cysteamine combination subgroup and TAP Cysteamine combination subgroup. Conclusion:Cysteamine usage provides a synergistic antibacterial effect with other intracanal medications. CHX Cysteamine combination was the most effective antibacterial drug that provided effectiveness against E. faecalis.