Background: Fistula in ano is a public perianal illness and as a long-lasting inflammatory illness which does not cure naturally. There are several surgical procedures for treatment of fistula in ano, but these surgical interventions have little degrees of success, long time of wound healing after surgery and prolonged pain, especially in complex and difficult fistulas are observed. Novel sphincter-saving methods have been used in the management of perianal fistula in order to evade the risk of fecal incontinence. Among them, the fibrin adhesive method is popular because of its ease and repeatability. Objective: The objective of the present study is to evaluate the effect of fibrin-glue injection in the treatment of anal fistula (low/high as well as primary/secondary). Method: A prospective, planned experiment was conducted on 322 patients who were established to have fistulas in ano. They were assessed by sorting them into high fistula (172/322) and low anal fistula type (150/322). The fibrin glue was instilled in their anal tracts. The character of the anal tract, whether it was simple or complex and primary or secondary, was analyzed. The outcome in terms of a postoperative discharge (failure) was noted at 3 months, 6 months, 9 months, 1 year, and 2 years. Results: Total 322 patients were involved in this study. The general success rate was 275/322 (85.4%) after a mean follow-up of 1.5 years. All patients with a complex fistula (for low/high fistula, primary/secondary) had failure of healing (success rate 0%) either through first or second injection of fibrin-glue while all patients with simple fistula had successive rate of healing (for low/high fistula, primary/secondary) either through first or need the second injection of fibrin-glue. None of the patients had postoperative continence problems, and no other complications were noted. Conclusion: Fibrin glue is a novel attractive approach, easy, safe, minimally invasive, repeatable and cost effective for treatment of anal fistula simple (low/high and primary/secondary) and promising option for treatment of high fistula, and do not have a role for healing the complex anal fistula.