Abstract: Background : The endodontic access cavity design directly relate to the identification and negotiation of the canals and allow straight line accessibility and visibility of the canals facilitating proper preparation. By introducing dental operating microscope and the accompanying capability of inspecting the root canals – ‘both orthograde and retrograde.’ This study is to determine the difference between the location and the difference in the access cavity for the conservation of the tooth structure. Materials and methods: Thirty mandibular first molars were selected which were scheduled for the root canal treatment. The endodontic access cavity was prepared by using#4 carbide bur or #557 tapered bur. Three canals were located after the completion of the access opening. The samples were measured simultaneously with operating microscope and Vernier caliper. The inter orifice distance is measured and compared between the groups. Results: Detection rate between the difference among the groups by operating microscope and Vernier caliper showed a non-significant difference between the groups only the mesiolingual orifice showed a significant difference. The microscopic method could more accurately detect orifices statistically than the other method. Conclusion: Under the condition of this study access opening done under the microscope done shows more conservation of the tooth structure.