Aim: The aim of the study was to evaluate the prevalence and morphology of the middle mesial canal and middle distal canal in mandibular first molars in North Indian subpopulation using Cone Beam Computed Tomography. Methodology: 60 Cone Beam Computed Tomography (CBCT) scans were retrospectively analyzed out of which 40 mandibular first molars were included in this study as per inclusion and exclusion criteria. The collected CBCT scans were observed for the presence of middle mesial and middle distal canal in mandibular first molars. The root canal configuration of the middle mesial and middle distal canals in the mandibular first molars was then observed in axial and coronal sections of CBCT scans. Results: Out of total 40 examined mandibular first molars, the prevalence of middle mesial canal in mandibular first molar was found out to be 5% and the prevalence of middle distal canal in mandibular first molar was found out to be 10%. The confluent configuration was the most common root canal configuration type of both middle mesial and middle distal canal. Conclusion: Locating middle mesial and middle distal canal is imperative in order to achieve good endodontic prognosis for mandibular first molars. CBCT analysis remains as a non-invasive and clinically effective tool for determining root canal morphology. Knowing the incidence of variations in root canal anatomy is crucial for alerting doctors to the possibility of encountering them in clinical practice.