ABSTRACT Aim To evaluate and compare various nuclear morphometric parameters in different histopathological grades of oral squamous cell carcinoma (OSCC) by using computerized image analysis and also to correlate it with regional lymph node metastasis. Materials and methods This retrospective study was conducted on paraffin blocks of 40 tissue specimens of OSCC cases treated with neck dissection, which were retrieved from the archives of the Department of Oral Pathology and Microbiology, Mahatma Gandhi Mission’s Dental College and Hospital, Navi Mumbai, India. All cases were histopathologically graded as well, moderately, and poorly differentiated OSCC. Further, they were also categorized based on pathological lymph node status as with or without lymph node metastasis. Sections from tumor proper were subjected to Feulgen nuclear staining technique. Images of 10 microscopic fields at the deepest invading part of tumor were captured randomly and 100 nuclei of tumor cells with clear, complete, nonoverlapping outlines were selected in each case. Nuclear morphometric parameters, such as large diameter, small diameter, nuclear area, and nuclear perimeter were measured for each of the 100 cells. Results An increase in mean nuclear area coefficient of variation (NACV) was observed in OSCC cases with lymph node metastasis pN(+) than in those without lymph node metastasis pN(–), (p = 0.67). A significant increase in nuclear area and perimeter was observed in pN(+) cases (p < 0.01). A significant decrease in circular rate and increase in largest to smallest nuclear diameter (L/S) ratio (p < 0.01) was observed in pN(+) cases. On comparing the nuclear morphometric parameters with different histopathological grades of OSCC, we found an increase in mean NACV values from well-differentiated OSCC to moderately differentiated and poorly differentiated OSCC (p = 0.612). An increase in mean nuclear area and perimeter was noted as grades of OSCC advanced (p > 0.01). The mean circular rate was found to be lowest in poorly differentiated OSCC (p = 0.362). A significant increase in mean L/S ratio was observed within different histopathological grades of OSCC (p = 0.044), which when further confirmed using least significant difference (LSD) post hoc test, indicated a difference only between well-differentiated and poorly differentiated cases of OSCC (p = 0.132). Conclusion Our observations reveal that tumor cells with greater nuclear dimension and more elliptical shape tend to show increased incidence of nodal metastasis. Also, a positive inclination was observed in nuclear size and shape with increasing histopathological grades of OSCC. However, our data warrant a large-scale study to establish nuclear morphometry as a quantitative objective parameter and also for the rational application of the same. How to cite this article Narayanan N, Pathak J, Patel S, Swain N. Evaluation of Nuclear Morphometry in Oral Squamous Cell Carcinoma: A Retrospective Study. J Contemp Dent 2017;7(2):107-113.