Oral squamous cell carcinoma (OSCC), a prevalent malignancy in the head and neck region, presents a significant global health burden. Recent studies have highlighted the role of hematological markers and immunohistochemical (IHC) markers, such as P40 and Cytokeratin 5/6 (CK5/6), in diagnosing and prognosticating OSCC. This case-control study aimed to assess hematological alterations and the diagnostic value of P40 and CK5/6 in OSCC patients compared to healthy controls. A total of 100 participants were recruited, including 50 OSCC patients and 50 matched controls. Hematological parameters, including hemoglobin, white blood cell count, and platelet indices, were analyzed alongside IHC staining for P40 and CK5/6 in OSCC tissues. Results demonstrated statistically significant alterations in hematological parameters in OSCC patients (p < 0.05), coupled with high sensitivity and specificity of P40 and CK5/6 markers in identifying OSCC. The study underscores the diagnostic potential of combining hematological analysis with IHC markers in OSCC. These findings provide a novel perspective on early detection and clinical management of OSCC, emphasizing the utility of P40 and CK5/6 as reliable biomarkers. Future research should explore the prognostic implications of these findings in larger cohorts.