Early diagnosis and effective therapy are the fundamental challenge for modern oncology. Hence, many researchers focus on the search for new or improved biomarkers. Due to the great importance of nuclear factor kappa B (NF-κB) in physiological and pathological processes, we focused on assessing its usefulness as a biomarker in OPSCC. The purpose of the research presented here was to evaluate the prevalence and the level of NF-κB in the serum of OPSCC patients (ELISA). Serum NF-κB levels were also assessed depending on the degree of histological differentiation of the tumor and TN classification. Additionally, we considered the existence of a correlation between the concentration of NF-κB and EBV antibody titers, viral load and selected MMPs-MMP3 and MMP9. Taken together, the obtained results demonstrated that NF-κB level was significantly higher among patients with EBV-related OPSCC than among those without EBV. In addition, the level of NF-κB was significantly higher in more advanced clinical stages. Moreover, a positive correlation was found between the concentration of NF-κB and the level of selected EBV antibodies, viral load and both tested MMPs. The diagnostic accuracy of NF-κB was confirmed by ROC analysis.