Background: The objective of this study is to examine the differential expression of EphB4, EphrinB2 and EGFR genes in papillary thyroid carcinoma (PTC), and evaluate their association with lymph node metastasis. Methods: EphB4, EphrinB2 and EGFR expression in 21 matched tumor and surrounding normal thyroid tissue were evaluated by complementary DNA (cDNA) microarray, Western blot and immunohistochemistry (IHC). Results: We noted a statistically significant overexpression of EphB4, EphrinB2 and EGFR in tumor versus normal tissue based on cDNA microarray, Western blot and IHC analysis. EphB4 and EphrinB2 overexpression were significantly associated with presence of lymph node disease. Conclusion: Overexpression of EphB4, EphrinB2 and EGFR are associated with PTC, while EphB4 and EphrinB2 overexpression are associated with lymph node metastases. These genes may be potential biomarkers for identification of sub-clinical lymph node involvement in PTC and potential small-molecule targets for pharmacotherapy research. Head Neck, 2014.