Focal Adhesion Kinase (FAK) and Src have been reported to regulate tumor growth, invasion, metastasis and angiogenesis. The present study aimed to evaluate by immunohistochemistry the clinical significance of FAK and Src expression in 108 patients with benign and malignant thyroid lesions. Total FAK expression provided a distinct discrimination between malignant and benign (p = 0.00001), as well as between papillary carcinoma and hyperplastic nodules thyroid lesions (p = 0.00005), being also associated with follicular cells' proliferative capacity (p = 0.0003). In malignant thyroid lesions, total FAK expression was associated with tumor size (p = 0.0455), and presence of capsular (p = 0.0102) and lymphatic (p = 0.0173) invasion. Total Src expression was borderline increased in cases of papillary carcinoma compared to hyperplastic nodules (p = 0.0993), being also correlated with tumor size (p = 0.0169). FAK and Src expression was ascribed to a significant extent to the phosphorylated forms of the enzymes, which provided a better discrimination between malignant and benign thyroid lesions. The current data revealed that FAK and to a lesser extent Src expression could be considered of clinical utility in thyroid neoplasia with potential use as therapeutic targets.