Squamous cell carcinomas in the head and neck (HNSCC) frequently develop within precursor fields of genetically altered cells that can reach dimensions of multiple centimeters. These fields are sometimes macroscopically visible as leukoplakia and erythroplakia lesions. Using genetic analyses we showed that in approximately 25% of surgically treated HNSCC patients the fields remain behind in the surgical margins. Recently we also showed that the presence of a genetically defined field is a risk factor for local relapse. Under the microscope these fields may be recognized as dysplasia, but it is unclear whether its presence and grading in mild, moderate and severe dysplasia is associated with the risk for malignant transformation. To detect these fields in surgically treated patients, the resection margins should be genetically analyzed. This is laborious and not easily implemented in routine daily practice. We therefore performed a proteomics screen to identify protein biomarkers that could predict local relapse using a simple immunostaining procedure. The proteomes of genetically characterized normal, precursor and tumor tissues of eight patients were compared by two-dimensional difference in-gel electrophoresis (2D-DIGE) and proteins with significantly differential expression levels were identified by mass spectrometry (LC-FT-ICR-MS/MS). Forty proteins showed a highly significant differential level of expression (FDR-corrected p<0.05). Most discriminative markers suited for immunostaining were keratin 4 and cornulin. The prognostic value of these two candidate protein biomarkers was evaluated by immunohistochemical analysis of 222 surgical margins taken from the specimen of 46 HNSCC patients who developed local relapse or remained disease-free. Low expression in the surgical margins of keratin 4 (p¼0.002, RR¼3.8), cornulin (p¼0.025, RR¼2.7), and their combination (p¼0.0005, RR¼8.8) showed a highly significant association with the development of local relapse. Immunostaining for mutated p53 also showed outcome associations. Dysplasia grading had no prognostic relevance. Immunohistochemical assessment of keratin 4 and cornulin expression as well as mutant p53 in surgical margins of HNSCC patients outperforms histopathological grading in predicting the risk for local relapse. These markers can be used to initiate more frequent and lifelong surveillance of patients at high risk for local relapse, and enable selection for adjuvant treatment or prevention trials.