Purpose: Cervical cancer is the second most prevalent malignancy of women. Our aim was toidentify additional marker protein patterns for objective diagnosis of squamous cervical cancer(SCC).Experimental design: Collected tissue biopsies of SCC, squamous vaginal cancer (SVC), normalcervical and vaginal mucosa were subjected to 2-DE, SameSpot analysis, MALDI-TOF-MSprotein identification, and analysis of the expression of selected proteins by immunohistochemistry.Results: In 148 protein spots selected by the difference in expression 99 proteins were identified.A differential protein pattern for SCC was, e.g. over-expressed (OE) eukaryotic translationinitiation factor 3-2b, neutrophil cytosolic factor 2, annexin A6 (ANXA6), for SVC it was OEcathepsin D, g-catenin, RAB2A, for both cancers it was OE apolipoprotein E, tropomyosin 3,HSPA8, and underexpressed cytokeratin 13, osteoglycin. In SCC nuclear expression ofneutrophil cytosolic factor 2, PRDX2, HSP27 (nine of ten cases), ANXA6 (nine of ten cases) wasobserved while tropomyosin 4 was expressed only in two of ten cases. There was 81.1% (43/53)agreement between the expression of protein spots and the immune expression of proteins(www.proteinatlas.org).Conclusions and clinical relevance: SCC is characterized by specific tissue marker proteinpatterns that allow objective detection of the disease. They can become a basis for objectiveautomated cytology-based screening and improve current diagnostics of SCC.