In the present study, the use of a robust and sensitive mass spectrometry based protein profiling analysis was tested as diagnostic tools for women with an ovarian tumor. The potential additional diagnostic value of serum protein profiles independent of the information provided by CA125 were also investigated. Protein profiles of 113 serum samples from women with an ovarian tumor (54 malign and 59 benign) were generated using MALDI-TOF MS. A total of 98 peaks with a significant difference (p< 0.01) in intensity between women with benign tumors/cysts and malignant ovarian tumors were identified. After average linkage clustering, a profile of 46 statistical significant mass peaks was identified to distinguish malignant tumors and benign tumors/cysts. In the subgroup of women with normal CA125 values (< or =35 U/mL) (62 patients) 36 of the 504 mass peaks showed significant (p< 0.05) differences in intensity between benign and malignant disease. After average linkage clustering, 25 statistical significant mass values were identified in this clinical difficult and important subgroup presenting with normal CA125 values. The current study demonstrates the potential of mass spectrometry based serum protein profiling as a diagnostic tool in discrimination between benign ovarian tumors/cysts and malignant ovarian tumors. Additionally, the method provided diagnostic information independent of CA125.