A variety of prefractionation methods (including a novel reversed-phase solid-phase-extraction (RP-SPE) combined with SDS-PAGE and proteomic based approaches (e.g., 2-dimensional gel electrophoresis (2DE) and MALDI-TOF mass spectrometry combined with Artificial Neural Network (ANN) bioinformatic tools) were used to investigate the protein/peptide signatures in patients with Polycystic Ovary Syndrome (PCOS). Four potential PCOS biomarkers were identified (complement C4alpha3c and C4gamma and haptoglobin alpha and beta chains).