Clinical depression is one of the most common and debilitating psychiatric disorders and contributes to increased risks of disability and suicide. Differentially expressed serum proteins may serve as biomarkers for diagnosing depression. In this study, samples from depressed patients are aggregated into a pool (22×100μL serum was used) and samples from healthy volunteers are aggregated into the other pool (20×100μL serum was used). Isobaric tag for relative and absolute quantitation (iTRAQ) technology and tandem mass spectrometry were employed to screen for differentially expressed serum protein in two separate pools. We identified 472 proteins in the serum samples, and 154 of these presented differences in abundance between the depression and control groups. Ingenuity pathway analysis (IPA) was employed to identify the highest scoring proteins in signaling pathway networks. Finally, four differentially expressed proteins were validated by enzyme-linked immuno sorbent assay (ELISA). Proteomic studies revealed that levels of c-reaction protein (CRP), inter-alpha-trypsin inhibitor heavy chain H4 (ITIH4), serum amyloid A1 (SAA1) and angiopoietin-like 3 (ANGPTL3) were substantially increased in depressed patients compared with the healthy control group. Therefore, these differentially expressed proteins may represent potential markers for the clinical diagnosis of depression.