Gel-based liquid chromatography-tandem mass spectrometry (GeLC-MS/MS), a couple of sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) and LC-MS/MS has emerged as a promising tool for profiling protein expression in saliva and tissue of canine oral tumors, not yet in serum samples. This study aimed to identify serum biomarkers for canine oral tumors, specifically oral melanoma (OM) and oral squamous cell carcinoma (OSCC), compared with non-oral cancer conditions (benign tumors (BN) and controls (a healthy and periodontitis group (CTRL)) using GeLC-MS/MS.