Gastric cancer (GC) is one of the most commonly occurring cancers and has the highest overall mortality rate worldwide. Unfortunately, the vast majority of GC patients present with advanced stage disease and the overall prognosis remains very poor. In GC diagnosis, the non-invasive tests are limited due to their lack of sensitivity and specificity. Glycosylation changes have been reported in a wide variety of human diseases, including immune disorders and cancers, and even is associated with malignant transformation. Recently, a targeted glycoproteomic approach has gained considerable attention as a novel method for biomarker discovery to improve the specificity and sensitivity for clinical use. Here, we comprehensively investigated the indirect and direct glycomic profile of a target glycoprotein, serum haptoglobin (Hp) by chip-based nano LC-QTOF MS and MS/MS analysis following antibody-assisted purification. From the results of significant N-glycan variations between GC patients and healthy controls, we conclusively suggest that aberrant glycans of serum Hp are associated with patients with gastric cancer and might be a promising marker for GC screening.