2-Aminobenzoic acid (2-AA) is widely used as a labeling reagent to derivatize released N-glycans at their free reducing terminus by reductive amination. 2-AA-labeled glycans have increased mass spectrometric sensitivity for their identification and enable fluorescence-chromatography-based glycan quantification. Drawbacks are that the labeling process is labor intensive and time consuming. Clean up of labeled glycans via removal of excess of labeling reagents often leads to sample losses. Here, we report use of 2-AA for labeling N-glycans on a MALDI target through nonreductive amination, while simultaneously functioning as a matrix in MALDI-MS glycan analysis. Coupling 2-AA to glycans results in significant increases of glycan anionic signals as compared to that using the traditional 2,5-dihydroxybenzoic acid (2,5-DHB) matrix. The on-MALDI-target sample preparation is a single-step protocol with high derivatization efficiency. It is also noticed that 2-AA-labeled glycan generated dominant deprotonated molecular anions with much fewer and low-intensity sodium adducts and therefore greatly simplified glycan profiles. We further explored its application in the N-glycan profile of a biotherapeutic monoclonal antibody and was able to achieve sensitive glycan identification at a low microgram level of glycoprotein. This 2-AA on-MALDI-target glycan derivatization eliminates tedious sample preparation and avoids sample loss. It is generally applicable for other applications (e.g., glycomics), where limited amounts of glycoproteins are available for analysis.