The development of robust high-performance liquid chromatography (HPLC) technologies continues to improve the detailed analysis and sequencing of glycan structures released from glycoproteins. Here, we present a database (GlycoBase) and analytical tool (autoGU) to assist the interpretation and assignment of HPLC-glycan profiles. GlycoBase is a relational database which contains the HPLC elution positions for over 350 2-AB labelled N-glycan structures together with predicted products of exoglycosidase digestions. AutoGU assigns provisional structures to each integrated HPLC peak and, when used in combination with exoglycosidase digestions, progressively assigns each structure automatically based on the footprint data. These tools are potentially very promising and facilitate basic research as well as the quantitative high-throughput analysis of low concentrations of glycans released from glycoproteins.