Glycosylation is one of the most commonly observed post-translational modifications (PTMs) in eukaryotes. It is believed that more than 50% eukaryotic proteins are glycosylated. To reveal the biological functions of protein-linked glycans involved in numerous biological processes, the high-throughput identification of both glycoproteins and the attached glycan structures becomes fundamentally important. Tandem mass spectrometry (MS/MS) is an effective method for glycoproteomic analysis because of its high sensitivity and selectivity. Two experimental approaches exist to obtain MS/MS spectral data of glycopeptides. One consists of isolating glycans from glycopeptides and generating MS/MS spectra of the glycans and peptides separately. The other approach produces spectra directly from intact glycopeptides. The latter approach has the advantage of retaining the glycosylation site information. However, the spectral data cannot be readily analyzed because of the lack of software specifically designed for the identification of intact glycopeptides. To address this need, we developed a novel software tool, GlycoMaster DB, to assist the automated and high-throughput identification of intact N-linked glycopeptides from MS/MS spectra. The software simultaneously searches a protein sequence database and a glycan structure database to find the best pair of peptide and glycan for each input spectrum. GlycoMaster DB can analyze mass spectral data produced with HCD/ETD mixed fragmentation, where HCD spectra are used to identify glycans and ETD spectra are used to determine peptide sequences. When only HCD spectra are available, GlycoMaster DB can still help to identify the glycans, and a list of possible peptide sequences are reported according to the accurate precursor mass and the N-linked glycopeptide sequon. GlycoMaster DB is freely accessible at http://www-novo.cs.uwaterloo.ca:8080/GlycoMasterDB .