A commercial prebiotic galacto-oligosaccharide mixture (Vivinal GOS) was extensively characterized using a combination of analytical techniques. The different techniques were integrated to give complementary information on specific characteristics of the oligosaccharide mixture, ranging from global information on degree of polymerization (DP) to the identity and concentration of individual oligosaccharides. The coupling of high-performance anion-exchange chromatography (HPAEC) to mass spectrometry (MS) was determined to be especially suitable to assign the DP of individual oligosaccharides on the basis of their m/z values as well as their quantification using external standards. The combination of NMR spectroscopy and methylation analysis after isolation using size exclusion chromatography (SEC) and hydrophilic interaction liquid chromatography (HILIC) was used for identification. All DP2 compounds could be identified and quantified in this way as well as the main DP3 compounds.