Capillary electrophoresis (CE) played an important role in developments in the life sciences. The technique is nowadays used for the analysis of both large and small molecules in applications where it performs better than or is complementary to liquid chromatographic techniques. In this review, principles of different electromigration techniques, especially capillary isoelectric focusing (CIEF), capillary gel (CGE) and capillary zone electrophoresis (CZE), are described and recent developments in instrumentation, with an emphasis on mass spectrometry (MS) coupling and microchip CE, are discussed. The role of CE in the life sciences is shown with applications in which it had a high impact over the past few decades. In this context, current practice for the characterization of biopharmaceuticals (therapeutic proteins) is shown with CIEF, CGE and CZE using different detection techniques, including MS. Subsequently, the application of CGE and CZE, in combination with laser induced fluorescence detection and CZE-MS are demonstrated for the analysis of protein-released glycans in the characterization of biopharmaceuticals and glycan biomarker discovery in biological samples. Special attention is paid to developments in capillary coatings and derivatization strategies for glycans. Finally, routine CE analysis in clinical chemistry and latest developments in metabolomics approaches for the profiling of small molecules in biological samples are discussed. The large number of CE applications published for these topics in recent years clearly demonstrates the established role of CE in life sciences.