The coupling between open channel-based microchip electrophoresis and mass spectrometry via electrostatic spray ionization is proposed for in situ detection of fractionated analytes. Electrophoretic separation is performed in an open channel fabricated in a plastic substrate. The solvent of background electrolyte is evaporated from the open channel because of Joule heating during electrophoresis, leaving the dried electrophoretic bands to be directly analyzed by mass spectrometry via scanning electrostatic spray ionization. Proof-of-concept results are obtained with fluorescent dyes and antibiotics as the test samples, demonstrating an efficient on-chip detection platform based on the electrophoresis and electrostatic spray ionization mass spectrometry.