Abstract: Coupling mass spectrometry ( MS ) with microdialysis ( MD ) has an unsubstitutable role for the on-line detection of biological sample in living animal, because MD is the best in-vivo sampling technology and MS has characteristics of minimum sample, high sensitivity and time-saving. In this review the basic principle and experimental method of microdialysis is introduced with thirty-five references, and some application progress in coupling MS with MD for the sample desalting, on-line detection of active biological component and pharmaceutical metabolism is reviewed.