Abstract: Time-of-flight mass spectrometry (TOF-MS) is widely used in protein chemistry and proteomics research due to its high sensitivity，high resolution and high accuracy. Recently two Q-TOF-MS analytical systems are developed. Both systems are equipped with electrospray ionization source which could be directly hyphenated with HPLC as an outstanding detector. The ESI-triple quadrupoleorthogonal acceleration-reflecting TOF mass spectrometer has some promising features, such as a ESI source with heatable nebulizer settled on a smart triaxiality moving stage, an excellent ion-optics system composed with triplequadruple, double-pulse acceleration, and a double stage reflector with homemade grids. The GramicidinS sample concentration below to 3 fmol/μL can be identified by these instruments under the optimized conditions. The mass range of these instruments is from 50 u to 6 000 u with a general ESI interface to the HPLC. The optimal mass resolution exceeds 11 000 (Full Width at Half Maximum, FWHM) and the signal/noise ratio of reserpine (10 pg/μL) is greater than 100.