Abstract In order to improve the operational accuracy of microsurgical instruments and increase the success rate of surgery, this paper carries out the design and analysis of six-degree-of-freedom (6-DOF) microsurgical instruments based on rigid-flexible coupling multi-body system. Firstly, the kinematic modeling method of the 6-DOF parallel mechanism with flexible hinges is improved based on the pseudo-rigid body theory in this paper. Secondly, a rigid-flexible coupling simulation system is built to analyze the error sources in terms of the remote center of motion (RCM), preload and side load. Then, the function of motion scaling, the accuracy of kinematic modeling and the validity of the workspace are demonstrated by analyzing the workspace. Finally, the maximum stress and modal analysis are solved to ensure the safety and reliability of the application. The analysis results show that the improved kinematic modeling method improves the positioning accuracy by more than two times, the root mean square error (RSME) at the end of the microsurgical instrument does not exceed 10μm in the workspace. And the microsurgical instrument can withstand a side load of 0.1 N at the RCM. This study will provide a reference for the structural design and control algorithm optimization of the 6-DOF parallel microsurgical instruments.