In order to improve the accuracy and reliability of micropipetting, a method of micro-pipette detection and calibration combining the dynamic pressure monitoring in pipetting process and quantitative identification of pipette volume in image processing was proposed. Firstly, the normalized pressure model for the pipetting process was established with the kinematic model of the pipetting operation, and the pressure model is corrected by the experimental method. Through the pipetting process pressure and pressure of the first derivative of real-time monitoring, the use of segmentation of the double threshold method as pipetting fault evaluation criteria, and the pressure sensor data are processed by Kalman filtering, the accuracy of fault diagnosis is improved. When there is a fault, the pipette tip image is collected through the camera, extract the boundary of the liquid region by the background contrast method, and obtain the liquid volume in the tip according to the geometric characteristics of the pipette tip. The pipette deviation feedback to the automatic pipetting module and deviation correction is carried out. The titration test results show that the combination of the segmented pipetting kinematic model of the double threshold method of pressure monitoring, can effectively real-time judgment and classification of the pipette fault. The method of closed-loop adjustment of pipetting volume can effectively improve the accuracy and reliability of the pipetting system.