Abstract: In order to test and analyze the mode Ⅱ inter-layer fracture toughness of additive manufacturing carbon fiber reinforced polymer (CFRP), and quantitatively evaluate the influence of printing parameters on the mode Ⅱ fracture toughness, then promote the application of additive manufacturing CFRP technology in bridge structure, experimental and simulation methods were used to carry out the relevant explorations, during this study. Firstly, the printing process was optimized and a novel method for preparing inter-layer pre-cracks was proposed, namely missing layer replacement method. Meanwhile, the influence of two types of key printing parameters (Printing temperature and printing speed) on the mode Ⅱ inter-layer fracture toughness of additive manufacturing CFRP was studied. Secondly, based on the cohesive zone theory, simulation models of the end notched flexure (ENF) test specimens with pre-cracks under various printing conditions were established. In addition, a comparative analysis between simulation results and test data was carried out. The results indicate that the influence of two key printing parameters on the mode II inter-layer fracture toughness of additive manufacturing CFRP are significant, and the influence of printing temperature is stronger. When the printing temperature increases from 245℃ to 285℃, the variation range of peak force test data is 18%~27%, and the variation range of inter-layer fracture toughness is 14%~32%. When the speed increases from 20 mm/s to 60 mm/s, the variation range of peak force test data is 4%~31%, and the variation range of inter-layer fracture toughness is 4%~16%. Moreover, the relative errors between simulation results and test data are controlled within 10%, which indicates that the test data in this study is reasonable and stable, so the missing layer replacement method has strong practicality for preparing additive manufacturing CFRP specimens with pre-cracks. In addition, the simulation method of traditional process composites is also suitable for the simulation analysis of additive manufacturing CFRP. Therefore, this method can provide technical support for the subsequent quantitative evaluation of inter-layer mechanical properties of additive manufacturing CFRP bridge structures.