Currently, the network security accidents occur frequently, and traditional passive defense technologies have no possible response to unknown network security threats. In response to this problem, a multi-stage evolutionary signal game model was constructed. And with the background that the defender actively launches inductive signals for security defense, a Moving Target Defense (MTD) decision-making algorithm based on the multi-stage evolutionary signal game model was proposed. Firstly, the basic elements of the model were defined and the overall model was analyzed theoretically based on the assumptions of incomplete information constraints and complete rationality of both sides of the game. Then, a method for quantifying the benefits of offensive and defensive strategies was designed, and a detailed optimal strategy solving process for equilibrium was given. Finally, the MTD method was introduced to analyze the evolution trends of both sides’ strategies in multi-stage attack and defense. Experimental results show that the proposed algorithm can predict the optimal defense strategies at different stages accurately, and has guiding significance for the research of new network active defense technology. At the same time, the results of comparing the proposed algorithm with the traditional random uniform strategy selection algorithm through Monte Carlo simulation experiment verify the effectiveness and safety of the proposed algorithm.