The surface of 40CrNiMo high strength steel is quenched with CO2 laser, and the fatigue limits of 40CrNiMo before and after laser quenching are obtained through the tension-tension fatigue experiments based on Locati theory. The fracture morphologies of 40CrNiMo before and after laser quenching are investigated by scanning electron microscope. The mechanism of laser quenching on 40CrNiMo′s fatigue strength is analyzed. The results show that a layer of hardened zone, martensite and compressive residual stress after laser quenching is formed on the sample surface, and the fatigue strength of 40CrNiMo is increased significantly from 626 MPa to 715 MPa, increasing by 14.2%. The fatigue crack sources of the sample in quenching and tempering state are produced in the surface, while that are transferred to the subsurface after laser quenching. The fatigue crack sources expand slowly to be fracture. Grain refinement and compressive residual stress by laser quenching are the main mechanism.