To determine the optimal process parameters for the preparation of a Co-based alloy cladding layer, the experimental research of laser cladding Co-based alloy was carried out based on the optimal process window and grey relational analysis methods with 42CrMo as the substrate. The analysis of variance (ANOVA) was used to explore the influence laws of laser process parameters on the forming characteristics of the cladding layer within the optimal process window range. Furthermore, the optimal process parameter combination was obtained by grey relational analysis, and the experimental verification of the optimization results was conducted. It was found that the process parameter interval determined by the optimal process window was laser power 1300–2100 W, scanning speed 6–14 mm/s, and powder feeding rate 17.90–29.84 g/min. The influence order of each process parameter was: laser power > scanning speed > powder feeding rate. The optimal process parameters of laser power 2100 W, scanning speed 6 mm/s, and powder feeding rate 17.90 g/min were obtained. The experimental verification results of optimal process parameters proved that the grey correlation grade of the optimized parameters was improved by 0.260 compared with the initial parameters and agreed well with the prediction value with an accuracy of 96%. After optimization, the cross-sectional area, the ratio of the width to height, cladding efficiency, and powder utilization rate of the cladding track increased by 4.065 mm2, 1.031, 19.032, and 70.3%, respectively, and the fluctuation ratio decreased by 60.9%. The optimal cladding track was well bonded to the substrate without cracks, holes, and evident element segregation, and included the phases of Cr3C7, CoCx, fcc-Co, and WC.