Nylon 66 is an engineering plastic with the advantageous characteristics of light weight, heat resistance, high strength, and high rigidity; it has been used in automobile parts as a substitute for aluminum alloys. There have also been many studies on the mechanical properties of nylon 66. However, the flow stress in nylon 66 after necking has not been thoroughly investigated yet due to the difficulty in identifying local stress. In this study, a flow stress optimization method based on the digital image correlation (DIC) method and response surface method, is proposed. The averaged true stress–true plastic strain curve is first obtained by a quasi-static tensile test via DIC. Subsequently, the flow stress is optimized via the response surface method to obtain the actual flow stress state after necking. The accurate flow stress in the specimen can be obtained based on the relationship between the global load force and distance between the two shoulders of the specimen grip parts that is unaffected by the cross-sectional area change during necking. The results indicate that the average true stress recorded from the experiment overestimates the flow stress after necking. The optimized flow stress exhibits the maximum decrease of 9%. The accuracy of the optimized flow stress is evaluated by comparison with the true strain distribution on the specimen that is accurately measured via DIC.