A virtual impactor for submicron particles is prerequisite for bio-aerosol optical online monitoring and it is a hot research topic. Based on aerodynamics and past research work, a 0.4-μm-cutpoint virtual impactor is designed with the help of a commercial computational fluid dynamics (CFD) package, Fluent. Discrete phase model (DPM) is applied to simulate and analyze the effect of nozzle-to-collection probe distance, collection probe diameter and minor-to-major flow ratio. The virtual impactor with optimum design parameters is fabricated and tested. According to the experimental results, the virtual impactor can collect 0.37 μm, 0.5 μm and 0.7 μm PSL microspheres efficiently as desired, which proves that the CFD method is practical. Cutoff diameter of the virtual impactor can reach 0.4 μm, which is fit for practical applications.