Cell migration through anisotropic microenvironment is critical to a wide variety of physiological and pathological processes. However, adequate analytical tools to derive motile parameters to characterize the anisotropic migration are lacking. Here, we proposed a method to obtain the four motile parameters of migration cells based on the anisotropic persistent random walk model which is described by two persistence times and two migration speeds at perpendicular directions. The key process is to calculate the velocity power spectra of cell migration along intrinsically perpendicular directions respectively, then to apply maximum likelihood estimation to derive the motile parameters from the power spectra fitting with double exponential decay. The simulation results show that the averaged persistence times and the corrected migration speeds can be good estimations for motile parameters of cell migration.