A tightly focused, linearly polarized laser beam, so-called optical tweezers, is proven to be a useful micromanipulation tool. It is known that there is a stiffness asymmetry in the direction perpendicular to the optical axis inherited from the polarization state of the laser. In this Letter, we report our experimental results of stiffness asymmetry for different bead sizes measured at the optimal trapping condition. We also provide the results of our generalized Lorenz-Mie based calculations, which are in good agreement with our experimental results. We also compare our results with previous reports.