In this paper, electron beam (e-beam) reduction method is applied for the catalyst layer preparation in the growth of carbon nanotubes (CNTs). A hot cathodic electron beam facility was employed to electron bombarding of catalyst layer before stage of CNTs growth. This new method leads to reducing the diameter of particles via sputtering and evaporating the surface of catalyst. The growth of CNTs was performed on the Fe catalyst layer with SiO2 substrate in an environment of different mixed gases (H-2, NH3 and C2H2) by thermal chemical vapor deposition (TCVD) system. The morphology of the electron beam reduced catalyst particles were probed by atomic force microscopy (AFM). All samples were analyzed by scanning electron microscopy (SEM) before and after growth of CNTs. SEM analyses clarified that the catalyst grains have been smaller under effect of electron beam bombardment.