We have studied the stochastic parameters of surface iron oxide nanograin chains, 97 nm in diameter and 2.4 mu m in length, prepared at different annealing temperatures, using atomic force microscopy (AFM) spectral analysis. In this regard, the roughness of the thin films including self-assembled twisted nanograin chains has been analysed and characterized using the height-height correlation function, the roughness exponent as well as the power spectrum density of the AFM profiles and their gradient, for the different annealing temperatures. The tip convolution effect on the stochastic parameters under study has also been investigated. The kinetics of the formation of nanograins on the film surface has been obtained using the AFM spectral analysis of the profiles and their gradient. The activation energy needed for the formation of surface nanograin chains was found to be 0.55 eV. It has been shown that the tip-surface interaction affects mainly the diffusion parameters obtained by using the surface roughness analysis of the profiles, while use of the surface roughness analysis of the gradient of the profiles results in a nearly independent tip convolution effect on the diffusion parameters. Hence, this work also provides a method for calculating the required activation energy for the formation of self-assembled nanostructures affecting the roughness of a surface.