Graphene oxide (GO)/chitosan composite membrane with a thickness of several hundred nanometers was prepared through an interfacial self-assembly process. Thermogravimetric analysis (TGA), Fourier transform infrared (FTIR) spectra and X-ray photoelectron spectroscopy (XPS) were employed to study the interactions that induced the assembly of GO nanosheets and chitosan and identified that the results showed that the major interactions were from electrostatic interactions. The surface morphology of synthesized membrane can be further tuned by adjusting the concentration of the chitosan solution. This new method is very effective to improve the GO/chitosan composite membrane production and may promote its broad potential applications in battery industry and life sciences.