In this study, highly stable SF-Au NPs were obtained using regenerated protein silk fibroin (SF) to stabilize gold nanoparticles (Au NPs), which were prepared through citrate reduction method. The stability of SF-Au NPs in buffer solutions with different pH conditions and different KCl concentrations were systematically investigated. Ultraviolet-visible spectroscopy (UV-Vis)、 transmission electron microscopy (TEM) and fourier transform infrared spectroscopy (FTIR) were employed to characterize the SF-Au NPs system. The results revealed that SF afforded the Au NPs with remarkable stability in the pH range of 3 to 11 and salt concentrations range of 50 mM to 1000 mM. Taking citrate and BSA stabilized gold nanoparticles as controls, the results indicated that steric hindrance may play an dominated role for the stability of SF-Au NPs. The proposed method was simpler, more efficient, and more cost effective than the conventional modification strategies for Au NPs; thus, SF–Au NPs can be potentially used in biomedical applications.