Multi-walled carbon nanotubes (MWCNTs) were homogeneously dispersed in silk fibroin (SF) solutions at different compositions, and a simple solvent-casting method was used to fabricate SF/MWCNT films. Structure, viscosity, and mechanical properties of the SF/MWCNT nanocomposites were characterized by FTIR (Fourier Transform Infrared Spectroscopy), viscometry, and tensile testing. Fibroblast cells were used to examine cell viability and attachment to nanocomposite films. Compared to a pure SF film, adding just 0.5 % (w/w) of MWCNT to the SF matrix could enhance the Young’s modulus and ultimate tensile strength by approximately 24 % and 39 %, respectively. In addition, with increasing MWCNTs content, the percentage of β-sheet structure increased significantly. When the content of MWCNTs increased from 0 to 1 % (w/w), the β-sheet content improved from 17.8 % to 53.6 %. Furthermore, adding up to 1 % (w/w) MWCNT increased the viscosity of the SF solution ten fold. From a biological point of view, no toxic effect was observed on fibroblast cells in the presence of MWCNTs after 3 and 7 days. This investigation suggests that SF/MWCNT nanocomposite films with improved mechanical properties and biocompatibility are well-suited as novel biomaterials.