In this study, a kind of nanocomposite film was fabricated via combining silk fibroin, polyvinyl alcohol (SF/PVA) and AgNP/polydopamine-modified Montmorillonite (AgNP/PDA-Mt). The structural characteristics and properties of the SF/PVA/AgNP/PDA-Mt nanocomposites films were identified using X-ray diffraction (XRD), Thermal gravimetric analyzer (TGA), Fourier transform infrared spectroscopy (FTIR), EDS-mapping analyses and Scanning electron microscope (SEM). The results indicated enhanced thermal performance of SF/PVA/AgNP/PDA-Mt nanocomposites with increased AgNP/PDA-Mt weight. The nanocomposite film exhibited excellent antibacterial activity against E. coli and S. aureus. The 2 % SF/PVA/AgNP/PDA-Mt film showed the highest zone of inhibition with an average inhibition circle diameter of 26.1 mm against E. coli and 20.61 mm against S. aureus. Cytotoxicity test results indicated that the nanocomposites films were biocompatible with L929 cells with a 100 % survival rate, which can be considered as one of the advantages of new nanocomposites films. These findings suggest that SF/PVA/AgNP/PDA-Mt films have potential clinical applications in wound dressing and antibacterial biomedical applications.