Nanoparticles are being increasingly used in day-to-day life. Therefore, concerns have been raised regarding their interactions with the surrounding environment. This study focused on a simple green method for synthesizing silver nanoparticles (Ag-NPs) in an autoclave at 15 psi (103 kPa) and 121°C. An aqueous solution of AgNO3 as a precursor of Ag-NPs and gelatin (type B) reducing and/or stabilizing (capping) agent were used. The effect of various AgNO3 concentrations of certain gelatin concentration and various gelatin concentrations at constant AgNO3 concentration, and autoclaving time, was studied. UV-Vis spectra ascribed that the presence of localized surface plasmon resonance (SPR) of the synthesized Ag-NPs. TEM images and the selected area of electron diffraction confirmed, the formation of Ag-NPs with a diameter of approximately 5 ±0.35 nm. Furthermore, FT-IR revealed that a gelatin polymer matrix stabilized the synthesized Ag-NPs. The Well diffusion assay was used to test the effect of Ag-NPs on six clinical bacterial isolates, where Gram positive bacteria were more susceptible to Ag-NPs than Gram negative bacteria. Therefore, Ag-NPs capped by gelatin have remarkable potential effect as an antibacterial agent, and they not only have various medical applications but can also be used in biological, pharmaceutical and industrial fields.