Silver nanoparticles have been shown to exert antiviral activity against HIV-1 at non-cytotoxic concentrations, but the underlying mechanism has not been fully elucidated. This study investigates the mode of action of silver nanoparticles against HIV-1 using various in vitro assays. The results suggest that silver nanoparticles act as virucidal agents or inhibitors of viral entry, preventing CD4-dependent virion binding, fusion, and infectivity. Silver nanoparticles bind to gp120, inhibiting the initial stages of the HIV-1 life cycle and effectively inactivating both cell-free and cell-associated virus. The antiviral activity of silver nanoparticles is broad-spectrum, effective against a wide range of HIV-1 strains, including drug-resistant strains, and does not appear to induce resistance. These findings highlight the potential of silver nanoparticles as a preventive tool against HIV-1 transmission, acting at early stages of viral replication and inhibiting post-entry stages of the viral life cycle.Silver nanoparticles have been shown to exert antiviral activity against HIV-1 at non-cytotoxic concentrations, but the underlying mechanism has not been fully elucidated. This study investigates the mode of action of silver nanoparticles against HIV-1 using various in vitro assays. The results suggest that silver nanoparticles act as virucidal agents or inhibitors of viral entry, preventing CD4-dependent virion binding, fusion, and infectivity. Silver nanoparticles bind to gp120, inhibiting the initial stages of the HIV-1 life cycle and effectively inactivating both cell-free and cell-associated virus. The antiviral activity of silver nanoparticles is broad-spectrum, effective against a wide range of HIV-1 strains, including drug-resistant strains, and does not appear to induce resistance. These findings highlight the potential of silver nanoparticles as a preventive tool against HIV-1 transmission, acting at early stages of viral replication and inhibiting post-entry stages of the viral life cycle.