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Herein, we report the use of a cell-free extract for the extracellular synthesis of silver nanoparticles (AgNPs) and their potential to address the growing threat of multidrug-resistant (MDR) pathogenic bacteria. The reproducibility of AgNP synthesis was good and AgNP formation kinetics were monitored as a function of various reaction factors via ultraviolet-visible absorption spectroscopy. This green method was dependent on the alkaline pH of the reaction mixture. With the addition of dilute sodium hydroxide, well-dispersed AgNPs could be produced in large quantities via the classical nucleation and growth route. The new biosynthetic route enabled the production of AgNPs within a narrow size range of 4 to 17 nm. The AgNPs were characterized using various techniques and their antibacterial activity against MDR pathogenic bacteria was evaluated. Field-emission scanning electron microscopic imaging revealed prominent morphological changes in <i>Staphylococcus aureus</i> cells due to mechanical damage, which led to cell death. <i>Escherichia coli</i> cells showed signs of contraction and intracellular fluid discharge as a consequence of disrupted cell membrane function. This new biologically-assisted extracellular strategy is potentially useful for the decontamination of surfaces and is expected to contribute to the development of new products containing AgNPs.