In the 21st century, a great interest is devoted to biomedical application of various nanoparticles, particularly, as a means of improving the effectiveness of therapy for different diseases. Silver nanoparticles (AgNPs) are the most studied and investigated type of nanoparticles. Due to the wide spectrum of their action, silver nanoparticles may be used both to influence pathogenic microorganisms and to improve the treatment of cancer. The basic physico-chemical characteristics and stabilizing agents play an important modifying role in the pharmacokinetics and pharmacodynamics of nanoparticles, determining the severity of the caused effect and their potential toxicity. This review summarizes the main physico-chemical properties of AgNPs and their impact on the biological effects. Additionally, biochemical and pathophysiological mechanisms of silver nanoparticles activity against various microorganisms and tumor cells are considered. Finally, we address the problems, associated with determining the optimal characteristics of nanoparticles, in order to increase their efficiency and reduce their toxicity for the macroorganism.