Silver nanoparticles (AgNPs), as a material with attractive properties, has received interest worldwide by the implementation of synthesis particles at nano-size. AgNPs has triggered intense research on the production of nanoparticles using different synthesis methods due actual and future application. The increasing attention of AgNPs resulted in the growth and development of nanoparticles synthesis methods. Recently, various approaches, including chemical and biological, have been used for them preparation. In order to obtain information about size, shape, or optical properties of nanoparticles, a broad range of analytical methods—microscopy, spectroscopy, and spectrometry techniques—have been used. AgNPs exhibit attractive antimicrobial properties against bacteria (Gram-positive and Gram-negative), fungi, and show also selective toxicity toward normal and cancerous cells. This chapter reviewed the methods of AgNPs synthesis, their characteristics, antimicrobial activity, and cytotoxicity against normal and tumor cells.