Silver nanoparticles (AgNPs) are the most commonly used nanoparticles owing to their antimicrobial properties. The motivation of the present study was (1) to analyze the effect of silver particle size on rat tissue distribution at different time points, (2) to determine the accumulation of AgNPs in potential rat target organs, (3) to analyze the intracellular distribution of AgNPs and (4) to examine the excretion of AgNPs by urine and feces. AgNPs were characterized by dynamic light scattering (DLS), zeta potential measurements, BET surface area measurements, transmission and scanning electron microscopy. AgNPs (20 and 200 nm) were administered intravenously (i.v.) to male Wistar rats at a dose of 5 mg kg–1 of body weight. Biological material was sampled 24 h, 7 and 28 days after injection. Using inductively coupled plasma-mass spectrometry (ICP-MS) and transmission electron microscopy (TEM) it was observed that AgNPs translocated from the blood to the main organs and the concentration of silver in tissues was significantly higher in rats treated with 20 nm AgNPs as compared with 200 nm AgNPs. The highest concentration of silver was found in the liver after 24 h. After 7 days, a high level of silver was observed in the lungs and spleen. The silver concentration in the kidneys and brain increased during the experiment and reached the highest concentration after 28 days. Moreover, the highest concentration of AgNPs was observed in the urine 1 day after the injection, maintained high for 14 days and then decreased. The fecal level of silver in rats was the highest within 2 days after AgNPs administration and then decreased. Copyright © 2012 John Wiley & Sons, Ltd.