Recently, Fe3O4 nanoparticles, due to their unique physico-chemical and magnetic properties have attracted considerable interest for different applications. Due to their minimal toxecicity they are of particular interest in biomedicine. The success of nanoparticles applications depends on stability, i.e. agregation in a specific medium. Stability of nanoparticles is affected by concentration of the particles themselves, pH, ionic strength and composition of the medium (presence of the ions and organic matter). The stabiliy of particles is usually determined by measuring their size. Different methods are available for determination of size, but for most of them, nanoparticles should be deposited onto a surface and dried if a suspension is to be characterized. Sizes obtained with these methods do not reflect the influence of solvent. A method enabling determination of medium influence is dynamic light scattering (DLS). It is a nondestructive, fast and reliable method for determining the size of particles in the suspensions. Stability of Fe3O4 nanoparticles in Mili Q water and in biological media (Phosphate Buffered Saline (PBS), Dulbecco 's Modified Eagle Medium (DMEM) and DMEM/ITS (insulin, transferrin and selenium)) has been investigated. Magnetite nanoparticles were synthesized by wet microemulsion technique and characterized by powder X-ray diffraction and scanning electron microscopy. Stability of Fe3O4 nanoparticles has been monitored by dynamic light scattering during 192 h for Mili Q water, 96 h for PBS i DMEM/ITS and 168 h for DMEM. The results have shown that the main factors that affect the stability of Fe3O4 nanoparticle suspensions are ionic strength, which is in accordance with Derjaguin–Landau– Verwey–Overbeek (DLVO) theory, and the presence of proteins that may stabilize nanoparticles by adsorption.