A facile method of stabilizing magnetic iron oxide nanoparticles (MNPs) in biological media (RPMI-1640) via surface modification with fetal bovine scrum (FBS) is presented herein. Dynamic light scattering (DLS) shows that the size of the MNP aggregates can be maintained at 190 +/-2 nm for up to 16 h in an RPMI 1640 culture medium containing >= 4 vol % FBS. Under transmission electron microscopy (TEM), a layer of protein coating is observed to cover the MNP surface following treatment with FBS. The adsorption of proteins is further confirmed by X-ray photoelectron spectroscopy (XPS). Gel electrophoresis and LC-MS/MS studies reveal that complement factor I-I, antithrombin, complement factor I, alpha-1-antiproteinase, and apolipoprotein E are the proteins most strongly attached to the surface of all MNP. These surface-adsorbed proteins serve as a linker that aids the adsorption of other serum proteins, such as albumin, which otherwise adsorb poorly onto MNPs. The size stability of FBS-treated MNPs in biological media is attributed to the secondary adsorbed proteins, and the size stability in biological media can be maintained only when both the surface-adsorbed proteins and the secondary adsorbed proteins are present on the particle's surface.