AbstractThe synthesis and characterization of small monodisperse multimodal iron oxide nanoparticles (IONPs), functionalized with boron‐dipyrromethene (BODIPY) derivatives is described. Optical and relaxometric properties of the nanoparticles have been evaluated, suggesting a potential applicability of the particles as bimodal contrast agent for magnetic resonance and optical imaging. The BODIPY funcionalized IONPs showed bright fluorescence and high transverse relaxivity r2 which was in the range 47‐49 mM−1 s−1. Two‐photon fluorescence microscopy experiments performed at 1100 nm revealed that BODIPY‐functionalized IONPs showed significant increase in two‐photon fluorescence compared to the bare nanoparticles, suggesting their potential for applications in multimodal imaging. Cell viability studies on rat tumor (AR42J) and human ovarian cancer (SKOV3) cells showed no significant cytotoxicity at any of the tested concentrations, while greater sensitivity was observed for human hepatic stellate (GRX), human umbilical vein (HUVEC) and rat insulinoma (INS1) cells at higher nanoparticle concentrations.