•The folic acid-conjugated Fe3O4 nanoparticles were synthesized by amidation method.•The conjugation of folic acid was verified qualitatively by UV–vis spectrum.•We confirmed the exothermic effect of the nanoparticles in alternating magnetic field.•The nanoparticles showed targeting effect to KB cells with FA receptors over-expression. The folic acid (FA)-conjugated Fe3O4 magnetic nanoparticles (MNPs) were synthesized by co-precipitation of Fe3+ and Fe2+ solution followed by surface modification with carboxymethyl dextran (CMD) to form carboxymethyl group terminated MNPs, then FA was conjugated with the carboxyl group functionalized MNPs. The morphology and properties of obtained nanoparticles were characterized by X-ray powder diffraction (XRD), Fourier transform infrared spectroscopy (FT-IR), UV–visible spectra (UV–vis), transmission electron microscopy (TEM), dynamic light scattering (DLS), vibrating sample magnetometer (VSM) and thermogravimetric analysis (TGA). The FA-conjugated MNPs exhibited relatively high saturation magnetization and fast magneto-temperature response which could be applied to hyperthermia therapy. To determine the accurate targeting effect of FA, we chose FA-conjugated MNPs as MRI contrast enhancement agent for detection of KB cells with folate receptor over-expression in vitro and in vivo. The results show that these magnetic nanoparticles appear to be the promising materials for local hyperthermia and MRI.