Objective：To prepare and characterize magnetic albumin nanospheres conjugated with cetuximab（C225），which is anti-epithelial growth factor receptor（EGFR） monoclonal antibody，as well as their in vitro anti-lung cancer effect when combined with magnetic fluid hyperthermia. Methods：Fe3O4 nanoparticles（NPs）were prepared by chemical co-precipitation，and magnetic albumin nanospheres（MANs） were prepare by desolvation-crosslinking method，then C225 was further conjugated to synthesize C225-MANs using N-succinimidyl-3-（2-pyridyldithio）propionate（SPDP）. Their morphology，mean particle size，zeta potential，the antibody conjugation efficiency，iron content，specific cell-binding ability，and magnetothermal dynamic profiles were characterized. Lastly，the therapeutic effect of C225-MANs induced double targeted magnetic fluid hyperthermia on lung adenocarcinoma in vitro was evaluated using CCK-8 and flow cytometry（FCM）assay. Results：TEM micrographs showed that the prepared C225-MANs were approximately spherical. Fe3O4 NPs were well incorporated into the core of the nanospheres and the iron content was about 2.0 mg/mL. The mean hydrodynamic diameter was about 140 nm with negative charge. When placing in alternating magnetic field with the output frequency of 200 kHz and the output current of 20 A，the corresponding C225-MANs with different Fe concentrations could rise to a steady temperature ranging from 39.3 ℃ to 52.0 ℃，and the temperature could keep stable without any change after 30 min heating. The fluorescence percentage detected by FCM analysis revealed that the antibody conjugation efficiency was 79.74%. The ability of nanospheres targeting GLC-82 cells in vitro was confirmed by immunofluorescence experiments and magnetic resonance imaging. The proliferation rate of double C225 and magnetic-targeted combined group was significantly lower than any other therapy group（P＜0.05），while the apoptotic rate of the double C225 and magnetic-targeted combined treatment group were significantly higher than any other therapy group（P < 0.05）. Conclusion：C225-MANs had good targeting effect on GLC-82 cells at cellular level. Moreover，dual-targeted magnetic fluid hyperthermia combined with molecular targeted therapy can effectively combat lung cancer in vitro.