It is currently a very active research area to develop multifunctional nanocomposites (NCs) which integrate the novel properties from various nanomaterials for multimodal imaging and simultaneous therapy. These theranostic nanoplatforms can provide complementary information from each imaging modality for accurate diagnosis and, at the same time, afford an imaging-guided focused tumor therapy. Among them, core/shell Fe₃O₄@Au NCs have attracted wide attention due to their unique advantages in magnetic targeting, multimodal imaging and photothermal therapy. This study developed a layer-by-layer assembling approach to synthesize Fe₃O₄@Au NCs with high photothermal conversion efficiency. The as-synthesized NCs showed significant photothermal ablation capability to HeLa cells in vitro under near infrared laser irradiation. To ensure the safety for medical applications, the bio-effects of Fe₃O₄@Au NCs on RAW264.7 cells were carefully assessed, in terms of cell viability, oxidative stress and apoptosis. We have demonstrated that Fe₃O₄@Au NCs had good biocompatibility in RAW264.7 cells and no significant cytotoxicity was found. Therefore, the Fe₃O₄@Au NCs synthesized in this study have great potential as an ideal candidate for CT/MR imaging and photothermal therapy.