Abstract There is an urgency to prepare non-neurotoxic nanoparticles (NPs) as MRI contrast agent. We prepared ZnFe2O4 NPs with monocrystal structure in the template of silk-fibroin (SF) peptide at room temperature. Analyses of their crystallite size and NPs diameter after the growth of 1d and 8d indicated that SF peptide could promote nucleation, growth and dispersion of NPs. Results of EDS, Raman and TG proved that there was 4wt% of the chemical composition of SF in as-prepared NPs. A possible growth mechanism of ZnFe2O4 NPs in the template of SF is proposed. The saturation magnetizations of NPs with SF (SF-NPs) were about 15 and 30emu/g for 1 and 8 d, respectively. SF-NPs obviously promoted the viability of PC12 cells at NPs concentration of 0.0625mg/mL in 2d and 5d of co-culture. Analysis of neurite length indicated no significant decreases of the experimental groups at different SF-NPs concentrations (from 0.0625 to 0.5mg/mL) after 5d cell co-culture, because SF peptide coating could slow the release of iron/zinc ions from NPs. Furthermore, SF-NPs did not destroy the organelles, karyotheca and the neurite using the observation in the cell ultrathin sections. Based on their good magnetic property and neuro-cytocompatibility, the potential application of ZnFe2O4 NPs with SF as MRI contrast agents would be envisioned.