other Pharmaceutical Biotechnology Pharmaceutical Biotechnology مقاله کامل Full Length/Original Article <strong>Background: </strong>Protein purification is the most complicated issue in the downstream processes of recombinant protein production; therefore, improved selective purification methods are important. Affinity-based protein purification method using polyhistidine-tag (His-tag) and nickel-<em>nitrilotriacetic acid</em> (Ni-NTA) resins is one of the most common strategies. Magnetic nanoparticles (MNPs) can be used as a beneficial alternative for Ni-NTA resins. However, there is no data on the capability of MNPs for protein purification from inclusion bodies; this issue is studied here.<strong> Methods: </strong>Recombinant His-tagged proteins of enhanced green fluorescent protein (EGFP)-His and streptokinase (SK)-His were expressed in <em>E. coli</em> BL-21 (DE3) in soluble and inclusion body forms, respectively. MNPs including Fe₃O₄ magnetic core, SiO₂ shell, and Ni²⁺ on the surface were synthesized by sol-gel and hydrothermal reactions and then characterized by X-ray powder diffraction, vibrating sample magnetometer, and scanning electron microscopy imaging. Both synthesized Fe₃O₄@NiSiO₃ and Fe₃O₄@Ni_xSiO_yMNPs were employed to purify EGEP-His and SK-His under native and denaturing conditions, respectively. The quantity and purity of purified proteins were analyzed by micro-Bradford assay and SDS-PAGE, respectively. <strong>Results: </strong>Both synthesized MNPs were spherical and well-dispersed with the size ranging from 290 to 415 nm. Synthesized MNPs contained Fe₃O_4, SiO₂ shell, and Ni²⁺ on their structures with suitable magnetization properties. Using Fe₃O₄@NiSiO₃and Fe₃O₄@Ni_xSiO_yyielded 192 and 188 &micro;g/mg of SK-His, as compared to 207 and 195 &micro;g/mg of EGFP-His, respectively. <strong>Conclusion: </strong>MNPs containing magnetic Fe₃O₄core, SiO₂ shell, and Ni²⁺on their surface are versatile alternatives for Ni-NTA resins in protein purification for proteins expressed in both soluble and inclusion body forms. Inclusion body, His-tag, Magnetic nanoparticle, Protein purification 192 200 http://ibj.pasteur.ac.ir/browse.php?a_code=A-10-1-806&slc_lang=other&sid=1 Masoud Seyedinkhorasani 0000-0002-4734-9424 No Nano-Biotechnology Department, Pasteur Institute of Iran, Tehran, Iran Reza Ahangari Cohan 0000-0002-0490-4184 No Nano-Biotechnology Department, Pasteur Institute of Iran, Tehran, Iran Saeid Taghavi Fardood 0000-0002-0645-1393 No Department of Chemistry, University of Zanjan, Zanjan, Iran Farzin Roohvand 0000-0002-9597-3555 No Virology Department, Pasteur Institute of Iran, Tehran, Iran Dariush Norouzian 0000-0002-3553-3048 No Nano-Biotechnology Department, Pasteur Institute of Iran, Tehran, Iran Malihe Keramati 0000-0002-7196-8751 Yes Nano-Biotechnology Department, Pasteur Institute of Iran, Tehran, Iran