Fe3O4@PAM@NTA-Ni2+ Magnetic Composite Nanoparticles for Highly Specific Separation of His-tagged Proteins

A facile approach has been developed to synthesize Fe 3 O 4 @PAM (polyacrylamide) nanoparticles (NPs) with carboxyl groups on the surfaces by copolymerization with acrylamide and acrylic acid in Fe 3 O 4 NPs aqueous suspension. Nitrilotriacetic acid (NTA) was conjugated to the magnetic NPs via well-known carboniimide chemistry using EDC and NHS. The Ni 2+ ions loaded on the surface of NPs provide abundant docking sites for immobilization of His-tagged green fluorescent proteins (His-tagged GFP). The high magnetic property of Fe 3 O 4 @PAM@NTA-Ni 2+ allows an easy separation of the NPs from solution under an external magnetic field, with high His-tagged protein binding capacity (42 μg protein/mg of NPs). The NPs can be recycled for at least four times without significant loss of binding capacity to proteins. These materials show great potential to separate His-tagged protein with low-cost purification at industrial scale.