Morphology-engineered nickel-iminodiacetic acid functionalized agarose magnetic microspheres for enhanced...

Morphology-engineered nickel-iminodiacetic acid (Ni-IDA)-functionalized agarose magnetic composite microspheres (MAMs/Ni-IDA) were rationally designed and synthesized via emulsion-solidification strategy for the efficient and selective histidine (His)-tagged proteins purification. The composite microspheres were constructed by incorporating large-sized system Fe₃O₄ magnetic nanoparticles (MNPs) into an agarose matrix, followed by Ni²⁺-chelated IDA functionalization. Optimal performance was achieved at 8% MNPs loading, which effectively suppressed microsphere agglomeration and maximized the epoxy group density (72.4 μmol/g), laying a robust foundation for subsequent IDA immobilization. Comprehensive characterizations including optical microscopy, laser particle sizing, vibrating sample magnetometry (VSM), Fourier-transform infrared spectroscopy (FT-IR), and elemental analysis (EDS) confirmed that the MAMs/Ni-IDA microspheres exhibited successful MAMs/Ni-IDA formation, well-defined structures, and excellent magnetic responsiveness. The Irregular@MAMs/Ni-IDA displayed a specific saturation magnetization of 16 emu/g, ensuring rapid and efficient magnetic separation. Zeta potential changes verified IDA/Ni²⁺ grafting. Importantly, the Irregular@MAMs/Ni-IDA maintained exceptional selectivity and recyclability for target His-tagged proteins (28.3 kDa), retaining over 95% of their initial adsorption capacity after six consecutive purification cycles, outperforming commercial counterparts. These findings collectively highlighted the significant potential of the morphology-engineered Irregular@MAMs/Ni-IDA as a cost-effective and high-performance adsorbent for His-tagged protein purification in academic research and industrial bioprocessing.