Synthesis and optimization of nickel-iron nano-composite for Grp-۷۸ immunoglobulin purification

Background: Protein separation and purification are widely used in all fields of life sciences and biotechnology. The development of new techniques that can replace current common methods increases efficiency and reduces production costs. Magnetic separation is one of the fast and easy methods for separating proteins, cells, and various molecules from the initial raw sample. Objectives: In the present study, Ni-Fe magnetic nano-composite was synthesized and applicate for separation and collection of recombinant GRP-۷۸ immunoglobulin protein.Materials and Methods: In this study, Ni-Fe nanocomposites are prepared by oxidation and reduction method. Then, the synthesized nanocomposites in each method are examined by different imaging and spectroscopy methods to measure the size of the nanocomposite and its surface morphology of Nano-composite, the crystal structure, and its stability. In the final stage, these nanocomposites are exposed to bind to protein histidine tag, and then the protein is collected, concentrated, and purified with an external magnet. In this study, the effect of independent variables including protein concentration, solution pH, and reaction time was simultaneously investigated on the dependent variable (size, protein loading) of iron/nickel oxide nanoparticles prepared by precipitation method. Simultaneous study of variables allows for more complete information on how different factors affect the dependent variable. Results: The present study, the results of the morphology of nanoparticles showed that Ni-Fe Nano-composite had a size smaller than ۱۰۰ nm and their shape is spherical. Vibrational Magnetometry indicated that the nanocomposite has good superparamagnetic properties and is well attracted by a permanent magnet. Therefore, it is suitable for collecting and purifying proteins from solution. Also, FTIR showed that nickel is abundant on the surface of the nanocomposite and is available for binding to the histidine tag of the protein. Continuing the research, protein loading on Ni-Fe nanocomposites was Optimization for several parameters such as time, temperature, pH, and concentration. Nano-composite can be loaded protein in a wide range of pH from ۵.۸ to ۸.۰. The protein loading rate by Ni-Fe Nano-composite is ۲۰ minutes. Temperature analysis also did not show a significant difference in the loading rate. Finally, the evaluation of the efficiency of Iron-Nickel Nano-composite for protein adsorption under optimal conditions indicated the efficiency and ability of the nanocomposite to adsorb a high percentage of protein. Conclusions: The results indicate that Iron-Nickel Nano-composite can be used as an efficient alternative in the purification of a high percentage of protein. Magnetic affinity and ion exchange separation have been successfully used in various fields such as molecular biology, biochemistry, analytical chemistry, environmental chemistry, etc.

کلیدواژه ها:

Immunoglobulin protein ، Purification ، Nano-Composites