Simplified in vitro refolding and purification of recombinant human granulocyte colony stimulating factor using protein folding cation exchange chromatography

• Optimized chromatographic conditions facilitated the efficient mass recovery, purity and specific activity of rhG-CSF.
• PF-SCX is an alternative technique for refolding and purification of rhG-CSF in the presence of additives.
• Cooperative effect of additives enhanced the protein refolding with simultaneous purification.

Protein folding-strong cation exchange chromatography (PF-SCX) has been employed for efficient refolding with simultaneous purification of recombinant human granulocyte colony stimulating factor (rhG-CSF). To acquire a soluble form of renatured and purified rhG-CSF, various chromatographic conditions, including the mobile phase composition and pH was evaluated. Additionally, the effects of additives such as urea, amino acids, polyols, sugars, oxidizing agents and their amalgamations were also investigated. Under the optimal conditions, rhG-CSF was efficaciously solubilized, refolded and simultaneously purified by SCX in a single step. The experimental results using ribose (2.0 M) and arginine (0.6 M) combination were found to be satisfactory with mass yield, purity and specific activity of 71%, ≥99% and 2.6 × 108 IU/mg respectively. Through this investigation, we concluded that the SCX refolding method was more efficient than conventional methods which has immense potential for the large-scale production of purified rhG-CSF.