Lab-Scale Production of Biologically Active Form of Immunotoxin Targeted against Granulocyte Colony Stimulating Factor

This study aimed to produce recombinant hybrid protein, DT389GCSF, in E. coli BL-21(DE3) in a soluble and active form and to purify it and evaluate its cytotoxicity.

The protein, His6-tagged DT389GCSF was expressed in E. coli BL-21(DE3) in a soluble form at 28 °C. Then it was purified by affinity chromatography on nickel sepharose column. Finally, the function of purified recombinant protein was evaluated by MTT assay on HL-60 cancer cell line.

The yield of expression and purification of DT389GCSF were determined at about 12 and 80 percent, respectively, using Image J software. The IC50 value upon 48 hours of exposure of DT389GCSF toward cell line was about 0.00017±0.000019 M.

By reducing the temperature and using the intracellular mechanism, the refolding of hybrid protein, DT389GCSF, can be observed in a proper and active form. As a result, in vitro production of relevant immunotoxins, by using this method, the steps of inclusion body production can be neglected.