Eukaryotic expression and biological activity identification of fusion protein ABD-Fc-IL-2 / 中国生物制品学杂志

ABSTRACT

@#Objective To express the fusion protein ABD-Fc-IL-2 in eukaryotic cells and detect its biological activity. Methods The target gene SP-ABD-Fc-IL-2 was amplified by direct and overlapping PCR,and then ligated to vector pcDNA3. 1(+).The obtained recombinant plasmid pcDNA3. 1/SP-ABD-Fc-IL-2 was transiently transfected into CHO-S cells to express the fusion protein ABD-Fc-IL-2,which was purified by Protein A beads affinity chromatography. The specificity of the purified fusion protein was detected by Western blot,the biological activity was detected by CTLL-2/MTT cell proliferation colourimetry,and the interaction between ABD fragment and human serum albumin(HSA)was detected by pull down/Western blot. Results The recombinant plasmid pcDNA3. 1/SP-ABD-Fc-IL-2 was constructed correctly as identified by restriction analysis and sequencing. The purified fusion protein ABD-Fc-IL-2 showed a purity of 90% and bound specifically to mouse anti-IL-2 monoclonal antibody with the biological activity of 3. 29 × 108IU/mL. The ABD of fusion protein and HSA bound to each other. Conclusion The eukaryotic fusion protein ABD-Fc-IL-2 had high biological activity,which promoted the proliferation of CTLL-2 cells and maintained the binding ability of ABD fragment to HSA.