Purification and Activity of Recombinant Human IL-12(CHO Cells)

Objective To develop a purification procedure for recombinant human IL-12(rhIL-12) expressed in CHO cells and determine the biological activity of purified rhIL-12. Methods The culture supernatant of CHO cells highly expressing rhIL-12 was purified by chromatography combined with ammonium sulfate precipitation. The content of target protein was determined by ELISA, and the recovery rate was calculated. The purified product was identified by SDS-PAGE, SEC-HPLC and Western blot. The biological activity of purified rhIL-12 was determined by induction of IFNγ in PBMCs. Results The total recovery rate of purified rhIL-12 was 56. 4%. SEC-HPLC showed that the purified rhIL-2 reached a purity of 99. 2%. SDS-PAGE proved that the relative molecular masses of two subunits were consistent with their theoretical values (40000 and 35 000)respectively. Western blot revealed specific binding of rhIL-12 to the corresponding antibody. The dose-response curve of rhIL-12 for induction of IFNγ was in a typical S shape, indicating the dose-dependent biological activity of rhIL-12. The potency of rhIL-12 was 8. 3 × 106 IU / mg. Conclusion The purification procedure of rhIL-12 expressed in CHO cells was developed, which was low-cost and simple to handle, and both the recovery rate and purity of purified product were high, indicating that the developed procedure was suitable for industrial production.