Purification Of Recombinant Human Interferon α2a

Studies on the purification process of interferon α2a from E.coli. PBE918/BMH7118 were described in this thesis.In this experiment three purification proceses without McAb affinity chromatography were established. The first purification process consisted of IEC, RP-HPLC and GFC. The target protein was purified by 55 folds. The purified product with a specific activity of 1.75 ×10~8 IU/mg protein,appeared as a single band on SDS-PAGE electrophoretogram and a single peak on the HPLC chromatotogram. The purity of the product was above 95% analyzed by SDS-PAGE and 97.00% by HPLC. The yield was 38.6%.In the second process, the extracted interferon was purified by HIC, IEC, RP-HPLC and GFC. The target protein was purified by 70 folds. Its specific activity was 2.22 × 10~8 IU/mg protein. SDS-PAGE of the purified protein showed a single band corresponding to an apparent molecular mass of 19kD. The purity of the product v/as above 95% detected by SDS-PAGE and 97.75% by HPLC. The recovery of rHuIFNa2a was 23.9%.In the third process, interferon was extracted using 7 mol/L guanidine hydrochloride and then purified by IMAC, IEC, RP-HPLC and GFC. The target protein was purified by 180.2 folds. Its specific activity reached 2.36×10~8 IU/mg protein. The purified product exhibited a single band on SDS-PAGE and its molecular weigh was about 19kD. The purity of the product was above 97% analyzed by SDS-PAGE and 97.83% by HPLC. The yield was 32.0%.