Abstract: To obtain an excellent antibody purification medium, affinity chromatographic packing with recombinant staphylococcal protein A (rProtein A) was synthesized and verified. With E. coli cells harboring the recombinant plasmid, the rProtein A was expressed and purified, then was conjugated to epichlorohydrin-activated Sepharose 4 Fast Flow to prepare an affinity chromatographic packing. The performances of the packing were validated with rabbit anti-urate oxidase. After the reaction, the concentration of rProtein A coupled to Sepharose 4 Fast Flow was 1.5×10~4 mol/L. Scatchard analysis of the binding isotherm for IgG showed excellent binding capacity on the adsorbent, giving a dissociation constant (Kd) of 2.28×10~7 mol/L and a theoretical maximum adsorption capacity of 20.697 g/L. The identification showed the packing was stable in 0.1 mol/L NaOH solution at 1 h. By using the packing, the pure antibody exhibited on sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) was obtained from rabbit serum after one-step elution, with 96.1% of yield and 19 mg IgG for one milliliter of gel. The research laid the foundation of the localization of rProtein A affinity packing.

Key words: affinity packing, antibody, preparation, purification, recombinant staphylococcal protein A (rProtein A)