Purification of monoclonal antibodies from whole hybridoma fermentation broth by fluidized bed adsorption

Abstract

To achive the coarse purification of a monoclonal antibody from whole hybridoma fermentation broth a fluidized bed cation exchange process was used. The procedure consisted of application of the crude sample and washing of the bed in a fluidized mode and elution in a fixed bed mode. A completely clarified eluate was obtained with purification factors between 4 and 8 and a concentration of the desired product (monoclonal antibody) by a factor of more than 3 was achived. Thus, a combination of the three early steps of the downstream process clarification, concentration and coarse purification was possible. Two different materials were tested: a commercially available agarose-based matrix (Stream-line-SP), and a self-derivatized material based on controlled-pore glass (Bioran). Initial experiments were performed to describe the fluidization of the glass material. Comparison with the agarose material showed several differences, the agarose matrix allowing liquid flow closer to plug flow than the glass material. Increased backmixing in the liquid phase was detected when fluidizing the glass adsorbent compared with the agarose-based matrix. Despite this fact, comparison of the two materials with respect to antibody binding and elution demonstrated a similar performance. (c) 1995 John Wiley & Sons, Inc.

Recovery and partial purification of penicillin G acylase from E. coli homogenate and B. megaterium culture medium using an expanded bed adsorption column

“…If the PGA adsorption could occur at a pH lower than 4.8, probably higher PGA dynamic capacities would have been determined. That may also happens for the ionic adsorption of other enzymes as the dynamic capacities values reported in the literature for total protein adsorption (q Prot ) capacities are comparable with the adsorption capacities obtained in this work, with crude broths and using the same cation exchanger [14,[27][28][29].…”

Development of operating conditions for protein purification using expanded bed techniques: The effect of the degree of bed expansion on adsorption performance

“…However, feedstocks containing particulates can be processed directly using fluidized or expanded bed adsorption procedures which are able to operate efficiently in the presence of particulates. 5,[8][9][10][11][12]22,23,27,37,44,48 G enerally, the behavior of expanded beds is different from conventional liquid-solid fluidized beds where both liquid and adsorbent phases are well mixed, which results in poor adsorption performance and low prod~ctivity. '~ In our work, a stable bed of adsorbent beads can be easily established without need to stabilize a fluidized bed using magnetic Previous work has described the operating principles of expanded bed ad~orption.~.…”

Purification of recombinant nucleocapsid protein of Newcastle disease virus from unclarified feedstock using expanded bed adsorption chromatography

“…EBA processes employ the flow of fluid through an initially packed bed structure to achieve a contactor with a high-fluid voidage. A stable bed is formed with adsorbent having a well-defined distribution of bead sizes [14]. Under these circumstances, the larger/heavier adsorbent beads will locate near the bottom of the bed, whereas the smaller/lighter adsorbent beads will locate near the top [15].…”