Integrative Antibody Purification from whole Broth by Fluidized Bed Adsorption

Abstract

The first operation during purification of proteins produced by mammalian cells usually is a solid-liquid separation of the cell containing culture broth, which is supposed to deliver a cell free supernatant. The protein concentration then is enhanced by a concentration step prior to further processing. This initial sequence of purification steps is referred to as primary recovery. Clarification of mammalian cell broths is performed by centrifu-gation (1) or crossflow filtration (2) successfully on a very large scale, but both methods may have certain disadvantages. Shear sensitive cells may be damaged in the centrifugal field of a large scale separator, thus contaminating the cell free supernatant with intracellu-lar proteins or nucleic acids. Conventional crossflow filtration suffers from low flux rates as well as from fouling of the membrane by lipids, cell debris, and protein precipitates. Additionally the protein of interest may be adsorbed irreversibly to the membrane leading to a reduction in product yield during primary recovery. Advanced methods of dynamic filtration indicate solutions to circumvent these problems (3). In addition to some drawbacks of conventional technologies for primary recovery the number of sequential steps in a protein purification correlates with the overall time requirements of the process and thus with the total costs of protein production. Furthermore each additional purification step reduces the overall yield of product from the isolation procedure due to the inherent product loss of the individual operation. As the downstream process may contribute up to 80 % of the total cost of a protein production (4), a reduction of the number of steps using an integrative purification technology especially in the primary recovery phase, may prove to be beneficial to process economics.