Medium format enhancement: design of an immobilized heparin affinity media for industrial bioprocessing.
Affinity chromatography is one of the most powerful and selective separation methods available. Recently, affinity methods are being incorporated into industrial processes with some frequency. One of the reasons for this is that affinity media robust enough for industrial bioprocessing are now available. By robust is meant that the media meet stringent requirements for cleanability, sanitization, physical and chemical stability, regulatory and technical support, batch-to-batch reproducibility and reliability of supply. This paper describes a medium format enhancement program to adapt a widely known group-specific affinity medium, Heparin Sepharose CL6B, to the requirements for industrial bioprocessing. The new medium, Heparin Sepharose 6 Fast Flow was designed for the recovery of antithrombia 3 (AT3) at industrial scale. The medium is based upon a highly cross-linked 6% agarose, which is produced in very large scale and is familiar to regulatory agencies. The ligand, heparin, is attached to the matrix by a reductive amination chemistry. The resulting linkage is stable in 0.1 N NaOH for 150 h, showing no decrease in AT3 binding affinity at that time. Heparin has a broad biological functionality and thus is useful chromatographically for the purification of a number of proteins which have an affinity for heparin. Heparin, as a complex sugar, is also a highly charged polyanion and thus has interesting ion-exchange properties. Because of its broad applicability to a number of purification problems, immobilized heparin is a useful case study in medium format enhancement.
Kennedy
R M
RM
Bioprocess Technology, Pharmacia Biotech, Inc., Piscataway, NJ 08854-1327, USA.
D000991:Antithrombins / Q000737:chemistry; D002846:Chromatography, Affinity; D006493:Heparin / Q000737:chemistry*; D008024:Ligands; D012685:Sepharose / Q000737:chemistry*
D000991:Antithrombins; D008024:Ligands; D006493:Heparin; D012685:Sepharose