Method of weak partitioning chromatography

Abstract

This invention relates to methods of using weak partitioning chromatography for the purification of a product from a load fluid. Further, the invention relates to methods of weak partitioning chromatography defined by operating conditions which cause a medium to bind least 1 mg of product per mL of medium, or alternatively, defined by a partition coefficient of at least 0.1.

Description

RELATED APPLICATIONS [0001] This application is a continuation of U.S. Application No. 11 / 372,054, filed on Mar. 10, 2006, which claims the benefit under 35 U.S.C. § 119(e) of U.S. Application No. 60 / 660,437, filed on Mar.11, 2005, the disclosures of which are incorporated by reference herein in their entirety.FIELD OF THE INVENTION [0002] The invention relates to methods of recovering a purified product from a load fluid. In certain embodiments of the invention, the methods comprise passing the load fluid through a medium at operating conditions which cause the medium to bind at least 1 mg of product per mL of medium, and recovering the purified product from the column effluent. In other embodiments of the invention, the methods comprise passing the load through a medium at operating conditions defined by a partition coefficient of at least 0.1. BACKGROUND OF THE INVENTION [0003] Within the biotechnology industry, the purification of proteins on a commercial scale is an important c...

AI Technical Summary

Benefits of technology

[0025] The methods of the invention can be optionally combined with one or more purification steps. The optional step(s) can be performed either prior to or following the practice of the inventive method. For example, the methods of the invention can optionally be combined with a Protein A chromatography step as an initial step.

[0026] In one embodiment of the invention, a product-containing fluid is eluted from a Protein A column using an elution buffer of low ionic strength; the pH and conductivity of the product-containing fluid is adjusted using a neutralization buffer which results in no more than 20 mM of the ionic strength of the product-containing fluid, resulting in the load fluid; and the load fluid is passed through an anion exchange medium under the operating conditions of the invention.

Problems solved by technology

Within the biotechnology industry, the purification of proteins on a commercial scale is an important challenge to the development of recombinant proteins for therapeutic and diagnostic purposes.

Problems related to yield, purity, and throughput plague the manufacturing sector.

However, drawbacks to using Protein A chromatography for protein purification include leakage of the Protein A capture agent, leading to contamination of the eluted protein product.

None of the conventional bind-elute or flow-through methods in the prior art, however, is able to meet the needs of the biotechnology industry in terms of all the requirements of throughput, yield, and product purity.

Bind-elute methods and displacement methods are limited by, among other factors, the capacity limit of the separation medium for the desired protein.

Flow-through methods, on the other hand, do allow for higher load challenges than bind-elute methods but are limited by the capacity of the separation medium for the impurities.

With flow-through methods, no substantial binding of the product to the column occurs; any substantial product binding is seen as negatively impacting product recovery.

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