Optimized method for antibody capturing by mixed mode chromatography

Patent metadata

Field	Value
Publication number	US-9422329-B2
Application number	US-201113883243-A
Country	US
Kind code	B2
Filing date	Nov 2, 2011
Priority date	Nov 5, 2010
Publication date	Aug 23, 2016
Grant date	Aug 23, 2016

Abstract

Herein is reported a method for the purification of an antibody directly captured from clarified cell culture supernatants using Streamline CST and/or Capto MMC, wherein especially product related (aggregates and fragments) and process related impurities (host cell protein, media components) could efficiently be removed, resulting in a preparation with a purity comparable to classical protein A affinity chromatography.

First claim

The invention claimed is:

1. A method for producing an anti-IGF-1R antibody comprising the following steps:
a) applying a crude mammalian cell culture cultivation supernatant to a multimodal weak cation exchange chromatography material,
b) recovering the anti-IGF-1R antibody by applying a buffered solution comprising ethylene glycol and an inorganic salt to the multimodal weak cation exchange chromatography material and thereby producing an anti-IGF-1R antibody.

2. The method according to claim 1 , wherein the method comprises the following additional step a-1) prior to step a):
a-1) applying a buffered solution comprising an inorganic salt to the multimodal weak cation exchange chromatography material.

3. The method according to claim 1 wherein the method comprises the following additional step a-b) after step a) and prior to step b):
a-b) applying a buffered solution to the multimodal weak cation exchange chromatography material, whereby the anti-IGF-1R antibody is not recovered from the multimodal weak cation exchange chromatography material.

4. The method according to claim 3 , wherein the step a-b) comprises two steps a-b1) and a-b2):
a-b1) applying a buffered solution comprising an inorganic salt to the multimodal weak cation exchange chromatography material, and
a-b2) applying a buffered solution comprising a denaturant to the multimodal weak cation exchange chromatography material,
whereby the anti-IGF-1R antibody is not recovered from the multimodal weak cation exchange chromatography material.

5. The method according to claim 4 wherein the denaturant is selected from guanidinium hydrochloride, and urea.

6. The method according to claim 1 wherein the inorganic salt is selected from sodium, chloride, potassium chloride, and ammonium chloride.

7. The method according to claim 1 wherein the buffered solution in step b) comprises 20 mM to 30 mM Tris, 1050 mM to 1350 mM sodium chloride, and about 20% (w/v) ethylene glycol at a pH value of from pH 7.1 to pH 7.3.

8. The method according to claim 2 , wherein the buffered solution in step a-1) comprises 20 mM to 30 mM Tris, and 80 mM to 120 mM sodium chloride at a pH value of from pH 7.1 to pH 7.3.

9. The method according to claim 4 , wherein the buffered solution in step a-b1) comprises 20 mM to 30 mM Tris, and 80 mM to 120 mM sodium chloride at a pH value of from pH 7.1 to pH 7.3.

10. The method according to claim 4 wherein the buffered solution in step a-b2) comprises 110 mM to 140 mM Tris, 80 mM to 120 mM sodium chloride, and 30 mM to 40 mM arginine at a pH value of from pH 7.1 to pH 7.3.

11. The method according to claim 1 wherein the multimodal weak cation exchange chromatography material comprises cross-linked agarose to which a carboxylic acid, ether, thioether and aromatic functional group containing multimodal weak cation exchange ligand is covalently attached.

Assignees

• Falkenstein Roberto
• Lemm Thorsten
• Strasser Markus
• Yamada Hidenari
• Hoffmann La Roche

Inventors

• Falkenstein Roberto
• Lemm Thorsten
• Strasser Markus
• Yamada Hidenari

Classifications

• C07K16/065 Primary

  Purification, fragmentation · CPC title

• C07K1/18 Primary

  Ion-exchange chromatography · CPC title

• C07K16/2863

  against receptors for growth factors, growth regulators · CPC title

• C07K16/28 Primary

  against receptors, cell surface antigens or cell surface determinants · CPC title

• C07K16/06

  from serum · CPC title