Protein a chromatography

Patent metadata

Field	Value
Publication number	US-2016193633-A1
Application number	US-201414912440-A
Country	US
Kind code	A1
Filing date	Jun 19, 2014
Priority date	Sep 4, 2013
Publication date	Jul 7, 2016
Grant date	—

Abstract

The present invention provides methods for cleaning a Protein A chromatography column employing a media comprising a Protein A ligand derived from the C domain of Staphylococcus aureus , such that the column can be cleaned using both acidic and alkaline solutions.

First claim

What is claimed is:

1 . A method of preserving the binding capacity of an affinity chromatography column over one or more affinity purification cycles, the method comprising cleaning the chromatography column after one or more affinity purification cycles with an acidic solution having a pH lower than 3.0, wherein the affinity chromatography column comprises a Protein A media comprising a Protein A ligand derived from the C domain of Staphylococcus aureus Protein A immobilized onto a solid support comprising a polymer selected from the group consisting of polyvinylether, polyvinylalcohol, polymethacrylate, polyacrylate, polystyrene, polyacrylamide, polymethacrylamide and polycarbonate.

2 . The method of claim 1 , wherein the Protein A ligand comprises an amino acid sequence selected from SEQ ID NO:3 and SEQ ID NO:4.

3 . The method of claim 1 , wherein the acidic solution comprises a pH of 2.5.

4 . The method of claim 1 , wherein the acidic solution comprises a pH of 2.0.

5 . The method of claim 1 , wherein the acidic solution comprises a pH of 1.5.

6 . The method of claim 1 , wherein the binding capacity is preserved over 10 or more cycles.

7 . The method of claim 1 , wherein the binding capacity is preserved over 50 or more cycles.

8 . The method of claim 1 , wherein the binding capacity is preserved over 100 or more cycles.

9 . The method of claim 1 , wherein the binding capacity is preserved over 200 or more cycles.

10 . The method of claim 1 , wherein the solid support comprises a polyvinylether polymer.

11 . The method of claim 1 , wherein the acidic solution comprises phosphoric acid.

12 . A method of sanitizing an affinity chromatography column after a campaign while maintaining the binding capacity of the column, wherein the method comprises contacting the affinity chromatography column with a solution comprising phosphoric acid, acetic acid and benzyl alcohol for at least three hours, and wherein the affinity chromatography column comprises a Protein A media comprising a Protein A ligand derived from the C domain of Staphylococcus aureus Protein A immobilized onto a solid support comprising a polymer selected from the group consisting of polyvinylether, polyvinylalcohol polymethacrylate, polyacrylate, polystyrene, polyacrylamide, polymethacrylamide and polycarbonate.

13 . The method of claim 12 , wherein the Protein A ligand comprises an amino acid sequence selected from SEQ ID NO:3 and SEQ ID NO:4.

14 . The method of claim 12 , wherein the solid support comprises a polyvinylether polymer.

15 . A method of cleaning an affinity chromatography column using both acidic and alkaline solutions, the method comprising:
a. contacting the column with both acidic and alkaline solutions between affinity purification cycles; or b. contacting the column with either an acidic solution after a cycle or an alkaline solution after a cycle, such that the acidic and alkaline solutions are used in an alternating manner,
wherein the affinity chromatography column comprises a Protein A media comprising a Protein A ligand derived from the C domain of Staphylococcus aureus Protein A immobilized onto a solid support.

16 . The method of claim 15 , wherein the solid support comprises a polymer selected from the group consisting of polyvinylether, polyvinylalcohol, polymethacrylate, polyacrylate, polystyrene, polyacrylamide, polymethacrylamide and polycarbonate.

17 . The method of claim 15 , wherein the acidic solution comprises phosphoric acid.

18 . The method of claim 15 , wherein the alkaline solution comprises sodium hydroxide.

19 . The method of claim 15 , wherein the method results in a synergistic removal of impurities.

20 . The method of claim 15 , wherein the ligand comprises an amino acid sequence selected from SEQ ID NO:3 or SEQ ID NO:4.

Assignees

• Emd Millipore Corp

Inventors

• Bian Nanying
• Mehtani Sapna
• Hubbard John Dana

Classifications

• B08B3/08

  the liquid having chemical or dissolving effect · CPC title

• A61L2/18

  Liquid substances · CPC title

• C07K1/22 Primary

  Affinity chromatography or related techniques based upon selective absorption processes · CPC title

• B01D15/3804

  Affinity chromatography · CPC title

• B08B9/027 Primary

  Cleaning the internal surfaces; Removal of blockages · CPC title