Methods of purifying Fc region containing proteins
First Claim
1. A method for purifying a protein having an Fc region from a source liquid comprising the protein and one or more impurities, comprising the steps of:
- adsorbing the protein to an Fc binding agent;
washing the Fc binding agent bound to the protein with a buffer solution containing CaCl2 at a concentration from about 0.5 M to about 3 M to reduce the one or more impurities; and
recovering the protein from the Fc binding agent in an elution solution, wherein the protein having an Fc region is purified from the source liquid.
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Abstract
The present application provides methods of purifying polypeptides having a Fc region, for example, antibodies or antibody fusions, by adsorbing the polypeptides to a Fc binding agent, such as, for example, Protein A or Protein G, followed by a wash with a divalent cation salt buffer to remove impurities and subsequent recovery of the adsorbed polypeptides. The present application also features methods of eluting the purified polypeptide as well as the incorporation of the methods within a purification train. Kits comprising components for carrying out the methods and instructions for use are also provided.
123 Citations
86 Claims
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1. A method for purifying a protein having an Fc region from a source liquid comprising the protein and one or more impurities, comprising the steps of:
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adsorbing the protein to an Fc binding agent; washing the Fc binding agent bound to the protein with a buffer solution containing CaCl2 at a concentration from about 0.5 M to about 3 M to reduce the one or more impurities; and recovering the protein from the Fc binding agent in an elution solution, wherein the protein having an Fc region is purified from the source liquid. - View Dependent Claims (2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41)
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42. A method for purifying a protein having an Fc region from a source liquid comprising the protein and one or more impurities, wherein the one or more impurities comprise one or more intron read through variant species (IRT), the method comprising the steps of:
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adsorbing the protein having an Fc region to an affinity ligand which is an Fc binding agent; washing the affinity ligand with a buffer solution containing CaCl2 at a concentration from about 0.5 M to about 3 M to reduce the IRT; and recovering the protein having an Fc region from the affinity ligand in an elution solution, wherein the protein having an Fc region is purified from the source liquid. - View Dependent Claims (43, 44, 45, 46, 47, 48, 49, 50, 51, 52, 53, 54, 55, 56, 57, 58, 59, 60)
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61. A method for purifying a protein having an Fc region from a source liquid comprising the protein and one or more impurities, wherein the protein having an Fc region is an antibody, the method comprising the steps of:
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adsorbing the protein having an Fc region to an affinity ligand which is an Fc binding agent; washing the affinity ligand with a buffer solution containing CaCl2 at a concentration from about 0.5 M to about 3 M to reduce the one or more impurities; and recovering the protein having an Fc region from the affinity ligand in an elution solution, wherein the protein having an Fc region is purified from the source liquid. - View Dependent Claims (62, 63, 64, 65, 66, 67, 68, 69, 70, 71, 72, 73, 74, 75, 76, 77, 78, 79, 80, 81, 82, 83, 84, 85, 86)
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Specification