Methods of improving protein titer in cell culture

A method for selecting a defined cell culture medium for use in cell culture to improve cell culture performance, the method comprising:

• (a) providing a defined cell culture medium comprising a 4-hydroxyethyl piperazine ethanesulfonic acid (HEPES) buffer;
• (b) analyzing the defined cell culture medium comprising the HEPES buffer to determine the amount of a HEPES related impurity having a molecular weight of 267.07 and the amount of a HEPES related impurity having a molecular weight of 221.06 present in the defined cell culture medium;
• (c) selecting the defined cell culture medium comprising the HEPES buffer for use in cell culture if the defined cell culture medium comprising the HEPES buffer is determined to have less than about 4000 μmol of a HEPES related impurity having a molecular weight of 267.07 per mole of total HEPES, and less than about 400 μmol of a HEPES related impurity having a molecular weight of 221.06 per mole of total HEPES;
• wherein the use of the defined cell culture medium comprising the HEPES buffer having less than about 4000 μmol of a HEPES related impurity having a molecular weight of 267.07 per mole of total HEPES, and less than about 400 μmol of a HEPES related impurity having a molecular weight of 221.06 per mole of total HEPES improves cell culture performance, as compared to cell culture performance in non-HEPES related impurity reduced media.

A method for selecting a HEPES buffer for use in cell culture to improve cell culture performance, the method comprising:

• (a) providing a 4-hydroxyethyl piperazine ethanesulfonic acid (HEPES) buffer;
• (b) analyzing the HEPES buffer to determine the amount of a HEPES related impurity having a molecular weight of 267.07 and the amount of a HEPES related impurity having a molecular weight of 221.06 present in the HEPES buffer;
• (c) selecting the HEPES buffer for use in cell culture if the HEPES buffer is determined to have less than about 4000 μmol of a HEPES related impurity having a molecular weight of 267.07 per mole of total HEPES, and less than about 400 μmol of a HEPES related impurity having a molecular weight of 221.06 per mole of total HEPES;
• wherein the use of the HEPES buffer having less than about 4000 μmol of a HEPES related impurity having a molecular weight of 267.07 per mole of total HEPES, and less than about 400 μmol of a HEPES related impurity having a molecular weight of 221.06 per mole of total HEPES to be used in connection with the cell culture improves cell culture performance, as compared to cell culture performance in the presence of HEPES buffer having higher amounts of said impurities.

A cell culture comprising (i) at least one recombinant eukaryotic cell that can express a recombinant protein and (ii) a cell culture medium, wherein the cell culture is produced by a method comprising the steps of:

• (a) providing a defined cell culture medium having reduced impurities, the defined cell culture medium having less than about 4000 μmol of a HEPES related impurity having a molecular weight of 267.07 per mole of total HEPES, and less than about 400 μmol of a HEPES related impurity having a molecular weight of 221.06 per mole of total HEPES;
• (b) culturing said recombinant eukaryotic cells in said defined cell culture medium having reduced impurities;
• (c) expressing a recombinant protein of interest from said recombinant eukaryotic cells;
• and
• (d) producing a higher titer of the recombinant protein in the defined cell culture medium having reduced impurities relative to the that of similar or identical cells cultured in non-reduced impurity media.

A recombinant protein produced in a cell culture comprising (i) at least one recombinant eukaryotic cell that can express said recombinant protein and (ii) a cell culture medium, wherein the recombinant protein is produced by a method comprising the steps of:

• (a) providing a defined cell culture medium having reduced impurities, the defined cell culture medium having less than about 4000 μmol of a HEPES related impurity having a molecular weight of 267.07 per mole of total HEPES, and less than about 400 μmol of a HEPES related impurity having a molecular weight of 221.06 per mole of total HEPES;
• (b) culturing said recombinant eukaryotic cells in said defined cell culture medium having reduced impurities;
• (c) expressing a recombinant protein of interest from said recombinant eukaryotic cells;
• and
• (d) producing a higher titer of the recombinant protein in the defined cell culture medium having reduced impurities relative to the that of similar or identical cells cultured in non-reduced impurity media.