CELL CULTURE PROCESSES
Culturing heterologous protein-secreting mammalian cells, such as CHO or BHK cells, at 35.1-36.5° C. and/or at pH 7.15-7.20 and/or at a dissolved CO 2 concentration of 10% or lower. Preferred heterologous proteins are Factor VIII, ADAMTS-13, furin or Factor VII.
1 - 51 . (canceled)
52 . A method comprising:
expressing a heterologous protein in a mammalian cell in continuous cell culture, wherein the continuous cell culture has a dissolved carbon dioxide concentration of from 1% to 10% and is maintained at a cell culture density of from 1.4×10 6 to 2.8×10 6 cells/ml.
53 . The method of claim 52 , wherein the continuous cell culture has a dissolved carbon dioxide concentration of from 4% to 9%.
54 . The method of claim 52 , wherein the continuous cell culture has a dissolved carbon dioxide concentration of from 5.5% to 8.5%.
55 . The method of claim 52 , wherein the continuous cell culture is maintained at a temperature of from 34.2° C. to 36.9° C.
56 . The method of claim 52 , wherein the continuous cell culture is maintained at a temperature of 36±0.9° C.
57 . The method of claim 54 , wherein the continuous cell culture is maintained at a temperature of 36±0.9° C.
58 . The method of claim 52 , wherein the continuous cell culture is maintained at a pH of from 7.10 to 7.25.
59 . The method of claim 52 , wherein the continuous cell culture is maintained at a pH of 7.20±0.05.
60 . The method of claim 54 , wherein the continuous cell culture is maintained at a pH of 7.20±0.05.
61 . The method of claim 56 , wherein the continuous cell culture is maintained at a pH of 7.20±0.05.
62 . The method of claim 57 , wherein the continuous cell culture is maintained at a pH of 7.20±0.05.
63 . The method of claim 52 , wherein the continuous cell culture is buffered with bicarbonate.
64 . The method of claim 52 , wherein the dissolved CO 2 concentration of the continuous cell culture is maintained by sparging the continuous mammalian cell culture with air.
65 . The method of claim 52 , wherein the heterologous protein is a blood protein.
66 . The method of claim 52 , wherein the heterologous protein is Factor VIII.
67 . The method of claim 66 , wherein the Factor VIII is co-expressed with von Willebrand Factor.
68 . The method of claim 52 , wherein the heterologous protein is ADAMTS-13, Furin, or Factor VII.
69 . The method of claim 52 , wherein the continuous cell culture comprises copper at a concentration of at least 5 parts per billion (ppb).
70 . The method of claim 52 , wherein the continuous cell culture comprises copper at a concentration of at least 7 parts per billion (ppb).
71 . The method of claim 52 , wherein the mammalian cell is a rodent cell.
72 . The method of claim 52 , wherein the mammalian cell is a CHO cell.
73 . The method of claim 52 , wherein the mammalian cell is a BHK cell.
74 . The method of claim 52 , wherein the continuous cell culture is operated under chemostatic conditions.
75 . The method of claim 52 , wherein the continuous cell culture is operated under turbostatic conditions.