PH adjustment to improve thaw recovery of cell banks

1. A method of improving thaw recovery of cell banks comprising freezing CHO cells for banking in a freezing medium, wherein the freezing medium comprises a buffered solution and a cryoprotective agent, and wherein the freezing medium has a pH of 7.7 to 8.3 prior to freezing.

2. A method of freezing CHO cells for storage comprising freezing the CHO cells in a freezing medium, wherein the freezing medium comprises a buffered solution and a cryoprotective agent, and wherein the freezing medium has a pH of 7.7 to 8.3 prior to freezing.

3. The method of claim 1 , wherein the freezing medium has a pH of 7.8 to 8.3 prior to freezing.

4. The method of claim 1 , wherein the cells are combined with a freezing medium before or after pH adjustment.

5. The method of claim 1 , wherein the adjusted pH is a target pH or a measured pH.

6. The method of claim 5 , wherein the target pH is 7.8 to 8.3.

7. The method of claim 5 , wherein the measured pH is 7.8 to 8.3.

8. The method of claim 1 , further comprising a step of measuring an initial pH of the freezing medium containing the cells prior to adjusting pH of the freezing medium.

9. The method of claim 1 , further comprising a step of measuring the adjusted pH of the freezing medium.

10. The method of claim 9 , wherein if the measured pH of the freezing medium is below a target pH, repeating the adjusting step and measuring step until the adjusted pH of the freezing medium is 7.8 to 8.3.

11. The method of claim 1 , wherein the pH is adjusted by adding a base.

12. The method of claim 11 , wherein the base is selected from the group consisting of sodium carbonate, sodium bicarbonate, HEPES sodium salt, sodium hydroxide, and potassium hydroxide.

13. The method of claim 11 , wherein the pH of the freezing medium is adjusted by adding sodium carbonate to the freezing medium according to the following formula V Na2CO3 =0.0085V p (pH t −pH i ), wherein V Na2CO3 is a volume of 1M sodium carbonate to add to the freezing medium, V p is the volume of the freezing medium, pH t is the target pH, and pH i is the initial pH.

14. The method of claim 13 , wherein the target pH is 7.8 to 8.3.

15. The method of claim 1 , wherein the CHO cells are in a medium having a pH of about 6.2 to about 6.6 before the CHO cells are combined with a freezing medium.

16. The method of claim 1 , wherein the cryoprotective agent is DMSO, glycerol, propanediol, ethylene glycol, or a sugar.

17. The method of claim 16 , wherein cryoprotective agent is DMSO or glycerol, and the DMSO or glycerol in the freezing medium prior to freezing is at a concentration of about 5% to about 12.5% by volume.

18. The method of claim 17 , wherein the DMSO or glycerol in the freezing medium prior to freezing the cells is at a concentration of about 5% to about 10% by volume.

19. The method of claim 1 , wherein the freezing medium containing the CHO cells has a cell density of about 8% to about 28% packed cell volume (PCV) prior to freezing.

20. The method of claim 1 further comprising a step of cooling cell culture fluid during cell harvest and concentration process before the CHO cells are combined with a freezing medium.

21. The method of claim 20 , wherein the cell culture fluid is cooled to a temperature at or below about 20° C.

22. The method of claim 20 , wherein the cell culture fluid is cooled to a temperature at or below about 10° C.

23. A method of freezing CHO cells for storage comprising (a) adjusting the pH of a freezing medium containing CHO cells to a pH of 7.7 to 8.3, wherein the freezing medium comprises a buffered solution and a cryoprotective agent; and (b) freezing the CHO cells.

24. The method of claim 23 , wherein the pH is adjusted to a pH of 7.8 to 8.3.

25. The method of claim 23 , wherein the adjusted pH is a target pH or a measured pH.

26. The method of claim 25 , wherein the target pH is 7.8 to 8.3.

27. The method of claim 25 , wherein the measured pH is 7.8 to 8.3.

28. The method of claim 23 , further comprising a step of measuring an initial pH of the freezing medium containing the cells prior to adjusting pH of the freezing medium.

29. The method of claim 23 , further comprising a step of measuring the adjusted pH of the freezing medium.

30. The method of claim 29 , wherein if the measured pH of the freezing medium is below a target pH, repeating the adjusting step and measuring step until the adjusted pH of the freezing medium is 7.8 to 8.3.

31. The method of claim 23 , wherein the pH is adjusted by adding a base.

32. The method of claim 31 , wherein the base is selected from the group consisting of sodium carbonate, sodium bicarbonate, HEPES sodium salt, sodium hydroxide, and potassium hydroxide.

33. The method of claim 31 , wherein the pH of the freezing medium is adjusted by adding sodium carbonate to the freezing medium according to the following formula V Na2CO3 =0.0085V p (pH t −pH i ), wherein V Na2CO3 is a volume of 1M sodium carbonate to add to the freezing medium, V p is the volume of the freezing medium, pH t is the target pH, and pH i is the initial pH.

34. The method of claim 33 , wherein the target pH is 7.8 to 8.3.

35. The method of claim 23 , wherein the CHO cells are in a medium having a pH of about 6.2 to about 6.6 before the CHO cells are combined with a freezing medium.

36. The method of claim 23 , wherein the cryoprotective agent is DMSO, glycerol, propanediol, ethylene glycol, or a sugar.

37. The method of claim 36 , wherein the cryoprotective agent is DMSO or glycerol and the DMSO or glycerol in the freezing medium containing the CHO cells prior to freezing is at a concentration of about 5% to about 12.5% by volume.

38. The method of claim 37 , wherein the DMSO or glycerol in the freezing medium containing the CHO cells prior to freezing the cells is at a concentration of about 5% to about 10% by volume.

39. The method of claim 23 , wherein the freezing medium containing the CHO cells has a cell density of 8% to 28% packed cell volume (PCV) prior to freezing.

40. The method of claim 23 further comprising a step of cooling cell culture fluid during cell harvest and concentration process before the CHO cells are combined with a freezing medium.

41. The method of claim 40 , wherein the cell culture fluid is cooled to a temperature at or below about 20° C.

42. The method of claim 40 , wherein the cell culture fluid is cooled to a temperature at or below about 10° C.

43. The method of claim 1 , wherein the CHO cells comprise a nucleic acid encoding a polypeptide.

44. The method of claim 43 , wherein the polypeptide is a therapeutic protein.

45. The method of claim 44 , wherein the therapeutic protein is selected from the group consisting of an antibody, an antibody fragment, an enzyme, and a receptor fusion protein.