Methods of Improving Yield in Recombinant Protein Production
OwnerJackie Shek Kei Chan, Brett Harold Jorgensen +1inventors
InventorsJackie Shek Kei Chan, Brett Harold Jorgensen, John Brian Mumm
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Abstract
Methods of enhancing production of cytokines such as IL-10 by, for example, optimizing refolding conditions, are described. The methods provide an efficient, cost-effective means of manufacturing IL-10 on a commercial scale.
Claims36
A method of generating refolded Interleukin-10 (IL-10), comprising:
- (a) obtaining a mixture comprising unfolded IL-10 monomers, and (b) contacting the mixture with a refold buffer to produce an admixture comprising refolded IL-10;
- wherein the concentration of unfolded IL-10 monomers in the refold buffer is 0.05 g/mL to 0.3 g/mL.
The method of any one of the preceding claims, wherein the mixture is produced by combining a plurality of inclusion bodies comprising IL-10 with a suspension buffer.
The method of any one of the preceding claims, further comprising adding a redox system to the refold buffer.
The method of any one of the preceding claims, wherein at least one naturally occurring or non-naturally occurring amino acid is added to the refold buffer.
The method of any one of the preceding claims, wherein a wash clarification is performed on the mixture prior to step (b).
The method of any one of the preceding claims, wherein an ultrafiltration/diafiltration (UFDF) is performed on the admixture.
The method of any one of the preceding claims, wherein the pH of the refold buffer is about pH 8.3.
An IL-10 refold buffer, comprising:
- (a) a mixture comprising unfolded IL-10 monomers in a concentration of from 0.05 g/mL to 0.3 g/mL;
- and
- (b) arginine in a molarity of from 0.005 to 0.3 M.
Dependencies are inferred from the claim text.