Feed media

Abstract

The invention provides stable feed media containing pyruvate and methods for stabilizing feed media by adding pyruvate. The invention further provides methods for producing proteins using such media and proteins produced through the use of such methods.

What is claimed is: 
     
         1 . A method for stabilizing a concentrated feed medium for feeding a mammalian cell culture comprising including in the feed medium at least about 9 mM pyruvate,
 wherein the feed medium comprises cysteine and/or cystine, wherein the sum of the concentrations of cysteine and/or cystine is at least about 7.9 mM,   wherein the pH of the feed medium is from about 5.8 to about 7.4,   wherein the feed medium comprises tyrosine and the tyrosine concentration is not more than about 4.4 mM, and   wherein the feed medium with the included pyruvate is stable for at least about 1 week at room temperature.   
     
     
         2 . The method of  claim 1 , comprising including in the feed medium at least about 18 mM pyruvate. 
     
     
         3 . The method of  claim 2 , comprising including in the feed medium at least about 30 mM pyruvate. 
     
     
         4 . The method of  claim 1 , wherein the pH of the feed medium is from about 6.0 to about 7.2. 
     
     
         5 . The method of  claim 1 , wherein the feed medium comprises at least about 6.0 mM cysteine. 
     
     
         6 . The method of  claim 5 , wherein the feed medium comprises at least about 12.0 mM cysteine. 
     
     
         7 . The method of  claim 6 , wherein the feed medium comprises not more than about 40 mM cysteine. 
     
     
         8 . The method of  claim 1 , wherein the feed medium comprises at least about 3 mM tyrosine. 
     
     
         9 . The method of  claim 8 , wherein the feed medium comprises at least about 4 mM tyrosine. 
     
     
         10 . The method of  claim 1 , wherein the feed medium is serum free. 
     
     
         11 . The method of  claim 1 , wherein the feed medium comprises a protein hydrolysate. 
     
     
         12 . The method of  claim 1 , wherein the feed medium has an osmolarity from about 200 mOsm to about 1000 mOsm. 
     
     
         13 . The method of  claim 12 , wherein the feed medium has an osmolarity from about 500 mOsm to about 1000 mOsm. 
     
     
         14 . The method of  claim 1 , wherein the feed medium comprises at least about 0.5 mM cystine. 
     
     
         15 . The method of  claim 14 , wherein the feed medium comprises at least about 1.0 mM cystine. 
     
     
         16 . The method of  claim 4 ,
 wherein at least about 30 to about 40 mM sodium pyruvate is included in the feed medium,   wherein the feed medium comprises from about 3 mM to about 4 mM tyrosine,   wherein the feed medium is stable for at least about 2 weeks at room temperature,   wherein the feed medium does not comprise cystine, and   wherein the feed medium comprises at least about 12 mM cysteine.   
     
     
         17 . A feed medium for a mammalian cell culture comprising at least about 9 mM pyruvate and at least about 5 mM cysteine,
 wherein the feed medium may or may not comprise cystine,   wherein the sum of the concentrations of cysteine and cystine is at least about 7.9 mM,   wherein the pH of the feed medium is from about 5.8 to about 7.4,   wherein the feed medium comprises tyrosine and the tyrosine concentration is not more than about 4.4 mM, and   wherein the medium is stable for at least about 1 week at room temperature.   
     
     
         18 . The feed medium of  claim 17 , wherein the pH of the feed medium is from about 6.0 to about 7.2. 
     
     
         19 . The feed medium of  claim 17 , comprising at least about 18 mM pyruvate. 
     
     
         20 . The feed medium of  claim 19 , comprising at least about 30 mM pyruvate. 
     
     
         21 . The feed medium of  claim 17 , comprising at least about 12.0 mM cysteine. 
     
     
         22 . The feed medium of  claim 21 , comprising at least about 16.0 mM cysteine. 
     
     
         23 . The feed medium of  claim 17 , wherein the feed medium comprises not more than about 40 mM cysteine. 
     
     
         24 . The feed medium of  claim 17 , comprising at least about 3 mM tyrosine. 
     
     
         25 . The feed medium of  claim 24 , comprising at least about 4 mM tyrosine. 
     
     
         26 . The feed medium of  claim 17 , wherein the feed medium is serum free. 
     
     
         27 . The feed medium of  claim 17 , comprising a protein hydrolysate. 
     
     
         28 . The feed medium of  claim 17 , wherein the feed medium is chemically defined. 
     
     
         29 . The feed medium of  claim 17 , wherein the feed medium has an osmolarity from about 200 mOsm to about 1000 mOsm. 
     
     
         30 . The feed medium of  claim 29 , wherein the feed medium has an osmolarity from about 500 mOsm to about 1000 mOsm. 
     
     
         31 . The feed medium of  claim 17 , wherein the mammalian cell culture is a CHO cell culture, and wherein the pyruvate is sodium pyruvate. 
     
     
         32 . The feed medium of  claim 17 , comprising at least about 0.5 mM cystine. 
     
     
         33 . The feed medium of  claim 32 , comprising at least about 1.0 mM cystine. 
     
     
         34 . The feed medium of  claim 17 , comprising
 (a) from about 3 mM to about 4 mM tyrosine,   (b) from about 30 mM to about 40 mM sodium pyruvate, and   (c) at least about 12 mM cysteine and not more than about 20 mM cysteine,   wherein the feed medium is stable for at least about 2 weeks at room temperature,   wherein the pH of the feed medium is from about 6.0 to about 7.4, and   wherein the feed medium does not comprise cystine.   
     
     
         35 . A method for producing a protein comprising
 culturing mammalian cells that produce the protein in a base medium,   feeding the culture with the feed medium of  claim 17 , and   recovering the protein from the culture medium.