Compounded media powder formulation and method of preparation of liquid medium for cell culture

Patent metadata

Field	Value
Publication number	US-2017191025-A1
Application number	US-201515302488-A
Country	US
Kind code	A1
Filing date	Apr 7, 2015
Priority date	Apr 10, 2014
Publication date	Jul 6, 2017
Grant date	—

Abstract

The present invention provides a compounded cell culture medium powder formulation comprising: a basal medium powder and a cell culture media supplement, wherein the cell culture media supplement comprises and one or more salts; one or more growth factors; one or more inorganic ions; an amino acid supplement comprising one or more of asparagine, glutamine, histidine, and serine; one or more buffers; and one or more anti-foaming agents. The invention further provides methods of making a compounded cell culture medium powder formulation methods of making a cell culture medium for growing mammalian cells and methods of producing a protein of interest by culturing cells in the cell culture medium and isolating the protein of interest.

First claim

What is claimed is:

1 . A compounded cell culture medium powder formulation comprising a basal medium powder and a cell culture media supplement, wherein the cell culture media supplement comprises
i) one or more salts; ii) one or more growth factors; iii) one or more inorganic ions; iv) an amino acid supplement comprising one or more of asparagine, glutamine, histidine, and serine; v) one or more buffers; and vi) one or more anti-foaming agents.

2 . The compounded cell culture medium powder formulation of claim 1 , wherein the basal medium powder comprises Dulbecco's Modified Eagle's Medium, Ham's Medium F12, Eagle's Minimal Essential Medium, RPMI 1640 Medium, and Dulbecco's Modified Eagle's Medium/Ham's F12 Medium (DMEM/F-12; 1:1 ratio).

3 . The compounded cell culture medium powder formulation of claim 1 , wherein the basal medium powder comprises one or more of the following components or a combination thereof: biotin, calcium chloride, choline chloride, cyanocobalamin (B12), D+ mannose, D-calcium pantothenate, dextrose (anhydrous), DL-alpha-lipoic acid, ferric nitrate 9H 2 O, ferrous sulfate 7H 2 O, folic acid, glycine, hypoxanthine, I-inositol, L-alanine, L-arginine, L-asparagine, L-aspartic acid, L-cysteine HCl H 2 O, L-cystine 2HCl, L-glutamic acid (anhydrous), L-glutamine, L-glutathione, L-histidine FB, L-histidine HCl, L-isoleucine, L-leucine, L-lysine, L-methionine, L-phenylaline, L-proline, L-serine, L-threonine, L-tryptophan, L-tyrosine, L-valine, magnesium chloride, magnesium sulfate anhydrous, niacinamide, O-phosphoryl-ethanolamine, potassium chloride, putrescine 2HCl, pyridoxal HCl, pyridoxine HCl, riboflavin, sodium chloride, sodium phosphate monobasic H 2 O, sodium phosphate dibasic anhydrous, sodium pyruvate, thiamine HCl, thymidine, zinc sulfate 7H 2 O, cupric sulfate, selenium dioxide, linoleic acid, beta-mercaptoethanol and ethanolamine free-base FB.

4 . The compounded cell culture medium powder formulation of claim 3 , wherein the basal medium powder has a concentration of 8-30 g/L upon hydration to form a cell culture medium.

5 . The compounded cell culture medium powder formulation of claim 4 , wherein the basal medium powder has a concentration of about 13 g/L upon hydration to form a cell culture medium.

6 . The compounded cell culture medium powder formulation of claims 1 - 5 further comprising a pH indicator.

7 . The compounded cell culture medium powder formulation of claim 6 , wherein the pH indicator is Phenol Red Na, wherein the Phenol Red Na is present at a concentration of about 0.001 to about 0.02 g/L upon hydration to form a cell culture medium.

8 . The compounded cell culture medium powder formulation of claim 7 , wherein the Phenol Red Na is present at a concentration of about 0.0069 g/L upon hydration to form a cell culture medium.

9 . The compounded cell culture medium powder formulation of claim 3 , wherein the basal medium powder components have the following final concentration upon hydration to form a cell culture medium.
i) 0.0003-0.003 g/L biotin; ii) 0.035-0.33 g/L calcium chloride; iii) 0.003-0.03 g/L choline chloride; iv) 0.0002-0.002 g/L cyanocobalamin (B12); v) 1-10 g/L D+ mannose; vi) 0.001-0.01 g/L D-calcium pantothenate; vii) 0.3-3.0 g/L dextrose (anhydrous); viii) 0.00003-0.0003 g/L DL-alpha-lipoic acid; ix) 0.00001-0.00015 g/L ferric nitrate; x) 0.00005-0.0015 g/L ferrous sulfate; xi) 0.001-0.01 g/L folic acid; xii) 0.007-0.20 g/L glycine; xiii) 0.001-0.01 g/L hypoxanthine 2Na; xiv) 0.005-0.05 g/L I-inositol; xv) 0.003-0.03 g/L L-alanine; xvi) 0.08-1.4 g/L L-arginine; xvii) 0.006-0.16 g/L L-asparagine; xviii) 0.005-0.10 g/L L-aspartic acid; xix) 0.005-0.05 g/L L-cysteine HCl; xx) 0.02-0.2 g/L L-cystine 2HCl; xxi) 0.005-0.15 g/L L-glutamic acid (anhydrous); xxii) 0.02-1.5 g/L L-glutamine; xxiii) 0.0003-0.003 g/L L-glutathione; xxiv) 0.02-0.2 g/L L-histidine HCl; xxv) 0.03-0.9 g/L L-isoleucine; xxvi) 0.03-0.9 g/L L-leucine; xxvii) 0.05-1.5 g/L L-lysine; xxviii) 0.01-0.3 g/L L-methionine; xxix) 0.02-0.6 g/L L-phenylaline; xxx) 0.008-0.25 g/L L-proline; xxxi) 0.009-0.25 g/L L-serine; xxxii) 0.03-0.9 g/L L-threonine; xxxiii) 0.006-0.16 g/L L-tryptophan; xxxiv) 0.02-0.9 g/L L-tyrosine 2Na; xxxv) 0.03-0.9 g/L L-valine; xxxvi) 0.01-0.18 g/L magnesium chloride; xxxvii) 0.02-0.12 g/L magnesium sulfate anhydrous; xxxviii) 0.001-0.01 g/L niacinamide; xxxix) 0.0005-0.005 g/L O-phoshphoryl-ethanolamine; xl) 0.1-1.0 g/L potassium chloride; xli) 0.00002-0.0002 g/L putrescine 2HCl; xlii) 0.001-0.01 g/L pyridoxal HCl; xliii) 0.00001-0.0001 g/L pyridoxine HCl, xliv) 0.0001-0.001 g/L riboflavin, xlv) 2.0-15 g/L sodium chloride, xlvi) 0.01-0.2 g/L sodium phosphate monobasic, xlvii) 0.02-0.2 g/L sodium phosphate dibasic anhydrous, xlviii) 0.015-0.15 g/L sodium pyruvate, xlix) 0.001-0.01 g/L thiamine HCl, l) 0.0001-0.001 g/L thymidine, li) 0.00006-0.0015 g/L zinc sulfate, lii) 0.0000003-0.000006 g/L cupric sulfate, liii) 0.0000005-0.000008 g/L selenium dioxide, liv) 0.00001-0.0001 g/L linoleic acid, lv) 0.0001-0.001 g/L beta-mercaptoethanol; and lvi) 0.0003-0.005 g/L ethanolamine.

10 . The compounded cell culture medium powder formulation of claim 9 , wherein the basal medium powder components have the following final concentration upon hydration to form a cell culture medium:
i) about 0.001 g/L biotin; ii) about 0.11665 g/L calcium chloride; iii) about 0.00998 g/L choline chloride; iv) about 0.00068 g/L cyanocobalamin (B12); v) about 3 g/L D+ mannose; vi) about 0.00312 g/L D-calcium pantothenate; vii) about 1 g/L dextrose (anhydrous); viii) about 0.000103 g/L DL-alpha-lipoic acid; ix) about 0.00005 g/L ferric nitrate 9H 2 O; x) about 0.000417 g/L ferrous sulfate 7H 2 O; xi) about 0.00366 g/L folic acid; xii) about 0.02626 g/L glycine; xiii) about 0.0027 g/L hypoxanthine 2Na; xiv) about 0.01451 g/L I-inositol; xv) about 0.01336 g/L L-alanine; xvi) about 0.27435 g/L L-arginine; xvii) about 0.0225 g/L L-asparagine; xviii) about 0.01995 g/L L-aspartic acid; xix) about 0.01756 g/L L-cysteine HCl H 2 O; xx) about 0.06256 g/L L-cystine 2HCl; xxi) about 0.02206 g/L L-glutamic acid (anhydrous); xxii) about 0.73 g/L L-glutamine; xxiii) about 0.001 g/L L-glutathione; xxiv) about 0.07348 g/L L-histidine HCl; xxv) about 0.1057 g/L L-isoleucine; xxvi) about 0.11096 g/L L-leucine; xxvii) about 0.16385 g/L L-lysine; xxviii) about 0.03224 g/L L-methionine; xxix) about 0.06748 g/L L-phenylaline; xxx) about 0.02875 g/L L-proline; xxxi) about 0.03676 g/L L-serine; xxxii) about 0.10156 g/L L-threonine; xxxiii) about 0.01902 g/L L-tryptophan; xxxiv) about 0.10771 g/L L-tyrosine 2Na 2H 2 O; xxxv) about 0.09866 g/L L-valine; xxxvi) about 0.028 g/L magnesium chloride; xxxvii) about 0.04884 g/L magnesium sulfate anhydrous; xxxviii) about 0.00302 g/L niacinamide; xxxix) about 0.0014 g/L O-phoshphoryl-ethanolamine; xl) about 0.31182 g/L potassium chloride; xli) about 0.000081 g/L putrescine 2HCl; xlii) about 0.003 g/L pyridoxal HCl; xliii) about 0.000031 g/L pyridoxine HCl, xliv) about 0.000319 g/L riboflavin, xlv) about 6.1234 g/L sodium chloride, xlvi) about 0.0625 g/L sodium phosphate monobasic H 2 O, xlvii) about 0.07099 g/L sodium phosphate dibasic anhydrous, xlviii) about 0.055 sodium pyruvate, xlix) about 0.00317 g/L thiamine HCl, l) about 0.000364 g/L thymidine, li) about 0.000432 g/L zinc sulfate 7H 2 O, lii) about 0.00000125 g/L cupric sulfate 5H 2 O, liii) about 0.00000222 g/L selenium dioxide, liv) about 0.000042 g/L linoleic acid, lv) about 0.00039065 g/L beta-mercaptoethanol; a

Assignees

• Bayer Healthcare Llc

Inventors

• Shimoni Yuval
• Möhrle Volker

Classifications

• C12N2500/22

  Zinc; Zn chelators · CPC title

• C12N5/0031

  Serum-free culture media · CPC title

• C12N2500/38

  Vitamins · CPC title

• G01N31/221

  for investigating pH value · CPC title

• C12N2501/10

  Growth factors · CPC title