Uniform stable-isotope labeling in mammalian cells: formulation of a cost-effective culture medium.
Uniform stable-isotope labeling of mammalian cells is achieved via a novel formulation of a serum-free cell culture medium that is based on stable-isotope-labeled autolysates and lipid extracts of various microbiological origin. Yeast autolysates allow complete replacement of individual amino acids and organic acids in a chemically defined medium (DMEM/F12), enabling a cost-effective formulation of a stable-isotope-labeled culture medium for mammalian cells. In addition, biomass-derived hydrolysates, autolysates, and lipid extracts of various classes of algae were explored as cell culture components, both separately and in combination with yeast autolysates. Optimal autolysate concentrations were established. Such novel medium formulations were tested on mammalian cell lines, often used for recombinant protein production, i.e., Chinese hamster ovary (CHO) and human embryonic kidney (HEK 293). Special attention was paid to the adaptation of these mammalian cell lines to serum-free media. Formulation of the novel proprietary cell culture medium PLIm, based on yeastolates instead of individual amino acids and organic acids, allows a four- to eightfold cost reduction for (15)N and (13)C,(15)N stable-isotope-labeling, respectively, in CHO cells and a three- to sixfold cost reduction in HEK 293 cells. A high level of stable-isotope enrichment of mammalian cells (>90%) was achieved within four passages by complete replacement of carbon and nitrogen sources in the medium with their stable-isotope-labeled analogs. These conditions can be used to more cost-effectively produce labeled recombinant proteins in mammalian cells.
Protein Labelling Innovation (PLI), BioScience Park, Archimedesweg 27, 2333 CM, Leiden, The Netherlands. proteinlabelling@zonnet.nl
Tatiana A
TA
Egorova-Zachernyuk
Giel J C G M
GJ
Bosman
Willem J
WJ
Degrip
D000818:Animals; D016466:CHO Cells; D002247:Carbon Isotopes / Q000032:analysis / Q000378:metabolism; D018929:Cell Culture Techniques / Q000191:economics / Q000295:instrumentation; D049109:Cell Proliferation; D002477:Cells / Q000737:chemistry* / Q000166:cytology / Q000378:metabolism; D006224:Cricetinae; D003412:Cricetulus; D016895:Culture Media, Serum-Free / Q000737:chemistry* / Q000191:economics* / Q000378:metabolism; D057809:HEK293 Cells; D006801:Humans; D007553:Isotope Labeling; D009587:Nitrogen Isotopes / Q000032:analysis / Q000378:metabolism
D002247:Carbon Isotopes; D016895:Culture Media, Serum-Free; D009587:Nitrogen Isotopes