Role of iron and sodium citrate in animal protein‐free CHO cell culture medium on cell growth and monoclonal antibody production

Abstract

Chemically defined iron compounds were investigated for the development of animal protein-free cell culture media to support growth of CHO cells and production of monoclonal antibodies (mAb). Using a multivessel approach of 96-well plates, shake flasks, and bioreactors, we identified iron and its chemical partner citrate as critical components for maintenance of continuous cell growth and mAb production. The optimized iron concentration range was determined to be 0.1-0.5 mM and that for citrate 0.125-1 mM. This complete formulation is able to maintain cell growth to similar levels as those supplemented with iron compounds alone; however, mAb productivity was enhanced by 30-40% when citrate was present. The addition of sodium citrate (SC) did not affect product quality as determined by size exclusion chromatography, ion exchange chromatography, reversed phase and normal phase-HPLC. No significant changes in glucose and lactate profiles, amino acid utilization, or mAb heavy and light chain expression ratios were observed. Cellular ATP level was ∼30% higher when SC was included suggesting that SC may have a role in enhancing cellular energy content. When cell lysates were analyzed by LC-MS to assess the overall cellular protein profile, we identified that in the SC-containing sample, proteins involved in ribosome formation and protein folding were upregulated, and those functions in protein degradation were downregulated. Taken together, this data demonstrated that iron and citrate combination significantly enhanced mAb production without altering product quality and suggested these compounds had a role in upregulating the protein synthetic machinery to promote protein production.

Novel Cell-Ess ® supplement used as a feed or as an initial boost to CHO serum free media results in a significant increase in protein yield and production

“…The same concept is generally used as a base media today but as technology has improved so have the methods of analysis. Assays testing metabolic byproducts and investigating proteomics are used today to better optimize a media for CHO cell lines [3], [4], [5], [6], [7]. The use rate of amino acids or glucose can be determined to identify the constituents rapidly used during a run and may need replenishment, usually in the form of a feed and or feeds.…”

Rational development of a serum-free medium and fed-batch process for a GS-CHO cell line expressing recombinant antibody

“…Our results demonstrated that the iron compound FAC was an important factor for the increased antibody titer. Recently, it has been reported that FAC supplementation of the culture medium can increase monoclonal antibody production of CHO cell (Bai et al 2011). We speculate that better cell morphology and larger cell size observed under microscope contributed to the enhanced titer.…”

Establishment of Reverse Direct ELISA and Its Application in Screening High-Affinity Monoclonal Antibodies

“…Conventionally an antibody of superior quality was selected by the antibody titer, which, in spite of a high degree of dilution, yielded a positive result. (2) In this study, our results indicate some discrepancy between the titer and affinity of MAbs; some MAbs existed with a lower titer but higher affinity. This was easily missed by antibody titer determination by the traditional method.…”