Developement of serum-free media in CHO-DG44 cells using a central composite statistical design

Ananth Parampalli, Kent M Eskridge, Leonard Smith, Michael M. Meagher, Mark C. Mowry, Anuradha Subramanian

Research output: Contribution to journalArticle

37 Citations (Scopus)

Abstract

A serum free medium was developed for the production of recombinant antibody against Botulinum A (BoNTA) using dihydrofolate reductase deficient Chinese Hamster Ovary Cells (CHO-DG44) in suspension culture. An initial control basal medium was prepared, which was similar in composition to HAM's F12: IMDM (1:1) supplemented with insulin, transeferrin, selenium and a lipid mixture. The vitamin concentration of the basal medium was twice that of HAM's F12: IMDM (1:1). CHO-DG44 cells expressing S25 antibody grew from 2 × 105 cells to maximum cell density of 1.04 × 106 cells/ml after 5 days in this control medium. A central composite design was used to identify optimal levels and interaction among five groups of medium components. These five groups were glutamine, Essential Amino Acids (EAA), Non Essential Amino Acids (NEAA), Insulin, Transferrin, Selenium (ITS), and lipids. Fifty experiments were carried out in four batches, with two controls in each batch. There was little effect of ITS and Lipid concentrations over the range studied, and glutamine concentration showed a strong interaction with EAA. The optimal concentrations of the variables studied were 2.5 mM Glutamine, 7.4 mM (2×) EAA, 1.4 mM (0.5×) NEAA, 1× ITS supplement, 0.7× Lipids supplement. The maximum viable cell density attained in the optimized medium was 1.4 × 106 cells/ml, a 35% improvement over the control culture, while the final antibody titer attained was 22 ± 3.4 μg/mL, a 50% improvement.

Original languageEnglish (US)
Pages (from-to)57-68
Number of pages12
JournalCytotechnology
Volume54
Issue number1
DOIs
StatePublished - May 1 2007

Fingerprint

Essential Amino Acids
CHO Cells
Serum-Free Culture Media
Insulin
Selenium
Amino acids
Lipids
Transferrin
Glutamine
Antibodies
Composite materials
Cell Count
Tetrahydrofolate Dehydrogenase
Vitamins
Cricetulus
Antibody Formation
Ovary
Suspensions
Chemical analysis
Experiments

Keywords

  • Media optimization
  • Response surface method

ASJC Scopus subject areas

  • Biotechnology
  • Bioengineering
  • Biomedical Engineering
  • Clinical Biochemistry
  • Cell Biology

Cite this

Developement of serum-free media in CHO-DG44 cells using a central composite statistical design. / Parampalli, Ananth; Eskridge, Kent M; Smith, Leonard; Meagher, Michael M.; Mowry, Mark C.; Subramanian, Anuradha.

In: Cytotechnology, Vol. 54, No. 1, 01.05.2007, p. 57-68.

Research output: Contribution to journalArticle

Parampalli, Ananth ; Eskridge, Kent M ; Smith, Leonard ; Meagher, Michael M. ; Mowry, Mark C. ; Subramanian, Anuradha. / Developement of serum-free media in CHO-DG44 cells using a central composite statistical design. In: Cytotechnology. 2007 ; Vol. 54, No. 1. pp. 57-68.
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