Chemistry: molecular biology and microbiology – Animal cell – per se ; composition thereof; process of... – Culture medium – per se
Reexamination Certificate
2001-06-21
2003-12-30
Naff, David M. (Department: 1651)
Chemistry: molecular biology and microbiology
Animal cell, per se ; composition thereof; process of...
Culture medium, per se
C435S260000, C435S348000
Reexamination Certificate
active
06670184
ABSTRACT:
FIELD OF THE INVENTION
The present invention relates to fermentation and cell culture, and more particularly, to cell culture media and concentrated lipid emulsions for use therein.
BACKGROUND OF THE INVENTION
Many recent developments in biotechnology, such as the industrial production of viral insecticides, recombinant products and the like via insect cell lines, require in vitro cell culture on a large scale, which in turn demands considerable amounts of cell culture media. Unfortunately, the serum and serum albumin found in conventional cell culture media is problematic, and can be cost-prohibitive for large-scale tank fermentation. The serum-free alternatives described thus far in the prior art are also costly in large volumes.
The major problem with most commercially-available media is the need to supplement the media with a large serum component (typically 5-20%), which creates a significant limiting factor due to the high price and limited availability of the serum. Moreover, the use of animal serum and/or serum albumin in the media is also problematic from a production standpoint, since the presence of unidentified proteins in the sera can complicate downstream efforts at product purification and contaminating animal viruses can pose serious safety issues. Similarly, the unidentified proteins found in these materials introduce an unwanted variable into smaller-scale experimental efforts as well. Further, the quality of the sera itself can vary from lot to lot, introducing a contamination risk which must be investigated and resolved by the scientist or production engineer with each change in sera.
Another related problem with the use of serum and/or serum albumin in cell culture media involves their conventional role as the carrier for the lipid component, an essential requirement for most cell culture work. Since the direct addition of lipids to the media is impractical due to their low solubility, they are typically introduced along with the serum component in the form of water soluble lipoproteins. Alternatively, the lipids can be bound to the albumin component and then added to media. Given the difficulties noted above with respect to the use of these materials, however, it would be advantageous to provide a suitable lipid emulsion which does not require either serum or albumin. Iscove, “Culture of Lymphocytes and Hematopoietic Cells in Serum-Free Medium,” in Barnes et al.,
Methods for Serum
-
Free Culture of Neuronal and Lymphoid Cells
, pp. 169-85 (1984). Although microemulsions have been described in the art to supply the necessary lipids for conventional insect serum-free media, Maoiella et al,
Bio/Technology,
6:1406 (1988), these prior art emulsions have proven unsatisfactory for a variety of reasons, including high manufacturing costs and short-term stability.
What is needed, therefore, is a cell culture media which provides the essential nutritional, biological and biophysical requirements needed for cell growth at substantially less cost. For large-scale tank fermentation, an ideal culture media would provide superior performance in cell growth and maximum cell density, and still be easy to prepare from a relatively small number of low-cost ingredients. Preferably, the media should be serum-free. Also needed is a concentrated lipid emulsion which can supply critical lipids in a bioavailable form, but eliminates the need for lipid carriers such as serum, serum albumin, or other proteins from culture media.
SUMMARY OF THE INVENTION
The present invention solves the problems in the prior art through the provision of a low-cost, serum-free cell culture media, comprising a modified basal media having a novel formulation of free amino acids and vitamins, a peptone component which substitutes for the traditional serum component, and a lipid emulsion component.
REFERENCES:
patent: 5024947 (1991-06-01), Inlow et al.
patent: 5372943 (1994-12-01), Inlow et al.
Vaughn et al. “Differential Requirements of Two Insect Cell Lines for Growth in Serum-Free Medium” In Vitro Cell. Dev. Biol. vol. 33 (1997) pp. 479-482.
Donaldson et al. “Low-Cost Serum-Free Medium for the BTI-Tn5B1-4 Insect Cell Line” Biotechnology Prog. vol. 14 (1998) pp. 573-579.
Hink “A Serum-Free Medium for the Culture of Insect Cells and Production of Recombinant Proteins” In Vito Cell Dev. Biol. vol. 27A (1991) pp. 397-401.
Chiarello Ronald H.
Himmerich Sarah
BASF AG
Licata & Tyrrell P.C.
Naff David M.
Ware Deborah K.
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