Recombinant collagen for animal product-free dextran microcarriers |
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Authors: | Michael K Dame James Varani |
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Institution: | (1) Department of Pathology, The University of Michigan, 1301 Catherine Road /Box 0602, Ann Arbor, MI 48109, USA |
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Abstract: | Manufacturers of vaccines and other biologicals are under increasing pressure from regulatory agencies to develop production
methods that are completely animal-component-free. In order to comply with this demand, alternative cell culture substrates
to those now on the market, primarily collagen or gelatin, must be found. In this paper, we have tested a number of possible
substitutes including recombinant collagen, a 100-kDa recombinant gelatin fragment and a peptide derived from a cell-binding
region of type I collagen. The small 15-amino acid peptide did not support attachment of human fibroblasts in monolayer culture.
The 100-kDa gelatin fragment supported cell attachment in monolayer culture, but was significantly less active than intact
porcine gelatin. Recombinant type I collagen was as successful in promoting cell attachment as native collagen, and both were
more effective than porcine gelatin. Based on these data, dextran microspheres were treated with the same attachment proteins—porcine
gelatin, native collagen, or recombinant collagen. The same trends were observed as in monolayer culture. Concentrations of
the recombinant collagen (as well as native collagen) supported cell attachment on dextran microspheres at concentrations
as low as 0.01 μg/cm2. Treatment of the dextran with a low level of polyethylenimine, a cationic moiety, further enhanced attachment when used
in conjunction with the low concentration of recombinant collagen. Where there was increased cell attachment, increased proliferation
followed. We are confident, based on these findings, that a fully recombinant substitute could replace gelatin in current
microcarrier preparations without losing the cell growth benefits provided by the native protein. |
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Keywords: | Microcarrier Animal component-free Recombinant collagen Dextran PEI |
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