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View additional product information for Vitronectin (VTN-N) Recombinant Human Protein, Truncated - FAQs (A14700, A31804)
12 product FAQs found
We recommend using Vitronectin (VTN-N) Recombinant Human Protein, Truncated (Cat. No. A14700), but Geltrex LDEV-Free hESC-qualified Reduced Growth Factor Basement Membrane Matrix (Cat. No. A1413301 or Cat. No. A1413302) can also be used.
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Yes. PSCs cryopreserved from cultures of mTeSR Medium and BD Matrigel Basement Membrane Matrix may be thawed into Gibco Essential 8 Medium and plated on VTN-N. Certain lines may benefit from thawing into the medium and substrate they were growing in at the time of cryopreservation. Then at the next passage, use EDTA to passage the cells into Gibco Essential 8 Medium and VTN-N.
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Enzymes such as dispase and collagenase do not work well with cells cultured in Gibco Essential 8 Medium on VTN-N. Use of these enzymes for passaging cells results in compromised viability and attachment.
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Cells cultured in Gibco Essential 8 Medium and VTN-N need to be passaged with EDTA.
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You should expect to see normal pluripotent stem cell (PSC) morphology. The expected morphology of PSCs is demonstrated specifically by tightly packed colonies with defined borders and a high nucleus-to-cytoplasm ratio.
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Cells cultured in other feeder-free media systems, such as mTeSR Medium with Matrigel Basement Membrane Matrix, or StemPro hESC SFM with Geltrex Matrix, can be successfully cultured in Gibco Essential 8 Medium and VTN-N. In addition, PSCs grown on feeders with KnockOut SR have also been shown to be successfully cultured in Gibco Essential 8 Medium on VTN-N. However, when changing media systems, cells must be passaged either manually, or with EDTA prior to culturing in Gibco Essential 8 Medium on VTN-N.
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Gibco Essential 8 Medium and vitronectin have been shown to support PSC growth for >50 passages without any signs of karyotypic abnormalities, and maintain the ability of PSCs to differentiate into all three germ line lineages. As published by Chen et al (http://www.ncbi.nlm.nih.gov/pubmed/21478862) in the laboratory of James Thomson, the VTN-N variant of vitronectin supports human pluripotent stem cell attachment and survival better than wild-type vitronectin when used in conjunction with Gibco Essential 8 Medium.
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Yes. VTN-N is a defined, recombinant human protein.
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Here are three major differences to be taken into consideration when culturing cells in Gibco Essential 8 Medium on Gibco Vitronectin (VTN-N) compared to other feeder-free systems:
- Cells should be typically passaged ~24 hours sooner than they would be with other feeder-free media.
- Passaging should take place when cells are at ~85% confluency. If cells are passaged when they are more than 85% confluent, the health of the cells and final cell yield may be compromised.
- Cells must be passaged in EDTA. Collagenase and dispase are not recommended.
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Since VTN-N is a defined, recombinant human protein, variability is reduced in PSC cultures compared to human plasma-derived vitronectin and standard basement membrane extracts (BMEs). In addition, compared to full-length vitronectin and other defined substrates, VTN-N helps enable economical and scalable PSC culture.
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Gibco Vitronectin (VTN-N) is a recombinant, truncated human protein corresponding to the amino acid fragment 62-478 of human vitronectin expressed in E. coli. VTN-N is purified from inclusion bodies and refolded for use as a substrate for the feeder-free culture of human PSCs (http://www.ncbi.nlm.nih.gov/pubmed/21478862). When used with Gibco Essential 8 Medium, VTN-N has been proven to maintain pluripotency and normal growth characteristics in multiple PSC lines.
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Yes. Following 2 passages on the rhLaminin-521 matrix, Versene or EDTA passaging should be used to subculture PSCs.
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