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Vitronectin (VTN-N) Recombinant Human Protein, truncated, 1 mL = 0.5 mg - FAQs

Vitronectin (VTN-N) Recombinant Human Protein, truncated, 1 mL = 0.5 mg - FAQs

View additional product information for Vitronectin (VTN-N) Recombinant Human Protein, Truncated - FAQs (A14700, A31804)

21 product FAQs found

What substrate should I use with the PSC Definitive Endoderm Induction Kit?

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.

Find additional tips, troubleshooting help, and resources within our Cell Culture Support Center.

Why are the recommended working concentrations different for CTS Vitronectin (VTN-N) Recombinant Human Protein, Truncated vs. Vitronectin (VTN-N) Recombinant Human Protein, Truncated? Are they not the same protein?

These two proteins are the same but have different protein concentrations due to the different methods used to measure the protein concentrations.

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I am interested in using StemFlex Medium in somatic cell reprogramming. What protocol should I follow for blood cells?

We have evaluated the use of Cytotune-iPS 2.0 Sendai Reprogramming Kit (Cat. No. A16517) for somatic cell reprogramming of CD34+ blood cells. For CD34+ cells, follow the instructions provided in the Cytotune 2.0 reprogramming manual for feeder-free reprogramming (pgs. 39-44). On Day 3, you can utilize rhVTN-N (Cat. No. A14700), Geltrex (Cat. No. A1413302), or rhLaminin-521 (Cat. No. A29248 or A29249). From Day 8 onward, rather than feeding daily with Essential 8 Medium, reprogrammed CD34+ cells should be fed every-other-day with StemFlex Medium.

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I am interested in using StemFlex Medium in somatic cell reprogramming. What protocol should I follow for human fibroblasts?

We have evaluated the use of Cytotune-iPS 2.0 Sendai Reprogramming Kit (Cat. No. A16517) for somatic cell reprogramming of both neonatal and adult human dermal fibroblasts. For fibroblasts, follow the instructions provided in the Cytotune 2.0 reprogramming manual for feeder-free reprogramming (pgs. 16-20). On Day 7, you may use rhVTN-N (Cat. No. A14700), Geltrex matrix (Cat. No. A1413302), or rhLaminin-521 (Cat. No. A29248 or A29249). From Day 8 onward, rather than feeding daily with Essential 8 Medium, we recommend that you feed reprogrammed fibroblasts every-other-day with StemFlex Medium.

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You recommend Geltrex as the matrix for routine culture of StemFlex Medium. Can I also use Vitronectin or rhLaminin-521?

Yes. We recommend following the coating instructions for Vitronectin or rhLaminin-521 and using the culture and passaging recommendations in the StemFlex protocol.

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What temperature should be used to coat vitronectin (VTN-N) on plates?

We recommed coating the plate at room temperature for 1 hour.

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How should I thaw vitronectin (VTN-N)?

This product should be thawed at room temperature for 10-15 minutes. It should not be thawed at 4 degrees C overnight which might result in poor product performance.

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How does the Vitronection (VTN-N) Recombinant Human Protein, Truncated compare to wild type vitronectin?

The truncated form of vitronectin supports human pluriportent stem cells attachment and survival better than wild-type vitronectin. This truncated form has been optimized for use with Essential 8 Medium.

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Can pluripotent stem cells (PSCs) cryopreserved in a different culture condition be thawed and subsequently cultured in Gibco Essential 8 Medium and on vitronectin (VTN-N)?

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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What substrate should I use for cardiomyocyte differentiation?

We recommend using Gibco Geltrex LDEV-Free, hESC Qualified Reduced Growth Factor Basement Membrane Matrix (Cat. No. A1413301 or A1413302) or Gibco Vitronectin (VTN-N) Recombinant Human Protein, Truncated (Cat. No. A14700) for xeno-free applications.

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Can I use enzymes such as dispase and collagenase for passaging cells cultured in Gibco Essential 8 Medium and on VTN-N?

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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What is the recommended passaging method to use with Gibco Essential 8 Medium and on Gibco Vitronectin (VTN-N)?

Cells cultured in Gibco Essential 8 Medium and VTN-N need to be passaged with EDTA.

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What will my cells look like growing in Gibco Essential 8 Medium and on Gibco Vitronectin (VTN-N)?

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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Can pluripotent stem cells (PSCs) previously cultured in other media and on other substrates be cultured in Gibco Essential 8 Medium and on Gibco Vitronectin?

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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Can Gibco Essential 8 Medium and Gibco Vitronectin (VTN-N) support long-term growth of pluripotent stem cells (PSCs)?

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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Is Gibco Vitronectin (VTN-N) xeno-free (human or recombinant origin only)?

Yes. VTN-N is a defined, recombinant human protein.

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What are the main 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?

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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How does Gibco Vitronectin (VTN-N) compare to other basement membrane extracts?

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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What is Gibco Vitronectin (VTN-N)?

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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Can PSCs transitioned over into Essential 8 Medium on recombinant human laminin-521 (rhLaminin-521) from feeder-dependent cultures, then be transferred onto vitronectin (VTN-N)?

Yes. Following 2 passages on the rhLaminin-521 matrix, Versene or EDTA passaging should be used to subculture PSCs.

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Do you still recommend using Gibco Vitronectin as the matrix for Gibco Essential 8 Flex Medium?

Both Gibco Vitronectin (VTN-N) Recombinant Human Protein, Truncated (Cat. No. A14700) and Gibco Geltrex LDEV-Free, hESC Qualified Reduced Growth Factor Basement Membrane Matrix (Can. No A1413301) have been successfully used with Essential 8 Flex Medium.

Find additional tips, troubleshooting help, and resources within our Cell Culture Support Center.