Product FAQ

My real-time PCR run stopped before it completed. What are the possible causes?

Answer

1) Power outage and/or power fluctuation. It is recommended that your instrument be equipped with an uninterruptible power supply (UPS). Several companies manufacture such systems for scientific equipment, and they are listed in FAQ Answer ID 1935.

2) Launching other programs, setting up another run or analyzing the data while a run is in progress. The real-time data collection consumes a large amount of hardware resources. Other applications may interfere with the run and disrupt data collection causing the software to crash.

3) Hard drive fragmentation. When the SDS and other programs run, many small program files are automatically generated and deleted in the hard drive, creating blank areas between files. This can slow down your system performance by as much as 50% since programs have to access more areas of the hard drive to run. It is recommended to defragment the hard drive once a month. Please see FAQ Answer ID 2713 for more information about how to defragment a hard drive.

Answer Id: E2716

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Product FAQ

How do I prepare Gibco Essential 8 Medium?

Answer

To prepare 500 mL of complete Gibco Essential 8 Medium, thaw Gibco Essential 8 Supplement (50X) at room temperature for 1 hour or 2-8 degrees C overnight and then aseptically combine the components listed below:

- Gibco Essential 8 Basal Medium: 490 mL
- Gibco Essential 8 Supplement (50X): 10 mL

Answer Id: E12429

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Product FAQ

Can Rho-associated protein kinase (ROCK) inhibitors be used in Gibco Essential 8 Medium?

Answer

Yes. However, this isn't necessary, and we do not routinely use these inhibitors in house. If the use of a Rho-associated protein kinase (ROCK) inhibitor is desired, you should add the inhibitor only to the medium at passage. Inhibitors should not be present for routine feeding. Use of inhibitors is assay dependent and not required for routine cell culture.

Answer Id: E12434

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Product FAQ

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?

Answer

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.

Answer Id: E12422

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Product FAQ

What is Gibco Essential 8 Medium?

Answer

Gibco Essential 8 Medium is a xeno-free and feeder-free medium specially formulated for the growth and expansion of human pluripotent stem cells (PSCs). Originally developed by Chen et al (http://www.ncbi.nlm.nih.gov/pubmed/21478862) in the laboratory of James Thomson, and validated by Cellular Dynamics International, Gibco Essential 8 Medium has been extensively tested and is proven to maintain pluripotency in multiple PSC lines.

Answer Id: E12417

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Product FAQ

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

Answer

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.

Answer Id: E12425

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Product FAQ

Can I use other versions of DMEM/F-12 in place of the Gibco Essential 8 Basal Medium supplied with the kit?

Answer

Other catalog versions of DMEM/F-12 cannot be used in place of the Gibco Essential 8 Basal Medium in the preparation of Gibco Essential 8 Medium. Gibco Essential 8 Basal Medium supplied with the kit has a higher level of sodium bicarbonate.

Answer Id: E12430

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Product FAQ

What is the role of a Rho-associated protein kinase (ROCK) inhibitor or blebbistatin?

Answer

The inclusion of either a ROCK inhibitor (HA100 or Y27632) or blebbistatin improves initial survival and supports a high cloning efficiency, which is increased by the addition of transferrin and selenium. If cells are cultured routinely in medium containing a ROCK inhibitor, it may become necessary to include it for routine culture.

Answer Id: E12435

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Product FAQ

Is Gibco Essential 8 Medium xeno-free (human or recombinant origin only)?

Answer

Yes. Gibco Essential 8 Medium provides reliable and robust cultures with a xeno-free, eight-component medium

Answer Id: E12423

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Product FAQ

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

Answer

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.

Answer Id: E12428

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Product FAQ

Can I warm Gibco Essential 8 Medium in a 37 degrees C waterbath for daily use?

Answer

It is very important that complete Gibco Essential 8 Medium is prewarmed at room temperature and not in a 37 degrees C water bath. bFGF activity can decline rapidly with repeated temperature changes from 4 degrees C to 37 degrees C.

Answer Id: E12433

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Product FAQ

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

Answer

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.

Answer Id: E12438

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Product FAQ

Does Gibco Essential 8 Medium already contain bFGF?

Answer

Yes. Gibco Essential 8 Medium contains 100 ng/mL basic fibroblast growth factor (bFGF), and no additional bFGF is required.

Answer Id: E12419

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Product FAQ

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?

Answer

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.

Answer Id: E12427

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Product FAQ

What is the shelf life of complete Gibco Essential 8 Medium?

Answer

The shelf life of complete Gibco Essential 8 Medium is two weeks at 2-8 degrees C.

Answer Id: E12432

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