Geltrex™ LDEV-Free Reduced Growth Factor Basement Membrane Matrix, 1 mL - FAQs

What growth factors are reduced in the Geltrex LDEV-Free Reduced Growth Factor Basement Membrane Matrix (Cat. No. A1413202)?

Although the standard basement membrane extract (BME) contains growth factors such as EGF, IGF‐1, PDGF and TGF‐beta, the reduced growth factor basement membrane extract product (Geltrex matrix) contains below detectable levels of EGF, PDGF and NGF. It still has some TGF‐beta (about 2 ng/mL) and 3 times less IGF‐1 on average than regular BME, hence the designation "reduced". Neither standard nor reduced BME contain NGF or FGF‐2.

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What volume do you recommend for thin coating 35 mm dishes with Geltrex LDEV-Free Reduced Growth Factor Basement Membrane Matrix (Cat. No. A1413201, A1413202) or Geltrex LDEV-Free, hESC-Qualified, Reduced Growth Factor Basement Membrane Matrix (Cat. No. A1413301, A1413302)?

We recommend thin (non-gelling) coating 35 mm dishes with 1.5 mL of Geltrex LDEV-Free Reduced Growth Factor Basement Membrane Matrix (Cat. No. A1413201, A1413202) or Geltrex LDEV-Free, hESC-Qualified, Reduced Growth Factor Basement Membrane Matrix (Cat. No. A1413301, A1413302) prediluted 1/100 in DMEM/F-12 medium (1% final concentration).

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Can I refreeze and reuse Geltrex LDEV-Free Reduced Growth Factor Basement Membrane Matrix after having used it in an experimental setup?

Yes, you can refreeze and reuse Geltrex LDEV-Free Reduced Growth Factor Basement Membrane Matrix. However, you should avoid multiple freeze/thaw cycles. hence, we would recommend the following:

1. Thaw Geltrex Matrix in a refrigerator overnight at 2-8 degrees C.
2. Mix Geltrex Matrix solution by slowly pipetting up and down. Be careful not to introduce air bubbles.
3. If not using up the complete solution, prepare multiple aliquots, either of the original solution or the diluted one, and freeze.

How do I characterize human embryonic stem ( ES) cells?

Human ES cells are generally characterized by their typical morphology (they grow as tightly packed clusters of small cells with high ratio of nucleus to cytoplasm); surface marker expression; RT-PCR detection of stem cell-specific gene expression (such as Oct3/4, Sox2, and Nanog); alkaline phosphatase staining, and telomerase activity assay. The most commonly used ES specific surface markers include stage-specific embryonic antigens SSEA-3 and SSEA-4 for human ES cells. Other ES-specific surface antigens also include TRA-1-60 and TRA-1-81. (Science 282:1145 (1998).

How are human embryonic stem (ES) cells derived?

Human ES cells are derived from human blastocyst inner cell masses, isolated by immunosurgery with rabbit antiserum to BeWO cells (a human trophoblast cell line) (Science 282:1145 (1998)).

What are ES cells?

Embryonic stem (ES) cells are derived from the early mammalian embryo and are capable of unlimited, undifferentiated proliferation in vitro while maintaining their potential to differentiate into a wide of range of adult tissues including germ cells. The pluripotency of the ES cells is normally demonstrated in vitro by inducing ES cells to differentiate into embryoid bodies and checking lineage-specific markers for differentiated cells in three body layers (endo, meso, and ectoderm), or injecting them into immunodeficient mice and determining the cell types produced in the teratomas.

Can differentiation of cells occur when using Gibco Geltrex Matrix? Can this be prevented?

Yes, try a variety of dilutions to coat the plates. Some customers use a 1:100 dilution and have better results. The optimal dilution is cell-line dependent.

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Why can Gibco Geltrex Matrix be used for adhesion-related research, as opposed to Gibco AlgiMatrix 3D Culture System which cannot?

Geltrex Matrix is a basement membrane extract that contains collagen IV, laminin, and other matrix proteins and growth factors that are required for cell adhesion. AlgiMatrix Matrix contains only alginate which does not facilitate cell adhesion. There is nothing in the alginate sponges for the cells to interact with.

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Is it normal to see floating debris when pre-coating with Ready-to-Use Gibco Geltrex Matrix?

Yes, this is normal. It has been observed in product development. Remove the Gibco Geltrex Matrix solution after pre-coating and add cells as normal. H9 human embryonic stem cells have been cultured on dishes pre-coated with Ready-to-Use Gibco Geltrex Matrix with this observation in the pre-coating procedure.

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What is LDEV?

Lactate dehydrogenase-elevating virus (LDEV) infects mouse macrophages and induces persistent infections in laboratory mice. LDEV, an RNA virus, alters in vivo physiology, and is a common contaminant of biological materials derived from transplantable tumor cell lines.

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Can cells be removed from the Gibco Geltrex Matrix?

If you would like to culture the cells once recovered from the matrix, we recommend washing the cells first, then using trypsin/EDTA, TrypLE reagent, or a solution of 3,000 U/mL collagenase and 1,000 U hyaluronidase in low-glucose DMEM. If you would like to monitor cell surface markers, we recommend removal of the cells by mechanical means, such as a vigorous wash or a delicate scrape.

Note that the phenotype of the cells may change if they are cultured in 2D after their removal from the matrix.

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Has Gibco Geltrex Matrix been tested with StemPRO hESC serum-free Medium?

Yes, these products were designed to work together.

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What is Gibco Geltrex Matrix?

Gibco Geltrex Matrix is a soluble form of reduced growth factor (RGF) basement membrane extract (BME) purified from murine Engelbreth-Holm-Swarm tumor. Historically these preparations have contained the lactate dehydrogenase-elevating virus (LDEV), which does not impact the performance of the material but may be of concern in certain applications such as xenografts or other clinically focused research. New manufacturing methods are now available that allow for the removal of this virus. The resulting LDEV-free material is tested using PCR methods to confirm that the material is virus-free.

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What are some general differences between Gibco Geltrex Matrix, collagen, AlgiMatrix 3D Culture System, and CTS Cellstart Substrate?

Gibco Geltrex Matrix and collagen (rat tail and bovine) can be used as either a coating solution or a 3D gel matrix. CELLstart Substrate was developed to be used as a xeno-free coating matrix for only ESC applications (as a substitute for Gibco Geltrex Matrix or Matrigel Matrix, which are of animal origin). AlgiMatrix Matrix is a 3D scaffold-type matrix that does not support cell attachment but does provide a good environment for growing spheroids that can be easily harvested for downstream applications.

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