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  6. Gibco™ rhLaminin-521 

Gibco™ rhLaminin-521

Catalog No. A29249
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A29248 100μg
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Description

rhLaminin-521

Recombinant human Laminin 521 (rhLaminin-521) is a recombinant human protein that provides a defined surface for feeder-free culture of human pluripotent stem cells (PSCs). When used with Essential 8™ Medium, rhLaminin-521 has been proven to promote cellular survival under stressful conditions in the absence of small molecule inhibitors. rhLaminin-521 has been shown to support PSC growth for >20 passages without any signs of karyotypic abnormalities and to maintain the ability of PSCs to differentiate into all three germ line lineages. rhLaminin-521 also provides optimum reprogramming efficiency of human somatic cells as well as human pluripotent stem cell survival during feeder-dependent to feeder-free transitions when used in conjunction with Essential 8 Medium. For a kit that combines the medium and substrate, see the Essential 8™ Adaptation Kit.

rhLaminin-521 is:
  • Robust—transition cells between feeder-dependent and feeder-free culture systems with minimal effort
  • Efficient—easily reprogram human fibroblasts to pluripotent stem cells
  • Physiologically relevant—expand cells with minimal differentiation
  • Scalable—passage stem cell lines as single cells with no impact to genetic stability

    Trouble-free transition
    When used with Essential 8 Medium, rhLaminin-521 provides a robust culture matrix that supports pluripotent stem cell adherence and expansion during the transition from feeder-dependent culture systems.

    Efficient reprogramming
    rhLaminin-521 has been shown to increase the number of colonies obtained during reprogramming of human fibroblasts, which can be important for in vitro disease modeling.

    Confident expansion
    rhLaminin-521 is expressed in blastocysts and therefore provides an environment mimicking the human stem cell niche.

    Easy scale-up
    rhLaminin-521 promotes stable pluripotent stem cell maintenance as single cells in the absence of ROCK inhibitors.
    • TRUSTED_SUSTAINABILITY

    Specifications

    Content And Storage Store in freezer (-10 to -30°C)
    Cell Type Stem Cells
    Classification Xeno-free
    Culture Type Stem Cell (Human, iPS - Induced Pluripotent Stem, Embryonic)
    Product Type Extracellular Matrix
    pH 6 to 8
    Purity or Quality Grade hESC-Qualified
    Quantity 1 mg
    Shipping Condition Dry Ice
    Source Xeno-Free

    Frequently Asked Questions (FAQs)

    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.

    You recommend Geltrex matrix 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.

    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. A400457 or A400458), Geltrex Flex matrix (Cat. No. A4000046801, A4000046802, or A4000046803), 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.

    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. A400457 or A400458), Geltrex Flex matrix (Cat. No. A4000046801, A4000046802, or A4000046803), 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.

    What is the alternative for rhLaminin-521 (Cat. No. A29249)?

    The appropriate alternative for rhLaminin-521 (Cat. No. A29249) would be either Laminin Mouse Protein, Natural (Cat. No. 23017015) or Vitronectin (VTN-N) Recombinant Human Protein, Truncated (Cat. No. A14700).

    Can I use the Poly-D-Lysine matrix with other coatings?

    iPSC-derived neurons have been cultured successfully on Poly-D-Lysine with a secondary coating of mouse and human laminin.
    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.

    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.

    Can I use Geltrex matrix instead of rhLaminin-521 for clonal expansion workflows where cells are seeded at 1-5 cells per well of a 96-well plate?

    We have not yet evaluated the Geltrex matrix system for clonal expansion in the presence of StemFlex Medium. However, rhLaminin-521 does provide optimal survival of cells following single-cell passaging and thus this matrix is recommended for such critical applications.
    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.

    Do I need to use ROCK inhibitor during single-cell passaging in StemFlex Medium?

    The use of ROCK inhibitor is not required when culturing in StemFlex Medium on rhLaminin-521. However, supplementation with RevitaCell Supplement can provide additional support to PSCs during stressful transitions such as single-cell passaging.
    Can I use EDTA or versene for passaging of stem cells?

    Yes, you may use EDTA or versene. However, we do not recommend using dispase or collagenase as it can lead to differentiation.
    Does the recombinant human laminin-521 (rhLaminin-521) have to be on ice before use?

    We recommend that you leave the product on ice while prepping to use it.
    Do recombinant human laminin-521 (rhLaminin-521) pre-coated plates have to be covered with liquid at all times?

    Yes, do not let the laminin dry out on the plates.
    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.
    Can recombinant human laminin-521 (rhLaminin-521) be used at coating concentrations lower than 0.5 ug/cm2?

    The optimal working concentration of rhLaminin-521 is cell line dependent and must be determined empirically. However, for some cell lines, coating concentrations as low as 0.3 µg/cm2 can be used with no decrease in performance. Additionally, coating plates overnight at 4 degrees C can support coating concentrations as low as 0.1 µg/cm2.
    Can recombinant human laminin-521 (rhLaminin-521) be used with Essential 8 Flex Medium?

    Yes. To ensure optimum recovery of PSCs following single-cell passaging, PSCs should be fed with Essential 8 Flex Medium the day before passaging.

    For Research Use Only. Not for use in diagnostic procedures.

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