Biolaminin 521 CTG

Biolaminin 521 CTG (CT521)

Human recombinant laminin 521

 

A CELL CULTURE SUBSTRATE DESIGNED FOR CLINICAL RESEARCH

Culturing human cells is about growing human cells outside the human body and recreating the natural cell niche is key to successful cultivation. Whatever your stem cell vision is, you need the right tools to succeed in your research. High-quality products and proper documentation are essential for the successful transition of basic research to cell therapy.

As a complement to our portfolio of defined and animal component-free laminin stem cell substrates, we now offer a cell therapy grade (CTG) Biolaminin 521 cell culture substrate For Research Use or Non-commercial Manufacturing of Cell, Gene, or Tissue-Based Products. Biolaminin 521 CTG (CT521) has been developed and manufactured to allow customers to qualify the material for use in the manufacturing of cells for clinical research. USP Chapter 1043: Ancillary materials for cell, gene and tissue-engineered products has been considered in the design of the product. The product is animal origin component free and has supporting documentation, such as Certificate of Analysis, Animal Origin Free Statement and Bill of Material provided with every lot to support regulatory filings.

The type of cell culture substrate used in your experiments will have a great impact on your research. The CT521 substrate only contains components of human origin and provides a defined surface for feeder-free culture of human embryonic stem cells (hESCs) and induced pluripotent stem cells (hiPSCs). The substrate can also be used for mesenchymal stem cell culture and most anchorage-dependent progenitor cell types.

 

SEAMLESS TRANSITION FROM BENCH TO CLINIC

CT521 is a full-length, human, recombinant laminin 521 substrate, the only one of its kind on the market, providing an optimal environment for feeder-free culture of human PSCs, MSCs and most anchorage-dependent progenitor cell types. With this new clinical grade product, scientists are supported throughout their cell therapy development process – from concept to commercialized therapy.

 

BIOLAMININ 521 PRODUCT SHEET

 

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Amount
Size
Price
Oty
500 ug 1 x 5 ml €315
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Estimated shipping time
1-5 working days

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SIZE GUIDE

Not sure how much laminin you need? To make it easy, we have created a tool where you can calculate the amount needed for your experiments. Just choose culture well format and fill in the desired coating concentration to see the amount required.

FORMAT
CONC. (ug/cm2).
NO OF WELLS
AMOUNT (uq)

Features and Specifications:

  • Cell therapy grade substrate

  • Designed for clinical research - USP Chapter 1043

  • Animal component–free to the secondary level

  • Manufacturing control and traceability

  • Consistent and reliable performance

  • Biologically relevant culture environment

  • Easy and flexible culture system

  • Homogenous and genetically stable hPSC cultures

  • Easily adaptable to automation platforms

  • Efficient differentiation and enhanced cell maturation, polarization and organization


CT521 cell culture application note

 

DESIGNED FOR CLINICAL RESEARCH

Biolaminin 521 CTG is the only laminin product labelled with "For Research Use or Non-Commercial Manufacturing of Cell, Gene, or Tissue-Based Products”. The product has been developed and manufactured to allow customers to qualify the material for use in the manufacturing of cells for clinical research. USP Chapter 1043: Ancillary materials for cell, gene and tissue-engineered products has been considered in the design of the product. The product has supporting documentation such as Certificate of Analysis (CoA), Animal Origin Free Statement and Bill of material provided with every lot to support regulatory filings.

 
MANUFACTURING CONTROL AND TRACEABILITY

Biolaminin 521 CTG is manufactured in compliance with ISO9001:2015 quality management system. Full traceability documentation is available for every lot.

 
DEFINED AND ANIMAL ORIGIN-FREE SYSTEM

Biolaminin 521 CTG is animal-origin free to the secondary level according to the ISCT guidance document ˮISCT Animal-Free Origin Survey Results-Summary”. All materials used in the production process have full traceability.

 
SEAMLESS TRANSITION FROM BENCH TO CLINIC

With this new cell therapy grade product, scientists are supported throughout their cell therapy development process. Human PSCs cultured on the Biolaminin 521 research grade substrate, Biolaminin 521 MX, can seamlessly be transferred to Biolaminin 521 CTG. hPSCs cultured on other feeder or on other feeder-free substrates can also be transitioned to Biolaminin 521 substrate with minimal adaptation.

EASY, FLEXIBLE AND OPERATOR-INDEPENDENT CULTURE SYSTEM

Biolaminin 521 CTG works well in any protocol using any commercially available medium. The substrate supports attachment and robust expansion of hPSCs seeded as single cells and small colonies, without the need for artificial apoptosis inhibitors. Biolaminin 521 CTG provides a robust and forgiving culture environment and allows for an operator-independent culture maintenance that is reliable, standardized and easy to monitor and control.

A BIOLOGICALLY RELEVANT CULTURE ENVIRONMENT

Biolaminin 521 CTG is a full-length, human, recombinant laminin 521 substrate, the only one of its kind on the market, providing an optimal environment for feeder-free culture of hPSCs. Laminin 521 is a key cell adhesion protein of the natural stem cell niche, expressed and secreted by hPSCs in the inner cell mass of the embryo. The Biolaminin 521 substrate recreates a biologically relevant hPSC milieu in vitro, promoting attachment, high survival, and robust long-term self-renewal of hPSCs. In addition, the more authentic culture environment primes the pluripotent stem cells for more efficient differentiation, enhanced cell maturation, polarization and organization of specialized cell types. The laminin 521 cell culture substrates have a positive effect on stabilizing and homogenizing pluripotent gene expression profiles between hES cell lines, providing a more controllable and robust culture system compared to feeder and many other feeder-free culture systems. Via the interaction with relevant cell surface receptors, laminins activate cell signalling cascades (e.g. PI3K/Akt pathway), giving consistent cellular responses and improved cell functionality, ultimately saving researchers both time and cost.

The CT521 stem cell substrate enables high expansion rate, monolayer homogenous growth, and maintained pluripotency. Cells are genetically stable without karyotypic abnormalities. hESC and iPSC grow faster on CT521 compared to cells cultured on feeders or on another feeder-free matrice, facilitating automation. Human pluripotent stem cells can routinely be cultured as single cells without the addition of ROCKi. CT521 also supports efficient clonal survival and expansion of hES and iPS cells, greatly facilitating the study of genomic alterations and manipulations in pluripotent cells. Furthermore, renders more efficient reprogramming of specific tissue cells to iPSCs.


The Biolaminin 521 substrate is more flexible, easier to use and less laborious compared to feeder-based culture systems and is compliant with any commercially available serum-free medium. The cells grow in a homogeneous monolayer and are easy to monitor, with maintained pluripotency and genetic integrity. The robust and flexible culture environment allows an operator-independent culture maintenance and reliable, standardized protocols, which can easily be adapted to automation platforms.


After birth, laminin 521 represent one of the most common laminins in the body and together with the other laminin isoforms, it supports efficient expansion and differentiation of many diverse tissue cell types.

Read more about different laminin cell culture applications


Download cell culture applications poster

biorelevant stem cell culture

Human stem cell culture

Easy culturing of human ES and iPS cells in a 
biorelevant and animal component-free environment 

effective clonal culture

Clonal cell culture

Efficient clonal culture and derivation of
human pluripotent stem cells 

  Homogenous hPSC monolayer 

Homogenous monolayer hPSC culture on CT521

PSCs plated as single-cell suspension on CT521 grow as a homogenous monolayer without any abnormal genetic aberrations. The specific CT521 binding of the α6β1 integrin enables high cell survival and self-renewal and hPSCs can routinely be cultured as single cells without the addition of ROCKi. CT521 works well in any protocol using any commercially available medium and both enzyme and EDTA-based cell dissociation agents can be used. LN521 also support clonal survival and is a good substrate for derivation and gene editing.


Rapid cell expansion

Rapid proliferation on CT521

Left: Cells passaged as single cells (wo ROCKi) on CT521 and LN521 in iPS-Brew medium have similar proliferation rate and accumulate significantly faster compared to cells cultured on Vitronectin. Fibronectin substrates could not support cell growth. Right: Accumulation rate (10 p) for hESC cell lines HS181 and H980 and iPSC cell line iPSC3.
 


Maintained differentiation capacity after long-term culture


 Maintained differentiation capacity  

Gene expression analysis of markers for pluripotency (Oct4 and Nanog) and germline-specific markers (Brachyury and Pax6) expressed in hESC cultured as single cells on CT521 for 10 passages (left) and embryoid body (EB) differentiated hESC cells (P10, right) show that hESC cultured on CT521 retain the capacity to differentiate.

Pluripotent and with a normal karyotype 

Pluripotent and genetically stable hPSCs on CT521

With CT521 hPSCs can be seeded as single cells at low densities and cultured long-term without introduction of genetic abnormalities. hESC cells (HS181) cultured on CT521 as single-cells in iPSC-Brew medium for 10 passages remain pluripotent (Oct4+; pink). The HS181 also have a normal karyotpe after 13 passages on CT521.


No spontaneous differentiation  

No spontaneous differentiation on CT521

Flow cytometry analysis shows that HS181 cells cultured on CT521 iPSC-Brew remain pluripotent after 10 passages and with retained capacity to differentiate (EB). Pluripotency markers; SSEA-4, Tra-1-60 and Oct3/4. Differentiation marker; SSEA-1.


Easy and flexible protocols

easy and flexible cell culture protocols

The single-cell split is easy, fast and forgiving, making anyone suitable to culture hPSCs. When using CT521, your cell culture protocol can easily be made totally defined and xeno-free with your choice of culture medium and dissociation reagent.

  1. Coat plates with CT521.

  2. Wash the cells with PBS and add dissociation reagent of choice - incubate

  3. Centrifuge the single-cell suspension and resuspend the pellet in fresh medium of choice

  4. Seed as single-cell suspension on fresh CT521 coated plates

Biological relevance

 natural substrate for stem cell culture

Laminin 521 is a key cell adhesion protein of the natural stem cell niche, expressed and secreted by hPSCs in the inner cell mass of the embryo. CT521 recreates the biologically relevant milieu in vitro and therefore supports robust expansion of human ES and iPS cells.


The impact of culture substrates on cell behaviour

Impact of cell culture matrix

The type of substrate used for cell culture will influence cell behaviour and have a great impact on the research results. Yet 40% of scientists use undefined cell culture substrates where 60% are facing issues, such as low cell yield and bad cell quality, and are struggling with laborious, inconsistent and operator-dependent procedures. Moreover, almost 90% of these scientists have issues maintaining a homogenous stem cell population, having to spend time manually removing differentiated cell areas. With the CT521 substrate, you will get a robust and reliable culture method with high cell quality and yield.

COATING PLATES

See instructions for coating protocol, recommended coating volumes and concentrations.


hPSC CULTURE ON CT521

See instructions for stem cell culture on Biolaminin 521, including:
- transfer
- passaging
- thawing
- cryopreservation of hPSCs on CT521. 


Embryoid body formation

Protocol for embryoid body formation from hPSCs cultured on Biolaminin 521 substrates

 

 

IMPORTANT NOTES

  • "For Research Use or Non-Commercial Manufacturing of Cell, Gene, or Tissue-Based Products”. 

  • Biolaminin 521 CTG (CT521) has been developed and manufactured to allow customers to qualify the material for use in the manufacturing of cells for clinical research.

  • CT521 is animal-component free to the secondary level according to the ISCT guidance document ˮISCT Animal-Free Origin Survey Results-Summary”**. All materials used in the production process have full traceability.

  • The product has supporting documentation such as Certificate of Analysis (CoA), Animal Origin Free Statement and Bill of material provided with every lot to support regulatory filings.

  • Avoid long exposure of the protein to ambient temperatures.

  • The laminin stock solution is stable for 3 years when stored at -20°C to -80°C. Thawed, undiluted laminin stock is stable for at least 3 months when stored at +2°C to +8°C under aseptic conditions.

  • Repeated freeze thawing should be avoided. If desired, the laminin stock can be dispensed into working aliquots and stored at -20°C to -80°C.

  • For your convenience, the coated plates can be kept for up to 4 weeks when stored aseptically at +2°C to +8°C.

 

Troubleshooting

Biolaminin plate coating

An uneven cell spread, it’s often a coating issue and could be caused by the following:

  • To low coating concentration used. Increase the coating concentration is high enough to support an even cell growth.
  • Bad coating coverage/the plate has dried out. Ensure that the entire surface is covered by the laminin coating solution when preparing fresh plates. Also, do not let the plate dry out as this will inactivate the laminin coating. Too long time in the incubator or long storage without sealing could cause too much evaporation so that part of the plate dries out (often in the center).


hPSC splitting and seeding

  • We recommend single-cell passage or passage as small clumps. 
  • Stem cells are sensitive and when using enzyme, do not treat the cells too long as that will damage the cells. Cells attach tighter on laminin compared to other matrices and scraping or pipetting without first loose up cells can affect cell integrity and viability which could result in less attachment next day. Less confluent cells need shorter treatment time whereas more confluent cells might need longer treatment time. The cells should detach easily without too much pipetting needed. Do not use too much mechanical force (extensive pipetting or scraping) as that will damage the cells. Increase the dissociation reagent incubation time rather than increasing the force.  If the cells stick very hard to the LN521 surface, try to lower the coating concentration.
  • After seeding: most cells should have attached after 1 hour and the cells should be evenly distributed over the entire plate. If there is a lot of cell death after seeding, the cells have most likely been treated too harsh during splitting.
  • The day after seeding: the cell has migrated and should have formed small colonies and will continue to expand as a homogenous monolayer. Cells cultured on the LN521 matrix are ready to be passaged when cell culture is 60-99% confluent. Depending on the cell line, seeding density and on the medium used, cultures are usually passaged 3-6 days after seeding.

Product name

Biolaminin 521 CTG

 


Product code

CT521

 


Declaration

For research use or non-commercial manufacturing of cell, gene, or tissue-based products

 

Documents

MSDS

CoA

Storage

-20°C to -80°C

 


Concentration

0.1mg/ml

 


Appearance

Clear, colourless, solution with a pH of 7.5 with
20mM HEPES, 0.25M NaCl, 10% glycerol

 


Shipping Condition

Dry Ice

Protein name

Laminin 521 (Laminin-11)

 


Classification

Animal component-free, human recombinant protein 

 

Product application

Culture of human embryonic and induced pluripotent stem cells (hESC and hiPSC), mesenchymal stem cells (MSC) and most anchorage-dependent progenitor cell types. Differentiation and maintenance of specialized cells, such as hepatocytes, cardiomyocytes and neurons.

Clonal culturing of human embryonic stem cells on laminin-521/E-cadherin matrix in defined and xeno-free environment

Rodin S., Antonsson L., Niaudet C., Simonson O.E., Salmela E., Hansson E.M., Domogatskaya A., Xiao Z., Damdimopoulou P., Sheikhi M., Inzunza J., Nilsson A.S., Baker D., Kuiper R., Sun Y., Blennow E., Nordenskjöld M., Grinnemo K.H., Kere J., Betsholtz C., Hovatta O., Tryggvason K.
Nature Communications, 2014


Monolayer culturing and cloning of human pluripotent stem cells on laminin-521 based matrices under xeno-free and chemically defined conditions

Rodin S., Antonsson L., Hovatta O., Tryggvason K.
Nature Protocols, 2014


a-5 Laminin Synthesized by Human Pluripotent Stem Cells Promotes Self-Renewal

Laperle A., Hsiao C., Lampe M., Mortier J., Saha K., Palecek S.P., and Masters K.S.
Stem Cell Reports, 2015

Laminin 521 stabilizes the pluripotency expression pattern of human embryonic stem cells initially derived on feeder cells

Albalushi H., Kurek M., Karlsson L., Landreh L., Rós Kjartansdóttir K., Söder O., Hovatta O., Stukenborg J-B.
Stem Cell International, 2017


Niche-derived laminin-511 promotes midbrain dopaminergic neuron survival and differentiation through YAP

Zhang D., Yang S., Toledo E.M., Gyllborg D., Saltó C., Villaescusa J.C., Arenas E.
Sci Signal. 2017