Laminin-511

LN511
Human recombinant laminin-511


Human recombinant laminin-511 (LN-511) is the natural laminin for mouse embryonic stem cells with sustained pluripotency without the need to use feeder cells or differentiation inhibitors like LIF.

Laminin-511 is together with laminin-521 (LN-521) the most widely expressed laminins in the body. LN-511 is especially recommended for cell culture of murine embryonic stem cells or induced pluripotent stem cells while LN-521 is recommended for human pluripotent stem cells.

Culture of mouse pluripotent stem cells on LN-511, which is already expressed at the 4‑cell stage during embryonic development, maintains the pluripotency without the use of LIF. Using LN-511 for adherent murine stem cell cultures makes culturing easy and efficient. 

 

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100µg 1 x 1ml €69
1mg 10 x 1ml €610
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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.

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Mouse ES and iPS cells grow on laminin-511 (LN-511) in a completely defined matrix, without the need of using time-consuming and cumbersome feeder cells or LIF since the laminin-511 protein creates a biological niche keeping the ES cells in a pluripotent state for months. BioLamina's LN-511 is a full-length human recombinant laminin that is used as a mouse ESC or iPSC culture substrate. 

Stem cells grow as monolayers on LN-511 enabling equal contact to both the matrix and cell culture medium, which leads to a more homogenous cell population. Since LN-511 is a recombinant protein, lot-to-lot variation is minimal and therefore fewer replicates are needed for reliable results (Domogatskaya, 2008; Rodin, 2010). 

The need for matrices that support pluripotent stem cell renewal in a reproducible manner is clear. Stem cell biologists have long experienced problems with the repeatability of results and with maintenance of pluripotency when working with ES cell cultures.

 

CULTURE OF MURINE STEM CELLS ON LAMININ-511


Additives like the cytokine LIF have thus been added to the cell culture medium to minimize spontaneous cell differentiation or one has needed to culture the cells on feeder cell layers.

Now, however, LN-511 allows the proliferation of pluripotent mouse ES cells without such differentiation inhibitors or feeder cells. Importantly, the ES cells grow as a homogenous monolayer, which ensures equal availability to both substrate and soluble factors and, thus, creates a more homogenous cell population (Domogatskaya, 2008).

LN-511 and other laminins can also be applied to glass, and as stem cells grow as a monolayer on LN-511 and LN-521 these laminins have been used for improved imaging of stem cells grown on glass slides.

The LN-511 matrix provides better opportunities for designing experiments and interpreting obtained results.

LN-511 can also be used for culture of many other cell types as seen under laminin applications.

laminin-511 cell culture

Murine stem cell culture

Self-renewal of mouse pluripotent stem cells 
without leukemia inhibitory factor

mESC expansion without LIF

laminin murine stem cell culture

LN-511 support the self-renewal of mouse ES cells for over 5 months without the presence of LIF or feeder cells, when other known matrices are unable to do so for longer than a couple of weeks.


 

Homogenous monolayer

Naive mouse ES cell culture

Mouse ES cells adhere to LN-511 with three to five-fold higher affinity than to other commonly used matrices. Pluripotent mouse ES cells grow even at low cell density as monolayers on the surface of LN-511, which is different from the conventional clusters in the cultures, and, therefore, have equal contact with the matrix and medium, allowing a homogeneous and defined growth environment.


 

Long-term pluripotent mouse ESC

Naive mouse ES cells germline transmission

Mouse ES cells grown on LN-511 for over 3 months are indeed pluripotent since they give rise to germline transmission (chimeric mice) after injection into mouse blastocysts.


 

PASSAGE MOUSE PSC ON LAMININ-511

1. Aspirate old medium from wells.

2. Wash the cells gently with 1xDPBS (Ca++/Mg++) and aspirate, add TrypLE to wells and incubate at room temperature for max 30 seconds.

3. Aspirate TrypLE, and wash the cells twice with pre-warmed medium.

4. Add fresh medium to wells. Scrape cells with a pipette tip, then gently pipette up and down to achieve a clump size of approximately 0.5 -1 mm in diameter. Avoid making single-cell suspension.

5. Seed cells with a ratio of 1:2, 1:3 or 1:4 on LN511 coated wells. Optimal seeding densities will vary from one cell line to another and can be determined empirically for your system.

6. Swing plates side-to-side to distribute cell aggregates evenly, then place them into an incubator (+37°C, 5% CO2).

 

Instructions for culture mouse ES cell on laminin

COATING PROCEDURE

1. Slowly thaw recombinant laminins at +4°C before use. Thawed laminin stock is stable for at least 3 months when stored at +2°C to +8°C under aseptic conditions. For your convenience, the coated plates and diluted coating solution can be kept for up to 4 weeks when stored aseptically at +2°C to +8°C.

2. Dilute the thawed laminin stock solution with 1xDPBS containing Ca2+ and Mg2+.

3. Add the diluted laminin solution to tissue culture-treated cultureware for a final coating concentration of 0.5-2 ug/cm2. The optimal coating concentration is cell line-dependent. See the laminin coating instruction for recommended coating volumes and concentrations.

4. Seal the plate (e.g. with Parafilm®) to prevent evaporation and incubate at +2°C to +8°C overnight. If a more rapid coating is required, incubate at +37°C for 2 hours. Make sure the laminin solution is spreaded evenly across the surface. Note that the laminin matrix will be inactivated if let dry.

IMPORTANT NOTES

  • The laminin stock solution is stable for 2 years when stored at -20°C. If desired, the laminin stock can be dispensed into working aliquots and stored at -20°C. Repeated freeze thawing should be avoided. 

  • When culturing mouse PSCs on LN-511 no treatment with differentiation inhibitors, such as leukemia inhibitory factor (LIF), is needed.

  • The protocol can easily be made totally defined and xeno-free with your choice of culture medium and enzyme.

  • Before start, all solutions used for cell passaging should be aliquoted in sufficient amounts and pre-warmed at +37°C, 5% CO2.

  • Cells are ready to be passaged when cell culture is ≥60% confluent. Optimal seeding densities will vary from one cell line to another and can be determined empirically for your system. With optimal media conditions and seeding density, most cell lines will reach confluence within 4-6 days and expand 10-25 fold.

  • When moving your cells from another feeder-free matrix (e.g. Matrigel) we recommend you to start smaller well format (e.g. 96-well or 48-well format) and a higher seeding density (50,000-100,000 cells/cm2) for the first number of passages to let the cells adapt to the laminin matrix before increasing the well format and lowering the seeding density.

  • When moving your cells from feeders to LN-511, follow the protocol here

Product name

LN511

 

Product use

Cell culture

 

Product application

Murine embryonic and induced pluripotent stem cells


 

Documents

MSDS

Storage

-20°C

 

Stability

Three years from date of receipt

 

Stock concentration

0.1mg/ml

 

Use concentration

0.5-2 ug/cm2 
Optimal concentration must be determined by the end user

Protein name

Laminin-511 (LN-511)

 

Synonyms

Laminin-10

 

Expressed in vivo

Expressed and secreted by human pluripotent stem cells and can, in addition, also be found in epithelia and dermal papillae, endothelial cells, pancreas, peripheral nerves, and placenta.

Laminin-511 but not -332, -111, or -411 enables mouse embryonic stem cell self-renewal in vitro.

Domogatskaya A, Rodin S, Boutaud A, Tryggvason K.

Stem Cells, 2008

 

Long-term self-renewal of human pluripotent stem cells on human recombinant laminin-511.

Rodin S, domogatskaya A, Ström S, Hansson EM, Chien KR, Inzunza J, Hovatta O, Tryggvason K.

Nature Biotechnology, 2010

Xeno-free culture of human pluripotent stem cells.

Bergström R, Ström S, Holm F, Feki A, Hovatta O.

Methods in Molecular Biology, 2011

 

Compositional and structural requirements for laminin and basement membranes during mouse embryo implantation and gastrulation.

Miner JH, Li C, Mudd JL, Go G, Sutherland AE.

Development, 2004

 

Trophoblastspecific expression and function of the integrin alpha 7 subunit in the peri-implantation mouse embryo.

Klaffky E,Williams R,Yao CC, Ziober B, Kramer R, Sutherland AE.

Developmental Biology, 2001