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MESC 18

MESC 18

Catalogue No.

11120816

Cell Line Name

MESC 18

Cell Line Description

The germ-line competent cell line MESC 18 was established from the inner cell mass of a 3.5 day male pre-implantation mouse embryo (strain 129/Olac). These pluripotent cells retain the ability to participate in normal embryonic development. Differentiation of the cells is inhibited by Leukaemia Inhibiting Factor (LIF). Addition of LIF allows culture without the use of a feeder layer.

Characteristics

Tissue of Origin

Embryo

Morphology

Spheroidal

Karyotype

XY, diploid

Applications

Embryonic stem (ES) cell-based genetic manipulations

Disease

None Stated

Culture Conditions

Cell Type

Stem cell

Subculture Routine

The MESC lines can be grown without the use of mitotically inactivated feeder cells (Brown et al., 1992 PMID: 1483967). However, the cells supplied by ECACC have been grown on mitomycin treated primary mouse embryonic fibroblasts to ensure the cells are maintained in an undifferentiated state. Mouse embryonic fibroblasts, STO (ECACC 86032003) or SNL 76/7 (ECACC 07032801) can be used. At ECACC plastic ware is pre-coated with gelatine prior to plating feeder cells.Porcine gelatine (Sigma G1890) is dissolved in sterile water (0.5g/500ml) at 56°C. The 0.1% solution is sterilized by filtration (0.22µm). Add 0.1% gelatine to plastic ware to cover bottom, and incubate for 20 minutes at room temperature. Remove gelatine, wash with PBS once and replace with appropriate culture medium. The flask/dish must not be allowed to dry out.Feeder layers are prepared on the gelatinized flasks at least 24 hours in advance of being required. An ampoule is thawed in 37°C water bath and the contents quickly transferred to a 15ml centrifuge tube. MEF medium is added drop wise to 5ml. Cells are centrifuged at 150 x g for 5 minutes at Room Temperature (RT). Cells are resuspended in 5ml of MEF medium. Cells are counted and added to flasks containing the correct medium at 1-3 x 10⁴ cells/cm².An ampoule of ES cells is thawed in 37°C water bath and the contents quickly transferred to a 15ml centrifuge tube. KSR medium is added drop wise to 5ml. Cells are centrifuged at 150 x g for 5 minutes. Cells are resuspended in 5ml of KSR medium. The prepared feeder flask is washed once with PBS and KSR medium added. ES cells should be plated at 4-5 x 10⁴ cells/cm². Cultures must be incubated in a humidified 5% CO₂/95% air incubator at 37°C. A 100% media change must be performed every day and cells passaged every 2-3 days. Colonies must not be allowed to touch each other as overgrowth will result in differentiation.

Culture Medium

MEF medium consists of Advanced DMEM/F12 (Invitrogen 12634010), 10% FBS (Perbio SH30070.03E), 2 mM Glutamine (Invitrogen 25030024) and 0.1 mM β-mercaptoethanol (Sigma M6250).KSR medium consists of KO-DMEM (Gibco 10829), 20% Knock-Out Serum Replacer (Gibco 10828), 2 mM Glutamine (Invitrogen 25030024), NEAA (Invitrogen 11140035), 0.1 mM β-mercaptoethanol (Sigma M6250) and LIF 1000 Units/ml (ESGRO ESG1106).

Growth Mode

Adherent

Additional Info

Depositor

DG Brown, MRC Toxicology Unit, Hodgkin Building, Leicester LE2 4UE

Country of Origin

United Kingdom

Hazard Group (ACDP)

Hazard Group (ACDP) 2

Applications

References

Brown DG, Willington MA, Findlay I, Muggleton-Harris AL.1992 Criteria that optimize the potential of murine embryonic stem cells for in vitro and in vivo developmental studies. In Vitro Cell Dev Biol.28A(11-12):773-8. PMID: 1483967.

Bibliography

Crolla JA, Brown D, Whittingham DG. 1990 Spontaneous induction of an homologous robertsonian translocation, Rb(11.11) in a murine embryonic stem cell line. Genet Res. 55(2):107-10. PMID: 2370006.

Available Formats

  • Frozen
  • DNA-5µg (100ng/µl)

If use of this culture results in a scientific publication, it should be cited in the publication as: MESC 18 (ECACC 11120816).

Unless specified otherwise, at ECACC we routinely handle all of our cell lines at containment level 2 in accordance with the ACDP guidelines (Advisory Committee on Dangerous Pathogens) (UK). All cell cultures have the potential to carry as yet unidentified adventitious agents. It is the responsibility of the end user to ensure that their facilities comply with biosafety regulations for their own country. ACDP Guidance: Biological agents: Managing the risks in laboratories and healthcare premises.

The Culture Collections represent deposits of cultures from world-wide sources. While every effort is made to ensure details distributed by Culture Collections are accurate, Culture Collections cannot be held responsible for any inaccuracies in the data supplied. References where quoted are mainly attributed to the establishment of the cell culture and not for any specific property of the cell line, therefore further references should be obtained regarding cell culture characteristics. Passage numbers where given act only as a guide and Culture Collections does not guarantee the passage number stated will be the passage number received by the customer.

Cultures supplied by Culture Collections are for research purposes only. Enquiries regarding the commercial use of a cell line are referred to the depositor of the cell line. Some cell lines have additional special release conditions such as the requirement for a material transfer agreement to be completed by the potential recipient prior to the supply of the cell line. Please view the Terms & Conditions of Supply for more information.