MEF (CF-1) (ATCC® SCRC-1040)

Organism: Mus musculus, mouse  /  Cell Type: Fibroblast  /  Tissue: Embryo  / 

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Organism Mus musculus, mouse
Tissue Embryo
Cell Type Fibroblast
Product Format frozen
Morphology Fibroblast
Culture Properties Adherent
Biosafety Level 1
Age 14 days gestation embryo
Gender male and female mixed
Strain CF-1, non-inbred mouse strain (non-agouti albino) from Carworth Farms
Applications
ATCC has successfully irradiated (SCRC-1040.1) and treated the cells with Mitomycin C (SCRC-1040.2a) for use as a feeder layer.
The cells can be used as a feeder layer to support the growth of embryonic stem (ES) cells and for the maintenance of ES cells in the undifferentiated state.
Storage Conditions liquid nitrogen vapor phase
Derivation
The cell line was established by ATCC in 2003 from embryonic day 14 (E14) CF-1 mouse embryos.
Clinical Data
male and female mixed
Comments

The growth of these cells should be arrested before being used as a feeder layer. ATCC has successfully irradiated (SCRC-1040.1) and treated the cells with Mitomycin C (SCRC-1040.2a) for use as a feeder layer. If the MEFs are being used as a feeder layer for ES cells, it is not recommended to use them past passage no. 6 (P6). 

Complete Growth Medium The base medium for this cell line is ATCC-formulated Dulbecco's Modified Eagle's Medium, Catalog No. 30-2002. To make the complete growth medium, add the following components to the base medium:
  • fetal bovine serum to a final concentration of 15%

  • This medium is formulated for use with a 5% CO2 in air atmosphere. (Standard DMEM formulations contain 3.7 g/L sodium bicarbonate and a 10% CO2 in air atmosphere is then recommended).
    Subculturing

    To insure the highest level of viability, be sure to warm media and Trypsin / EDTA to 37ºC before using it on the cells. Cells should be split when they reach confluency. Split cells at approximately 0.4 X 104 cells/cm2

    1. Remove and discard culture medium.
    2. Briefly rinse the cell layer with 1XPBS (SCRR-2201) solution to remove all traces of serum, which contain trypsin inhibitor.
    3. Add 5 mL of Trypsin-EDTA (0.25% (w/v) Trypsin-0.53 mM EDTA solution, ATCC# 30-2101) solution to flask and incubate for 1 minute, gently tapping the flask observe cells under an inverted microscope until cells detach (usually within 1 to 2 minutes).
    4. Add 6.0 to 8.0 mL of complete growth medium and rinse surface of the flask to detach all cells. Gently pipetting up and down will break cell clumps.
    5. Transfer all cells into a centrifuge bottle or tube and centrifuge at 271x g for 5 minutes.
    6. Remove and discard the supernatant
    7. Add 10 mL complete growth medium to cell pellet and with 10 mL pipette resuspend the cells gently (create a single-cell suspension).
    8. Add more complete growth medium to cell suspension as needed to plate cells.
    9. Place flasks in incubator @ 37°C with a 5% CO2 in air atmosphere.

     Note: For more information on enzymatic dissociation and subculturing of cell lines consult Chapter 13 in Culture of Animal Cells, a manual of Basic Technique by R. Ian Freshney, 5th edition, published by Alan R. Liss, N.Y., 2005.   

     Flask/Plate

     

    Growth Area (cm2

     

    1xPBS (mL) 

     

    Trypsin/EDTA (mL) 

     

    Equal vol. Complete Growth Medium (mL) 

     

    Growth Medium (mL) 

     

     T225

     

     225

     

     10 ± 0.2

     

    6 ± 0.2 

     

    6 ± 0.2 

     

    30 

     

     75

     

     75

     

     5 ± 0.1

     

    3 ± 0.1 

     

    3 ± 0. 1 

     

     12

     

     T25

     

     25

     

    3 ± 0.1 

     

    1.5 ± 0.1 

     

    1.5 ± 0.1 

     

     6

     

     6 well

     

     9.5

     

     1 ± 0.1

     

     1 ± 0.1

     

     1 ± 0.1

     

     3

     

     

    Subcultivation Ratio: Plate the cells at approximately of 0.4 X 104 cells/cm2.
    Medium Renewal: Twice a week or when pH decreases.
    Cryopreservation

    Complete growth medium supplemented with an additional 40% FBS and 10% (v/v) DMSO

    Cell culture tested DMSO is available as ATCC Catalog No. 4-X.

    Culture Conditions
    Atmosphere: air, 95%; carbon dioxide (CO2), 5%
    Temperature: 37°C
    Name of Depositor ATCC
    Year of Origin 2003
    References

    Amit M, et al. Human feeder layers for human embryonic stem cells. Biol. Reprod. 68: 2150-2156, 2003. PubMed: 12606388

    Hovatta O, et al. A culture system using human foreskin fibroblasts as feeder cells allows production of human embryonic stem cells. Hum. Reprod. 18: 1404-1408, 2003. PubMed: 12832363

    Andrews P, et al. Human embryonic fibroblast feeder cells. International Patent Application WO 03/078611 A1

    Notice: Necessary PermitsPermits

    These permits may be required for shipping this product:

    • Customers located in the state of Hawaii will need to contact the Hawaii Department of Agriculture to determine if an Import Permit is required. A copy of the permit or documentation that a permit is not required must be sent to ATCC in advance of shipment.
    Basic Documentation
    References

    Amit M, et al. Human feeder layers for human embryonic stem cells. Biol. Reprod. 68: 2150-2156, 2003. PubMed: 12606388

    Hovatta O, et al. A culture system using human foreskin fibroblasts as feeder cells allows production of human embryonic stem cells. Hum. Reprod. 18: 1404-1408, 2003. PubMed: 12832363

    Andrews P, et al. Human embryonic fibroblast feeder cells. International Patent Application WO 03/078611 A1