Serum-free medium for culturing mouse ES cells

mESF Basal Medium / mESF Supplement Set

mESF Basal Medium is a serum-free medium optimized for the culture of mouse ES cells. When used in combination with our mESF Supplement Set and LIF, this medium supports the growth of mouse ES cells without requiring feeder cells. It also eliminates the need for FBS or a serum replacement.

Product Lines

  • mESF Basal Medium (Cat. No. 136-17805)
  • mESF Supplement Set (Cat. No. 299-73301)

mESF Supplement A: 5 mL x 1

mESF Supplement B: 5 mL x 1 (contains 0.0078% 2-mercaptoethanol)

Advantages

  • Sustains cell proliferation without the use of FBS or a serum replacement (serum-free medium).
  • Promotes cell growth under feeder-free condition.
  • Maintains the undifferentiated state of mouse ES cells at low concentrations of LIF.

Instructions for Use

Please add each mESF Supplement and LIF to mESF Basal Medium before use.

* Note: When cells are directly transferred to mESF medium from medium currently in use, the number of live cells may decrease in the new culture environment. To overcome this problem, please seed more cells than those used for routine subcultures. If direct transferring does not go well, please use the following adaptation protocol.

(1) Grow cells in their standard medium. Then, subculture the cells into a 1:1 mixture of mESF medium and the standard medium.

(2) During the subsequent successive cultures, allow the cells to gradually adapt to their new serum-free environment by increasing the ratio of mESF medium to the standard medium stepwise (3:1 followed by 7:1), until the cells are transferred into 100% mESF medium.

Culture of Mouse ES Cells D3 Strain

Mouse ES cells D3 strain were grown in mESF Basal Medium containing the required supplements. After the cells were passaged five times, their morphology and growth potential were analyzed. Using the same cells, we also checked the expression of undifferentiation markers.



«Cell Morphology»

We confirmed these cells could form colonies with a shiny appearance that is typical of mouse ES cells.

00557_img01.jpg



«Cell Growth Curve»

The growth curve of these cells is shown below.

00557_img02.jpg

<Growth Medium>
mESF Basal Medium containing mESF Supplement A, mESF Supplement B, and 1,000 units/ml StemSure LIF

<Seeding Density>
7,000 cells/well
(Cultured in collagen-coated 12-well plate)

«Expression of Undifferentiation Markers»

We confirmed that these cells expressed the following undifferentiation markers: Nanog, Sox2, Oct3/4, and SSEA-1.

00557_img03.jpg

Culture of Mouse ES Cells D3 Strain in the Presence of Low Concentrations of LIF

Using the mouse ES cells D3 strain, we checked the effects of low concentrations (250 and 500 units/mL) of LIF on the proliferation and differentiation of cells cultured in our mESF medium.



«Morphologies of ALP-Stained Colonies and Cell Growth Curves»

The results of our cell morphology and growth curve are shown below.

00557_img04.jpg

<Growth Medium>

mESF Basal Medium containing mESF Supplement A, mESF Supplement B, and StemSure LIF (250, 500, or 1,000 units/mL)

<Seeding Density>

7,000 cells/well

(Cultured in collagen-coated 12-well plate)



«Percentage of Colonies Positive for ALP Staining»

Virtually all colonies were stained positive for ALP activity at all concentrations of LIF tested.

00557_img05.jpg

<Growth Medium>

mESF Basal Medium containing mESF Supplement A, mESF Supplement B, and StemSure LIF (250, 500, or 1,000 units/mL)

<Seeding Density>

500 cells/well

(Cultured in collagen-coated 6-well plate)

<Incubation Time>

10 days

References

  • Furue, M., et al.: In Vitro Cell Dev. Biol. Anim., 41, 19-28, (2005).
  • Hayashi,Y., et al.: Stem Cells, 25, 3005-3015, (2007).

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