CF6-Neo Mouse Embryonic Fibroblasts, MitC-treated - FAQs

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5 个常见问题解答

Is it better to use irradiated mouse embryonic fibroblasts (MEFs) or Mitomycin C-treated MEFs to grow embryonic stem cells?

Either method will work in arresting cell division. However, the irradiation process will ensure that cell division will cease regardless of cell aggregation. Cell clumping can potentially not inactivate all cells when using mitomycin C, as cells within clumps may not be exposed to the mitomycin C. Irradiated cells are preferred by those who have concerns about chemical treatment. Mitomycin C-treated cells are preferred by those who have concerns about DNA damage from irradiation.

What are the differences between CF1, CF6, and DR4 mouse embryonic fibroblasts?

CF1 mouse embryonic fibroblasts do not have drug resistance. CF6 mouse embryonic fibroblasts are resistant to Neomycin/geneticin (G418). DR 4 mouse embryonic fibroblasts are resistant to geneticin (G418), puromycin, hygromycin, and 6-thioguanine.

What is the recommended seeding density for mouse embryonic fibroblasts? Why is the seeding density critical?

We recommend seeding these cells at densities ranging from 2 x 10E4 to 5.5 x 10E4 cells/cm2. A good starting point is 3 x 10E4 cells/cm2. If the feeder cells are too sparse, they may not maintain the pluripotent cells without differentiation, and the pluripotent cells may not attach well. If the feeder cells are too dense, the feeder layer may detach from the plate, and the culture will be lost.

When should mouse embryonic fibroblasts be plated and how long can these cells be used?

These cells should be plated 24 hours prior to plating the ESCs or iPSCs and should be used for only 7-10 days.

Does the culture vessel need to be coated prior to seeding mouse embryonic fibroblasts?

Yes, the culture vessel needs to be coated with Attachment Factor protein (Cat. No. S006100) at 37 degrees C for 30 mins or at room temperature for 2 hours. The coated vessels can be used immediately or stored at room temperature for up to 24 hours.