Currently I am using high glucose DMEM with 20% horse serum to culture 6-23 (Rat medullary thyroid carcinoma) cells. After passage 7 or so, the cell proliferation has dramatically decreased (therefore I increased horse serum from 15 to 20%). Additionally, when I seed the cells into a new dish, they sometimes don't attach or they aggregate into clusters and float away. The morphology of the cells seemed to have changed. Seeding into coverslips have been a challenge as well.
Could anyone kindly suggest me protocol for 6-23 (Rat medullary thyroid carcinoma) cell culture (both on petri dish and coverslips)? Thank you!
Hi Rakshya,
ATCC is recommends Dulbecco's modified Eagle's medium with 4.5 g/L glucose, 85%; horse serum, 15% which basically you use. They also suggest to subculture every 10 to 12 days in a ratio of 1:10 to 1:20. I don't know how you do it?
If cells don't attach after seeding they might have been longer under trypsin treatment (or whatever you use to detach them from the plate while sub-culturing) and have difficulties to attach again due to ''destabilization'' of integrins on the cell surface. Try shorter time of incubation or less concentrated trypsin. Also, you could try to coat you dishes and cover slips with collagen or fibronectin or other matrix your cells might like. This could help them to attach better, especially if you are seeding on glass cover slips.
Check in some publications if people were using some matrix to coat plates while seeding this type of cells.
Best of luck,
Milica
Hi Rakshya,
ATCC is recommends Dulbecco's modified Eagle's medium with 4.5 g/L glucose, 85%; horse serum, 15% which basically you use. They also suggest to subculture every 10 to 12 days in a ratio of 1:10 to 1:20. I don't know how you do it?
If cells don't attach after seeding they might have been longer under trypsin treatment (or whatever you use to detach them from the plate while sub-culturing) and have difficulties to attach again due to ''destabilization'' of integrins on the cell surface. Try shorter time of incubation or less concentrated trypsin. Also, you could try to coat you dishes and cover slips with collagen or fibronectin or other matrix your cells might like. This could help them to attach better, especially if you are seeding on glass cover slips.
Check in some publications if people were using some matrix to coat plates while seeding this type of cells.
Best of luck,
Milica
I agreed with Milica Bozic Stanojevic, but I would rather recommend Thawing a new tube of MCF-7 cells. in my experience, if the cells are less confluent or we sub-culture very few cells in the plate sometimes they do exhibit above mentioned characteristics.
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