*Tears of joy and relief flowing down my face*
Its back!
Anyway,
I have some cells that I need to culture with a feeder layer- I have irradiated fibroblasts and keratinocyte cell line. Since the fibroblasts are irradiated I have to replace them every 2-3 weeks, but my keratinocytes should be confluent before then.
Do any of you think I can seed my fibroblasts in a few different flasks- to act as a stock for the keratinocyte cell line or will I have to seed fresh fibroblasts each time before spliting my keratinocytes?
Any advice- I've never used a feeder layer before and can't afford to buy these cells for a while so I don't want to mess things up.
Lost
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2 replies to this topic
#1
LostintheLab
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Posted 28 January 2009 - 05:01 PM
I knew it! I knew it! Well, not in the sense of having the slightest idea, but I knew there was something I didn't know.
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gugo
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Posted 30 January 2009 - 05:10 AM
I have work a little with feeder layers. I didn't irradiated the cells, I used mitomycin to inactivate their growth. I froze little aliquots of my fibroblast cell line and I only treated the cells when I need, after thaw them and seed.
I supose your cells irradiated didn't growth, but I don't know if you could mantain them in culture for long time.
If I were you I would buy a fibroblast cell line (I used 3T3-J2 from mouse) and then you work with them as usual and only irradiate o treat with mitomycin when necesary . After that, you add your keratinocytes over your feeder layer until confluence.
Best,
Gugo
I supose your cells irradiated didn't growth, but I don't know if you could mantain them in culture for long time.
If I were you I would buy a fibroblast cell line (I used 3T3-J2 from mouse) and then you work with them as usual and only irradiate o treat with mitomycin when necesary . After that, you add your keratinocytes over your feeder layer until confluence.
Best,
Gugo
#3
LostintheLab
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Posted 01 February 2009 - 04:35 PM
gugo, on Jan 30 2009, 11:10 PM, said:
I have work a little with feeder layers. I didn't irradiated the cells, I used mitomycin to inactivate their growth. I froze little aliquots of my fibroblast cell line and I only treated the cells when I need, after thaw them and seed.
I supose your cells irradiated didn't growth, but I don't know if you could mantain them in culture for long time.
If I were you I would buy a fibroblast cell line (I used 3T3-J2 from mouse) and then you work with them as usual and only irradiate o treat with mitomycin when necesary . After that, you add your keratinocytes over your feeder layer until confluence.
Best,
Gugo
I supose your cells irradiated didn't growth, but I don't know if you could mantain them in culture for long time.
If I were you I would buy a fibroblast cell line (I used 3T3-J2 from mouse) and then you work with them as usual and only irradiate o treat with mitomycin when necesary . After that, you add your keratinocytes over your feeder layer until confluence.
Best,
Gugo
Hi thanks,
Unfortunately the cell line I got came with the already irradiated cells so I figured I should use them.
But I might buy the fibroblast cells and carryon that way.
Lost
I knew it! I knew it! Well, not in the sense of having the slightest idea, but I knew there was something I didn't know.
