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Message | User | Date(yyyy-mm-dd) |
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Adult astrocytes culture | luda | 2008-01-07 | Click here to register. |
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| Dear colleagues, is there anyone who has experience of culturing adult astrocytes? I tested number of protocols from different books and articles but have no results. Would you please, help me? Thank you very much. | | |
| Fulvio | 2008-01-08 | Click here to register. |
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 | Dear Luda. what protocol are you using? when you say "adult" you mean from adult animal?If you can give more details is easier to help you.. :)
Fulvio |
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| luda | 2008-01-08 | Click here to register. |
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| Dear Fulvio, thank you for reply!
Yes, I am using adult rat cortex for astrocytes isolation. I know (according to the literature) that it is much easier to obtain the astrocytes culture from newborn or embryonic brain; however for my future experiments I need the astrocytes from adult rat brain. I quite experienced in culture of adult rat Schwann cells, DRG neurons, olfactory ensheathing cells, fibroblasts and bone marrow stromal cells, but completely stack with astrocytes. I will be very appreciated if you can help me!
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| Fulvio | 2008-01-09 | Click here to register. |
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| yes from adult can be quite tricky...I made from p2 rats but some colleague of mine have done from adult and we use the same protocols...but some tricks..
1-make the dissection as quick as possible
2-be sure to clean out all the meninges..the adult brain has a lot..use a cotton wool stick to remove it
3-plate on poly-l-lysine coated flasks to make it better adhere...
tell me if you need a complete protocol! :)
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| luda | 2008-01-09 | Click here to register. |
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| Dear Fulvio,
Thank you for the tips!
I will be very appreciated if you can send me a complete working protocol!
I will try to describe my way of brain harvesting; might be I am doing something wrong from the very beginning:
1-make the dissection as quick as possible
anesthesia wait until rat is not breathing (1-2 min) cut the hart to remove the main blood (1 min) opening the cranium and brain harvesting (1-2 min) brain in medium to clean out the meninges and dissect the cortex (max 5 min). The total time about 10 min; is it too long?
2-be sure to clean out all the meninges..the adult brain has a lot..use a cotton wool stick to remove it
I usually use fine forceps to remove the pia matter and blood vessels from the brain surface but not cotton wool.
3-plate on poly-l-lysine coated flasks to make it better adhere...
I always use poly-l-lysine (50µg/ml) coated tissue culture flasks.
Thank you. |
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