Hello everyone,
I have been trying to establish primary rat DRG neuron culture for a couple of months already. I am using the protocol I found somewhere during the Google search, apparently coming from materials for Biology classes ( https://nanopdf.com/download/preparation-of-rat-dorsal-root-ganglion-neurons_pdf# ) with some modifications:
The rest is following the above-mentioned protocol.
My problem is, even though I managed to get quite a number of cells from the preparation, they start to die off very quickly (around DIV3-4 - pics attached). I noticed, that even if I restrict the initial seeding surface to around 1 cm^2, when the restricting ring is removed from the culture, the cells disperse on the whole glass. Do you think restriction of the surface for a longer time would be helpful? Or could the mitosis inhibition be an issue? Or do you see anything else that could hamper my trials?
Thank you very much for all your answers - I am a total newbie to cell culture and in my lab not many people deal with neurons in general.
Best regards,
Marta
Hello Marta,
The survival of P1-P2 DRG neurons will depend on the cell culture media you use and the presence of trophic support. Adding NGF to this cultures will help you to keep these neurons alive, but may not really fit your experimental protocol. More mature DRGs (e.g. P30) are less dependent on trophic factors for the survival.
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