Protocol Online logo
Top : Forum Archives: : Cell Biology

How to culture rat dorsal root ganglia cell? - (Dec/01/2005 )

unsure.gif I am culturing newborn rat dorsal root ganglia(DRG).I have tried many times but failed absolutly.
The culture media is DMEM/F12 .15%FBS is contained . DRG were dissected out,they were incubated at 37℃ for 30min in DMEM/F12media(containing 0.125%trypsin).Then DMEM/F12 containing FBS was added to terminate disgestion.And then centrifugated at 1000rpm for 10min.But I could not see the pellet. When they were planted the cells kept circular ,no dendrite or axon growed out .Could anyone tell me the reason or tell me the protocol in detail?

-tangstatlight-

QUOTE (tangstatlight @ Dec 1 2005, 08:33 AM)
unsure.gif I am culturing newborn rat dorsal root ganglia(DRG).I have tried many times but failed absolutly.
The culture media is DMEM/F12 .15%FBS is contained . DRG were dissected out,they were incubated at 37℃ for 30min in DMEM/F12media(containing 0.125%trypsin).Then DMEM/F12 containing FBS was added to terminate disgestion.And then centrifugated at 1000rpm for 10min.But I could not see the pellet. When they were planted the cells kept circular ,no dendrite or axon growed out .Could anyone tell me the reason or tell me the protocol in detail?


hi,

I am culturing newborn mouse DRG's and they have axon/dendrit outgrowth like hell biggrin.gif

the step where you could be wrong is the dissociation step with the trypsin(0.125% for about 30-35 minutes). I DONT do them in the DMEM:F12, I do it in the HBS -Mg -Ca ! After that I stop it like you do, also the centrifugation. There should be a very tiny pellet. After a day you should already see axon outgrowth. p.s. : the laminin coating works, but as well the collagen.

Good Luck
swiz1

-swiz1-