In article <IAN-MU.91.30736F11 at med.monash.edu.au>,
IAN-MU at med.monash.edu.au (Ian Musgrave) wrote:
> ... questions on dissociating neurons ...
Don't know if this'll work - but may be worth a shot. It's what I used for
acutely dissociated retinal neurons from turtle.
Original Ref:
Le Dain, A.C., Anderton, P.J., Martin, D.K., Millar, T.J. (1994) J. Membrane
Biol. 141, 239-245.
Tissue pieces incubated in incubation solution containing 15U/mL papain and
2.5mM cysteine. Following 30min @ RT preincubation of enzyme soln, tissue
pieces added on oscillating (2Hz) culture dish for 45min @ RT. Pieces
transferred to recording soln and triturated through very narrow bore pastuer
pipette (fire-polished). Stored at 6 deg for max 6 hours.
Incubation soln (mM) 120 NaCl, 2.5 KCl, 0.5 MgCl2, 0.5 MgSO4, 2.5 CaCl2, 1.0
NaHCO3 0.5 NaH2PO4, 16 glucose, 4.0 HEPES, 0.02mg/ml BovineSerumAlbumin.
pH7.6 (NaOH)
Recording soln (mM) 125 NaCl, 3 KCl, 2 CaCl2, 1 MgCl2, 6 NaHCO3, 0.5 NaH2PO4,
10 HEPES.
pH7.4 (NaOH)
Cells plated in chamber dish (plastic) precoated with 0.25/mL concanavalin A.
(60min with at least three H20 washes to remove excess - then allowed to
air dry).
Any questions, drop me a line: aledain at receptor.pharm.uwa.edu.au
