Membranefractionsarepreparedfrombrainhomogenates,andFAAHactivityisassayedusing[3H]anandamide(anandamide[ethanolamine-3H],60Ci/mmol)asasubstrate.Ratbra…
Membrane fractions are prepared from brain homogenates, and FAAH activity is assayed using [3H]anandamide (anandamide[ethanolamine-3H], 60 Ci/mmol) as a substrate. Rat brain membranes (50 μg protein) are incubated for 30 min at 37 °C in buffer containing [3H]anandamide and varying concentrations of URB597. At the end of the incubation period, we stopp the reactions with a mixture of chloroform/methanol and measured [3H]ethanolamine in the aqueous phase by liquid scintillation counting.
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