A radiochemical method for the estimation of choline acetyltransferase |
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Authors: | F Fonnum |
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Institution: | Biochemistry Department, Agricultural Research Council Institute of Animal Physiology, Babraham, Cambridge |
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Abstract: | 1. A radiochemical method for the estimation of choline acetyltransferase (choline acetylase) has been devised which involves the formation of labelled acetylcholine from labelled acetate. 2. 1-(14)C]Acetate and coenzyme A are pre-incubated in the presence of non-rate-limiting concentrations of acetyl-coenzyme A synthetase to give 1-(14)C]acetyl-coenzyme A, which then reacts with choline in the presence of the acetyltransferase to give (14)C]acetylcholine. 3. Any (14)C]-acetyl-coenzyme A remaining at the end of the reaction is destroyed by the addition of excess of hydroxylamine, and (14)C]acetylcholine is freed from other labelled compounds by precipitation with sodium tetraphenylborate (Kalignost). 4. The washed precipitate is dissolved in acetonitrile-benzyl alcohol and estimated by scintillation counting. 5. Advantages over other methods are discussed. |
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