REVIEW Acetylthiocholine iodide is used as a synthetic substrate for the assay of acetylcholine esterase (E.C. 184.108.40.206), replacing the natural substrate acetylcholine. This enzyme hydrolyses the substrate to yield acetate and thiocholine. The free thiol group of thiocholine reacts with 5,5'-dithiobis-2-nitrobenzoic acid (DTNB, Ellman's reagent, producing the yellow 4-nitrothiolate anion. The release of this yellow anion is followed at 412 nm. The assay may be performed at 30°C in 0.1 M sodium phosphate buffer with 10 mM DTNB and 12.5 mM acetylthiocholine iodide.
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