Invitrogen™ Acetylcholinesterase Fluorescent Activity Kit

The Acetylcholinesterase Fluorescent Activity research-use-only kit is a fluorescent activity assay designed for the quantification and detection of acetylcholinesterase activity in serum, plasma, and erythrocyte membrane samples.

This complete, ready-to-use kit includes black 96-well plate(s), acetylcholinesterase standard (1,000 mU/mL), acetylcholinesterase substrate, acetylthiocholine iodide, and other components to perform the assay. A fluorescence 96-well microplate reader capable of reading fluorescent emission at 510 nm, with excitation at 390 nm, is required for use of this kit.

Performance characteristics
• Assay type: fluorescent activity kit
• Sample types: serum, plasma, and erythrocyte membrane samples
• Sensitivity: 0.218 mU/mL
• Standard curve range: 6.25–100 mU/mL
• Reactivity: species independent

Background
Acetylcholine (ACh) is an essential neurotransmitter in the central and peripheral nervous systems. In the brain multiple areas exist where cholinergic neurons are concentrated. Nicotinic and muscarinic ACh receptors are recognized as binding and effector proteins to mediate chemical neurotransmission at neurons, ganglia, heart, and smooth muscle fibers and glands.

Acetylcholinesterase is encoded by the single AChE gene; the structural diversity in the gene products arises from alternative mRNA splicing and post translational associations of catalytic and structural subunits. The major form of acetylcholinesterase found in brain, muscle, and other tissues is the hydrophilic species, which forms disulfide-linked oligomers with collagenous, lipid-containing structural subunits. The other alternatively-spliced form, expressed primarily in the erythroid tissues, is structurally different at the C-terminal end and contains a cleavable peptide with a GPI anchor. It associates with membrane receptors through phosphoinositide moieties added post-translationally. This assay has been validated for serum, EDTA and heparin plasma, and solubilized RBC ghosts from a variety of species.

Assay principle
The Acetylcholinesterase Activity kit is designed to quantitatively measure acetylcholinesterase activity in a variety of samples. A human AChE standard is provided to generate a standard curve for the assay and all samples should be read off the standard curve. The kit utilizes a proprietary non-fluorescent molecule, that covalently binds to the thiol product of the reaction between the AChE Substrate and AChE in the standards or samples, yielding a fluorescent product read at 510 nm in a fluorescent plate reader with excitation at 390 nm. The kit is suitable for measuring AchE activity in appropriately diluted serum, plasma, and RBC ghosts from a number of species. It will also measure AChE in extracted tissue samples and cell lysates.

AChE unit definition
One unit will hydrolyze 1.0 μmol of acetylthiocholine iodide per minute at pH 7.4 and 37°C.