Nori Human Acetylcholinesterase ELISA Kit Summary

Alternative names for acetylcholinesterase: ACHE

Standard Curve

Kit Components
1. Pre-coated 96-well Microplate
2. Biotinylated Detection Antibody
3. Streptavidin-HRP Conjugate
4. Lyophilized Standards
5. TMB One-Step Substrate
6. Stop Solution
7. 20 x PBS
8. Assay Buffer

Other Materials Required but not Provided:
1. Microplate Reader capable of measuring absorption at 450 nm
2. Log-log graph paper or computer and software for ELISA data analysis
3. Precision pipettes (1-1000 µl)
4. Multi-channel pipettes (300 µl)
5. Distilled or deionized water

Protocol Outline
1. Prepare all reagents, samples and standards as instructed in the datasheet.
2. Add 100 µl of Standard or samples to each well and incubate 1 h at RT.
3. Add 100 µl of Working Detection Antibody to each well and incubate 1 h at RT.
4. Add 100 µl of Working Streptavidin-HRP to each well and incubate 20 min at RT.
5. Add 100 µl of Substrate to each well and incubate 5-30 min at RT.
6. Add 50 µl of Stop Solution to each well and read at 450 nm immediately.

Background: 

Acetylcholinesterase (HGNC symbol ACHE; EC 3.1.1.7), also known as AChE or acetylhydrolase is encoded by ACHE gene^[1] and is the primary cholinesterase in the body. It catalyzes the breakdown of acetylcholine and of some other choline esters that function as neurotransmitters. AChE is found at mainly neuromuscular junctions and in chemical synapses of the cholinergic type, where its activity serves to terminate synaptic transmission. It belongs to carboxylesterase family of enzymes. It is the primary target of inhibition by organophosphorus compounds such as nerve agents and pesticides.

During neurotransmission, ACh is released from the presynaptic neuron into the synaptic cleft and binds to ACh receptors on the post-synaptic membrane, relaying the signal from the nerve. AChE, also located on the post-synaptic membrane, terminates the signal transmission by hydrolyzing ACh. The liberated choline is taken up again by the pre-synaptic neuron and ACh is synthesized by combining with acetyl-CoA through the action of choline acetyltransferase. An endogenous inhibitor of AChE in neurons is Mir-132 microRNA, which may limit inflammation in the brain by silencing the expression of this protein and allowing ACh to act in an anti-inflammatory capacity.^[2] It has also been shown that the main active ingredient in cannabis, tetrahydrocannabinol, is a competitive inhibitor of acetylcholinesterase.^[3]

AChE is found in many types of conducting tissue: nerve and muscle, central and peripheral tissues, motor and sensory fibers, and cholinergic and noncholinergic fibers. The activity of AChE is higher in motor neurons than in sensory neurons.^[4] Acetylcholinesterase is also found on the red blood cell membranes, where different forms constitute the Yt blood group antigens.^[33] Acetylcholinesterase exists in multiple molecular forms, which possess similar catalytic properties, but differ in their oligomeric assembly and mode of attachment to the cell surface.

References

1. Soreq H, et al. (1991). Proc Natl Acad Sci USA. 87 (24): 9688–92.
2. Shaked I, et al. (2009). Immunity. 31 (6): 965–73.
3. Eubanks LM, et al. (2006). Mol. Pharm. 3 (6): 773–7.
4. Massoulié J, et al. (1993). Progress in Neurobiology. 41 (1): 31–91.
5. Bartels CF, et al. (1993). Am. J. Hum. Genet. 52 (5): 928–36.