Amplite® Fluorimetric Acetylcholinesterase Assay Kit *Red Fluorescence*
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Telephone | 1-800-990-8053 |
Fax | 1-800-609-2943 |
sales@aatbio.com | |
International | See distributors |
Bulk request | Inquire |
Custom size | Inquire |
Shipping | Standard overnight for United States, inquire for international |
Excitation (nm) | 571 |
Emission (nm) | 584 |
H-phrase | H303, H313, H333 |
Hazard symbol | XN |
Intended use | Research Use Only (RUO) |
R-phrase | R20, R21, R22 |
UNSPSC | 12171501 |
Amplite® Fluorimetric Acetylcholinesterase Assay Kit *Green Fluorescence* |
Overview | ![]() ![]() |
Excitation (nm) 571 | Emission (nm) 584 |
Platform
Fluorescence microplate reader
Excitation | 540 nm |
Emission | 590 nm |
Cutoff | 570 nm |
Recommended plate | Solid black |
Components
Example protocol
AT A GLANCE
Protocol summary
- Prepare AChE working solution (50 µL)
- Add AChE standards or AChE test samples (50 µL)
- Incubate at room temperature for 10 - 30 minutes
- Monitor fluorescence intensity at Ex/Em =540/590 nm (Cutoff = 570 nm)
Important notes
Thaw all the kit components at room temperature before starting the experiment.
PREPARATION OF STOCK SOLUTION
1. AmpliteTM Red stock solution (250X):
Add 40 µL of DMSO (Component G) into the vial of AmpliteTM Red (Component A) to make 250X AmpliteTM Red stock solution. Note: Avoid exposure to light.
2. Acetylcholinesterase standard solution (50 units/mL):
Add 100 µL of Assay Buffer (Component E) into the vial of Acetylcholinesterase Standard (Component D) to make 50 Units/mL Acetylcholinesterase standard solution.
3. Acetylcholine stock solution (1000X):
Add 100 µL of Assay Buffer (Component E) into the vial of Acetylcholine (Component C) to make a 1000X Acetylcholine stock solution.
PREPARATION OF STANDARD SOLUTION
For convenience, use the Serial Dilution Planner: https://www.aatbio.com/tools/serial-dilution/11402
Add 20 µL of 50 Units/mL Acetylcholinesterase standard solution to 980 µL of Dilution Buffer (Component F) to generate 1000 mU/mL Acetylcholinesterase standard solution. Take 1000 mU/mL Acetylcholinesterase standard and perform 1:10 in Dilution Buffer (Component F) to get 100 mU/mL Acetylcholinesterase standard (AS7). Then take 100 mU/mL Acetylcholinesterase standard (AS7) and perform 1:3 serial dilutions to get serially diluted Acetylcholinesterase standard (AS6 - AS1) with Dilution Buffer (Component F). Note: Diluted Acetylcholinesterase standard solution is unstable and should be used within 4 hours.
PREPARATION OF WORKING SOLUTION
1. Add 5 mL of Assay Buffer (Component E) to the bottle of Acetylcholinesterase Probe (Component B) and mix well.
2. Add 5 μL of 1000X Acetylcholine stock solution into the bottle of Acetylcholinesterase Probe mixture and mix well.
3. Add 20 μL of 250X AmpliteTM Red stock into this bottle of Acetylcholinesterase Probe mixture to make Acetylcholinesterase (AChE) working solution. Note: This Acetylcholinesterase (AChE) working solution should be used promptly and kept from light. The assay background would increase with longer storage time.
For guidelines on cell sample preparation, please visit
https://www.aatbio.com/resources/guides/cell-sample-preparation.html
SAMPLE EXPERIMENTAL PROTOCOL
Table 1. Layout of Acetylcholinesterase standards and test samples in a solid black 96-well microplate. AS= Acetylcholinesterase Standards (AS1 - AS7, 0.14 to 100 mU/mL); BL=Blank Control; TS=Test Samples.
BL | BL | TS | TS |
AS1 | AS1 | ... | ... |
AS2 | AS2 | ... | ... |
AS3 | AS3 | ||
AS4 | AS4 | ||
AS5 | AS5 | ||
AS6 | AS6 | ||
AS7 | AS7 |
Table 2. Reagent composition for each well.
Well | Volume | Reagent |
AS1 - AS7 | 50 µL | Serial Dilutions (0.14 to 100 mU/mL) |
BL | 50 µL | Dilution Buffer (Component F) |
TS | 50 µL | test sample |
- Prepare Acetylcholinesterase standards (AS), blank controls (BL), and test samples (TS) according to the layout provided in Tables 1 and 2. For a 384-well plate, use 25 µL of reagent per well instead of 50 µL. Note: Treat cells or tissue samples as desired.
- Add 50 µL of AChE working solution to each well of Acetylcholinesterase standard, blank control, and test samples to make the total Acetylcholinesterase assay volume of 100 µL/well. For a 384-well plate, add 25 µL of AChE working solution into each well instead, for a total volume of 50 µL/well.
- Incubate the reaction at room temperature for 10 to 30 minutes, protected from light.
- Monitor the fluorescence increase with a fluorescence microplate reader at Ex/Em = 540/590 nm (Cutoff = 570 nm).
Product Family
Name | Excitation (nm) | Emission (nm) |
Amplite® Fluorimetric Acetylcholinesterase Assay Kit *Green Fluorescence* | 510 | 525 |
Images
Citations
Authors: Xu, Tuan and Li, Shuaizhang and Li, Andrew J and Zhao, Jinghua and Sakamuru, Srilatha and Huang, Wenwei and Xia, Menghang and Huang, Ruili
Journal: Journal of Chemical Information and Modeling (2023)
Authors: Nishio, Takahiro and Taura, Kojiro and Iwaisako, Keiko and Koyama, Yukinori and Tanabe, Kazutaka and Yamamoto, Gen and Okuda, Yukihiro and Ikeno, Yoshinobu and Yoshino, Kenji and Kasai, Yosuke and others, undefined
Journal: Journal of Gastroenterology (2017): 1--12
Authors: Li, Shuaizhang and Huang, Ruili and Solomon, Samuel and Liu, Yitong and Zhao, Bin and Santillo, Michael F and Xia, Menghang
Journal: Biotechnology journal (2017): 1600715
Authors: Elsawy, H and Al-Omair, MA and Sedky, A and Al-Otaibi, L
Journal: Journal of Chemical Neuroanatomy (2017)
Authors: Koh, Eun-Jeong and Kim, Kui-Jin and Song, Ji-Hyeon and Choi, Jia and Lee, Hyeon Yong and Kang, Do-Hyung and Heo, Ho Jin and Lee, Boo-Yong
Journal: International Journal of Molecular Sciences (2017): 2401
Authors: Chen, Zhuo and Huang, Chao and Ding, Wenbin
Journal: Neurochemical Research (2016): 3322--3332
Authors: Kameshima, S and Yamada, K and Morita, T and Okada, M and Yamawaki, H
Journal: Acta Physiologica (2016): 203--210
Authors: Dasuri, Kalavathi and Zhang, Le and Kim, Sun OK Fern and ez , undefined and Bruce-Keller, Annadora J and Keller, Jeffrey N
Journal: Biochimica et Biophysica Acta (BBA)-Molecular Basis of Disease (2016): 274--283
Authors: Reddy, Eeda Koti and Remya, C and Sajith, Ayyiliath M and Dileep, KV and Sadasivan, C and Anwar, Shaik
Journal: RSC Advances (2016): 77431--77439
Authors: Lee, Jin-Seok and Hong, Sung-Shin and Kim, Hyeong-Geug and Lee, Hye-Won and Kim, Won-Yong and Lee, Sam-Keun and Son, Chang-Gue
Journal: PloS one (2016): e0159823
Application notes
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