PhosphoWorks™ Fluorimetric ATP Assay Kit
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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 |
H-phrase | H303, H313, H333 |
Hazard symbol | XN |
Intended use | Research Use Only (RUO) |
R-phrase | R20, R21, R22 |
UNSPSC | 12352200 |
Overview | ![]() ![]() |
Platform
Fluorescence microplate reader
Excitation | 540 mn |
Emission | 590 nm |
Cutoff | 570 nm |
Recommended plate | Solid black |
Components
Example protocol
AT A GLANCE
Protocol summary
- Prepare ATP working solution (50 µL)
- Add ATP standards or test samples (50 µL)
- Incubate at room temperature for 10 - 30 minutes
- Monitor fluorescence increase at Ex/Em = 540/590 nm (Cutoff = 570 nm)
Important notes
Thaw the kit components at room temperature before starting the experiment.
PREPARATION OF STOCK SOLUTION
1. AmpliteTM Red Substrate stock solution (200X):
Add 30 µL of DMSO (Component E) into the vial of AmpliteTM Red Substrate (Component A) to make 200X AmpliteTM Red Substrate stock solution.
2. ATP standard solution (10 mM):
Add 0.5 mL of ddH2O into the vial of ATP Standard (Component D) to make 10 mM ATP standard solution.
PREPARATION OF STANDARD SOLUTION
For convenience, use the Serial Dilution Planner: https://www.aatbio.com/tools/serial-dilution/21620
Add 10 µL of 10 mM ATP standard solution into 990 µL 1X PBS buffer to generate 100 µM ATP standard solution (AS7). Take 100 µM ATP standard solution (AS7) and perform 1:3 serial dilutions to get serially diluted ATP standards (AS6-AS1) with 1X PBS buffer.
PREPARATION OF WORKING SOLUTION
1. Add 5 mL of Assay Buffer (Component C) into Enzyme Mix bottle (Component B), and mix well.
2. Add 25 µL of 200X AmpliteTM Red Substrate stock solution to the Enzyme Mix bottle, mix well to make APT working solution.
SAMPLE EXPERIMENTAL PROTOCOL
Table 1. Layout of ATP standards and test samples in a solid black 96-well microplate. AS= ATP Standards (AS1 - AS7, 0.14 to 100 µM), 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 µM) |
BL | 50 µL | 1 X PBS Buffer |
TS | 50 µL | test sample |
- Prepare ATP 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.
- Add 50 µL of ATP working solution to each well of ATP standard, blank control, and test samples to make the total ATP assay volume of 100 µL/well. For a 384-well plate, add 25 µL of ATP working solution into each well instead, for a total volume of 50 µL/well.
- Incubate the reaction at room temperature for 10 - 30 minutes, protected from light.
- Monitor the fluorescence increase with a fluorescence plate reader at Ex/Em=540/590 nm (Cutoff = 570 nm).
Images
Citations
Authors: Nowak, E., Kammerer, S., Kupper, J. H.
Journal: Clin Hemorheol Microcirc (2018): 327-336
Authors: Tahara, H., Matsuda, S., Yamamoto, Y., Yoshizawa, H., Fujita, M., Katsuoka, Y., Kasahara, T.
Journal: J Pharmacol Toxicol Methods (2017): 92-99
Authors: Wang, Mengqiao and Xu, Miao and Long, Yan and Fargue, Sonia and Southall, Noel and Hu, Xin and McKew, John C and Danpure, Christopher J and Zheng, Wei
Journal: Scientific Reports (2016)
Authors: Zhang, Lu and Han, Jianjun and Jackson, Am and a L , undefined and Clark, Leslie N and Kilgore, Joshua and Guo, Hui and Livingston, Nick and Batchelor, Kenneth and Yin, Yajie and Gilliam, Timothy P and others, undefined
Journal: Journal of Hematology & Oncology (2016): 91
Authors: Chen, Ya-Hui and Chen, Yi-Chun and Liu, Chin-San and Hsieh, Ming-Chia
Journal: Journal of Diabetes Research (2016)
Authors: Yuan, Lingqin and Sheng, Xiugui and Willson, Adam K and Roque, Dario R and Stine, Jessica E and Guo, Hui and Jones, Hannah M and Zhou, Chunxiao and Bae-Jump, Victoria L
Journal: Endocrine-related cancer (2015): 577--591
Authors: Jiang, Ying and Xia, Wei and Yang, Jie and Zhu, Yingshuang and Chang, Huailong and Liu, Juan and Huo, Wenqian and Xu, Bing and Chen, Xi and Li, Yuanyuan and others, undefined
Journal: Toxicology (2015): 21--31
Authors: Gonneaud, Alexis and Turgeon, Naomie and Boisvert, Frančois-Michel and Boudreau, Frančois and Asselin, Claude
Journal: FEBS letters (2015): 2776--2783
Authors: Qiu, Haifeng and Jackson, Am and a L , undefined and Kilgore, Joshua E and Zhong, Yan and Chan, Leo Li-Ying and Gehrig, Paola A and Zhou, Chunxiao and Bae-Jump, Victoria L
Journal: Oncotarget (2015): 6915
Authors: Pohl, Kerstin and Hayes, Elaine and Keenan, Joanne and Henry, Michael and Meleady, Paula and Molloy, Kevin and Jundi, Bakr and Bergin, David A and McCarthy, Cormac and McElvaney, Oliver J and others, undefined
Journal: Blood (2014): 999--1009
References
Authors: Hanshaw RG, Lakshmi C, Lambert TN, Johnson JR, Smith BD.
Journal: Chembiochem (2005): 2214
Authors: Koulov AV, Stucker KA, Lakshmi C, Robinson JP, Smith BD.
Journal: Cell Death Differ (2003): 1357
Authors: Saurel O, Cezanne L, Milon A, Tocanne JF, Demange P.
Journal: Biochemistry (1998): 1403
Authors: Hall MP, Burson KK, Huestis WH.
Journal: Biochim Biophys Acta (1998): 101
Authors: Hanada K, Pagano RE.
Journal: J Cell Biol (1995): 793
Authors: Mancheno JM, Gasset M, Lacadena J, Ramon F, Martinez del Pozo A, Onaderra M, Gavilanes JG.
Journal: Biophys J (1994): 1117
Authors: Arnhold J, Wiegel D, Hussler O, Arnold K.
Journal: Biochim Biophys Acta (1994): 375
Authors: Walter A, Siegel DP.
Journal: Biochemistry (1993): 3271
Authors: Kobayashi T, Arakawa Y.
Journal: J Cell Biol (1991): 235
Authors: Wrzosek A, Famulski KS, Lehotsky J, Pikula S.
Journal: Biochim Biophys Acta (1989): 263
Application notes
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