AAT Bioquest

Amplite® Fluorimetric Coenzyme A Quantitation Kit *Green Fluorescence*

CoA dose response was measured in a 96-well solid black plate with Amplite® Fluorimetric Coenzyme A Quantitation Assay Kit using a NOVOstar microplate reader (BMG Labtech).
CoA dose response was measured in a 96-well solid black plate with Amplite® Fluorimetric Coenzyme A Quantitation Assay Kit using a NOVOstar microplate reader (BMG Labtech).
CoA dose response was measured in a 96-well solid black plate with Amplite® Fluorimetric Coenzyme A Quantitation Assay Kit using a NOVOstar microplate reader (BMG Labtech).
Ordering information
Catalog Number
Unit Size
Add to cart
Additional ordering information
InternationalSee distributors
Bulk requestInquire
Custom sizeInquire
ShippingStandard overnight for United States, inquire for international
Request quotation
Spectral properties
Excitation (nm)510
Emission (nm)525
Storage, safety and handling
H-phraseH303, H313, H333
Hazard symbolXN
Intended useResearch Use Only (RUO)
R-phraseR20, R21, R22


Excitation (nm)
Emission (nm)
Coenzyme A (CoA) is a universal and essential cofactor in all forms of cellular life acting as a principal acyl carrier in numerous biosynthetic, energy-yielding, and degradative pathways. It plays important roles in the synthesis and oxidation of fatty acids, pyruvate oxidation and the citric acid cycle. Measurement of CoA is one of the essential tasks for investigating biological processes and events in many biological systems. There are a few reagents or assay kits available for quantitating CoA content in biological systems. However, the existing commercial kits either lack sensitivity or have tedious procedures. Our Amplite® Fluorimetric CoA Qutitation Assay Kit provides an ultrasensitive fluorimetric assay to quantitate CoA content by detection of -SH group in CoA. Our proprietary fluorogenic CoA Green™ dye used in the kit becomes strongly fluorescent upon reacting with -SH. The assay kit can detect as little as 4 picomole of CoA in a 100 µL assay volume (40 nM). It can be performed in a convenient 96-well or 384-well microtiter-plate format and easily adapted to automation without a separation step.


Fluorescence microplate reader

Excitation490 nm
Emission520 nm
Recommended plateSolid black


Example protocol


Protocol summary

  1. Prepare CoA working solution (50 µL)
  2. Add CoA standards or test samples (50 µL)
  3. Incubate at RT for 10 minutes - 1 hour
  4. Monitor the fluorescence increase at Ex/Em = 490/520 nm

Important notes
Thaw all the kit components at room temperature before starting the experiment.


Unless otherwise noted, all unused stock solutions should be divided into single-use aliquots and stored at -20 °C after preparation. Avoid repeated freeze-thaw cycles.

1. CoA standard solution (1 mM):
Add 200 µL of ddH2O into the CoA standard vial (Component C) to make 1 mM (1 nmol/µL) stock solution. Note: It is highly recommended to use the ddH2O that has been sparged with nitrogen to remove oxygen for preparing coenzyme A standard solution. The aqueous solution is not stable and will degrade rapidly. It should be stored at 2 - 8 °C and used within the day.

2. CoA Green™ stock solution (100X):
Add 100 µL of DMSO (Component D) into the vial of CoA Green™ (Component A) to make 100X stock solution.


CoA standard

For convenience, use the Serial Dilution Planner: https://www.aatbio.com/tools/serial-dilution/15270

Add 30 µL of CoA standard solution to 970 µL of Assay Buffer (Component B) to generate 30 µM (30 pmol/µL) CoA standard. Take the 30 µM CoA standard solution to perform 1:3 serial dilutions with Assay buffer (Component B) to get serial dilutions of CoA standard (CoA1 - CoA7). Note: Diluted CoA standard solution is unstable, and should be used within 4 hours. 


Add 50 μL of CoA Green™ stock solution (100X) into 5 mL of Assay Buffer (Component B) and mix well.


Table 1. Layout of CoA standards and test samples in a solid black 96-well microplate. CoA = Coenzyme A standard (CoA1 - CoA7, 0.01 to 10 µM); BL = blank control; TS = test sample.


Table 2. Reagent composition for each well.

CoA1-CoA750 µLserial dilution (0.01 to 10 µM)
BL50 µLAssay Buffer (Component B)
TS50 µLsample
  1. Prepare coenzyme A standards (CoA), blank controls (BL), and test samples (TS) according ot the layout provided in Tables 1 and 2. For a 384-well plate, use 25 µL of reagent per well instead of 50 µL.

  2. Add 50 µL of CoA working solution to each well of the CoA standard, blank control, and test sample to make the total CoA assay volume of 100 µL/well. For a 384-well plate, add 25 µL of CoA working solution into each well instead, for a total volume of 50 µL/well.

  3. Incubate the reaction at room temperature for 10 minutes to 1 hour, protected from light.

  4. Monitor the fluorescence increase at Ex/Em = 490/520 nm with a fluorescence plate reader.


Open in Advanced Spectrum Viewer

Spectral properties

Excitation (nm)510
Emission (nm)525



View all 2 citations: Citation Explorer
Structural basis of the acyl-transfer mechanism of human GPAT1
Authors: Johnson, Zachary Lee and Ammirati, Mark and Wasilko, David Jonathan and Chang, Jeanne S and Noell, Stephen and Foley, Timothy L and Yoon, Hyejin and Smith, Kathleen and Asano, Shoh and Hales, Katherine and others,
Journal: Nature Structural \& Molecular Biology (2022): 1--9
Methods for measuring CoA and CoA derivatives in biological samples
Authors: Tsuchiya, Yugo and Pham, Uyen and Gout, Ivan
Journal: Biochemical Society Transactions (2014): 1107--1111


View all 87 references: Citation Explorer
A spectrophotometric assay for measuring acetyl-coenzyme A carboxylase
Authors: Kroeger JK, Zarzycki J, Fuchs G.
Journal: Anal Biochem (2011): 100
A continuous assay for alpha-methylacyl-coenzyme A racemase using circular dichroism
Authors: Ouazia D, Bearne SL.
Journal: Anal Biochem (2010): 45
An optimized luciferase bioluminescent assay for coenzyme A
Authors: Marques SM, da Silva JC.
Journal: Anal Bioanal Chem (2008): 2161
A simple homogeneous scintillation proximity assay for acyl-coenzyme A:diacylglycerol acyltransferase
Authors: Seethala R, Peterson T, Dong J, Chu CH, Chen L, Golla R, Ma Z, Panemangalore R, Lawrence RM, Cheng D.
Journal: Anal Biochem (2008): 144
A fluorescence-based thiol quantification assay for ultra-high-throughput screening for inhibitors of coenzyme A production
Authors: Chung CC, Ohwaki K, Schneeweis JE, Stec E, Varnerin JP, Goudreau PN, Chang A, Cassaday J, Yang L, Yamakawa T, Kornienko O, Hodder P, Inglese J, Ferrer M, Strulovici B, Kusunoki J, Tota MR, Takagi T.
Journal: Assay Drug Dev Technol (2008): 361
CE assay of methylmalonyl-coenzyme-a mutase activity
Authors: Carlucci F, Rosi F, Tommassini V, Tabucchi A.
Journal: Electrophoresis (2007): 1921
Dephospho-CoA kinase provides a rapid and sensitive radiochemical assay for coenzyme A and its thioesters
Authors: Wadler C, Cronan JE.
Journal: Anal Biochem (2007): 17
Development of a semiquantitative degenerate real-time pcr-based assay for estimation of numbers of butyryl-coenzyme A (CoA) CoA transferase genes in complex bacterial samples
Authors: Louis P, Flint HJ.
Journal: Appl Environ Microbiol (2007): 2009
Discovery of acetyl-coenzyme A carboxylase 2 inhibitors: comparison of a fluorescence intensity-based phosphate assay and a fluorescence polarization-based ADP Assay for high-throughput screening
Authors: Liu Y, Zalameda L, Kim KW, Wang M, McCarter JD.
Journal: Assay Drug Dev Technol (2007): 225
Assay of the activity of malonyl-coenzyme A decarboxylase by gas chromatography-mass spectrometry
Authors: Wang X, Stanley WC, Darrow CJ, Brunengraber H, Kasumov T.
Journal: Anal Biochem (2007): 169