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Acetoacetate Colorimetric Assay Kit

based on 2 citations in multiple journalsAcetoacetate Colorimetric Assay Kit24.1 4
Only kit on the market to measure acetoacetate
Catalog #: K650

Product Details

Cat # +Size K650-100
Size 100 assays
Detection Method Absorbance (550 nm)
Applications Measurement of acetoacetate in various biological samples.
Analysis of acetoacetate in pathological conditions such as Type I diabetes, alcoholism, etc.
Features & Benefits • Specific for acetoacetate and does not detect 3-β-hydroxybutyrate
• The assay can detect less than 25 µM of acetoacetate in various sample types.
Kit Components • Acetoacetate Assay Buffer
• Acetoacetate Standard (Lyophilized)
• Acetoacetate Substrate
Storage Conditions -20°C
Shipping Conditions Gel Pack
USAGE For Research Use Only! Not For Use in Humans.


Acetoacetate (AcAc), a β-ketoacid, is one of the three ketone bodies and is formed via condensation of two molecules of acetyl-CoA in liver mitochondria. AcAc can be enzymatically reduced to 3-β-hydroxybutyrate (β-HB), or decarboxylated producing acetone (CH3)2CO). Ketone bodies (βHB: 78%; AcAc: 20% & CH3)2CO: 2%) are mainly used as an alternative energy source when glucose cannot be delivered to the system. Excessive concentration of ketone bodies (ketoacidosis) is observed in patients with Type I diabetes, severe starvation or alcoholism. Traditionally, AcAc levels have been qualitatively detected using dipsticks that use sodium nitroferricyanide as a chromophore. BioVision’s Acetoacetate Assay Kit has adapted that principle with a modification that provides a sensitive method to quantitate endogenous levels of AcAc in human blood, and urine. In this non-enzymatic assay, AcAc reacts with a substrate to generate a colored product that can be measured at 550 nm. The reaction is specific for AcAc and does not detect 3-β-hydroxybutyrate. The assay kit can detect samples containing acetoacetate as low as 25 µM.

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Can you share the protocol for preparing cell or tissue samples?
We have not tested cells or tissue samples with K650. However, here is a general protocol: Homogenize 1 X 10^6 cells or 10 mg tissue in 100 µL of Assay Buffer using a dounce homogenizer on ice. Once lysis is completed, centrifuge the lysate at 10,000 g for 5 min at 4 °C and use the supernatant for the assay.
Is it ok to use blood collected with EDTA for the assay?
It is ok to use blood collected with EDTA.
Can EDTA be used for blood collection for this assay?
Yes, it is ok to use EDTA for blood collection for this assay.
Does this kit also detect acetoacetyl-CoA?
Unless the acetoacetyl CoA in solution forms acetoacetic acid or acetoacetate, it will not be detected by this assay.
What are a few key protocol/reagent-related tips that are important to get good results?
1. The substrate is light sensitive. Exposure to light can result in color change irrespective of amount of acetoacetate.
2. The standard is unstable if not kept on ice at all times during use. This is also valid for the samples.
3. Samples should be deproteinized to remove all proteins/enzymes to ensure acetoacetate is not broken down quickly.
4. Endogenous interference may result in color for all samples. Spiking the samples with known amounts of the standard might help.
How should samples be handled if acetoacetate is very unstable?
Samples should be tested while fresh and kept on ice at all times.
What is the exact volume of sample required for this assay?
There is no specific volume we can recommend for the amount any sample to be used since it is completely sample concentration and quality based. It is recommended to do a pilot expt with multiple sample volumes to determine the optimal volume which gives a reading within the linear range of the standard curve. Please refer to the citations for this product to see what other clients have used with similar sample types.
Are trial sizes of this kit available?
Unfortunately, we do not have trial sizes of this kit available. However,for customers based in the US or Canada, a 10% off list price introductory discount can be offered on the list price. For all other customers based out of this area our regional BioVision distributor should be contacted.
Can individual components of this kit be purchased separately?
Yes, any of the kit's components can be purchased separately without having to buy the whole kit. Please refer to the component Cat #s mentioned on the datasheet for ordering.
Desiree Abdurrachim . A new hyperpolarized 13C ketone body probe reveals an increase in acetoacetate utilization in the diabetic rat heart , Sci Rep . Apr 19. 30940842
Manuela Verri, Plasma energy substrates at two stages of Alzheimer’s disease in humans. Int J Immunopathol Pharmacol, Dec 2018; PMC6309029.
Garg, Manisha et al. (2017) Liraglutide acutely suppresses glucagon, lipolysis and ketogenesis in type 1 diabetes, Diabetes Obes Metab. 2017 Mar 17.
Goldgof et al., The Chemical Uncoupler 2,4-Dinitrophenol (DNP) Protects against Diet-induced Obesity and Improves Energy Homeostasis in Mice at Thermoneutrality. J. Biol. Chem., Jul 2014; 289: 19341 - 19350.
For more citations of this product click here